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Journal of Jilin University(Medicine Edition)
Bimonthly
ISSN 1671-587X
CN 22-1342/R
Director: LI Xinxin
Editor:JIANG Jinqiu 
    HAN Hongzhi
    GUAN Xin
    CHEN Sihan 
    LI Xinwei
Phone:0431-85619279
E-mail:xuebao@jlu.edu.cn
Address:No. 828 Xinmin Street, Changchun, Jilin, China
Postcode:130021
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Table of Content
28 September 2024, Volume 50 Issue 5
Research in basic medicine
Effect of cold exposure on nociception and transient receptor potential ion channels in sensory neurons in rats
Ding JIANG,Yuelong CAO,Qinguang XU,Anping SHEN,Nan WANG,Fengxi QIU,Yan XUE
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1189-1195.  DOI: 10.13481/j.1671-587X.20240501
Abstract ( 59 )   HTML ( 4 )   PDF (802KB) ( 30 )  

Objective To discuss the effect of cold exposure on nociception in the rats and its regulatory mechanism on transient receptor potential (TRP) ion channels in the sensory neurons, and to provide the basis for clarifying the biological mechanism of cold-sensitive pain. Methods Sixteen female SD rats were divided into control group (n=8) and cold group (n=8). The rats in control group were exposed to the environment of (24±2)℃, and the rats in cold group were exposed to low temperature (4 ℃±1 ℃) in an artificial intelligence climate chamber for 4 h daily, for one week. Von Frey filaments were used to detect the mechanical withdrawal threshold (MWT) of the rats in two groups; immunofluorescence staining was used to observe the expression levels of TRPA1, TRPM8, TRPV1, and TRPV4 in dorsal root ganglion (DRG) tissue of the rats in two groups, the expression levels of calcitonin gene-related peptide (CGRP) and substance P (SP) in DRG tissue of the rats in two groups, and the expression levels of TRPA1, TRPM8, TRPV1, and TRPV4 in synovial tissue of the rats in two groups. Results Compared with control group, the MWT of the rats in cold group was significantly decreased (P< 0.05), the expression levels of TRPA1 and TRPM8 in DRG tissue were significantly increased (P< 0.05), the expression level of TRPV1 was significantly decreased (P<0.05), there was no significant difference in the expression level of TRPV4 (P>0.05), and the expression levels of CGRP and SP were significantly increased (P<0.05). Compared with control group, the expression level of TRPA1 in synovial tissue of the rats in cold group was significantly increased (P<0.05), while the expression levels of TRPM8, TRPV1, and TRPV4 were significantly decreased (P<0.05). Conclusion Short-term cold exposure can induce the hyperalgesia of the rats, and its mechanism may be associated with the changes in the expression of TRP ion channels in DRG and synovial tissues. TRPA1 sensory neurons play an important role in local joint cold pain.

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Alleviative effect of ginsenoside Rg1 on brain injury induced by chronic intermittent hypoxia in mice and its mechanism
Yan MENG,Hongxin WANG,Yuhong YANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1196-1204.  DOI: 10.13481/j.1671-587X.20240502
Abstract ( 50 )   HTML ( 7 )   PDF (1487KB) ( 22 )  

Objective To discuss the alleviative effect of ginsenoside Rg1 on chronic intermittent hypoxia (CIH)-induced brain injury in the mice,and to clarify its possible mechanism. Methods Forty male C57BL/6 mice were randomly divided into control group, model group, inhibitor group (treated with calpain-1 inhibitor), low dose of ginsenoside Rg1 group (treated with 10 mg·kg-1 ginsenoside Rg1), and high dose of ginsenoside Rg1 group (treated with 20 mg·kg-1 ginsenoside Rg1). Except for the control group, the mice in all other groups were placed in a hypoxic chamber with automatically regulated oxygen concentration to induce hypoxic brain injury.The peripheral blood oxygen saturation (SpO2) of tail of the mice in various groups was detected; the escape latencies and path lengths and the frequency of swimming route crossing the target quadrant of the mice in various groups were determined by Morris water maze test; the levels of blood urea nitrogen (BUN), lactate (LA), malondialdehyde (MDA), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) and the activities of superoxide dismutase (SOD) and lactate dehydrogenase (LDH) in serum of the mice in various groups were detected by kits; the degrees of brain tissue injury of the mice in various groups were observed by HE staining. The levels of reactive oxygen species (ROS) in CA1 region of hippocampus tissue of the mice in various groups were detected by dihydroethidium(DHE) probe; the expression levels of calpain-1, IL-6, and TNF-α proteins in brain tissue of the mice in various groups were detected by Western blotting method. Results Compared with control group, the SpO2 of the mice in model group was significantly decreased (P<0.01), indicating that the model was successfully established. Compared with model group, the SpO2 of the mice in inhibitor group, low dose of ginsenoside Rg1 group and high dose of ginsenoside Rg1 group were significantly increased (P<0.01). The Morris water maze test results showed that compared with control group, the escape latency and path length of the mice in model group were significantly prolonged (P<0.01), and the frequency of swimming route of crossing the target quadrant was significantly decreased; compared with model group, the escape latencies and path lengths of the mice in inhibitor group, low dose of ginsenoside Rg1 group and high dose of ginsenoside Rg1 group were significantly shortened(P<0.01), and the frequency of swimming route of crossing the target quadrant was significantly increased. Compared with control group, the levels of BUN, LA, MDA, IL-6, and TNF-α in serum of the mice in model group were significantly increased (P<0.01), while the activity of LDH was significantly increased (P<0.01), and the activity of SOD was significantly decreased (P<0.01); compared with model group, the levels of BUN, LA, MDA, IL-6, and TNF-α in serum of the mice in inhibitor group, low dose of ginsenoside Rg1 group and high dose of ginsenoside Rg1 group were significantly decreased (P<0.01), while the activities of LDH were significantly decreased(P<0.01), and the activities of SOD were significantly increased (P<0.01). The HE staining results showed that compared with control group, the pyramidal neurons in CA1 region of hippocampus tissue of the mice in model group were loosely arranged, while some neurons were triangular, with nuclear pyknosis, cytoplasmic hyperchromasia, and a few neurons were lost, indicating obvious hypoxic neuronal injury; compared with model group, the hypoxic neuronal injury in CA1 region in hippocampus tissue of the mice in inhibitor group, low dose of ginsenoside Rg1 group and high dose of ginsenoside Rg1 group was effectively alleviated. The DHE probe detection showed that compared with control group, the level of ROS in CA1 region in hippocampus tissue of the mice in model group was significantly increased (P<0.01); compared with model group, the levels of ROS in CA1 region in hippocampus tissue of the mice in inhibitor group, low dose of ginsenoside Rg1 group and high dose of ginsenoside Rg1 group were significantly decreased (P<0.01). The Western blotting results showed that compared with control group, the expression levels of calpain-1, TNF-α, and IL-6 proteins in hippocampus tissue of the mice in model group were significantly increased (P<0.01); compared with model group, the expression levels of calpain-1, TNF-α, and IL-6 proteins in hippocampus tissue of the mice in inhibitor group, low dose of GRg1 group and high dose of GRg1 group were significantly decreased (P<0.01). Conclusion Ginsenoside Rg1 can alleviate brain tissue injury of the mice induced by CIH; its mechanism may be related to the inhibition of brain tissue inflammatory response and oxidative stress, and the downregulation of calpain-1 expression.

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Effect of Aspergillus fumigatus on DNA damage and IL-33 expression in human bronchial epithelial cells and its mechanism
Qiao WANG,Ziling ZENG,Xing WANG,Ning MA,Zhibin WANG,Guofeng XU,Xiefang YUAN,Xiaoyun WANG,Yuejiao LI,Hongmei TANG,Yun ZHANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1205-1216.  DOI: 10.13481/j.1671-587X.20240503
Abstract ( 41 )   HTML ( 0 )   PDF (2879KB) ( 21 )  

Objective To discuss the effect of Aspergillus fumigatusAf) on DNA damage and interleukin (IL)-33 expression in the human bronchial epithelial cells, and to clarify its related mechanism. Methods Different concentrations (1, 5, and 10 mg·L?1) of Af were used to stimulate the bronchial epithelial BEAS-2B cells to select the appropriate stimulation concentration. When the BEAS-2B cells were treated with N-acetylcysteine (NAC) and Af, the cells were divided into control group, Af group, NAC group, and Af+NAC group. When the BEAS-2B cells were treated with DNA double-strand break repair inhibitor NU7441 and Af, the cells were divided into control group, Af group, NU7441 group, and Af+NU7441 group. The comet assay was used to detect the percentages of comet tail DNA of cells in various groups; immunofluorescence method was used to detect the expression levels of DNA damage-related protein phosphorylated H2AX(γH2AX) in the cells in various groups; 2,7-dichlorofluorescein diacetate (DCFH-DA) fluorescence probe was used to detect the levels of reactive oxygen species (ROS) in the cells in various groups; real-time fluorescence quantitative PCR (RT-qPCR) method was used to detect the expression levels of interleukih-33 (IL-33), thymic stromal lymphopoietin (TSLP), and interleukih-25 (IL-25) mRNA in the cells in various groups; Western blotting method was used to detect the expression levels of phosphorylated nuclear factor κB (p-NF-κB), phosphorylated ataxia telangiectasia mutated (p-ATM), and γH2AX proteins in the cells in various groups. Results Compared with control group, the percentage of comet tail DNA and the expression level of γH2AX in the cells in 1 mg·L?1 Af group showed no significant difference (P>0.05), while the percentage of comet tail DNA and the expression level of γH2AX in the cells in 5 mg·L?1 Af group were significantly increased (P<0.01); compared with 5 mg·L?1 Af group, the percentage of comet tail DNA and the expression level of γH2AX in the cells in 10 mg·L?1 Af group were significantly increased (P<0.01). Compared with control group, the ROS levels in the bronchial epithelial cells in 1 mg·L?1 Af group was significantly increased (P<0.05); compared with 1 mg·L?1 Af group, the ROS level in the cells in 5 mg·L?1 Af group was significantly increased (P<0.01); compared with 5 mg·L?1 Af group, the ROS level in the cells in 10 mg·L?1 Af group was significantly increased (P<0.05). After treatment of NAC, compared with Af group, the percentage of comet tail DNA (P<0.01), the expression level of γH2AX (P<0.05), and the ROS level (P<0.01) in the cells in Af+NAC group were significantly decreased; after treatment of NU7441, compared with Af group, the percentage of comet tail DNA and the expression level of γH2AX in the cells in Af+NU7441 group were significantly increased (P<0.01). The RT-qPCR results showed that after treatment of NAC, compared with control group, the expression level of IL-33 mRNA in the cells in Af group was significantly increased (P<0.05); compared with Af group, the expression level of IL-33 mRNA in the cells in Af+NAC group was significantly decreased (P<0.05); after treatment of NU7441, compared with Af group, the expression level of IL-33 mRNA in the cells in Af+NU7441 group was significantly increased (P<0.05). The Western blotting results showed that after treatment of NAC, compared with control group, the expression levels of p-NF-κB, p-ATM, and γH2AX proteins in the cells in Af group were significantly increased (P<0.05); after treatment of NU7441, compared with Af group, the expression levels of p-NF-κB, p-ATM, and γH2AX proteins in the cells in Af+NAC group were significantly decreased (P<0.05); After treat ment of NU7441, compared with Af group, the expression levels of p-NF-κB, p-ATM, and γH2AX proteins in the cells in Af+NU7441 group were significantly increased (P<0.05). Conclusion Af promotes the IL-33 expression in the human bronchial epithelial cells by causing DNA damage, and its mechanism may be related to the activation of ATM/NF-κB signaling pathway.

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Effect of azathioprine on ferroptosis in spermatocytes of mice induced by RSL3
Yanjue YE,Ziyi TANG,Yupei TAN,Shiying YANG,Yong LIU,Li YIN
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1217-1226.  DOI: 10.13481/j.1671-587X.20240504
Abstract ( 41 )   HTML ( 0 )   PDF (748KB) ( 12 )  

Objective To discuss the effect of azathioprine(AZA) on ferroptosis in spermatocytes of the mice induced by reduced glutathione(GSH) peroxidase 4 inhibitor RSL3,and to clarify the potential mechanism. Methods The spermatogonia GC-2 cells of the mice were randomly divided into control group (no treatment), RSL3 group (treated with 10 nmol·L-1 RSL3 for 24 h), RSL3+ferroptosis inhibitor (Ferrostatin-1, Fer-1) group (treated with 10 nmol·L-1 RSL3 for 24 h+2 μmol·L-1 Fer-1 for 12 h), RSL3+low dose of AZA group (treated with 10 nmol·L-1 RSL3 for 24 h +5 μmol·L-1 AZA for 12 h), RSL3+medium dose of AZA group (treated with 10 nmol·L-1 RSL3 for 24 h+10 μmol·L?1 AZA for 12 h), and RSL3+high dose of AZA group (treated with 10 nmol·L-1 RSL3 for 24 h+20 μmol·L-1 AZA for 12 h). The MTT method was used to detect the activities of the GC-2 cells in various groups after treated with different concentrations of AZA and RSL3; the GSH and GSSG levels in the GC-2 cells were detected by GSH and oxidized glutathione(GSSG) detection kits; the malondialdehyde(MDA) levels in the GC-2 cells in various groups were detected by MDA detection kit; Western blotting method was used to detect the expression levels of long-chain acyl-CoA synthetase 4 (ACSL4), heme oxygenase-1(HO-1), and glutathione peroxidase 4(GPX4) proteins in the cells in various groups; immunofluorescence staining was used to detect the expressions of ACSL4 protein in the cells in various groups. Results Compared with control group, the differences in activities of the GC-2 cells in 5, 10, and 20 μmol·L?1 AZA groups had no significant differences(P>0.05), while the activities of the GC-2 cells in 30 and 40 μmol·L?1 AZA groups were significantly decreased(P<0.01); therefore, the AZA concentration was selected to be within 20 μmol·L?1. Compared with control group, the differences of the activities of the GC-2 cells in 1, 5, and 10 nmol·L?1 RSL3 groups had no significant differences(P>0.05), while the activities of the GC-2 cells in 50, 100, 500, and 1 000 nmol·L?1 RSL3 groups were significantly decreased(P<0.05 or P<0.01); therefore, the RSL3 concentration was set to be within 10 nmol·L?1. The GSH and MDA detection kits results showed that compared with control group, the levels of GSSG and MDA in the GC-2 cells in RSL3 group were significantly increased(P<0.05), while the GSH levels were significantly decreased(P<0.05); compared with RSL3 group, the levels of GSSG and MDA in the GC-2 cells in RSL3+Fer-1 group and RSL3+AZA group were significantly decreased (P<0.01), while the GSH levels were significantly increased(P<0.01). The Western blotting results showed that compared with control group, the expression levels of ACSL4 and HO-1 proteins in the GC-2 cells in RSL3 group were significantly increased(P<0.05), while the expression level of GPX4 protein was significantly decreased(P<0.01); compared with RSL3 group, the expression levels of GPX4 protein in the GC-2 cells in RSL3+Fer-1 group and RSL3+AZA group were significantly increased(P<0.05), while the expression levels of ACSL4 and HO-1 proteins were significantly decreased(P<0.01). The immunofluorescence staining results showed that compared with control group, the expression amount of ACSL4 protein in the GC-2 cells in RSL3 group was significantly increased, and compared with RSL3 group, the expression amounts of ACSL4 protein in the cells in RSL3+Fer-1 group and RSL3+AZA group were significantly decreased. Conclusion AZA can alleviate the ferroptosis-induced by RSL3 in spermatocytes of the mice.

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Alleviative effect of fluid resuscitation on damage of structure injury of vascular cells after blast injury complicated with hemorrhagic shock in rats by inhibiting ferroptosis of vascular tissue
Xiaoyong PENG,Yu ZHU,Shuangbo ZHANG,Yingguo ZHU,Tao LI,Liangming LIU,Jianmin WANG,Guangming YANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1227-1234.  DOI: 10.13481/j.1671-587X.20240505
Abstract ( 34 )   HTML ( 1 )   PDF (1063KB) ( 7 )  

Objective To discuss the effect of fluid resuscitation on the occurrence of ferroptosis in vascular tissue and the structure of vascular cells in the rats with blast injury complicated with hemorrhagic shock, and to clarify its mechanism. Methods A total of 54 healthy adult SD rats were randomly divided into normal group, blast injury complicated with hemorrhagic shock (model) group, and the fluid resuscitation (treatment) group, and there were 18 rats in each group. Among them,10 rats were randomly selected to observe the surival status and another 8 rats were selected to detect the other indexes. The average survival time(ST), 24 h and 72 h survival rates of the rats in various groups were observed; the blood pressure (BP), heart rate (HR), and respiratory rate (RR) of the rats in various groups were observed; the levels of serum creatinine (Scr), blood urea nitrogen (BUN), lactate (LAC), glucose (GLU), iron ions, glutathione (GSH), and malondialdehyde (MDA) and the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) in serum of the rats in various groups were detected;Western blotting method was used to detect the expression levels of ferroptosis marker proteins glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), and heme oxygenase 1 (HO-1) proteins in superior mesenteric artery tissue of the rats in various groups; the pathomorphology of the superior mesenteric artery of the rats in various groups was observed. Results All the rats in normal group survived for 72 h, while the longest ST of the rats in model group did not exceed 9 h. Compared with model group, the ST and 24 h survival rate (SR) of the rats in treatment group were significantly increased (P<0.05). Compared with normal group, the BP, HR, and RR of the rats in model group were significantly decreased (P<0.01). Compared with model group, the BP, HR, and RR of the rats in treatment group were significantly increased after fluid resuscitation (P<0.05). Compared with normal group, the activities of AST and ALT, and the levels of Scr and BUN in serum of the rats in model group were significantly increased (P<0.01). Compared with model group, the serum levels of LAC and GLU of the rats in treatment group were significantly decreased (P<0.01). Compared with normal group, the concentration of iron ion, GSH level, MDA level, LDH activity in serum of the rats in model group were significantly increased (P<0.05); compared with model group, the concentration of iron ion and LDH activity in serum of the rats in treatment group was significantly decreased (P<0.01). Compared with normal group, the expression levels of GPX4 and SLC7A11 in superior mesenteric artery tissue of the rats in model group were significantly decreased (P<0.05);compared with model group, the expression levels of GPX4 and SLC7A11 in superior mesenteric artery tissue of the rats in treatment group were significantly increased (P<0.05). Compared with normal group, the expression level of HO-1 protein in superior mesenteric artery tissue of the rats in model group was increased (P<0.01); compared with model group, the expression level of HO-1 protein in superior mesenteric artery tissue of the rats in treatment group was increased (P<0.01). The microscopic pathology results showed that the cell arrangement in the layers of the superior mesenteric artery tissue of the rats in model group was disordered, the swelling was significant and the thickness was increased; the pathological changes in superior mesenteric artery tissue of the rats in treatment group was alleviated. The ultramicroscopic pathology results showed that the endothelial cell structure of blood vessels of the rats in normal group was intact, and there was no swelling in the subendothelial matrix; the vascular endothelial cell membrane of the rats in model group was damaged, there were cytoplasmic dissolution and fragmentation, and the swelling of the subendothelial matrix was significant; the swelling of the vascular endothelial cells in treatment group was alleviated. Conclusion Ferroptosis occurs in vascular tissue of the rats with blast injury complicated with hemorrhagic shock, and fluid resuscitation can alleviate the structural damage of the vascular cells by inhibiting the vascular tissue ferroptosis.

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Effect of polygonatum odoratum polysaccharide on acute kidney injury in mice induced by cisplatin and its ferroptosis mechanism
Fangyang JIANG,Jing XIAO,He CHANG,Mingyang SUN,Wenjing ZHANG,Guangfu LYU,He LIN,Zhe LIN,Xiaowei HUANG,Yuchen WANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1235-1242.  DOI: 10.13481/j.1671-587X.20240506
Abstract ( 53 )   HTML ( 2 )   PDF (1389KB) ( 14 )  

Objective To discuss the protective effect of polygonatum odoratum polysaccharide (POP) on the mice with cisplatin-induced acute kidney injury(AKI), and to clarify its possible mechanism. Methods Forty male C57BL/6 mice were randomly divided into control group, model group, POP group,and ferroptosis inducer Erastin combined with POP(Erastin+POP) group,and there were 10 mice in each group.The mice in POP group and Erastin+POP group were given intragastric administration of POP (400 mg·kg-1), and on the 7th day,the mice in model group, POP group, and Erastin+POP group were intraperitoneally injected with cisplatin (20 mg·kg-1) to establish the AKI models,the mice in control group were injected with the same volume of normal saline, and the mice in Erastin+POP group were intraperitoneally injected with Erastin (40 mg·kg-1) one day in advance (on the 6th day of the experiment). After 9 d, the mice were killed and the serum and kidney tissue were collected, and the levels of serum creatinine (Scr) and blood urea nitrogen (BUN) and the levels of malondialdehyde (MDA) and glutathione (GSH) in kidney tissue of the mice in various groups were detected by kit; HE staining was used to observe the pathomorphology of kidney tissue of the mice in various groups; the expression levels of ferroptosis suppressor protein 1 (FSP1), ferritin heavy chain 1 (FTH1), and glutathione peroxidase 4 (GPX4) proteins in kidney tissue of the mice in various groups were detected by immunohistochemistry; Western blotting method was used to detect the expression levels of nuclear factor-E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) proteins in kidney tissue of the mice in various groups. Results Compared with control group, the levels of Scr and BUN of the mice in model group were significantly increased (P<0.01), the level of MDA in kidney tissue was significantly increased (P<0.01), and the level of GSH was significantly decreased (P<0.01); most kidney tubules were dilated, the epithelial cells were swollen,the vacuolar degeneration and epithelial cells fell off, and the protein-like tubules could be seen in the lumen; the expression levels of FSP1, FTH1, GPX4, Nrf2, and HO-1 proteins in kidney tissue were decreased significantly (P<0.05 or P<0.01). Compared with model group, the levels of Scr and BUN of the mice in POP group were significantly decreased (P<0.01), the level of MDA in kidney tissue was significantly decreased (P<0.01), and the level of GSH was significantly increased (P<0.01); the dilatation of kidney tubular lumen, epithelial cell swelling, vacuolar degeneration,and epithelial cell exfoliation were decreased; the expression levels of FSP1, FTH1, GPX4, Nrf2,and HO-1 proteins in kidney tissue of the mice in POP group were significantly increased (P<0.05 or P<0.01). Compared with POP group, the levels of Scr and BUN of the mice in Erastin +POP group were significantly increased (P<0.01), the level of MDA in kidney tissue was increased (P<0.05), and the level of GSH was significantly decreased (P<0.01); the pathological injury of kidney tissue was aggravated obviously; the expression levels of FSP1, FTH1, GPX4, Nrf2, and HO-1 proteins in kidney tissue were significantly decreased (P<0.05 or P<0.01). Conclusion POP can reduce the AKI in the mice induced by cisplatin, and its mechanism may be related to the inhibitory effect of POP on the ferroptosis induced by cisplatin.

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Ameliorative effect of novel antiepileptic drug Q808 on rats with temporal lobe epilepsy and its mechanism
Weiwei ZHENG,Fan GAO,Zhenlin YANG,Jiarui LI,Jingjing GUO,Jinzi LI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1243-1249.  DOI: 10.13481/j.1671-587X.20240507
Abstract ( 29 )   HTML ( 0 )   PDF (863KB) ( 75 )  

Objective To discuss the ameliorative effect of a novel antiepileptic drug Q808 on neuronal injury in temporal lobe epilepsy (TLE) rats, and to clarify its mechanism of action. Methods TLE rat model was prepared by intraperitoneal injection of the innovative antiepileptic drug candidate 6-(4-chlorophenoxy)- tetrazolo(5,1-a) phthalazine (Q808). Forty-five successfully modeled rats were randomly divided into model group, low dose of Q808 group, and high dose of Q808 group, and there were 15 rats in each group. The rats in low dose of Q808 group and high dose of Q808 group were gavaged with 20 and 80 mg·kg-1 Q808,respectively, and the rats in model group were gavaged with an equal amount of 0.3% sodium carboxymethyl cellulose. Another 15 healthy SD rats were selected as control group. After 4 weeks of continuous gavage treatment, the morphology of the rats in varioius groups was observed; PONEMAH 6.X experimental animal telemetry platform was used to record the electroencephalogram of the rats in various groups; Golgi staining was used to observe the morphology of dendritic and dendritic spine density of hippocampal CA1 neurons of the rats in various groups; Western blotting method was used to detect the expression levels of synaptic plasticity-specific protein calcium/calmodulin-dependent protein kinaseⅡ (CaMKⅡ) in hippocampus tissue of the rats in various groups. Results The rats in control group showed normal activity without convulsions or other abnormal manifestations. The rats in model group, low dose of Q808 group, and high dose of Q808 group showed varying degrees of reduced activity, trembling and nodding, loss of balance, muscle rigidity and forelimb convulsions, gradually transforming into whole-body muscle rigidity and standing, followed by falling backwards, and there were no convulsions during the interictal period. Compared with control group, the total durations of epileptic seizures of the rats in model group, low dose of Q808 group, and high dose of Q808 group were significantly prolonged (P<0.01). Compared with model group, the total durations of epileptic seizures in low dose of Q808 group and high dose of Q808 group were significantly shortened (P<0.01). The hippocampal CA1 neurons of the rats in control group showed regular distribution of dendrites with dense and orderly dendritic networks. The hippocampal CA1 neurons of the rats in model group showed disordered arrangement of dendrites with massive dendritic entanglement, forming thicker nerve fiber bundles. Compared with model group, the dendritic networks of hippocampal CA1 neurons of the rats in low dose of Q808 group and high dose of Q808 group were partially recovered with relatively regular arrangement. Compared with control group, the dendritic spine density of hippocampal CA1 neurons of the rats in model group was significantly decreased (P<0.01). Compared with model group, the dendritic spine densities of hippocampal CA1 neurons in low dose of Q808 group and high dose of Q808 group significantly increased (P<0.01). Compared with control group, the expression levels of CaMKⅡ protein in hippocampus tissue of the rats in model group, low dose of Q808 group, and high dose of Q808 group were significantly decreased (P<0.01). Compared with model group, the expression levels of CaMKⅡ protein in hippocampus tissue of the rats in low dose of Q808 group and high dose of Q808 group were significantly increased (P<0.01). Conclusion The novel antiepileptic drug Q808 has an ameliorating effect on the TLE model rats;its mechanism may be related to Q808’s ability to reduce the dendritic lesions in hippocampal CA1 neurons and increase the expression level of synaptic plasticity-related protein CaMKⅡ protein.

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Inhibitory effect of mesalazine on pro-inflammatory factors and peroxides in RAW264.7 cells and its therapeutic effect on periodontitis model rats
Haoyu WANG,Yuqi WANG,Bingqian WANG,Jinhan NIE,Jiaqing YAN,Min HU
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1250-1258.  DOI: 10.13481/j.1671-587X.20240508
Abstract ( 36 )   HTML ( 0 )   PDF (2393KB) ( 11 )  

Objective To discuss the anti-inflammatory and antioxidant effect of mesalazine (MSZ) in the RAW264.7 cell model,and to elucidate its therapeutic effect on periodontitis in the rats. Methods The proliferation rates of RAW264.7 cells stimulated by different concentrations (0, 62.5, 125.0, 250.0, 500.0, 1 000.0, and 2 000.0 mg·L-1)of MSZ were detected by CCK-8 method to determine the optimal concentration of MSZ for cell treatment. Porphyromonas gingivalis lipopolysaccharide (P.g-LPS) and MSZ were used to treat the RAW264.7 cells, and the cells were divided into control group, P.g-LPS group, and MSZ+P.g-LPS group. The levels of reactive oxygen species (ROS) in the cells in various groups were detected by the DCFH-DA fluorescent probe assay; the malondialdehyde (MDA) levels, glutathione (GSH) levels and superoxide dismutase (SOD) activities in the cells in various groups were detected by ELISA method; the expression levels of inflammatory factors interleukin-8 (IL-8) and interleukin-1β (IL-1β) mRNA in the cells in various groups were detected by real-time fluorescence quantitative PCR (RT-qPCR) method.The periodontitis rat model was established by the ligation method combined with the injection of P.g bacterial fluid. A total of 18 rats were randomly divided into control group (without treatment), model group (making periodontits model), and drug administration group(making periodontits model and given MSZ), and there were 6 rats in each group. Micro-CT was used to assess the alveolar bone destruction of the rats in various groups; HE staining was used to observe the morphology of periodontal tissue of the rats in various groups. Results Compared with control group, the proliferation rate of the cells in 500.0 mg·L-1 MSZ group was significantly increased (P<0.01), so 500.0 mg·L-1 MSZ was subsequently selected to treat the cells. Compared with control group, the levels of ROS and MDA in the cells in P.g-LPS group were significantly increased (P<0.01), and the level of GSH and activity of SOD were significantly decreased (P<0.01), and the expression levels of IL-1β and IL-8 mRNA were significantly increased (P<0.01); compared with P.g-LPS group, the levels of ROS and MDA in the cells in MSZ+P.g-LPS group were significantly decreased (P<0.01), the level of GSH and activity of SOD were significantly increased (P<0.01), and the expression levels of IL-1β and IL-8 mRNA were significantly decreased (P<0.01). The micro-CT assay results showed that compared with control group, the distance from the cemento-enamel junction to alveolar bone crest (CEJ-ABC) of the rats in model group was significantly increased(P<0.01), and the bone volume fraction (BV/TV) was significantly decreaced (P<0.05); compared with model group; the CEJ-ABC of the rats in drug administration group was decreased(P<0.01), and the BV/TV was increased (P<0.05). The HE staining results showed that the inflammatory cell infiltration in periodontal tissue of the rats in drug administration group was reduced, and epithelial attachment was restored. Conclusion MSZ effectively inhibits the production of pro-inflammatory factors and peroxides in the P.g-LPS-induced RAW264.7 cells, improves the cellular anti-inflammatory and antioxidant capacity, inhibits the alveolar bone resorption, and alleviates the inflammation of periodontal tissues in the periodontitis rats.

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CBCT and three-dimensional finite element analysis based on differences in dentin stress distribution before and after root canal preparation of mandibular first premolar teeth
Xinmiao JIANG,Zhibo XU,Yuqi ZHEN,Quzhen BAIMA,Xiuping MENG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1259-1265.  DOI: 10.13481/j.1671-587X.20240509
Abstract ( 32 )   HTML ( 0 )   PDF (927KB) ( 14 )  

Objective To analyze the root canal diameter of the mandibular first premolar by using finite element analysis to simulate the stress distribution of dentin under three different preparation methods,and to provide the basis for clinical root canal preparation strategies of the mandibular first premolars. Methods Twenty-one patients with complete cone beam computed tomography (CBCT) images were selected. The original DICOM format data from CBCT were imported into Mimics 21.0 software to measure the root canal diameter at 3, 6, 9, and 12 mm from the apex and the root canal taper was segmentally calculated. Based on this, three-dimensional finite element models of the dental and periodontal tissues were constructed. Control group, maximum diameter preparation group, uniform preparation group, and 0.06 taper instrument preparation group were designed. In ANSYS Workbench 17.0 finite element analysis software, a 200 N load was applied to the buccal, lingual, and occlusal surfaces in various groups, and the stresses on dentin in various groups were analyzed. Results The analysis of root canal taper at 3-6 mm, 6-9 mm, and 9-12 mm from the apex of mandibular first premolar teeth showed that the taper was similar in the mesial-distal direction at 3-6 mm from the apex. The average taper in the buccal-lingual direction at 6-9 mm from the apex was 0.29, which was greater than the taper in the apical 1/3 and coronal 1/3. Under the same load, the peak stress values in dentin of mandibular first premolar teech in various groups were increased sequentially: 4.693 6, 16.304 0, 14.278 0, and 18.682 0 MPa. The stress in maximum diameter preparation group concentrated on the canal wall with the highest stress value. The stress in uniform preparation group concentrated on the root surface,and the stress values on each section were lower than those in maximum diameter preparation group. The stress in 0.06 taper instrument preparation group concentrated on the apical 1/3 of the root surface. Conclusion The root canal of the mandibular first premolar has a unique elliptical taper shape, and there are significant differences in diameter and taper between the mesial-distal and buccal-lingual directions. Different preparation methods result in different stress distributions on the canal wall, and the uniform preparation is the best method for enlarging the canal.

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Effect of Wnt/β-catenin signaling pathway inhibitor MSAB on fibrogenic responses of human endometrial stromal cells
Feina WANG,Xuguang MI,Xiuying LIN,Jianhua FU,Lei LIU,Xinyue YU,Huanhuan ZANG,Linjun LIU,Shiling CHEN,Yanqiu FANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1266-1274.  DOI: 10.13481/j.1671-587X.20240510
Abstract ( 38 )   HTML ( 0 )   PDF (713KB) ( 10 )  

Objective To discuss the effect of Wnt/β-catenin signaling pathway inhibitor methyl 3-{[(4-methyl- phenyl)sulfonyl] amino} benzoate(MSAB)on the fibrogenic response of the human endometrial stromal cells (HESCs), and to provide the foundation for the application of MSAB in the target therapy of intrauteriue adhesion (IUA). Methods The normal HESCs were cultured in vitro and divided into two groups: control group and transforming growth factor β1 (TGF-β1) group; the HESCs from the adhesion part of the IUA patients were cultured in vitro, regarded as IUA group. Western blotting method was used to detect the expression levels of fibrotic marker protein type Ⅰ collagen α1 (COL1A1) in the cells in various groups at different time points (0, 12, 24, 48, and 60 h) after treated with TGF-β1. MTT assay was used to detect the proliferation activities of the cells in various groups. Western blotting method was used to detect the expression levels of the fibrotic marker protein COL1A1, stromal marker proteins such as N-cadherin and α-smooth muscle actin (α-SMA), and Wnt/β-catenin signaling pathway-related protein β-catenin in the cells in control and IUA groups. Based on the MSAB concentrations, the normal HESCs were divided into 0 (control), 0.25, 0.50, 0.75, and 1.00 μmol·L-1 MSAB groups, and MTT assay was used to detect the survival rates of the cells in various groups. After treated with MSAB, the normal HESCs were divided into control group (normal HESCs), TGF-β1 group (10 μg·L-1 TGF-β1 induced normal HESCs for 24 h then the drug was withdrawn, replaced with complete culture medium, and the cells continued to be cultured for 24 h), and MSAB group (10 μg·L-1 TGF-β1 induced normal HESCs for 24 h then the drug was withdrawn, replaced with a complete medium containing 0.75 μmol·L-1 MSAB and the cells continued to be cultured for 24 h). Real-time fluorescence quantitative PCR(RT-qPCR) method was used to detect the expression levels of epithelial-mesenchymal transition (EMT)-related transcription factors Snail, Slug, Smuc, ZEB1, and ZEB2, and COL1A1 mRNA in the cells in various groups. Western blotting method was used to detect the expression levels of COL1A1, N-cadherin, α-SMA, β-catenin, and c-myc proteins in the cells in various groups. Results Compared with control group (after treated with TGF-β1 for 0 h), the expression levels of COL1A1 proteins in the HESCs after treated with TGF-β1 for 12, 24, 48, and 60 h in TGF-β1 group were increased (P<0.05 or P<0.01).Compared with control group, there was no significant difference in the proliferation activity of the HESCs in IUA and TGF-β1 groups (P>0.05).Compared with control group, the expression levels of COL1A1, β-catenin, N-cadherin, and α-SMA proteins in the cells in IUA group were increased (P<0.05 or P<0.01). Compared with control group, the survival rates of the cells in 0.75 and 1.00 μmol·L-1 MSAB groups were decreased (P<0.05 or P<0.01). Compared with control group, the expression levels of Snail, Slug, and COL1A1 mRNA in the cells in TGF-β1 group were increased (P<0.05 or P<0.01); compared with TGF-β1 group, the expression levels of Snail, Slug, and COL1A1 mRNA in the cells in MSAB group were decreased (P<0.05 or P<0.01). Compared with control group, after treated with TGF-β1 for 24 h, the expression levels of COL1A1, N-cadherin, α-SMA, β-catenin, and c-myc proteins in the cells in TGF-β1 group were increased (P<0.01); compared with TGF-β1 group, the expression levels of COL1A1, N-cadherin, α-SMA, β-catenin, and c-myc proteins in the cells in MSAB group were decreased (P<0.05 or P<0.01). Conclusion MSAB can inhibit the fibrogenic responses of the HESCs in vitroand the results provide the theoretical basis for the application of MSAB in the target therapy of IUA.

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Damage effect of VSV on vascular endothelial barrier function in vitro and its mechanism
Yuxuan CAO,Wei CHEN,Chengbiao SUN,Na ZHAO,Yan WANG,Mingxin DONG,Na XU,Wensen LIU,Yongmei LI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1275-1285.  DOI: 10.13481/j.1671-587X.20240511
Abstract ( 36 )   HTML ( 0 )   PDF (3186KB) ( 13 )  

Objective To discuss the damage effect of vesicular stomatitis virus (VSV) on the vascular endothelial (VE) barrier, and to clarify its mechanism. Methods The canine kidney cells were used to amplify VSV. The half tissue culture infective dose (TCID50) of VSV was determined using mouse brain endothelial tumor bEnd.3 cells, and subsequent experiment was conducted using 300 times the TCID50. The bEnd.3 cells were divided into infection 0 h group, infection 4 h group, infection 8 h group, and infection 12 h group for VE barrier damage experiments due to VSV infection. The bEnd.3 cells were also divided into control group, infection group, and correction group for experiments to inhibit the VSV replication and restore the VE barrier. The bEnd.3 cells were inoculated into Transwell chambers to construct an in vitro VE barrier model. Cell voltage resistance meter was used to detect the transepithelial resistance (TER) in various groups after the bEnd.3 cells were infected with VSV at different time points;fluorescein isothiocyanate-dextran leakage assay was used to detect the permeability coefficients of the cells in various groups; immunofluorescence staining was used to observe the localization changes of VE-cadherin, β-catenin, and phosphorylated β-catenin (p-β-catenin) in cytoskeleton and adherens junctions (AJs) of the bEnd.3 cells after VSV infection; real-time fluorescence quantitative PCR (RT-qPCR) method was used to detect the expression levels of Wnt and β-catenin mRNA in the cells in various groups; Western blotting method was used to detect the expression levels of Wnt, β-catenin, and p-β-catenin proteins in the cells in various groups. Results The TCID50 of VSV was 10-4.5·100 μL-1. The Transwell chamber experiment results showed that compared with infection 0 h group, the TERs in the cells in the other groups were significantly decreased (P<0.05), and the permeability coefficients were significantly increased (P<0.05). The immunofluorescence staining results showed that compared with control group, the cytoskeleton of the bEnd.3 cells in infection group was disordered, the cell gaps was increased, the linear index of AJs was significantly decreased (P<0.05), and β-catenin and p-β-catenin translocated from the cell membrane to the perinuclear area. The RT-qPCR results showed that compared with infection 0 h group, the expression levels of Wnt mRNA in the cells in the other groups were significantly decreased (P<0.05), while the expression levels of β-catenin mRNA showed no statistically significant difference (P>0.05). The Western blotting results showed that compared with infection 0 h group, the expression levels of Wnt protein in the cells in the other groups were significantly decreased (P<0.05), the expression levels of β-catenin showed no statistically significant differences (P>0.05), and the expression levels of p-β-catenin were significantly increased (P<0.05).After inhibiting the VSV replication and correcting the low density lipoprotein receptor (LDLR) abnormalities, the Transwell chamber experiment results showed that compared with infection group, the TER in the cells in correction group was significantly increased (P<0.05), and the permeability coefficient was significantly decreased (P<0.05).The immunofluorescence staining results showed that compared with infection group, the gaps in the cells in correction group were reduced, and the perinuclear aggregation of β-catenin and p-β-catenin in the cells was restrained. The RT-qPCR results showed that compared with infection group, the expression level of Wnt mRNA in the cells in correction group was significantly increased (P<0.05). The Western blotting results showed that compared with infection group, the expression level of Wnt protein in the cells in correction group was significantly increased (P<0.05), the expression level of β-catenin showed no statistically significant difference (P>0.05), and the expression level of p-β-catenin was significantly decreased (P<0.05). Conclusion VSV infection can cause the LDLR inactivation, reduce the expression level of Wnt protein, increase the phosphorylation level of β-catenin and cause its internalization, disrupt the stability of AJs, and ultimately lead to VE barrier damage.

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Expression and localization of fever with thrombocytopenia syndrome virus nonstructural protein and screening and analysis of host-interacting proteins
Like LUO,Ziwen CHENG,Kuo CHENG,Yonggang LI,Dawei WANG,Baoling YANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1286-1296.  DOI: 10.13481/j.1671-587X.20240512
Abstract ( 37 )   HTML ( 0 )   PDF (1929KB) ( 26 )  

Objective To screen the host interaction proteins of the severe fever with thrombocytopenia syndrome virus (SFTSV) nonstructural protein (NSs) by immunoprecipitation combined with mass spectrometry analysis, to discuss the functions, subcellular localization, and biological pathways of these interaction proteins, and to provide the basis for clarifying the replication and pathogenic mechanism of SFTSV. Methods The eukaryotic expression vectors pSFTSV-NSs-Flag (experimental group) and Flag-CMV-3 (negative group) were transfected into the human embryonic kidney 293T cells, and contorl group (no treatment) was set up. The lysates of the cells in various groups were collected, and the expression and localization of SFTSV NSs in the host cells were verified by indirect immunofluorescence and Western blotting methods. The protein lysates were treated with protein A/G and immunoprecipitation was used to enrich host proteins binding to NSs. The captured interaction proteins were initially analyzed by silver staining and Coomassie brilliant blue staining to observe the differential protein bands in various groups; liquid chromatography-tandem mass spectrometry was used to obtain the information of protein sequences; the reliable proteins were retained and searched by UniProt database;Gene Ontology (GO) functional enrichment analysis, IPR, eukaryotic orthologous groups (KOGs) functional annotation, Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway enrichment analysis, subcellular localization, and transcription factor (TF) functional annotation were used to determine the subcellular structure, gene functions, and biological processes of the interaction proteins. Results The immunofluorescence results showed that the SFTSV NSs expressed a single specific band at relative molecular mass 33 000 and was localized in the cytoplasm in a granular inclusion body-like manner. The silver staining and Coomassie brilliant blue staining results showed there were significant differential protein bands between experimental group and negative group. The mass spectrometry results identified 46 potential interaction proteins. The GO functional enrichment analysis, KOGs functional annotation, and KEGG signaling pathway enrichment analysis results showed that the biological pathways related to viral translation, cellular metabolism, and protein transport were enriched with a considerable number of proteins. Eight annotated proteins had intermediate filament domains. The highest percentage of subcellular localization was cytoplasmic proteins, consistent with the NSs localization site. The TF functional annotation analysis results showed one protein from the NF-Y family. Conclusion The interaction proteins play roles in assisting the proper protein folding, participating in the cribosome translation, and forming the cytoskeleton, which may be involved in antiviral replication. These proteins can be used as candidate proteins for further study on the replication mechanism of SFTSV.

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Protective effect of carnosine against oxygen-glucose deprivation/reoxygenation-induced astrocyte injury through inhibition of autophagy by AMPK/mTOR signaling pathway
Yutong WANG,Ruili RAN,Jiang BIAN,Xiaohan JIANG,Junqiu SONG,Dewei WANG,Jing YANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1297-1304.  DOI: 10.13481/j.1671-587X.20240513
Abstract ( 29 )   HTML ( 0 )   PDF (1481KB) ( 9 )  

Objective To discuss the protective effect of carnosine(CAR) against oxygen-glucose deprivation/reoxygenation(OGD/R)-induced astrocyte(AS) injury,and to clarify its possible mechanism. Methods The AS were divided into control group, model group (OGD/R group), OGD/R+CAR group (CAR group), and OGD/R+CAR+AMP-activated protein kinase (AMPK) activator AICAR group(CAR+AICAR group). MTT assay and green cyanine staining method were used to detect the survival rates and green cyanine staining positive rates of the AS in various groups; Annexin Ⅴ-FITC/PI method and flow cytometry were used to detect the apoptotic rates of the AS in various groups; Western blotting method was used to detect the expression levels of AMPK, phosphorylated AMPK (p-AMPK), mammalian target of rapamycin (mTOR), phosphorylated mTOR (p-mTOR), microtubule-associated protein light chain 3B(LC3B), Beclin-1, and P62 proteins in the AS in various groups; immunofluorescence staining was used to observe the LC3B positive fluorescence intensities in the AS in various groups. Results Compared with control group, the survival rate and green cyanine staining positive rate of the AS in OGD/R group were decreased (P<0.01), the apoptotic rate of the AS was increased (P<0.01), the ratios of p-AMPK/AMPK and LC3BⅡ/LC3BⅠ and the expression level of Beclin-1 protein were increased (P<0.01), and the ratio of p-mTOR/mTOR and the expression level of P62 protein were decreased (P<0.01). Compared with OGD/R group, the survival rate and green cyanine staining positive rate of the AS in CAR group were increased (P<0.01), the apoptotic rate of the AS was decreased (P<0.01), the ratios of p-AMPK/AMPK and LC3B Ⅱ/LC3BⅠ and the expression level of Beclin-1 protein were decreased (P<0.01), and the ratio of p-mTOR/mTOR and the expression level of P62 protein were increased (P<0.01). Compared with CAR group, the survival rate and green cyanine staining positive rate of the AS in CAR+AICAR group were decreased (P<0.01), the apoptotic rate of the AS was increased (P<0.01), the ratios of p-AMPK/AMPK and LC3BⅡ/LC3BⅠ and the expression level of Beclin-1 protein were increased (P<0.01), and the ratio of p-mTOR/mTOR and the expression level of P62 protein were decreased (P<0.01). The LC3B immunofluorescence staining results were consistent with the Western blotting results. Conclusion CAR has the protective effect on injury of the AS induced by OGD/R, and its molecular mechanism may be related to the inhibition of the AMPK/mTOR signaling pathway, thereby inhibiting autophagy.

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Effect of silencing CDC20 on proliferation and cell cycle of endometrial cancer cells by inhibiting Wnt/β-catenin signaling pathway
Chunjing LIU,Yujie YANG,Wei ZHAO,Lijing LIU,Na WANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1305-1312.  DOI: 10.13481/j.1671-587X.20240514
Abstract ( 34 )   HTML ( 0 )   PDF (1328KB) ( 12 )  

Objective To discuss the effect of cell division cycle protein 20 (CDC20) on the proliferation and cell cycle of endometrial carcinoma (EC) cells, and to clarify its mechanism. Methods Real-time fluorescence quantitative PCR(RT-qPCR) and Western blotting methods were used to detect the expression levels of CDC20 mRNA and protein in human endometrial stromal T-HESC cell and EC cells (KLE, RL95-2, ZJB-ENC1, and ECC-1 cells). The RL95-2 cells were selected for the subsequent experiments. CDC20 shRNA interference lentivirus was transfected into the RL95-2 cells and the cells were divided into control group, sh-NC group (infected with negative control lentivirus), sh-CDC20 group (infected with CDC20 shRNA interference lentivirus), sh-NC+SM04690 group (infected with negative control lentivirus followed by treatment with 64 nmol·L?1 Wnt/β-catenin signaling pathway inhibitor SM04690 for 48 h), and sh-CDC20+SM04690 group (infected with CDC20 shRNA interference lentivirus followed by treatment with 64 nmol·L?1 SM04690 for 48 h). RT-qPCR and Western blotting methods were used to detect the expression levels of CDC20 mRNA and proteins in the cells in various groups; CCK-8 method was used to detect the proliferation activities of the RL95-2 cells in various groups; BrdU assay was used to detect the percentages of BrdU positive cells in various groups; flow cytometry was used to detect the percentages of the cells at G2/M stage in various groups; Western blotting method was used to detect the expression levels of β-catenin, oncogene c-Myc, and cyclin D1 proteins in the cells in various groups. Results Compared with T-HESC cells, the expression levels of CDC20 mRNA and protein in the KLE, RL95-2, ZJB-ENC1, and ECC-1 cells were significantly increased(P<0.05), and the highest expression levels of CDC20 mRNA and protein were observed in RL95-2 cells. Compared with sh-NC group, the proliferation activities and percentages of the BrdU positive cells in sh-CDC20 group and sh-NC+SM04690 group were significantly decreased (P<0.05), the percentages of the cells at G2/M phase were significantly increased (P<0.05), and the expression levels of β-catenin, c-Myc, and cyclin D1 proteins were significantly decreased (P<0.05). Compared with sh-CDC20 group, the proliferation activity and percentage of BrdU positive cells in sh-CDC20+SM04690 group were significantly decreased (P<0.05), the percentage of the cells at G2/M phase was significantly increased (P<0.05), and the expression levels of β-catenin, c-Myc, and cyclin D1 proteins in the cells were significantly decreased (P<0.05). Conclusion CDC20 is highly expressed in the EC cells. Silencing CDC20 may inhibit the cell proliferation by inducing G2/M phase arrest in the RL95-2 cells through the regulation of Wnt/β-catenin signal transduction.KEYWODS Endometrial cancer; Cell division cycle protein 20; Cell cycle; Cell proliferation; Wnt/β-catenin signaling pathway

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Screening of host proteins interacting with Nelson Bay orthoreovirus σNS based on yeast two-hybrid technology
Lyuyin SUN,Zhuping MA,Runlin LI,Yonggang LI,Xiaoli TAO
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1313-1321.  DOI: 10.13481/j.1671-587X.20240515
Abstract ( 35 )   HTML ( 0 )   PDF (1193KB) ( 7 )  

Objective To discuss the host proteins that interact with the Nelson Bay orthoreovirus (NBV) σNS protein in the fibroblasts L929 of the mice, and to clarify the effect of host proteins on the viral replication. Methods The bait plasmid pGBKT7-S3 expressing σNS protein was constructed, and sequencing technology was used to verify the accurate expression of the bait plasmid in the Y2H yeast cells. The pGBKT7-S3 and pGADT7 plasmids were separately and jointly transformed into the Y2HGold yeast cells, plated on solid medium, and the colony growth was observed to confirm that the σNS protein was non-toxic to the yeast cells and could not self-activate the reporter gene. The bait plasmid pGBKT7-S3 was hybridized with the cDNA library in fibroblasts L929 of the mice, and the plasmids encoding the interacting proteins were extracted from the positive clones. The positive sequencing results were screened for the proteins interacting with NBV σNS through the Uniprot database. Gene Ontology(GO) functional enrichment analysis, Kyoto Encyclopedia of Genes and Genomes(KEGG) signaling pathway enrichment analysis, and STRING bioinformatics analysis were performed on the interacting proteins. Results A total of 61 positive clones were successfully screened. The DNA sequencing analysis and BLAST alignment removed 23 positive clones that did not match the database or were similar in sequence. The positive sequencing results identified 38 proteins interacting with NBV σNS through the Uniprot database. Thirty-one proteins were involved in cellular biological processes; thirty-six proteins were cellular anatomical components; thirty-one proteins had binding functions. Five proteins were part of the mitochondrial respiratory chain; seven proteins were ribosomal proteins and components of the ribosomal subunits; two proteins were involved in iron metabolism homeostasis.The GO functional enrichment analysis results showed that the interacting proteins were enriched in biological processes(BP) such as cellular processes, metabolic processes, biological regulation, localization, and response to stimuli; the cellular components were mainly cellular anatomical components and protein-containing complexes; the molecular functions were concentrated in binding, catalytic activity, structural molecule activity, and transporter activity. The KEGG signaling pathway enrichment analysis results showed that the proteins were highly enriched in translation, folding, sorting, and degradation pathways of genetic information processing and were mainly associated with the digestive system in the organism; they were linked to various viral infections and cancers. The STRING analysis results showed that the interacting proteins included ribosomal proteins, protein modification proteins, metabolic proteins, and immune proteins. Conclusion The host proteins that interact with NBV σNS protein are successfully screened, and these host proteins play important roles in viral replication.

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Construction of dedicator of cytokinesis 4 over-expressed lentivirus vector and establishment of stable transfected Neuro-2a cells
Shengnan LI,Jiawen HE,Keqi LIAO,You LI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1322-1329.  DOI: 10.13481/j.1671-587X.20240516
Abstract ( 46 )   HTML ( 1 )   PDF (890KB) ( 18 )  

Objective To construct an over-expression lentiviral vector of the dedicator of cytokinesis 4 (DOCK4), and to establish DOCK4 stably over-expressing Neuro-2a cells. Methods The DOCK4 sequence was searched in the National Center for Biotechnology Information (NCBI) and primers were designed and synthesized; polymerase chain reaction (PCR) method was used to amplify the DOCK4 gene sequences. After digestion with BamHⅠ and AgeⅠ restriction endonucleases, the DOCK4 gene sequences were ligated with the digested lentiviral vector GV492 to construct the GV492-DOCK4 over-expression recombinant plasmid. The positive clones with a similar length to the target gene fragment were screened and identified by PCR method. The GV492-control plasmid and GV492-DOCK4 over-expression recombinant plasmid were transfected into the HEK293T cells, and the lentivirus was collected and titered 48 h after transfection. The Neuro-2a cells were divided into GV492-control group and GV492-DOCK4 group, and the cells were infected with GV492-control lentivirus and GV492-DOCK4 over-expression lentivirus,respectively, and the multiplicity of infection (MOI) was 100. After 72 h of infection, the successfully infected Neuro-2a cells were screened by using puromycin (10 mg·L-1). The growth status of Neuro-2a cells and the expression of green fluorescent protein in various groups were observed under fluorescence microscope. Real-time quantitative PCR (RT-qPCR) and Western blotting methods were used to detect the expression levels of DOCK4 mRNA and DOCK4 protein in the Neuro-2a cells in various groups. Results The PCR results showed that the gene fragment length of the GV492-DOCK4 over-expression recombinant plasmid was approximately 691 bp. The sequencing results showed that the gene sequence of the GV492-DOCK4 over-expression recombinant plasmid was consistent with the designed over-expression sequence of DOCK4. The titers of the lentiviruses in GV492-control group and GV492-DOCK4 over-expression group were 2.5×108 TU·mL-1 and 2.5×108 TU·mL-1, respectively. The fluorescence microscope observation results showed that Neuro-2a cells in various groups grew well and expressed green fluorescent protein. The RT-qPCR results showed that compared with GV492-control group, the expression level of DOCK4 mRNA in the Neuro-2a cells in GV492-DOCK4 group was significantly increased (P<0.01). The Western blotting results showed the specific bands near the relative molecular mass of 225 000 in various groups. Compared with GV492-control group, the expression level of DOCK4 protein in the Neuro-2a cells in GV492-DOCK4 group was significantly increased (P<0.01). Conclusion This study successfully constructs the DOCK4 over-expression lentiviral vector and establishes the Neuro-2a cells stably over-expressing DOCK4.

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Effects of 5-Aza-CdR on autophagy and apoptosis of papillary thyroid cancer cells in subcutaneous xenograft tumor tissue of nude mice and its mechanism
Yuxiao SHI,Meilan LIU,Meilin ZHU,Feng WEI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1330-1338.  DOI: 10.13481/j.1671-587X.20240517
Abstract ( 33 )   HTML ( 0 )   PDF (1199KB) ( 12 )  

Objective To discuss the effects of 5-aza-2'-deoxycytidine(5-Aza-CdR) on autophagy and apoptosis of the TPC-1 cells in subcutaneous xenograft tumor tissue of the nude mice, and to clarify its mechanism. Methods Sixteen female BALB/c nude mice were inoculated with human papillary thyroid carcinoma (PTC) TPC-1 cells in the right axilla to establish the xenograft tumor model. After tumor formation, the mice were randomly divided into control group and experiment group(n=8). The nude mice in control group were given an intraperitoneal injection of saline, while the nude mice in experiment group were given the intraperitoneal injection of 5-Aza-CdR, administered once every other day for four weeks. The growth status of xenograft tumor of the mice in both groups was observed, and the mice were sacrificed after the final administration and the tumor weights of the nude mice in two groups were detected. HE staining was used to observe the pathomorphology of xenograft tumor tissue of the nude mice in both groups;immunohistochemistry was used to detect the expression levels of microtubule-associated protein light chain 3 (LC3), B-cell lymphoma 2(Bcl-2), and Bcl-2-associated X protein(Bax) in xenograft tumor tissue of the nude mice in two groups;real-time fluorescence quantitative PCR(RT-qPCR) and Western blotting methods were used to detect the expression levels of LC3, Beclin-1, Bcl-2, Bax, mitogen-activated protein kinase kinase(MEK), extracellular signal-regulated kinase 1(ERK1), extracellular signal-regulated kinase 2(ERK2), phosphorylated EPK1(p-ERK1), and phosphorylated EPK2(p-ERK2) mRNA and proteins in xenograft tumor tissue of the nude mice in two groups. Results Compared with control group, the tumor volume and weight of the nude mice in experiment group were significantly decreased(P<0.01). The number of cancer cells of the nude mice in control group was high, and the cells were densely arranged, with irregular shapes, clear nuclear staining, large overlapping nuclei, and lobulation, showing significant pathological mitotic figures consistent with PTC pathological characteristics. The number of cancer cells of the nude mice in experiment group showed a significant decresing trend, and the cells were sparse arrangement, nuclear shrinkage, and less distinct nuclei, with a significant increase in connective tissue. Compared with control group, the expression levels of LC3B and Beclin-1 mRNA and proteins in xenograft tumor tissue of the nude mice in experiment group were significantly increased(P<0.05) and the ratio of and Bax/Bcl-2 was increased(P<0.05 or P<0.01), while the expression levels of MEK, ERK1/2, and p-ERK1/2 mRNA and proteins were significantly decreased(P<0.05 or P<0.01). Conclusion 5-Aza-CdR can inhibit the growth of TPC-1 cells in subcutaneous xenograft tumor tissue of the nude mice, induce the autophagy, and promote the apoptosis of the tumor cells. The mechanism may be related to the inhibition of the mitogen-activated protein kinase(MAPK)/MEK/extracellular signal-regulated kinase(ERK) signaling pathways.

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Alleviatory effect of curcumin on cardiomyocyte toxicity induced by doxorubicin by regulating SIRT3/SOD2 signaling pathway
Fengmei XIONG,Yuxiang CAI,Zhuo LIU,Na SUN,Yang LI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1339-1347.  DOI: 10.13481/j.1671-587X.20240518
Abstract ( 30 )   HTML ( 1 )   PDF (1000KB) ( 16 )  

Objective To discuss the effect of curcumin on ameliorating doxorubicin (DOX)-induced cytotoxicity in the H9c2 cardiomyocytes, and to clarify its mechanism. Methods The H9c2 cardiomyocytes were treated with DOX to establish the cardiotoxicity model. The H9c2 cells were divided into normal group, DOX group, DOX+curcumin group, and DOX+curcumin+silent information regulator 3 (SIRT3) inhibitor-3 (3-TYP) group. After 24 h, the morphology of the cells in various groups were observed; CCK-8 method was used to detect the viabilities of the cells in various groups; TUNEL staining was used to detect the apoptotic rates of the cells in various groups; enzyme-linked immunosorbent assay (ELISA) method was used to detect the activities of catalase (CAT) and superoxide dismutase (SOD) and the levels of malondialdehyde (MDA) in the cells in various groups; 2,7-dichlorofluorescein diacetate(DCFH-DA) staining was used to detect the levels of reactive oxygen species (ROS) in the cells in various groups; Western blotting method was used to detect the expression levels of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), nicotinamide adenine dinucleotide phosphate(NADPH) oxidase 2 (NOX2), NADPH oxidase 4 (NOX4), SIRT3, acetylated superoxide dismutase 2 (Ac-SOD2), and superoxide dismutase 2 (SOD2) proteins in the cells in various groups. Results Compared with normal group, the H9c2 cells in DOX group exhibited swelling, the activity of the cells was decreased (P<0.05), the CAT and SOD activities were decreased (P<0.05), the MDA and ROS levels were increased (P<0.05), the expression levels of NOX2, and NOX4 proteins were increased (P<0.05), the ratio of Bcl-2/Bax and the expression level of SIRT3 protein were decreased (P<0.05), and the expression levels of SOD2 and Ac-SOD2 proteins were increased (P<0.05). Compared with DOX group, the H9c2 cells in DOX+curcumin group showed improved morphology, the activity of the cells was increased (P<0.05),the CAT and SOD activities were increased (P<0.05), the MDA and ROS levels were decreased (P<0.05), the expression levels of NOX2, and NOX4 proteins were decreased (P<0.05), the ratio of Bcl-2/Bax and the expression level of SIRT3 protein were increased (P<0.05), and the expression levels of SOD2 and Ac-SOD2 proteins were decreased (P<0.05). Compared with DOX+curcumin group, the cells in DOX+curcumin+3-TYP group exhibited swelling, the activity of the cells was decreased (P<0.05), the apoptotic rate of the cells was significantly increased (P<0.05), the CAT and SOD activities were decreased (P<0.05), the MDA and ROS levels were significantly increased (P<0.05), the expression levels of NOX2 and NOX4 proteins were increased (P<0.05), the ratio of Bcl-2/Bax and the expression level of SIRT3 protein were decreased (P<0.05), and the expression levels of SOD2 and Ac-SOD2 proteins were increased (P<0.05). Conclusion Curcumin suppresses the oxidative stress and apoptosis by activating the SIRT3/SOD2 signaling pathway, improving the cell activity and alleviating the DOX-induced cytotoxicity in the H9c2 cells.

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Effect of bone marrow mesenchymal stem cell-derived exosomes on myocardial fibrosis in rats induced by isoproterenol and its mechanism
Junping WEI,Dajia FU,Qingwen MENG,Daofei LIN,Yanzai LIN
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1348-1357.  DOI: 10.13481/j.1671-587X.20240519
Abstract ( 37 )   HTML ( 0 )   PDF (6025KB) ( 39 )  

Objective To discuss the effect of bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (Exo) on isoproterenol (ISO)-induced myocardial fibrosis in the rats, and to clarify its mechanism. Methods The Exo was isolated from the BMSCs and characterized by transmission electron microscope, nanoparticle tracking analysis, and Western blotting methods. Forty SD rats were divided into control group, model group, BMSCs-Exo group, and BMSCs-Exo+ferroptosis activator (Erastin) group, and there were 10 rats in each group. The myocardial fibrosis models were established by subcutaneous injection of ISO in all the rats except control group. The rats in BMSCs-Exo and BMSCs-Exo+Erastin groups were given BMSCs-Exo, and the rats in BMSCs-Exo+Erastin group were additionally injected with Erastin intraperitoneally. After 4 weeks, echocardiography was used to detect the left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), left ventricular end diastolic diameter (LVEDD), and left ventricular end systolic diameter (LVESD) of the rats in various groups; HE staining was used to observe the pathomorphology of myocardium tissue of the rats in various groups; Masson staining was used to observe the fibrosis degrees of myocardium tissue of the rats in various groups; immunohistochemistry was used to detect the positive expression rates of α-smooth muscle actin (α-SMA), type Ⅰ collagen (COL-Ⅰ), and type Ⅲ collagen (COL-Ⅲ) in myocardium tissue of the rats in various groups; colorimetry was used to detect the Fe2+ level in myocardium tissue of the rats in various groups; Western blotting method was used to detect the expression levels of acyl-CoA synthetase long chain family member 4 (ACSL4), ferritin heavy chain 1 (FTH1), glutathione peroxidase 4 (GPX4), and solute carrier family 7 member 11 (SLC7A11) proteins in myocardium tissue of the rats in various groups. Results The particles isolated from BMSCs had a typical lipid bilayer structure, and most particle sizes distributed around 100 nm. High expression levels of CD63, CD9, tumor susceptibility gene 101 (TSG101), and heat shock protein 70 (HSP70) proteins confirmed the particles as Exo. Compared with control group, the LVEF and LVFS of the rats in model group were significantly decreased (P<0.05), LVEDD and LVESD were increased (P<0.05), the myocardial cell arrangement was disordered, some nuclear shrinkage and necrosis were seen, the collagen volume fraction (CVF) was significantly increased (P<0.05), the positive expression rates of α-SMA, COL-Ⅰ, and COL-Ⅲ were significantly increased (P<0.05), the Fe2+ level in myocardium tissue was significantly increased (P<0.05), the expression level of ACSL4 protein was significantly increased (P<0.05), and the expression levels of FTH1, GPX4, and SLC7A11 proteins were significantly decreased (P<0.05). Compared with model group, the LVEF and LVFS of the rats in BMSCs-Exo group were significantly increased (P<0.05), the LVEDD and LVESD were significantly decreased (P<0.05), the myocardial tissue damage was significantly alleviated, the CVF was significantly decreased (P<0.05), the positive expression rates of α-SMA, COL-Ⅰ, and COL-Ⅲ were significantly decreased (P<0.05), the Fe2+ level in myocardium tissue was significantly decreased (P<0.05), the expression level of ACSL4 protein was significantly decreased (P<0.05), and the expression levels of FTH1, GPX4, and SLC7A11 proteins were significantly increased (P<0.05). Compared with BMSCs-Exo group, the LVEF and LVFS of the rats in BMSCs-Exo+Erastin group were significantly decreased (P<0.05), the LVEDD and LVESD were significantly increased (P<0.05), the myocardial cell edema and necrosis were seen, the CVF was significantly increased (P<0.05), the positive expression rates of α-SMA, COL-Ⅰ, and COL-Ⅲ were significantly increased (P<0.05), the Fe2+ level in myocardium tissue was significantly increased (P<0.05), the expression level of ACSL4 protein was significantly increased (P<0.05), and the expression levels of FTH1, GPX4, and SLC7A11 proteins were significantly decreased (P<0.05). Conclusion BMSC-derived Exo can improve the myocardial fibrosis in the rats induced by ISO, and its mechanism may be related to the inhibition of ferroptosis.

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Research in clinical medicine
Effect of MEIS1 expression on survival in patients after radical gastrectomy and its value in prognostic evaluation
Jiaxin YI,Yangyu ZHANG,Yingli FU,Yuchen PAN,Yongjie HAN,Jing JIANG,Yanhua WU
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1358-1364.  DOI: 10.13481/j.1671-587X.20240520
Abstract ( 35 )   HTML ( 1 )   PDF (946KB) ( 5 )  

Objective To discuss the postoperative survival of the gastric cancer patients with different expression levels of myeloid ecotropic viral integration site 1 (MEIS1), and to analyze the predictive value of MEIS1 expression in the prognosis evaluation of gastric cancer. Methods In a gastric cancer survival cohort, 215 patients who underwent radical gastrectomy were selected. Immunohistochemical staining was used to detect the expression levels of MEIS1 in both gastric cancer and adjacent normal tissues. The relationship between expression level of MEIS1 and the clinicopathological characteristics of the patients were analyzed by χ2 test or Fisher’s exact probability method;survival curves were plotted by Kaplan-Meier method; the differences in survival of the patients between MEIS1 high expression group and MEIS1 low expression group were compared by Log-rank test; multivariate Cox proportional hazards regression model was used to calculate the hazard ratios (HR) and 95% confidence intervals (CI) to assess the relationship between MEIS1 expression level and the survival of the gastric cancer patients. Results The immunohistochemical staining result showed that the expression level of MEIS1 in gastric cancer tissue was decreased. The univariate analysis results showed that the patients with high MEIS1 expression had a longer overall survival than those with low expression (P=0.049), and had a better prognosis. The multivariate Cox proprotional hazards regression analysis results showed that the low MEIS1 expression and high TNM stage were the independent risk factors for poor prognosis of the patients with gastric cancer (HR=1.577, 95%CI: 1.011-2.460, P=0.045; HR=2.709, 95%CI: 1.708-4.297, P<0.001). Conclusion The gastric cancer patients with low expression of MEIS1 have a shorter postoperative overall survival; MEIS1 is a promising biomarker for prognosis assessment of the patients after radical gastrectomy.

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Expressions of EHD2, miRNA let-7c,and lncRNA FOXD2-AS1 in human laryngeal squamous cell carcinoma tissue and their relationship analyses
Meiheng GONG,Mo CHEN,Hui HAN,Tingting YU
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1365-1371.  DOI: 10.13481/j.1671-587X.20240521
Abstract ( 32 )   HTML ( 0 )   PDF (914KB) ( 16 )  

Objective To discuss the expression levels of Eps15 homology domain-containing protein 2 (EHD2), microRNA let-7c (miR-let-7c), and long non-coding RNA (lncRNA) FOXD2-AS1 in laryngeal squamous cell carcinoma (LSCC) tissue, and to clarify the association between EHD2/miR-let-7c/FOXD2-AS1 signaling axis and occurrence of LSCC. Methods Forty LSCC tissue samples were collected and classified into low grade group (moderately or high differentiated, 32 cases) and high grade group (poorly differentiated, 8 cases) according to the pathology results; according to the tumor node metastasis(TNM) clinical staging results, the samples were divided into TNM early stage group (stages Ⅰ-Ⅱ, 13 cases) and TNM late stage group (stages Ⅲ-Ⅳ, 27 cases); based on the lymph node metastasis results, the samples were divided into metastasis group (21 cases) and non-metastasis group (19 cases). Additionally, 40 corresponding normal adjacent tissue samples were collected as control group. Immunohistochemistry method was used to detect the expressions of EHD2 in various groups and its relationships with clinical pathoparameters of the LSCC patients were analyzed; bioinformatics method was used to confirm that the miR-let-7c was the candidate microRNA(miRNA) and FOXD2-AS1, which had binding sites in its promoter region, was a candidate lncRNA. Ten pairs of fresh LSCC tissue samples and adjacent normal tissue samples were collected. Real-time fluorescence quantitative PCR (RT-qPCR) method was used to detect the expression levels of EHD2 mRNA, miR-let-7c, and FOXD2-AS1 in the samples in two groups, and their associations were verified. Results Compared with adjacent normal tissue, the expression level of EHD2 in LSCC tissue was significantly decreased (P<0.01). The positive expression rate of EHD2 in LSCC tissue of the patients in TNM early stage group was significantly higher than that in TNM late stage group (P<0.05). There was no significant association between EHD2 expression and pathological type or lymph node metastasis (P>0.05). Compared with control group, the expression level of miR-let-7c in LSCC tissue was significantly decreased (P<0.05), while the expression level of FOXD2-AS1 was significantly increased (P<0.05). In LSCC tissue, the expression level of FOXD2-AS1 was negatively correlated with the expression level of miR-let-7c (r=-0.67,P<0.05), and the expression level of miR-let-7c was negatively correlated with the expression level of EHD2 mRNA(r=-0.83, P<0.01). Conclusion EHD2 and miR-let-7c both express at low levels in LSCC tissue and may be new tumor suppressor genes; FOXD2-AS1 is highly expressed in LSCC tissue and may be a new oncogene. FOXD2-AS1/miR-let-7c/EHD2 signaling axis may be involved in the occurrence and development of LSCC.

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Effects of different nickel-titanium archwires on orthodontic alignment efficiency and pain perception of patients with extraction orthodontic treatment and non-extraction orthodontic treatment
Jiyu SONG,Yi ZHANG,Qingyuan REN,Jinhan NIE,Min HU,Huichuan QI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1372-1380.  DOI: 10.13481/j.1671-587X.20240522
Abstract ( 44 )   HTML ( 1 )   PDF (696KB) ( 15 )  

Objective To discuss the effects of superelastic nickel-titanium archwires (SENT) and heat-activated nickel-titanium archwires (HANT) on the initial alignment efficiency of mandibular anterior teeth and pain levels of the patients with extraction orthodontic treatment and non-extraction orthodontic treatment by using low-friction self-ligating brackets, and to provide the basis for the selection of the most suitable archwire in orthodontic clinical practice. Methods Eighty patients underwent fixed orthodontic treatment with self-ligating brackets were randomly assigned to SENT or HANT subgroups, and there were 40 patients in extraction orthodontic treatment group and 40 patients in non-extraction orthodontic treatment group. A single operator fully engaged a 0.014-inch straight archwire into the brackets.The patients, operators, and data measurers were all blinded;the initial clinical alignment efficiency of the archwires of the patients in various groups was observed, and the Little’s index was calculated; the visual analog scale (VAS) scores and pain perception of the patients in various groups were recorded 4 h after initial bonding of the appliances and every day before breakfast during the first week of orthodontic treatment; multivariate regression analysis was performed for the initial Little’s index, gender, and age, and the influence factors of pain in the patients was analyzed. Results There were no significant differences in age, gender, and initial Little’s index among the patients in extraction group and non-extraction group (P>0.05). Compared with before treatment, the Little’s indexes of the patients in both HANT and SENT subgroups in non-extraction orthedontic treatment group were decreased 4 weeks after treatment,but the difference between groups was not significant (P>0.05). Compared with SENT subgroup in non-extraction orthodontic treatment group, the Little’s index of the patients in HANT subgroup was decreased (P<0.05). In extraction orthodontic treatment group, there were no significant differences in Little’s indexes of the patients between the HANT and SENT subgroups before and after treatment (P>0.05). The pain perception peaked on the first day after initial bonding and gradually decreased to baseline levels.The patients in SENT and HANT subgroups in both extraction and non-extraction orthodontic treatment groups showed the similar patterns of pain change. There were no significant differences in the average VAS scores and maximum pain intensity scores between the patients in SENT and HANT subgroups at different time points (P>0.05). The type of archwire had no significant effect on the degree of pain, while time significantly affected the degree of pain. The multivariate regression analysis results showed a significant correlation between initial Little’s index and maximum VAS scores of the patients in extraction orthodontic treatment group (b=0.359, P=0.033). Gender and age did not affect the degree of pain of the patients in either group. Conclusion When applying low-friction self-ligating brackets for orthodontic treatment, the initial alignment efficiency with 0.014-inch HANT archwires is superior to 0.014-inch SENT archwires in non-extraction treatment group, while both archwires show the same efficiency in extraction patients. SENT and HANT archwires do not affect the initial degrees of pain of the orthodontic patients.

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Mendelian randomization analysis based on causal relationship between gut microbiota and gestational diabetes mellitus
Zhifei LIU,Yaru BI,Chenglin SUN,Suyan TIAN
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1381-1389.  DOI: 10.13481/j.1671-587X.20240523
Abstract ( 47 )   HTML ( 0 )   PDF (668KB) ( 14 )  

Objective To analyze the causal relationship between gut microbiota and gestational diabetes, and to clarify its mechanism. Methods Two-sample Mendelian randomization(MR) analysis was conducted by using summary data from genome-wide association study (GWAS) for gut microbiota and gestational diabetes. The GWAS data of gut microbiota were obtained from a GWAS study from the MiBioGen consortium; the GWAS data on gestational diabetes were sourced from the FinnGen consortium’s publicly available R8 dataset;inverse variance weighted (IVW) method was used as the primary method to detect the causal association between the gut microbiota and the gestational diabetes. Sensitivity analysis was performed by Weighted Median and MR Egger methods; heterogeneity and pleiotropy were detected by Cochran’s Q, MR-PRESSO, Egger intercept tests and Leave-One-Out analysis; multivariable MR was used to adjust for the effect of body mass index (BMI); reverse MR was used to explore the presence of reverse causal associations; Gene Ontology (GO) fuctional and Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling enrichment analyses were used to explore the potential pathways through which gut microbiota may have impact on gestational diabetes. Results Four gut microbes were found to be causally associated with gestational diabetes: the genus Methanobrevibacter and the phylum Euryarchaeota displayed negative causal relationships with the risk of gestational diabetes, while the genus Olsenella and genus Lachnoclostridium exhibited positive causal associations. No significant heterogeneity or horizontal pleiotropy was detected in the analysis.The reverse MR analysis did not reveal any causal relationship. After adjusting for BMI, the multivariable MR analysis results showed there were the causal associations between the genus Olsenella and the phylum Euryarchaeota with the risk of gestational diabetes.The GO fuctional and KEGG signaling pathway enrichment analyses results showed that axon development, axon production, insulin secretion and other pathways were significantly enriched. Conclusion There are causal associations between four gut microbes and gestational diabetes. Among them, the significant correlations with gestational diabetes are still observed in phylum Euryarchaeota and genus Olsenella after adjusting for BMI.

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Predictive value of new thrombotic risk assessment model for venous thromboembolism in patients with malignant tumors
Honghong LI,Na YU,Minghao SHI,Ying SUN,Yao LI,Zhongjun SHEN,Xiaoyi LIU,Liyan ZHAO
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1390-1399.  DOI: 10.13481/j.1671-587X.20240524
Abstract ( 42 )   HTML ( 3 )   PDF (693KB) ( 13 )  

Objective To construct a new thrombus risk assessment model and evaluate its predictive ability for venous thromboembolism (VTE) in the patients with malignant tumors, and to provide the basis for the early predition of the malignant tumor patients with high risk for VTE. Methods A total of 128 untreated malignant tumor patients were included, of which 40 were diagnosed with VTE within 2 months of malignant tumor diagnosis and categorized as VTE group. A total of 88 patients who did not develop VTE were categorized as non-VTE group. The clinical risk factors and laboratory indicators of the patients in two groups were compared and analyzed; the types of thrombotic events of the patients were analyzed; the diagnostic values of thrombin-antithrombin-complex (TAT), α2-plasmin inhibitor-plasmin complex(PIC), D-dimer(D-dimer), and fibrin degradation products(FDP) in malignant tumors complicated by VTE were assessed using receiver operating characteristic (ROC) curve analysis; Multivariate Logistic regression analysis was used to analyze the correlations of the clinical risk factors and biomarkers with the malignant tumors complicated with VTE. A new thrombus risk assessment model was constructed, consisting of TAT≥0.70 μg·L-1, poor differentiation, and cardiovascular risk factors. The predictive probability of the model for malignant tumors complicated by VTE was evaluated based on the significance, goodness of fit, calibration curve, and C value of the model. The clinical application value of the new thrombus risk assessment model, COMPASS-CAT risk score (CRS), and Khorana risk score (KRS) in assessing malignant tumor patients complicated by VTE was compared using the C value and decision curve analysis (DCA). Results The plasma levels of TAT (P<0.001), PIC (P<0.001), D-dimer (P<0.05), and FDP (P<0.01) of the patients in VTE group were higher than those in non-VTE group. Compared with the patients without cardiovascular risk factors, poor differentiation, and lymphatic metastasis, the malignant tumor patients with cardiovascular risk factors (P<0.001), poor differentiation (P<0.001), and lymphatic metastasis (P<0.05) were more likely to develop VTE. Most VTE events (65%) were isolated deep vein thromboembolism(DVT). The ROC curve analysis showed that the area under the curve (AUC), sensitivity, and specificity of TAT and PIC were higher than those of D-dimer and FDP. TAT≥0.70 μg·L-1P<0.05), poor differentiation (P<0.01), and cardiovascular risk factors(P<0.01) were the independent risk factors for VTE in the malignant tumor patients. A new thrombus risk assessment model consisting of TAT≥0.70 μg·L-1, poor differentiation, and cardiovascular risk factors was constructed. The new risk assessment model had a high goodness of fit (P=0.805) and good predictive ability during internal validation (χ2=75.266, P<0.001). The ROC curve analysis results showed that the C values for the new thrombus risk prediction model, CRS, and KRS were 0.908, 0.676, and 0.541, respectively. The DCA curve analysis results showed that the new thrombus risk assessment model had a higher net benefit rate compared with CRS and KRS. Conclusion TAT and PIC have greater diagnostic efficiency than D-dimer in the early prediction of the malignant tumor patients with high-risk VTE. For the patients included in this study, the new thrombus risk assessment model, constructed from TAT≥0.70 μg·L-1, poor differentiation, and cardiovascular risk factors, has superior diagnostic efficiency and clinical predictive value compared with CRS and KRS.

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Association analysis on peripheral blood lymphocyte subsets and occurrence of carotid atherosclerosis
Jiayuan YU,Di ZHAO,Xin JIANG,Jing XU,Lili JIANG,Hongyu JIANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1400-1405.  DOI: 10.13481/j.1671-587X.20240525
Abstract ( 35 )   HTML ( 1 )   PDF (440KB) ( 14 )  

Objective: To analyze the association between the peripheral blood lymphocyte subsets and the occurrence of carotid atherosclerosis (CAS) and discuss the risk factors of CAS, and to provide the basis for risk prediction and early intervention of CAS. Methods A total of 419 subjects were selected according to the inclusion and exclusion criteria.All the subjects underwent carotid ultrasonography, and the detection of T lymphocyte, B lymphocyte, and natural killer(NK) cell subsets were performed. According to the carotid artery intima-media thickness(IMT), the subjects were divided into normal carotid artery group (n=166) and CAS group (n=253). The basic data and laboratory inspection indexes of the subjects in two groups were collected, including age, gender, blood pressure, medical history, smoking history, medication history, blood lipids, and percentages of T lymphocyte, B lymphocyte, and NK cell subsets. The risk factors for the occurrence of CAS of the subjects were analyzed using Logistic regression analysis. Results The univariate analysis results showed that compared with normal carotid artery group, the age, morbidity rate of hypertension, morbidity rate of diabetes, and smoking rate of the subjects in CAS group were increased (P<0.05), and the percentages of CD4+ T lymphocytes, CD4+ T lymphocyte count, CD4+/CD8+ ratio, CD16+CD56+NK cell count, and low-density lipoprotein cholesterol (LDL-c) level in peripheral blood were increased (P<0.05), while the percentage of CD8+T lymphocyte was decreased (P<0.05). The multivariable Logistic regression analysis results showed that age (OR=1.112,95%CI:1.083-1.142, P<0.001), smoking (OR=1.997,95%CI:1.192-3.346, P=0.009),LDL-c (OR=1.427, 95%CI:1.017-2.001, P=0.039), and percentage of CD4+T lymphocyte(OR=1.044,95%CI:1.002-1.087,P=0.039) were the independent risk factors for the occurrence of CAS. Conclusion The percentage of CD4+ T lymphocyte is associated with the occurrence of CAS and it is a predictive risk factor for CAS.

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Effects of different doses of esketamine on remifentanil-induced hyperalgesia in patients underwent thoracoscopic pulmonary lobectomy
Ruikun WANG,Zongsheng DUAN,Wenwen ZHANG,Hushan WANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1406-1413.  DOI: 10.13481/j.1671-587X.20240526
Abstract ( 34 )   HTML ( 0 )   PDF (501KB) ( 8 )  

Objective To compare the preventive effects of different doses of esketamine on remifentanil-induced hyperalgesia (RIH) in the patients underwent thoracoscopic pnlmonary lobectomy, and to provide the basis for the multimodal analgesia and rapid postoperative recovery. Methods The prospective, double-blind,parallel-designed randomized controlled trial (RCT) were conducted, and 107 patients who underwent visual-assisted thoracoscopic pulmonary lobectomy were included. The patients were randomly divided into normal control group,low dose of esketamine group,and high dose of esketamine group using random number methods. Except the patients who were eliminated and those who dropped out of the study,there were 31 patients in normal control group,33 patients in low dose of esketamine group,and 33 patients in high dose of esketamine group.The patients in low dose of esketamine group were given the intravenous injection of 0.25 mg·kg-1 esketamine (diluted to 5 mL) during anesthesia induction;the patients in high dose of esketamine group were given the intravenous injection of 0.50 mg·kg-1 esketamine (diluted to 5 mL), and the patients in normal control group were given 5 mL intravenous injection of saline. The mechanical pain thresholds of the non-dominant forearm skin and skin around the surgical incision at different time points, numeric rating scale (NRS) scores, Ramsay sedation scores, perioperative analgesic drug dosages, and the incidences of adverse reactions such as postoperative delirium, nausea, and vomiting of the patients in various groups were recorded. Results Compared with normal control group, the mechanical pain thresholds around the surgical incision skin of the patients in low and high doses of esketamine groups were increased (P<0.05); compared with low dose of esketamine group, the extubation time of the patients in high dose of esketamine group was increased(P<0.05). Two minutes after anesthesia induction administration, compared with normal control group, the mean arterial pressure(MAP) and heart rate(HR) of the patients in low and high doses of esketamine groups were increased (P<0.05), but there were no significant differences in the MAP and HR of the patients between low dose of esketamine and high dose of esketamine group (P>0.05);compared with normal control group, the incidences of hallucinations and delirium among the patients in high dose of esketamine group were increased (P<0.05), while there were no significant differences in the incidences of above adverse reactions in low dose of esketamine group (P>0.05); compared with low dose of esketamine group, the incidences of hallucinations and delirium among the patients in high dose of esketamine group were increased (P<0.05). Conclusion Intravenous administration of esketamine with a dosage of 0.25 mg·kg-1 during anesthesia induction improves the postoperative mechanical pain threshold of the patients undergoing thoracoscopic pulmonary lobectomy, which exhibits effective prevention of RIH without an increase in incidences of adverse reactions during the perioperative period.

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Clinical medicine
Reactive plasmacytosis caused by methimazole in patients with Graves’ disease: One case report and literature review
Shimeng LI,Xin QI,Sitong LIN,Xiangwen SAN,Ling JIN,Sitong ZHANG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1414-1419.  DOI: 10.13481/j.1671-587X.20240527
Abstract ( 37 )   HTML ( 1 )   PDF (2507KB) ( 35 )  

Objective To discuss the clinical manifestations and laboratory examination results of agranulocytosis and reactive plasmacytosis (RP) in the patient with Graves’ disease (GD) after treated with methimazole (MMI), and to provide the basis for the clinicians to differentiate RP from multiple myeloma (MM). Methods The clinical manifestations, laboratory examinations, diagnosis and treatment processes of one patient with GD agranulocytosis complicated with RP were analyzed, and the related literatures were reviewed. Results The patient had a history of GD and abdominal infection. Upon admission, a complete blood count revealed a significant decrease in white blood cell count accompanied by neutropenia, and a smear re-examination showed suspicious plasma cells.The bone marrow cytology examination results showed the percentage of bone marrow plasma cells was 33%, and the percentage of plasma cells in peripheral blood was 4%; the serum immunoglobulin results showed polyclonal hyperplasia; the serum immunofixation electrophoresis results were negative; the flow cytometry analysis results indicated the immunophenotype of the plasma cells was normal. Based on the medical history and laboratory results, MM was largely excluded, supporting the diagnosis of RP. Neutropenia was considered to be related to medication, so MMI was discontinued, granulocyte colony-stimulating factor was administered to increase the number of white blood cells, and specialized GD treatment was conducted after controlling the abdominal infection. The patient had a good prognosis, and his blood count was normal upon re-examination 6 months later. Conclusion Agranulocytosis complicated with RP in the GD patients is clinically rare. Serum immunofixation electrophoresis, blood cell morphology, and cell immunophenotype analysis are helpful for the accurate diagnosis. After actively treating the primary disease causing RP, the patient’s prognosis is favorable.

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Late-onset methylmalonic acidaemia CblC type:Two case reports and literature review
Meng SUN,Ran ZHOU,Xinying ZHANG,Yaying CHENG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1420-1425.  DOI: 10.13481/j.1671-587X.20240528
Abstract ( 38 )   HTML ( 0 )   PDF (1057KB) ( 6 )  

Objective To discuss the clinical phenotype and genotype characteristics of two pediatric patients with late-onset methylmalonic acidemia (MMA) cblC type, and to provide the basis for early clinical recognition of MMA. Methods The clinical data of two pediatric patients with late-onset MMA cblC type were collected, including clinical phenotypes, biochemical detection results, blood and urine organic acid analyses, neuroimaging, electroencephalograms, genotypes and so on. The characteristics of the disease were analyzed in combination with the related literature review. Results Both pediatric patients were female, with onset in adolescence. Patient 1 presented with psychiatric symptoms, while pediatric patient 2 presented with cognitive impairment. Both pediatric patients experienced weakness in both lower limbs and speech disorders. At initial diagnosis, the serum homocysteine (Hcy) levels were severely increased, the urine methylmalonic acid levels were increased, the brain magnetic resonance imaging results indicated brain atrophy, and the electroencephalogram results showed the increased slow wave activity in both cerebral hemispheres. The pediatric patient 2 exhibited epileptiform discharges in bilateral frontal and temporal regions. The genetic testing results showed the c.482G>A mutation in the MMACHC gene. Both two pediatric patients were treated with intramuscular injections of vitamin B12, along with oral folic acid, vitamin B6, levocarnitine, and betaine. The symptoms of two patierts were improved, the serum Hcy levels were decreased, and the urine methylmalonic acid levels returned to normal. Conclusion The phenotype of late-onset MMA cblC type is diverse, primarily involving neuropsychiatric impairment, with the c.482G>A mutation being the most common genotype. The increasing of serum Hcy levels and brain atrophy can serve as the biomarkers for the early recognition of late-onset cblC type pediatric patients.

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Diagnosis of bicuspid aortic valve malformation resulting in anterior mitral aneurysm by TTE combined with TEE: A case report and literature review
Yan YAN,Feng GUO,Sibao YANG,Shaomin SHI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1426-1431.  DOI: 10.13481/j.1671-587X.20240529
Abstract ( 28 )   HTML ( 0 )   PDF (636KB) ( 7 )  

Objective To analyze the clinical manifestations, imaging characteristics, treatment measures, and efficacy of mitral valve aneurysm (MVA), and to enhance the clinicians’ understandings of MVA. Methods The clinical data of one patient with aortic valve bicuspid malformation leading to mitral valve anterior leaflet aneurysm were collected. The clinical diagnosis was confirmed based on the clinical characteristics and imaging features, the treatment methods were selected, and the efficacy was analyzed. The relevant literatures were reviewed. Results The patient, a 68-year-old female, was admitted due to palpitations and shortness of breath for 13 years, and the symptoms worsened one month ago. Thirteen years ago, the patient experienced palpitations and shortness of breath without any inducement and was diagnosed with “heart valve disease” in the local hospital. The symptoms worsened one month ago, leading to hospitalization. The transthoracic two-dimensional echocardiography (TTE) results showed the left ventricular hypertrophy, bicuspid aortic valve with thickened and echogenic leaflets, the forward flow velocity was increased, and the aortic valve orifice area was 2.0 cm2; the mitral valve anterior leaflet margin was slightly thickened and echogenic, presenting a cystic “honeycomb-like” structure closely related to the aortic valve regurgitation jet, which appeared to enter the “sac”. The preoperative transesophageal echocardiography (TEE) results showed the bicuspid aortic valve malformation with the anterior leaflet prolapsing into the left ventricular outflow tract during diastole; a “sac-like” structure was detected on the atrial surface of the mitral valve anterior leaflet, changing shape with the cardiac cycle and communicating with left ventricular blood flow. The ultrasound diagnosis was bicuspid aortic valve malformation with transverse fissure, severe regurgitation with mild stenosis, and mitral valve anterior leaflet aneurysm. Intraoperatively, the aortic valve annulus was enlarged with the significant leaflet regurgitation. The leaflets were excised, and a 23 mm bioprosthetic valve was implanted in the aortic position. Upon exploration through the aortic valve orifice, a “sac-like” structure was found on the mitral valve anterior leaflet, which was not specially treated. The postoperative TEE results showed good echo and activity of the aortic bioprosthetic valve, and the “sac-like” structure on the mitral valve remained unchanged. The follow-up results at 10 d and 4 months after operation showed good echo and activity of the aortic bioprosthetic valve, and compared with before operation, there was no significant change in the nature and size of the mitral valve cystic lesion. Conclusion MVA is clinically rare. TTE is currently the most valuable imaging diagnostic method for MVA, especially when combined with TEE, which is the best diagnostic method and can assist in the treatment and efficacy evaluation.

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Parasitic leiomyoma of abdominal wall complicated with disseminated peritoneal leiomyomatosis : A case report and literature review
Jinping ZHANG,Lingling TONG,Lu GAO,Hongjing CHENG,Minjia SHENG
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1432-1437.  DOI: 10.13481/j.1671-587X.20240530
Abstract ( 38 )   HTML ( 0 )   PDF (661KB) ( 6 )  

Objective To discuss the diagnosis and treatment process of the patients with parasitic leiomyoma (PM) of the abdominal wall complicated with disseminated peritoneal leiomyomatosis (DPL) after laparoscopic myomectomy,and to improve the clinical understanding and management of this condition. Methods The clinical data of one patient with PM of the abdominal wall complicated with DPL after laparoscopic myomectomy were collected. The causes, clinical features, diagnosis, differential diagnosis and treatment process were analyzed, and the relevant literatures were reviewed. Results The patient, a 49-year-old woman, was admitted due to a self-discovered abdominal mass lasting for one year. The physical examination results showed a palpable mass, approximately 6 cm×4 cm, in the lower left abdominal wall with poor mobility, with clear borders, and without tenderness. Another palpable mass, approximately 7 cm×5 cm, was found in the lower right abdomen with fair mobility,with clear borders, and without tenderness. The gynecological ultrasonography results showed a hypoechoic area of approximately 6.6 cm×2.7 cm in the subcutaneous tissue below the left umbilicus and another hypoechoic area of approximately 7.6 cm×3.3 cm in the abdominal cavity below the umbilicus. The superficial ultrasonography of the local area showed a hypoechoic area of approximately 5.79 cm×2.55 cm×4.74 cm within the left lower abdominal rectus muscle, with smooth edges, located 1.97 cm from the skin at its shallowest point and 4.73 cm at its deepest point, without penetration of the rectus sheath but adjacent to the peritoneum. The patient was diagnosed as uterine leiomyoma, abdominal mass, and post-myomectomy status. The elective surgeries for uterine leiomyoma enucleation, abdominal wall leiomyoma excision, and peritoneal leiomyoma excision were performed under combined intravenous-inhalation anesthesia. The operation procedure was successful, and the patient recovered well and was discharged smoothly. Conclusion PM and DPL lack typical clinical features and require imaging examinations for diagnosis. Surgical exploration is the main treatment modality, and while PM and DPL are generally benign, there is a potential for malignant transformation, and the patients need further postoperative follow-up.

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Methodology
Improvement of isolation and culture methods for primary chondrocytes of neonatal rats
Dandan YANG,Jiaoyang CHEN,Xinheng WANG,Zetong ZHAO,Ying PAN,Baigong XUE,Changzhao GAO
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1438-1449.  DOI: 10.13481/j.1671-587X.20240531
Abstract ( 36 )   HTML ( 0 )   PDF (1246KB) ( 9 )  

Objective To discuss the improved methods for the isolation and culture of primary chondrocytes from the neonatal rats,and to establish an efficient and economical in vitro chondrocyte culture system. Methods The primary chondrocytes were isolated from the joints of neonatal rats and divided into overnight digestion (OD) group and rapid digestion (RD) group for separation. The chondrocytes in OD group were digested overnight by typeⅡ collagenase, while the chondrocytes in RD group were separated by the combination of pre-digestion with physical and chemical digestion methods. The chondrocytes were cultured in modified media containing 0% (blank group 1), 1%, 2%, 4%, and 10% fetal bovine serum (FBS), 0 (blank group 2), 0.1, 0.2, 0.4, 0.8, 1.0, and 2.0 g·L-1 vitamin C(VC), and 0 (blank group 3), 0.5, 1.0, 2.0, 4.0, 8.0, 10.0 μg·L-1 poly(lactic-co-glycolic acid) (PLGA) nanoparticles. The media containing different concentrations of FBS, VC, and PLGA were mixed with Dulbecco’s modified Eagle’s medium/nutrient mixture F-12(DMEM/F12), and were divided into related groups based on the concentrations of ingredients. Cell counter was used to count the chondrocytes in various groups and the survival rates and diameters of the chondrocytes in various groups were detected; Toluidine blue staining was used to detect the morphology of the chondrocytes in various groups; CCK-8 method was used to detect the proliferative activities of the chondrocytes in various groups; cell adhesion assay was used to detect the adhesion rates of the chondrocytes in various groups; Hoechst/propidium iodide(PI) staining was used to detect the apoptosis of the chondrocytes in various groups; MTT assay was used to detect the proliferation activities of the chondrocytes in various groups after treated with modified media.The cells were divided into DMEM/F12+10%FBS group, DMEM/F12+1%FBS group, and DMEM/F12+1% FBS+0.4 g·L-1 VC+1 μg·L-1 PLGA group. Real-time fluorescence quantitative PCR(RT-qPCR) method was used to detect the expression levels of sex-determining region Y-box 9 (SOX9), collagen type Ⅱ alpha 1 chain (Col2A1), collagen type Ⅹ alpha 1 chain (Col10A1), and matrix metallopeptidase 13 (MMP13) mRNAs in the chondrocytes in various groups after treated with modified media;immunofluorescence staining was used to detect the expressions of type Ⅱ collagen (COLⅡ) and SOX9 in the chondrocytes in various groups after treated with modified media. Results The survival rate of primary chondrocytes in OD group was lower than that in RD group, and the average cell diameter was larger than that in RD group. The primary chondrocytes in OD group were larger and spindle-shaped, and most cells exhibited pseudopodia; in RD group, the primary chondrocytes were smaller, mostly rhomboid in shape, with only a portion of the cells showing pseudopodia. The Toluidine blue staining results showed significant coloration in both groups, but the digestion time of the chondrocytes in RD group was shorter, and compared with OD group, the actual culture time of the chondrocytes was reduced by 9-13 h, and more immature morphology of the primary chondrocytes were observed. The proliferation activity of the primary chondrocytes in OD group was slow at 24 h of culture but increased at 48 h of culture, and the proliferation activity of the primary chondrocytes was significantly higher at 48 h of culture compared with 12 h of culture (P<0.01). Compared with 12 h of culture,the proliferation rates of the primary chondrocytes in RD group were increased at 24 and 48 h of culture (P<0.01). At 24 and 48 h of culture, compared with OD group, the proliferation rates of the primary chondrocytes in RD group were increased (P<0.05). The number of apoptotic chondrocytes in RD group was lower than that in OD group, and no necrotic chondrocytes were observed in either group. The proliferation activities of chondrocytes of the rats were increased with the rising of FBS concentration in the culture medium. Compared with blank group 1, the proliferation activities of chondrocytes of the rats after treated with culture mediums containing 1%, 2%, 4%, and 10% FBS were significantly increased (P<0.05). Compared with blank group 2, the proliferative activities of chondrocytes of the rats after treated with culture mediums containing 0.2-1.0 g·L-1 VC were significantly increased (P<0.05), and the highest proliferation activity was found when the concentration of VC was 0.4 g·L-1 P<0.01). Compared with blank group 3, the proliferation activities of chondrocytes of the rats after treated with culture mediums containing 1-4 μg·L-1 PLGA were significantly increased (P<0.05), and the highest proliferation activity was found after treated with culture medium containing 1 μg·L-1 PLGA (P<0.05). Compared with DMEM/F12+10%FBS group, the expression levels of SOX9 mRNA and Col2A1 mRNA in the chondrocytes in DMEM/F12+1%FBS group were significantly increased (P<0.05 or P<0.01). Compared with DMEM/F12+10%FBS group, the expression levels of SOX9 mRNA and Col2A1 mRNA in the chondrocytes in DMEM/F12+1%FBS+0.4 g·L-1 VC+1 μg·L-1 PLGA group were significantly increased (P<0.01). The immunofluorescence staining results showed that the green fluorescence signal of COLⅡ and the red fluorescence signal of SOX9 were observed in some chondrocytes in DMEM/F12+10%FBS group under fluorescence microscope, and the fluorescence intensity was weak. In DMEM/F12+1%FBS group, most chondrocytes exhibited COLⅡ green fluorescence signal and SOX9 red fluorescence signal, and the fluorescence intensity was significantly stronger than that in DMEM/F12+10% FBS group. In DMEM/F12+1% FBS+0.4 g·L-1 VC+1 μg·L-1 PLGA group, the COLⅡ green fluorescence signal and SOX9 red fluorescence signal were found in all the chondrocytes, and the fluorescence intensity was significantly higher than those in DMEM/F12+10%FBS and DMEM/F12+1%FBS groups. The expression levels of COLⅡ and SOX9 proteins in the chondrocytes in DMEM/F12+1%FBS group were significantly higher than those in DMEM/F12+10%FBS group, and the expression levels of COLⅡ and SOX9 proteins in the chondrocytes in DMEM/F12+1%FBS+0.4 g·L-1 VC+ 1 μg·L-1 PLGA group were significantly higher than those in DMEM/F12+10%FBS group. Conclusion The improved methods for the isolation and culture of primary chondrocytes of the rats can overcome the shortcomings of traditional methods, shorten the isolation time of primary chondrocytes, and improve the quality of in vitro culture of primary chondrocytes.

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Review
Research progress in molecular mechanism of hepatic fibrosis and related therapeutic targets
Zhaohui LIAO,Zhengyuan XIE
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1450-1456.  DOI: 10.13481/j.1671-587X.20240532
Abstract ( 46 )   HTML ( 2 )   PDF (422KB) ( 18 )  

Hepatic fibrosis (HF) is a common pathological repair response occurring after liver injury and is a critical stage in the progression of chronic liver diseases towards cirrhosis. The molecular mechanisms of HF occurrence are complex. Liver injury triggers the release of various cytokines by multiple cell types, and initiates the downstream signaling pathways to activate the hepatic stellate cells (HSCs) and transform them into myofibroblasts (MFBs). MFBs can release large quantities of extracellular matrix (ECM), thereby disrupt the normal liver architecture and lead to the occurrence and development of HF. The potential therapeutic targets for HF are still in the experimental animal phase, and there are no clinical applications yet. This review summarizes the signaling pathways and related factors involving HSCs and ECM in HF, such as the transforming growth factor-β (TGF-β)/Smad signaling pathway, platelet-derived growth factor (PDGF), matrix metalloproteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs), and connective tissue growth factor (CTGF). It also discusses the related therapeutic targets, and provids the theoretical basis for the development of new drugs for HF.

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Research progress in effect of histone acetylation modifications on occurrence and development of periodontitis
Yi XIE,Yang LIU,Hongyan LI,Xiaowei XU
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1457-1464.  DOI: 10.13481/j.1671-587X.20240533
Abstract ( 41 )   HTML ( 0 )   PDF (428KB) ( 13 )  

Periodontitis is a chronic inflammatory disease caused by plaque biofilm, and the homeostasis of the periodontal microenvironment is crucial for periodontal health. The epigenetics explores how the environmental and other non-genetic factors regulate the gene expression without changing the DNA nucleotide sequence, thereby affecting the occurrence and development of this disease. The histone acetylation modification is one of the common epigenetic modifications, primarily regulated by the histone acetyltransferases and histone deacetylases. Imbalance in this regulation can lead to the gene regulatory disorders, causing the chronic inflammation, autoimmune diseases and cancer. Recent studies have shown there is a clear correlation between histone acetylation modifications and the development of periodontitis. This paper discusses the changes in histone acetylation modifications in the gingival epithelial cells, immune cells, and periodontal ligament stem cells during periodontitis, elucidating the role and molecular mechanisms of histone acetylation modifications in the development of periodontitis and providing the basis for the epigenetic treatment of periodontitis.

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Research progress in mechanism of fibrosis regulated by macrophage polarization
Tan CHEN,Yan CHEN
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1465-1473.  DOI: 10.13481/j.1671-587X.20240534
Abstract ( 50 )   HTML ( 0 )   PDF (452KB) ( 11 )  

Organ structure destruction and functional decline leading to failure due to fibrosis pose severe threats to the human health and life. Macrophages are important immune cells present in various tissues and organs. Under the influence of numerous factors and pathways, such as transforming growth factor-β1(TGF-β1), Toll-like receptor 4(TLR4)/nuclear factor-κB(NF-κB), Janus kinase(JAK)/signal transducer and activator of transcription(STAT), Notch signaling pathway, peroxisome proliferator-activated receptor(PPAR), and cAMP response element-binding protein(CREB), the macrophages may undergo polarization. In visceral organ fibrosis, the macrophage polarization signaling network, composed of single or multiple factors and pathways, is one of the crucial mechanisms regulating fibrosis. After polarization, the macrophages produce the chemokines and matrix metalloproteinase(MMP), which are related to promoting fibrosis, leading to the occurrence and development of fibrosis. The current research conducted domestically and internationally mainly focuses on the roles of macrophages in infection, tumors, and fibrosis, with few summaries on the mechanisms of macrophage polarization in fibrosis. This review summarizes the pathways involved in macrophage polarization and the mechanisms of macrophage polarization in organ fibrosis, and provide the basis for the targeted therapy of fibrosis.

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Research progress in effect of CXC chemokine receptor 3 on occurrence and development of nervous system diseases
Wenhui LIU,Miao YU,Ying GUO,Yupeng LIU,Yang XING,Xinyu HONG,Jiale CUI
Journal of Jilin University(Medicine Edition). 2024, 50 (5):  1474-1480.  DOI: 10.13481/j.1671-587X.20240535
Abstract ( 32 )   HTML ( 0 )   PDF (410KB) ( 8 )  

Chemokines and their receptors play crucial roles in nervous system diseases, among which the CXC chemokine receptor 3 (CXCR3) is an important mediator of intercellular communication. CXCR3 not only participates in inflammatory chemotaxis and malignant behaviors of tumor cells but also regulates the neurologic diseases. CXCR3 and its ligands directly or indirectly contribute to neuroinflammation and neuroimmunity, and potentially serve as therapeutic targets for diseases like multiple sclerosis, Alzheimer’s disease, and glioma and so on. This review discusses the expression and impact of CXCR3 and its ligands in neurological diseases such as multiple sclerosis, neurologic tumors, neurodegenerative diseases, and neuropathic pain, as well as their associations with CXCR3 ligands. Understanding the mechanisms linking CXCR3 to these diseases could facilitate its potential as an early diagnostic biomarker and a target for drug intervention, and provide the basis of studying the occurrence and developmant of nervous system disease.

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