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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 May 2022, Volume 48 Issue 3
Research in basic medicine
Effect of pregnancy on cold defense-induced interscapular brown adipose tissue thermogenesis in famale rats and its mechanism
Xiaoyu HOU,Ya LI,Jiangyan XIE,Yian SONG,Jie ZHANG,Jianhui XU
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  553-560.  DOI: 10.13481/j.1671-587X.20220301
Abstract ( 378 )   HTML ( 9 )   PDF (966KB) ( 64 )  

Objective: To study the effect of pregnancy on the cold defense response in the female rats, and to analyze its mechanism.

Methods

A total of 42 adult female SD rats were selected and divided into non-pregnancy group (n=14), second trimester of pregnancy group (n=14) and third trimester of pregnancy group (n=14). The female rats in second trimester of pregnancy and third trimester of pregnancy groups were mated with the adult male rats, whereas the rats in non-pregnancy group were not. The rats in various groups were anesthetized with urethane and α-chloralose. The abdominal skin of the rats was cooled by ice-cold water for making skin cooling state to establish the cold defense response model. The abdominal skin temperature, rectum temperature and temperature of interscapular brown adipose tissue (iBAT) of the rats in various groups before and after skin cooling were recorded through multichannel temperature measuring system,and the abdominal skin temperature, rectum temperature, and the iBAT sympathetic nerve activity (iBAT SNA) of the rats in various groups before and after skin cooling were monitored by multichannel temperature measuring system and biological signal acquisition system.

Results

Compared with before skin cooling, the skin temperature of the rats in various groups after skin cooling was decreased (P<0.05), the rectum temperature of the rats in various groups after skin cooling had no significant differences(P>0.05). In addition, after skin cooling,there was no significant difference in the decreasing of skin temperature of the rats among various groups(P>0.05);there was no significant difference in the decreasing of rectum temperature of the rats among various groups(P>0.05).Compared with before skin cooling, the iBAT temperture of the rats in non-pregnancy and second trimester of pregnancy groups were increased after skin cooling (P<0.05), and there was no significant difference in iBAT temperature of the rats in third trimester of pregnancy group (P>0.05).After skin cooling, compared with non-pregnancy group, the increasing of iBAT temperature of the rats in second trimester of pregnancy group had no significant difference(P>0.05). After skin cooling, compared with non-pregnancy or second trimester of pregnancy groups, the increasing of iBAT temperature of the rats in third trimester of pregnancy group was decreased (P<0.05).Compared with before skin cooling,the iBAT SNA of the rats in non-pregnancy group and second trimester of pregnancy group after skin cooling were increased(P<0.05),and the iBAT SNA of the rats in third trimester of pregnancy group had no significant difference(P>0.05).After skin cooling,compared with non-pregnancy group,the increasing of iBAT SNA of the rats in second trimester of pregnancy group had no significant difference(P>0.05);compared with non-pregnancy group and second trimester of pregnancy group,the increasing of iBAT SNA of the rats in third trimester group was decreased(P<0.05).

Conclusion

The cold defense response in the non-pregnancy and second trimester of pregnancy rats is normal, whereas the cold defence response in the third trimester of pregnancy rats is reduced.This phenomenon is related to the normal cold defensive iBAT SNA in the non-pregnancy and second trimester of pregnancy rats, while the cold defensive iBAT SNA in the third trimester of pregnancy rats is inhibited.

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Effects of salvianolic acid B on atherosclerosis and efferocytosis of macrophages of mice and their mechanisms
Yifan ZHANG,Jie DING,Min DU,Xiaoteng FENG,Ping LIU
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  561-567.  DOI: 10.13481/j.1671-587X.20220302
Abstract ( 508 )   HTML ( 5 )   PDF (898KB) ( 194 )  
Objective

To investigate the effects of salvianolic arid B (Sal B) on atherosclerosis of the mice and oxidized low-density lipoprotein (ox-LDL)-induced efferocytosis of macrophages of the mice,and to clarify the mechanism of anti-atherosclerosis of Sal B.

Methods

Thirty-two male low-density lipoprotein receptor gene knockout(LDLR-/-) mice were randomly divided into control group, model group, Sal B group, and atorvastatin group,and there were 8 mice in each group. Except for control group (fed with ordinary feed), the mice in the other groups were fed with high-fat feed for 12 weeks.The mice in control group and model group were injected intraperitoneally with normal saline daily, the mice in Sal B group were injected intraperitoneally with Sal B solution, and the mice in atorvastatin group were administered with atorvastatin solution by gavage. The serum levels of total cholesterol(TC), triglyceride(TG),and low density lipoprotein-cholesterol(LDL-c)of the mice in various groups were detected. Oil red O staining method was used to observe the percentages of aortic atherosclerotic plaque area of the mice in various groups. The RAW 264.7 cells were divided into control group,ox-LDL induction group (induced by 40 mg·L-1 ox-LDL for 24 h), low, middle, and high doses of Sal B groups(given 1.25, 2.50,and 5.00 mg·L-1 Sal B). Real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting methods were used to detect the expression levels of AXL, MERTK, TYRO3,and milk fat globule epidermal growth factor 8(MFGE8)mRNA and proteins in arterial tissue of the mice and RAW264.7 cells in various groups.

Results

Compared with control group, the serum levels of TC, TG, and LDL-C of the mice in model group were increased (P<0.01), and the percentage of aortic atherosclerotic plaque area was increased (P<0.01),the expression levels of AXL, MERTK, TYRO3, and MFGE8 mRNA and proteins in arterial tissue were significantly decreased(P<0.05 or P<0.01). Compared with model group, the serum levels of TC, TG, LDL-C of the mice in Sal B and atorvastatin groups were decreased (P<0.01), and the percentages of aortic atherosclerotic plaque areas were decreased (P<0.01); the expression levels of AXL, MERTK, TYRO3 and MFGE8 mRNA and proteins in arterial tissue of the mice were increased (P<0.05 or P<0.01). Compared with control group, the expression levels of AXL, MERTK, TYRO3, MFGE8 mRNA and proteins in the RAW264.7 cells in ox-LDL group were decreased (P<0.05); compared with ox-LDL induction group, the expression levels of AXL, MERTK, TYRO3,and MFGE8 mRNA and proteins in the RAW264.7 cells in Sal B groups were increased (P<0.05 or P<0.01).

Conclusion

Sal B promotes the efferocytosis of the LDLR-/- mice and RAW264.7 cells by up-regulating the expressions of efferocytosis-related molecules, thereby exerting an anti-atherosclerotic effect.

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Improvement effect of recombinant human growth hormone on motor function in mice with focal cerebral ischemia- reperfusion and its mechanism
Tao LIN,Yinghua LI,Yawen LIAN,Xiaowei CHEN,Qi WAN
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  568-574.  DOI: 10.13481/j.1671-587X.20220303
Abstract ( 517 )   HTML ( 3 )   PDF (772KB) ( 149 )  
Objective

To observe the effect of recombinant human growth hormone on motor function in the mice with focal cerebral ischemia-reperfusion, and to explore its possible mechanism.

Methods

A total of 24 C57BL/6J mice were selected to establish the cerebral ischemia-reperfusion injury models by the modified Zea-Longa suture method, and they were randomly divided into growth hormone treatment group (cerebral ischemia-reperfusion+growth hormone, SG group, n=12) and control group (cerebral ischemia-reperfusion+saline, SS group, n=12). A total of 48 h after injury, the mice in SG group were injected subcutaneously with the recombinant human growth hormone at the dose of 1.4 mg·kg-1·d-1 for 14 d,and the mice in SS group were injected with saline, and the injection time, dose, location and times were consistent with the mice in SG group. Beam balance test was used to evaluate the motor coordination abilities of the mice before injury, 1 d after injury, and 16 d after injury;screen test was used to evaluate the muscle strengths of the mice, and the cylinder test was used to evaluate the utilization rates of the affected limb of the mice;the expression levels of synapsin1(SYN1) in peri-infarct region of the mice in two groups were detected by Western blotting method 16 d after injury.

Results

The beam balance test results showed that compared with before injury, the scores of motor coordination abilities of the mice in two groups 1 d after injury were decreased(P<0.01); the scores of motor coordination abilities of the mice in two groups 16 d after injury were higher than those 1 d after injury (P<0.01), and the score of motor coordination ability of the mice in SG group was higher than that in SS group (P<0.05). The screen test results showed that compared with before injury, the residence time when grasping the screen upside down of the mice in two groups was significantly shortened 1 d after injury (P<0.01), and the residence time of the mice in two groups 16 d after injury was longer than that 1 d after injury (P<0.01); compared with SS group, the residence time when grasping the screen upside down of the mice in SG group was significantly increased (P<0.05).The cylinder test results showed that the utilization rates of affected limb of the mice in two groups were decreased significantly 1 d after injury (P<0.01);compared with 1 d affter injury, the utilization rates of affected limb of the mice in two groups were increased 16 d after injury (P<0.01), and the utilization rate of the mice in SG group was higher than that in SS group (P<0.05). The Western blotting results showed that the expression level of SYN1 in peri-infarct region of the mice in SG group was significantly higher than that in SS group (P<0.05).

Conclusion

Recombinant human growth hormone can improve the motor function of the mice with focal cerebral ischemia-reperfusion, and its mechanism may be related to the promoting the expression level of SYN1 in peri-infarct region.

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Improvement effect of Panax ginseng glycoproteins on spermatogenesis of mice with oligoasthenozoospermia
Mengyao SHAN,Weigang WANG,Jinxiang DONG,Jianming TIAN,Lianlian SONG,Yinghong CHEN,Xiaoxue FANG,Zhidong QIU,Haoming LUO,Difu ZHU
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  575-583.  DOI: 10.13481/j.1671-587X.20220304
Abstract ( 548 )   HTML ( 2 )   PDF (1624KB) ( 74 )  
Objective

To investigate the improvement effect of Panax ginseng glycoproteins (PGG) on the spermatogenesis of the mice with oligoasthenozoospermia, and to preliminarily elucidate its mechanism.

Methods

Sixty healthy male mice were randomly divided into normal group, model group, positive control group (Shengjing capsule, 700 mg·kg-1), low dose of PGG group (100 mg·kg-1 PGG), medium dose of PGG group (200 mg·kg-1 PGG) and high dose of PGG group (400 mg·kg-1 PGG),and there were 10 mice in each group. Except for normal group, the mice in other groups were intraperitoneally injected with 30 mg·kg-1 cyclophosphamide to establish the oligoasthenozoospermia models.The sperm was collected from the mice in various groups. The total number, survival rate, malformation rate and motility of the sperm of the mice in various groups were examined under microscope. The morphology of the sperm of the mice in various groups was observed. The testis (double), prostate+seminal vesicle (double), preputial gland (double) and levator ani muscle of the mice in various groups were collected and weighed for the calculation of the organ coefficients. Hematoxylin-eosin (HE) staining was used to detect the pathomorphology of the testis tissue of the mice in various groups. The levels of serum testosterone (T), follicle stimulating hormone(FSH),and prolactin (PRL) of the mice in various groups were measured by ELISA method.

Results

Compared with normal group, the total number of sperm of the mice in model group was decreased (P<0.01), the survival rate of sperm was decreased (P<0.01), the malformation rate of sperm was increased (P<0.01), the motility of sperm was decreased (P<0.01), the organ coefficients of testis and prostate + seminal vesicle were decreased (P<0.05 or P<0.01), the organ coefficients of preputial gland and levator ani muscle had no significant differences (P>0.05), the spermatogenic cell layer thickness and spermatogenic cell layer cross-sectional area in seminiferous tubule were decreased (P<0.01),and the levels of serum T, FSH, and PRL were decreased (P<0.05 or P<0.01). Compared with model group, the total number of sperm of the mice in medium and high doses of PGG groups were increased (P<0.05), the survival rates of sperm were increased (P<0.01), the malformation rates of sperm were decreased (P<0.05 or P<0.01), the motilities of sperm were increased (P<0.05 or P<0.01), the organ coefficients had no significant differences(P>0.05), and the spermatogenic cell layer thickness and spermatogenic cell layer cross-sectional areas in seminiferous tubule were increased (P<0.05 or P<0.01); the levels of serum T of the mice in Shengjing capsule group and medium and high doses of PGG groups were increased (P<0.05 or P<0.01), the levels of serum FSH of the mice in Shengjing capsule group and different doses of PGG groups were increased (P<0.05 or P<0.01), and the levels of serum PRL of the mice in different doses of PGG groups were increased (P<0.01).

Conclusion

PGG has a dose-dependent effect; with the increasing of dosage, the efficacy is enhanced.PGG can obviously improve the spermatogenesis of the oligoasthenospermic mice, and its mechanism may be related to the increasing of serum sex hormone levels by PGG.

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Expression of androgen receptor co-regulator BAP18 in testicular cells of rats and its significance
Shiying SUN,Ge SUN,Lin LIN,Yue ZHAO
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  584-590.  DOI: 10.13481/j.1671-587X.20220305
Abstract ( 460 )   HTML ( 3 )   PDF (791KB) ( 72 )  
Objective

To observe the expression of bromodomin PHD domain transcription factor(BPTF) associated protein18( BAP18 )in testicular cells of the rats and its role in the androgen receptor(AR) signaling pathway, and to clarify the specific target gene of BAP18 directly regulating in the AR signaling pathway,and explore the biological capacity of BAP18 in the testicular cells.

Methods

The sertoli cells and leydig cells of the rats were collected. The expression levels of BAP18 and AR mRNA in the cells were detected by real time fluorescence quantitative PCR (RT- qPCR) method, the expression levels of BAP18 and AR proteins in the cells were detected by Western blotting method, and the interaction between BAP18 and AR in the testicular R2C cells was detected by protein immunoprecipitation assay (co-IP). The BAP18 over-expression plasmid with FLAG tag was used to over-express the BAP18 in the R2C cells(BAP18 over-expression group).Luciferase double gene report test was used to detect the luciferase activity in the cells. In control group, the pcDNA3 empty plasmid was used to transfect the R2C cells. The expression levels of BAP18, AR, STAR, HSD3B1, CYP17A1, and DDX25 mRNA in the R2C cells treated with testosterone(T) and dihydrotestosterone(DHT) before and after BAP18 over-expression were detected by RT-qPCR method. The expression levels of CYP17A1 protein in the R2C cells treated with T and DHT before and after BAP18 over-expression were detected by Western blotting method.

Results

BAP18 was expressed in the leydig cells and sertoli cells, but AR was mainly expressed in the leydig cells; BAP18 interacted with AR and up-regulated the AR-mediated gene transcription in the testicular R2C cells. Compared with control group, after over-expression of BAP18 under the action of T and DHT, the mRNA and protein expression levels of CYP17A1 in the R2C cells in BAP18 over-expression group were increased (P<0.01). After exogenous transfection of quantitative BAP18 expression plasmid in R2C cells at the same time, the luciferase activity in R2C cells was increased nearly 3 times under the action of DHT, while the luciferase activity in R2C cells was increased 1.5 times under the action of T. After BAP18 over-expression of in the R2C cells, the expression levels of AR, STAR, HSD3B1,and DDX25 mRNA in the R2C cells in BAP18 over-expression group were not significantly different from those before BAP18 over-expression(P>0.05), while the expression level of CYP17A1 mRNA was higher than that before BAP18 over-expression(P<0.05); after T or DHT treatment, the expression levels of CYP17A1 mRNA in the R2C cells were higher than before T or DHT treatment(P<0.05). After BAP18 over-expression in the cells, the expression levels of CYP17A1 protein in the cells after T or DHT treatment were significantly higher than before T or DHT treatment(P<0.05), but there was no significant difference in the expression level of DDX25 protein before and after T or DHT treatment(P>0.05).

Conclusion

BAP18 can interact with AR in the Leydig cells and regulate the important target gene CYP17A1 of AR signaling pathway, which may be involved in the physiological process of Leydig cells.

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Inhibitory effect of miR-152 on proliferation and invasion of endometrial carcinoma cells by reducing low-density lipoprotein receptor expression
Guowu WANG,Yuan YAO,Yu ZHANG,Na XU,Fang LIU
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  591-599.  DOI: 10.13481/j.1671-587X.20220306
Abstract ( 408 )   HTML ( 0 )   PDF (992KB) ( 158 )  

Objective: To investigate the expression and role of microRNA-152 (miR-152) in endometrial carcinoma (EC) tissue, and to provide the basis for study of pathogenesis and targeted treatment of EC.

Methods

The cancer tissues of 21 patients with EC (tumor group) and 39 patients with non-EC (control group) and human endometrial cancer RL95-2 cells and 293T cells were selected as the subjects. The clinical data such as age, height, body mass (BM), body mass index (BMI), systolic blood pressure (SBP), diastolic blood pressure (DBP), waist circumference (WC) and abdominal circumference (AC) of the patients in tumor group and control group were collected. The levels of triglyceride(TG),total cholesterol (TC), low-density lipoprotein (LDL), high density lipoprotein (HDL), Ki-67 and low-density lipoprotein receptor (LDLR) of the patients in two groups were measured.Bioinformatics method was used to predict the expression of target gene LDLR related to the proliferation and invasion downstream of miR-152. Real-time fluorescence quantitative PCR (RT-qPCR) method was used to detect the expression level of miR-152 in endometrial tissue of the patients in tumor group and control group, Western blotting method and immunohistochemistry were used to detect the expression levels of LDLR protein in endometrial tissue of the patients in tumor group and control group, and Person correlation analysis was used to analyze the correlations between the expression of LDLR mRNA and the general data, biochemical indexes of the patients and miR-152 expression. The RL95-2 cells were divided into empty plasmid group (transfected with miR-NC empty plasmid), miR-152 group (transfected with miR-152-mimics), miR-152-mimics+pcDNA3.1-vector group (simultaneously transfected with miR-152-mimics and pcDNA3.1-vector), and miR-152-mimics+pcDNA3.1-LDLR group (simultaneously transfected with miR-152-mimics+pcDNA3.1-LDLR). The proliferation rates of the EC cells in various groups were detected by CCK-8 method, and the number of invasive EC cells in various groups was detected by Transwell assay. The luciferase activities of the 293T cells in various groups were detected by double luciferase reporter gene experiment.

Results

The expression level of miR-152 in endometrial tissue of the patients in tumor group was significantly lower than that in control group (P<0.05), and the expression levels of LDLR mRNA and protein were higher than those in control group (P<0.05 or P<0.01). The results of dual-luciferase reporter gene assay showed that LDLR was the target gene of miR-152.The Person correlation analysis results showed that the expression of LDLR mRNA in endometrial tissue of the patients in tumor group had a positive correlation with Ki-67, BMI, TC, TG, BM,and WC (r=0.4490, r=0.4377, r=0.4472, r=0.4706, r=0.5882, r=0.5130, P<0.05), and it was negatively correlated with the expression of miR-152(r=-0.4378, P<0.05). Compared with empty plasmid group, the proliferation rate of the RL95-2 cells in miR-152 group was decreased (P<0.05), and the number of invasive cells was decreased (P<0.05); compared with miR-152-mimics+pcDNA3.1-vector group, the proliferation rate of the cells in miR-152-mimics+pcDNA3.1-LDLR group was increased (P<0.01), and the number of invasive cells was increased (P<0.01).

Conclusion

MiR-152 inhibits the proliferation and invasion of the EC cells by decreasing the expression of LDLR.

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Analgesic effect of baicalein on neuropathic pain of rats and its mechanism
Xiaohua LIU,Bingying FANG,Man HAN,Haifa QIAO,Yuanwang YU
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  600-605.  DOI: 10.13481/j.1671-587X.20220307
Abstract ( 429 )   HTML ( 2 )   PDF (822KB) ( 97 )  
Objective

To observe the inhibitory effect of baicalein on the pain behavior caused by chronic sciatic nerve compression in the rats and the regulation effect of glial cell-derived neurotrophic factor (GDNF) expression in the primary sensory neurons, and to clarify the analgesic mechanism of baicalein in the neuropathic pain.

Methods

A total of 48 healthy male SD rats were randomly divided into sham operation group, model group and baicalein group,and there were 16 rats in each group. After anesthesia,the sciatic nerve of the rats in sham operation group was exposed, and the chronic constricting nerve injury of the rats in model group and baicalein group was performed to establish the chronic constriction injury(CCI) models. The rats in baicalein group were injected with baicalein continuously for 7 d after successful modeling. The mechanical withdraw threshold (MWT) and thermal withdrawal latency (TWL) of the rats in various groups were measured at different time points before and after operation to evaluate the pain behavior caused by nerve injury. On the 11th day,6 rats were randomly selected from each group for immunofluorescence staining to determine the number of C-Fos positive neurons in spinal dorsal horn tissue of the rats in various groups. On the 28th day, the fluorescence intensities of GDNF in dosal root ganglion(DRG) of the rats in various groups were detected.

Results

There was no significant difference between MWT and TWL of the rats in various groups before operation (P>0.05). Compared with sham operation group, the MWT and TWL of the rats in model group and baicalein group were decreased significantly 1 d after operation (P<0.05), and they were increased in baicalein group 3 d after administration (P>0.05). Compared with sham operation group, the number of c-Fos positive neurons in spinal dorsal horn tissue of the rats in model group was increased (P<0.05); compared with model group, the number of c-Fos positive neurons in spinal dorsal horn tissue of the rats in baicalein group was decreased (P<0.05). Compared with sham operation group, the fluorescence intensity of GDNF in DRG of the rats in model group was decreased (P<0.05);compared with model group, and the fluorescence intensity of GDNF in DRG of the rats in baicalein group was increased (P<0.05).

Conclusion

Baicalein has the analgesic effect on neuropathic pain of the rats.The GDNF in primary sensory neurons is involved in the neuropathic pain caused by sciatic nerve injury. The analgesic effect of baicalein on the neuropathic pain may be related to its regulation of GDNF expression.

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Effect of autophagy induced by exosomes derived from bone marrow mesenchymal stem cells on survival of SH-SY5Y cells inhibited by MPP+ and its mechanism
Wentao WANG,Xuguang MI,Yang ZHOU,Wenxing PU,Jiaxu GAO,Meng JING,Fankai MENG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  606-614.  DOI: 10.13481/j.1671-587X.20220308
Abstract ( 590 )   HTML ( 0 )   PDF (1413KB) ( 70 )  
Objective

To investigate the effect of autophagy induced by exosomes (exo) derived from the bone marrow mesenchymal stem cells (BMSCs) on the survival of the human neuroblastoma SH-SY5Y cells inhibited by 1-methyl-4-phenylpyridine ion (MPP+), and to darify its mechanism.

Methods

The human neuroblastoma SH-SY5Y cells were given different concentrations(0,0.25,0.50,1.00,and 2.00 mmol·L-1)MMP+,and the survival rates of the SH-SY5Y cells were detected after 24 and 48 h.The experiment was divided into control group[given phosphate buffered saline(PBS)], 0.50 mmol·L-1 MPP+ group (given 0.50 mmol·L-1 MPP+),MPP++BMSCs supernatnt (BMSCs-Sup)group (given 0.50 mmol·L-1 MPP++BMSCs-Sup),0.50 mmol·L-1 MPP++BMSCs-Exo group (given 0.50 mmol·L-1 MPP+ +100 mg·L-1 BMSCs-Exo),MPP+ +rapamycin(Rap) group (given 0.50 mmol·L-1 MPP+ +2 mmol·L-1 Rap) and MPP+ +BMSCs-Exo+3-methyladenine(3-MA) group(given 0.50 mmol·L-1 MPP+ +100 mg·L-1 BMSCs-Exo +1 mmol·L-1 3-MA). The survival rates of the SH-SY5Y cells in various groups were detected by MTT assay.The fluorescence intensities of autophagic acid vesicles in the SH-SY5Y cells in various groups were detected by acridine orange staining under confocal microscope.The expression levels of CD63, TGS101,and Calnexin proteins in the SH-SY5Y cells in various groups were detected by Western blotting method.

Results

The MTT assay results showed that the survival rates of the SH-SY5Y cells were decreased gradually with the increasing of the MPP+ concentration or the extension of treatment time (P<0.05) in a concentration- and time-dependent manner. Compared with MPP+ group, the survival rates of the SH-SY5Y cells in MPP+ + BMSCs-Sup group, MPP+ +BMSCs-Exo group,and MPP+ + Rap group were increased (P<0.05). Compared with MPP+ +BMSCs-Exo group, the survival rate of the SH-SY5Y cells in MPP+ +BMSCs-Exo + 3-MA group was decreased (P<0.05).The acridine orange staining results showed that compared with control group, the fluorescence intensities of autophagic acid vesicles in the SH-SY5Y cells in MPP+ + BMSCs-Sup group, MPP+ +BMSCs-Exo group and MPP+ + Rap group were increased (P<0.05); compared with MPP+ +BMSCs-Exo group, the fluorescence intensity of autophagic acid vesicles in the SH-SY5Y cells in MPP+ +BMSCs-Exo + 3-MA group was decreased (P<0.05).The Western blotting results showed that the exosome marker proteins CD63 and TGS101 were highly expressed, while the endoplasmic reticulum marker protein Calnexin was lowly expressed.

Conclusion

BMSCs-Exo can resist the MPP+-induced decreasing of the survival of the SH-SY5Y cells by activating the autophagy.

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Repair effect of new nHA/PLGA nanocomposite microspheres on skull defects of rats
Jie PAN,Dezhou WANG,Wenzhi SONG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  615-621.  DOI: 10.13481/j.1671-587X.20220309
Abstract ( 472 )   HTML ( 2 )   PDF (955KB) ( 192 )  
Objective

To investigate the repair effect of new nano-hydroxyapatite (nHA)/polylactide-glycolide (PLGA) nanocomposite microspheres on the skull bone defects of the rats, and to provide the basis for the clinical application of the material.

Methods

Two symmetrical critical bone defect models with a diameter of 5 mm were prepared in the skulls of 9 SD rats with a dental low-speed handpiece, and they were randomly divided into experimental group (implanted with nHA/PLGA nanocomposite microspheres), control group (implanted with Bio-Oss particles) and blank group (only the defect was prepared without filling material),and there were 6 skull defect models in each group. The CT three dimensional reconstruction of skull was performed on the right postoperative day, 4 weeks after surgery and 8 weeks after surgery to check the repair effect of skull defect.The rats were sacrificed at the 8th week,and micro-computed tomography (micro-CT) scanning was used to scan the three-dimensional reconstruction, and the number of trabecular bone(Tb.N ), trabecular bone spacing (Tb.Sp), trabecular bone thickness (Tb.Th), and the ratios of new bone volume/total bone volume (BV/TV)in skull defect area of the rats in various groups were detected.

Results

The naked eye observation results showed that the boundary of the skull defect area of the rats in experimental group was clear, and new bone could be seen in the intergranular space; in control group, there was granular bone powder accumulation in the skull defect area without obvious absorption; in blank group, there was obvious depression in the central area of skull defect without obvious bone deposition.The CT three-dimensional reconstruction results showed that 8 weeks after sugery,compared with blank group, the rats in experimental group had obvious new bone formation; compared with control group, the rats in experimental group had obvious new bone deposition.The micro-CT detection results showed that 8 weeks after sugery, compared with control group, there were no significant differences in the Tb.N, Tb.Sp and Tb.Th in skull defect area of the rats in experimental group (P>0.05), and the BV/TV was decreased (P<0.05); compared with blank group, there were significant differences in the Tb.Sp, Tb.Th and BV/TV in skull defect area of the rats in experimental group and control group (P<0.05).

Conclusion

nHA/PLGA nanocomposite microspheres have a good repair effect on the bone defects.

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Inhibitory effect of valproic acid combined with X-ray irradiation on proliferation of breast cancer MDA-MB-231 cells and its mechanism
Guanhu LI,Qingxu LANG,Chunyan LIU,Qin LIU,Mengrou GENG,Xiaoqian LI,Zhenqi WANG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  622-629.  DOI: 10.13481/j.1671-587X.20220310
Abstract ( 364 )   HTML ( 2 )   PDF (1053KB) ( 401 )  
Objective

To observe the inhibitory effect of valproic acid (VPA), a histone deacetylase inhibitor (HDACi), on the proliferation of the human triple negative breast cancer (TNBC)MDA-MB-231 cells alone or combined with X-ray irradiation, and to analyze the effect of combined irradiation on the apoptosis and autophage of the MDA-MB-231 cells.

Methods

The MDA-MB-231 cells at logarithmic growth phase were divided into control group, different concentrations(2.5,5.0,10.0,20.0, and 40.0 mmol·L-1) of VPA groups 4 Gy, X-ray irradiation groups,and different concentrations (2.5,5.0,10.0,20.0, and 40.0 mmol·L-1) of VPA combined with 4 Gy X-ray irradiation groups. The proliferation rates of the cells in various groups were detected by CCK-8 method after treated for 24, 48, and 72 h.The cells were divided into control group, 4 Gy X-ray irradiation group, 5 mmol·L-1 VPA group, 5 mmol·L-1 VPA combined with 4 Gy X-ray irradiation group, the apoptotic rates and autophage rates of the cells in various groups were analyzed by fiow cytometry after treated for 24 and 48 h.

Results

Compared with control group, the proliferation rates of the MDA-MB-231 cells in VPA group and VPA combined with 4 Gy X-ray irradiation groups were decreased significantly in a concentration and time-dependent manner (P<0.01). Compared with 4 Gy X-ray irradiation group, the proliferation rates of the MDA-MB-231cells in VPA combined with 4 Gy X-ray irradiation groups were significantly decreased (P<0.01). Compared with 2.5 mmol·L-1 VPA group, the proliferation rate of the MDA-MB-231 cells in 2.5 mmol·L-1 VPA combined with 4 Gy X-ray irradiation group after treated for 24 h was decreased(P<0.05). After treated for 72 h, the proliferation rates of the MDA-MB-231 cells in 2.5 and 10.0 mmol·L-1 VPA combined with 4 Gy X-ray irradiation groups were lower than those in 2.5 and 10.0 mmol·L-1 VPA groups(P<0.05),respectively. After treated for 24 and 48 h, the apoptotic rates and autophagy rates of the MDA-MB-231 cells in 4 Gy X-ray group, 5 mmol·L-1 VPA group and 5 mmol·L-1 VPA combined with 4 Gy X-ray group were significantly higher than those in control group (P<0.05 or P<0.01). Compared with 4 Gy X-ray irradiation group, the apoptotic rate and autophagy rate of the MDA-MB-231 cells in 5 mmol·L-1 VPA combined with 4 Gy X-ray irradiation group were significantly increased (P<0.01). After treated for 48 h,compared with 5 mmol·L-1 VPA group,the autophagy rate of the MDA-MB-231 cells in 5 mmol·L-1 VPA combined with 4 Gy X-ray irradiation group was increased (P<0.05).

Conclusion

VPA can inhibit the proliferation of MDA-MB-231 cells in a concentration and time-dependent manner, and the combined inhibitory effect of VPA and X-ray is better, and the mechanism may be involved in the increasing of apoptosis and autophage of the MDA-MB-231 cells induced by the combination of VPA and X-ray.

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Resistance rates of Helicobacter pylori to clarithromycin, metronidazole, and levofloxacin and mutation characteristics of their related drug resistance genes
Shunjia SONG,Xinying WANG,Feifei JIANG,Huijian JIA,Yuan ZHAO,Zhiping YANG,Liyuan SUN,Yundong ZHAO
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  630-637.  DOI: 10.13481/j.1671-587X.20220311
Abstract ( 742 )   HTML ( 6 )   PDF (1241KB) ( 164 )  

Objective: To detect the drug resistance rates of Helicobacter pylori (Hp) to three common antibiotics,and to explore the mutation forms of drug resistance genes in order to provide the basis for elucidating the molecular mechanism of drug resistance.

Methods

The gastric mucosa samples from 35 patients with chronic gastritis or peptic ulcer were isolated and cultured to identify Hp.The minimal inhibitory concentrations (MIC) of Hp to clarithromycin, metronidazole and levofloxacin were detected by the agar dilution method and Hp was divided into sensitive group and resistant group according to the results of the drug sensitivity test.The clarithromycin resistance gene 23S rRNA, metronidazole resistance gene rdxA, and levofloxacin resistance gene gyrA were amplified by PCR method. The nucleotide and amino acid mutations were analyzed after the sequencing of PCR purified product.

Results

After cultured for 3-5 d, smooth, translucent, needle-like colonies could be seen on the plate of Hp strain. All 35 Hp strains were resistant to metronidazole and levofloxacin, the resistance rate to clarithromycin was the highest(80%). Compared with the sensitive strains, all the clarithromycin- resistant strains had gene mutations. The most common mutation site of the 23S rRNA gene was A2143G, followed by mutation sites A2223G and A2142G+A2164G. The rdxA gene of metronidazole resistant strains mainly had missense mutations, followed by frameshift mutations and nonsense mutations. The N87K mutation in the gyrA gene of levofloxacin resistant strains was the highest, followed by the D91N mutation, N87I, D91G, and N87K+D143E mutations,but 17.1% of the resistant strains did not have any site mutations.

Conclusion

The resistance rates of Hp to clarithromycin, metronidazole, and levofloxacin are high, and the mutations of related drug resistance genes are the main cause of the antibiotic resistance. Screening these mutations may help to prevent the antibiotic resistance and select the appropriate antimicrobials to eradicate Hp.

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Protective effect of lidocaine on PC12 cells in Parkinson’s disease model and its mechanism
Xiaochen HUANG,Hao LI,Baohua WANG,Kai LI
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  638-647.  DOI: 10.13481/j.1671-587X.20220312
Abstract ( 467 )   HTML ( 0 )   PDF (1299KB) ( 113 )  
Objective

To investigate the protective effect of lidocaine on the PC12 cells in Parkinson’s disease(PD) model induced by 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine (MPTP), and to elucidate its possible molecular mechanism.

Methods

CCK-8 method was used to determine the optimal concentration and time of MPTP with different concentrations (0.2, 0.4, 0.8, 1.6, and 3.2 mmol·L-1). CCK-8 method was used to determine the optimal concentration and time of lidocaine with different concentrations (0.001, 0.010, 0.100, 0.200, 0.400, 0.600, 0.800, and 1.000 g·L-1), and Western blotting method was used to further determine the optimal concentration of lidocaine. The PC12 cells were divided into control group, MPTP group(0.8 mmol·L-1 MPTP for 24 h), MPTP+ lidocaine group(given 0.8 mmol·L-1 MPTP for 24 h, followed by 0.6 g·L-1 lidocaine for 6 h), MPTP+ lidocaine +DKK1 group(given 0.8 mmol·L-1 MPTP for 24 h, followed by 0.6 g·L-1 lidocaine and 100 μg·L-1 DKK1 for 6 h), and MPTP+DKK1 group(given 0.8 mmol·L-1 MPTP for 24 h, followed by 100 μg·mL-1 DKK1 for 6 h). Flow cytometry was used to detect the levels of reactive oxygen species (ROS) in the PC12 cells in various groups. The expression levels of interleukin-1β(IL-1β), interleukin-6(IL-6),and tumor necrosis factor-α(TNF-α) in supernatant of the PC12 cells in various groups were detected by ELISA assay;real-time fluorescence quantitative PCR (RT-qPCR) method was used to detect the expression levels of Wnt ligand protein 1 (Wnt1),β-catenin,and glycogen synthase kinase-3β(GSK-3β) mRNA in the PC12 cells in various groups.The expression levels of Wnt1, β-catenin, GSK-3β, phosphorylated GSK-3β (p-GSK-3β), and cysteine aspartate protease 3 (caspase 3) proteins in the PC12 cells in various groups were detected by Western blotting method.

Results

Compared with control group, the levels of ROS, IL-1β, IL-6 and TNF-α,and the expression levels of GSK-3β mRNA, and the expression levels of GSK-3β, p-GSK-3β, caspase 3 proteins in the PC12 cells in MPTP group were significantly increased(P<0.05), and the expression levels of Wnt1, β-catenin mRNA and proteins were significantly decreased(P<0.05). Compared with MPTP group, the levels of ROS, IL-1β, IL-6, and TNF-α and the expression levels of GSK-3β mRNA, and the expression levels of GSK-3β, p-GSK-3β, and caspase 3 proteins in the PC12 cells in MPTP+lidocaine group were significantly decreased(P<0.05), however, the expression levels of Wnt1 and β-catenin mRNA and proteins were significantly increased(P<0.05).

Conclusion

Lidocaine can significantly reduce the production of ROS and inhibit the release of pro-inflammatory cytokines in the PC 12 cells, and also significantly down-regulate the expression of apoptotic protein caspase 3, thus playing a protective role in the MPTP-induced PD model PC12 cells; its mechanism may be related to the Wnt /β-catenin signaling pathway.

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Therapeutic effect of adipose-derived mesenchymal stem cells on premature ovarian failure model rats and its mechanism
Peiyu YAN,Aichen ZHANG,Hong ZHANG,Yang LI,Mengmeng ZHANG,Mengze LUO,Ying PAN
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  648-656.  DOI: 10.13481/j.1671-587X.20220313
Abstract ( 500 )   HTML ( 0 )   PDF (1721KB) ( 109 )  
Objective

To explore the biological characteristics of adipose-derived mesenchymal stem cells (ADMSCs), and to clarify the therapeutic effect of ADMSCs in the premature ovarian failure (POF) model rats.

Methods

The ADMSCs from SD rats were isolated and cultured, and the morphology of cells was observed under optical microscope. The osteogenic and adipogenic differentiations of ADMSCs were induced by osteogenic and adipogenic cell induction reagents. The expressions of surface antigen markers CD44, CD90,and CD45 of ADMSCs were detected by flow cytometry.The growth of ADMSCs was determined by CCK-8 assay and the growth curves were drawn. The models of PDF were established by intraperitoneal injection of cisplatin.Thirty SD rats were randomly divided into normal control group (n=10) and modeling group (n=20). The rats in modeling group were randomly divided into model group and ADMSCs treatment group, and there were 10 rats in each group.The rats in ADMSCs treatment group were injected with allogeneic ADMSCs through the tail vein every 3 d; the rats in normal control group and model group were injected with normal saline every 3 d through the tail vein. The body weights of the rats in various groups were detected, and the levels of estradiol (E2), follicle stimulating hormone (FSH), and luteinizing hormone (LH) in serum of the rats in various groups were detected by ELISA mehod. The pathomorphology of ovary tissue of the rats in various groups was observed by HE staining.

Results

The ADMSCs were adherent growth cells, which grew vigorously and could differentiate into the osteogenic and adipogenic cells. The expressions of CD44 and CD90 in the ADMSCs were highly, and the expression rates were 75.85% and 59.71%; the expression of CD45 was low, and the expression rate of 39.12%. Compared with normal control group, the body weight of the rats in model group was significantly decreased (P<0.05); the serum E2 level was significantly decreased (P<0.05), and the levels of FSH and LH were significantly increased (P<0.05), lacking of follicles at all levels, and the number of atretic follicles was increased. Compared with model group, the serum E2 level of the rats in ADMSCs treatment group was increased (P<0.05), the FSH and LH levels were decreased (P<0.05); the number of follicles was increased,and the number of atretic follicles was decreased.

Conclusion

ADMSCs can grow and reproduce in vitro, and have strong growth and proliferation abilities and multi-directional differentiation abilities; ADMSCs can improve the serum E2 level and ovarian function of the POF model rats, and have a certain therapeutic effect in the POF model rats.

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Effect of silencing helicase BLM gene on chemotherapy sensitivity of irinotecan in colorectal cancer cells and its mechanism
Qiuting CAO,Jingchun HAN,Xiaofei ZHANG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  657-667.  DOI: 10.13481/j.1671-587X.20220314
Abstract ( 643 )   HTML ( 2 )   PDF (2420KB) ( 417 )  
Objective

To investigate the effect of silencing the Bloom’s syndrome helicase(BLM) expression on the chemotherapy sensitivity of irinotecan (CPT-11) in the colorectal cancer (CRC) cells, and to elucidate its related mechanism.

Methods

The HT-29, Lovo, HCT-116, RKO, and DLD1 cells were cultured in vitro. The RKO and DLD1 cells with high expressions of BLM mRNA and protein were screened by real-time fluorescence quantitative PCR(RT-qPCR) and Western blotting methods. The cells were infected with lentivirus vector carrying shRNA to silence the expression of BLM in the RKO and DLD1 cells. The survival rates of the CRC cells and the half inhibitory concentration (IC50) values of CPT-11 in various groups after transfection were detected by CCK-8 assay. The RKO or DLD1 cells were divided into LV-shNC group (infected with LV-shNC), CPT-11+LV-shNC group ( infected with LV-shNC and treated with CPT-11) and CPT-11+LV-shBLM group (infected with LV-shBLM and treated with CPT-11). The percentages of the cells in various groups at different cell cycles were detected by flow cytometry. The expression levels of BCL2 associated death promoter(Bad),cleaved of cysteinyl aspartate specific proteinase(cleaved caspase-3), cyclin-dependent protein kinase 4(CDK4),cyclin-dependent protein kinase 6 (CDK6),and cyclin-dependent kinase inhibitor 1A(p21) proteins in the CRC cells in various groups were detected by Western blotting method; the apoptotic rates of the cells in various groups were detected by flow cytometry.The tumor volume and positive expression rates of BLM, Ki-67,and TUNEL in tumor tissue of the CRC-bearing mice were detected by constructing the allograft tumor model.

Results

Compared with the HT-29, Lovo or HCT-116 cells, the expression levels of BLM mRNA and protein in the RKO and DLD1 cells were increased (P<0.05);compared with RKO or DLD1 cells infected with LV-shNC, the expression levels of BLM mRNA in the RKO or DLD1 cells infected with LV-shBLM lentivirus were decreased(P<0.05). Compared with LV-shNC group, the survival rates and IC50 values of the cells in CPT-11+LV-shNC group and CPT-11+LV-shBLM group were decreased(P<0.05).Compared with LV-shNC group, the percentages of the cells at G0/ G1 phase and S phase and the apoptotic rates of the cells in CPT-11+LV-shNC group and CPT-11+LV-shBLM group were increased (P<0.05), while the percentages of the cells at G2 / M phase were decreased (P<0.05),and the expression levels of Cyclin D1, CDK4, CDK6 and Bcl-2 proteins were decreased(P<0.05), while the expression levels of p21, Bax, Bad,and cleaved caspase-3 proteins were increased (P<0.05);compared with CPT-11+LV-shNC group, the percentages of the cells at G0/G1 phase and S phase and the apoptotic rate of the cells in CPT-11+LV-shBLM group were increased (P<0.05), while the percentage of the cells at G2/M phase was decreased (P<0.05),and the expression levels of Cyclin D1,CDK4, CDK6,and Bcl-2 proteins were decreased(P<0.05), while the expression levels of p21, Bax, Bad,and cleaved caspase-3 proteins were increased (P<0.05). The nude mice transplanted tumor experiment results showed that compared with LV- shNC group, the tumor volumes of the nude mice and the positive expression rates of BLM and Ki-67 in the cells in CPT-11+LV-shNC group and CPT-11+LV-shBLM group were decreased (P<0.05), and the positive expression rates of TUNEL were increased(P<0.05); compared with CPT-11+LV-shNC group, the tumor volume of the nude mice in CPT-11+LV -shBLM group was decreased (P<0.05), and the positive expression rates of BLM and Ki-67 in tumor tissue were decreased (P<0.05), and the positive expression rate of TUNEL was increased(P<0.05).

Conclusion

Silencing BLM expression in the CRC cells may promote CPT-11-induced apoptosis by inhibiting the cycle progression of tumor cells, and ultimately improve chemotherapy resistance of CRC cells in vivo and in vitro.

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Protective effect of polyphyllin on acute lung injury in rats with severe acute pancreatitis by inhibiting NF-κB signaling pathway
Zhaohui WAN,Liang ZENG,Hui ZHOU,Jing LI,Changcheng LEI
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  668-675.  DOI: 10.13481/j.1671-587X.20220315
Abstract ( 560 )   HTML ( 0 )   PDF (1265KB) ( 117 )  
Objective

To observe the protective effect of polyphyllin Ⅶ (PP Ⅶ) on acute lung injury of the rats with severe acute pancreatitis (SAP), and to explore its possible mechanism.

Methods

Seventy-five rats were randomly divided into sham operation group, model group, dexamethasone group (positive control, given 2 mg·kg-1 dexamethasone ), low dose of PP Ⅶ group (given 50 mg·kg-1 PP Ⅶ), and high dose of PP Ⅶ group (given 150 mg·kg-1 PP Ⅶ), and there were 15 rats in each group. Except sham operation group,the SAP rat models were established by retrograde biliopancreatic tube injection of 5% sodium sulfonate solution in the rats in the other groups.The rats were immediately treated with the corresponding drugs. The arterial blood oxygenation index (OI) and wet weight/dry weight (W/D) ratio of lung tissue of the rats in various groups were measured. The levels of lipase and amylase in serum and the levels of interleukin1β (IL-1β),interleukin 6 (IL-6),andtumor necrosis factor(TNF-α)in bronchoalveolar lavage fluid (BALF) of the rats in various groups were detected by ELISA method; HE staining was used to observe the pathomophology of pancreas tissue and lung tissue of the rats in various groups,and the expression levels of nuclear factor-κB(NF-κB) signaling pathway related proteins in lung tissue of the rats in various groups were detected by Western blotting method.

Results

Compared with sham operation group, the pancreas tissue and lung tissue of the rats in model group were severely damaged,the OI value of arterial blood was significantly decreased (P<0.01), while the W/D ratio of lung tissue, levels of lipase and amylase in serum, levels of IL-1β, IL-6 and TNF-α in BALF, and the expression levels of inhibitor of nucler factor-κB α(IκBα) and phosphorylated NF-κB(p-NF-κB) p65(Ser536) proteins in lung tissue were significantly increased (P<0.01). Compared with model group, the damage degree of pancreas tissue and lung tissue of the rats in dexamethasone group and high dose of PP Ⅶ group was significantly improved, the OI values of arterial blood were significantly increased (P<0.01), and the W/D ratios of lung tissue, levels of lipase and amylase in serum,levels of IL-1β, IL-6 and TNF-α in BALF, and the expression levels of IκBα and p-NF-κB p65(Ser536) proteins in lung tissue were significantly decreased (P<0.01).

Conclusion

PP Ⅶ may play a protective role in acute lung injury of the SAP rats by inhibiting the NF-κB signaling pathway.

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Improvement effect of luteolin on acute respiratory distress syndrome by inhibiting ROS/TXNIP/NLRP3 signaling pathway activation in mice
Haixin QU,Erwei YUAN,Weiping GUO,Yajing ZHANG,Wenxia MA,Dan WU
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  676-683.  DOI: 10.13481/j.1671-587X.20220316
Abstract ( 444 )   HTML ( 5 )   PDF (866KB) ( 141 )  
Objective

To investigate the improvement effect of luteolin in the mice with acute respiratory distress syndrome (ARDS) by regulating the activation of reactive oxygen species (ROS)/thioredoxin binding protein (TXNIP)/NOD like receptor associated protein 3 (NLRP3) signaling pathway.

Methods

A total of 60 SD mice were randomly divided into control group, model group, luteolin (20 mg·kg-1) group, pyrogallol(PG) (75 mg·kg-1) group, luteolin (20 mg·kg-1) + PG(75 mg·kg-1) group, and there were 12 mice in each group.Besides control group, the mice in the other groups were instilled with lipopolysaccharide(LPS) solution through the trachea to establish the ARDS models, and the mice in control group were instilled with the same amount of normal saline through the trachea. The wet weights and dry weights of right lung tissue of the mice in various groups were weighed, and ratios of the wet weight/dry weight (W/D) of the mice in various were caculated; the arterial blood oxygen pressure (PaO2) was measured; the pathomorphology of lung tissue of the mice in various groups was observed by HE staining; the inspiratory resistance (Ri), minute ventilation (MV) and peak expiratory flow rate (PEF) of the mice in various groups were detected; the levels of ROS, glutathione peroxidase (GSH-Px), catalase (CAT) in lung tissue and levels of serum interleukin-18 (IL-18),interleukin-1β(IL-1β) and tumor necrosis factor α(TNF-α) of the mice in various groups were determined; the expression levels of TXNIP, caspase-1, caspase-4 and NLRP3 proteins in lung tissue of the mice in various groups were detected by Western blotting method.

Results

Compared with control group, the alveolar walls of the mice in model group became thicker, the alveoli collapsed and deformed, the pulmonary interstitial edema was accompanied by a large number of inflammatory cell infiltration, and the lung tissue had the severe pathological damage; the W/D ratio, Ri, serum IL-18, IL-1β and TNF-α levels, the level of ROS and the expression levels of TXNIP, caspase-1, caspase-4, and NLRP3 proteins in lung tissue were increased (P<0.05),and the MV, PEF, PaO2, levels of GSH-Px and CAT in lung tissue were decreased (P<0.05). Compared with model group, the pathological damage of lung tissue of the mice in luteolin group was improved, the W/D ratio, Ri, serum IL-18, IL-1β and TNF-α levels, ROS level and the expression levels of TXNIP, caspase-1, caspase-4 and NLRP3 proteins in lung tissue were significantly decreased (P<0.05), and the MV, PEF, PaO2, and GSH-Px and CAT levels in lung tissue were increased (P<0.05); the pathological damage of lung tissue of the mice in PG group was increased, the W/D ratio, Ri, serum levels of IL-18, IL-1β, and TNF-α,the ROS level and the expression levels of TXNIP, caspase-1, caspase-4,and NLRP3 proteins in lung tissue were increased (P<0.05),and the MV, PEF, PaO2,GSH-Px, and CAT levels in lung tissue were decreased (P<0.05).

Conclusion

Luteolin can reduce the lung inflammation and oxidative stress, improve the lung injury in the ARDS mice, and repair their lung functions by inhibiting the ROS/TXNIP/NLRP3 signaling pathway activation.

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Effect of Schwann cell-like cells on growth of neurites and expression of nerve growth factor in dorsal root ganglion cells of rats and their mechanisms
Yuanliang DU,Wang REN,Lin LIU,Shuodan HU,Mingyu LIU,Pengfei DU,Xiumei FU
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  684-691.  DOI: 10.13481/j.1671-587X.20220317
Abstract ( 625 )   HTML ( 4 )   PDF (1698KB) ( 156 )  
Objective

To investigate the effect of Schwann cell-like cells (SCLCs) on the growth of neurites and the expression of nerve growth factor (NGF) of dorsal root ganglion (DRG) cells in the rats, and to clarify the role of SCLCs on promoting the growth in the DRG cells and its mechanism.

Methods

The adipose-derived stem cells (ADSCs) were isolated and cultured,and the calcified nodules in the cells were detected by Alizarin red staining to observe their differentiation ability, and the ADSCs were induced to differentiate into the SCLCs.The DRG cells were isolated and cultured, and the expression of neuronal nuclear antigen (NeuN) protein in the DRG cells was detected by immunohistochemical staining. The co-culture system of SCLCs and DRG cells was established. The cells were divided into single culture group and co-culture group. HE staining was used to observe the morphology of the DRG cells in two groups, the number of DRG cells and the number of DRG cell processes in two groups were counted, immunohistochemical staining was used to detect the expression of NGF protein in the DRG cells in two groups, and Western blotting method was used to detect the expression level of NGF protein in the DRG cells in two groups.

Results

The primary ADSCs were cultured for 7 d, under the inverted microscope, a large number of cells with large volume and long spindle shape could be seen, which were similar to the arrangement of vortex or paving stone.The Alizarin red staining results showed that the calcified nodules can be seen under the microscope, and the cultured cells had the ability of differentiation. The immunofluorescence staining results showed that most cells were S100 staining positive, and the ADSCs could be successfully induced into the SCLCs. After cultured for 72 h, the processes of DRG cells were thin and long, arranged in a wreath-like arrangement. After 7-8 d, the number of cells was increased, the body of cells was large and round, and the processes were slender and connected like a fence. The immunohistochemical staining results showed that almost all the nuclei were marked as brown and NeuN staining positive. Compared with single culture group, the number of DRG cells and the number of DRG cell processes in co-culture group were increased significantly (P<0.05), and the expression level of NGF protein in the DRG cells in co-culture group was increased significantly (P<0.05).

Conclusion

SCLCs could promote the growth of DRG cells through increasing the number of DRG cells and neurites and upregulating the expression of NGF protein.

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Regulatory effect of over-expression of miRNA-138-5p on testicular sertoli cell injury of rats induced by cigarette smoking exposure
Chunxue ZHONG,Hua XU,Chen ZHANG,Lijuan HE
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  692-701.  DOI: 10.13481/j.1671-587X.20220318
Abstract ( 287 )   HTML ( 1 )   PDF (1427KB) ( 60 )  
Objective

To investigate the damage effect of cigarette smoking exposure on the testicular sertoli cells of the rats, and to clarify the protective effect of miR-138-5p in this process.

Methods

A total of 200 SPF male SD rats were randomly divided into control group and low, middle, and high doses of cigarette exposure groups (given 10, 20, and 30 cigarettes per rat per day). The rats were anesthetized at 2rd, 4th, 6th, 8th, and 12th weeks, respectively. The plasma inhibin B levels of the rats in various groups were detected, the expression levels of miR-138-5p in testicular tissue of the rats in various groups were detected by real-time fluorescence quantitative PCR(RT-qPCR) method, and the expression levels of vimentin in testicular TM4 cells of the rats in various groups were detected by immunohistochemistry. The testicular TM4 cells (control group) were cultured in vitro. The cigarette smoke extract (CSE) stock solution was diluted to 5% and 10% and to treat the TM4 cells,and used as 5% CSE group and 10% CSE group. The survival rates of the testicular TM4 cells and lactate dehydrogenase (LDH) activities in the testicular TM4 cells in various groups were detected by MTT method, and the apoptotic rates of the cells in various groups were detected by flow cytometry and TUNEL method. After transfected with miR-138-5p plasmid,the testicular TM4 cells were divided into control group, 5% CSE group, 5% CSE+Pri-mirna-138-5p Ctrl group, 5% CSE+Pri-mirna-138-5p group, 10% CSE group, 10% CSE+Pri-mirna-138-5p Ctrl group, and 10% CSE+Pri-mirna-138-5p group. The survival rates of the cells, LDH activities in the cells and apoptotic rates of the cells in various groups were detected by the above methods.

Results

Compared with control group, the plasma inhibin B levels of the rats in different doses of cigarette exposure groups were decreased; compared with control group, the plasma inhibin levels of the rats in high dose of cigarette exposure group at the 8 weeks and in middle and high doses of cigarette expasure groups at the 12 th week were decreased (P<0.05). The light microscope detection results showed that the cytoplasm of the testicular cells in control group showed positive yellow staining, and the expression levels of vimentin in the testicular TM4 cells of the rats in cigarette exposure groups were decreased gradually; compared with control group, the expression levels of vimentin in testicular TM4 cells of the rats in high dose of cigarette exposure group at different time points were decreased (P<0.05). Compared with control group, the expression levels of miR-138-5p in testicular tissue of the rats in middle and high doses of cigarette exposure groups at the 8 th week and in different doses of cigarette exposure groups at the 12th week were decreased (P<0.05). The survival rates of the cells in 5% CSE group and 10% CSE group were significantly lower than that in control group (P<0.01). Compared with control group, the activities of LDH in testicular TM4 cells of the rats in 5% CSE group and 10% CSE group were decreased (P<0.05 or P<0.01). After over-expression of miR-138-5p, compared with control group, the proliferation rates of testicular TM4 cells in 5% CSE group, 5% CSE+Pri-mirna-138-5p Ctrl group, 5% CSE+Pri-mirna-138-5p group, 10% CSE group, 10% CSE+Pri-mirna-138-5p Ctrl group,and 10% CSE+Pri-mirna-138-5p group were decreased(P<0.05 or P<0.01); compared with 5% CSE group, the proliferation rate of testicular TM4 cells in 5% CSE+Pri-mirna-138-5p group was increased (P<0.05 or P<0.01), and the apoptotic rate was decreased (P<0.05 or P<0.01); compared with 10% CSE group, the proliferation rate of testicular TM4 cells in 10% CSE+Pri-mirna-138-5p group was increased (P<0.05 or P<0.01), and the apoptotic rate was decreased (P<0.05).

Conclusion

Long-term exposure to heavy cigarette smoking can induce the irreversible damage of testicular sertoli cells, and miR-138-5p plays an important protective role in the process.

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Effect of adiponectin receptor agonist AdiopRon on biological behaviors of glioma cells and its mechanism
Cuilan LIU,Fengai HU,Jing LIU,Dan WANG,Changyun QIU,Dunjiang LIU,Di ZHAO
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  702-710.  DOI: 10.13481/j.1671-587X.20220319
Abstract ( 363 )   HTML ( 0 )   PDF (1635KB) ( 264 )  

Objective: To investigate the effect of adiponectin receptor agonist AdipoRon on the biological behavior of the glioma cells, and to clarify its possible mechanism.

Methods

The glioma cells U251 and U87 MG at the logarithmic phase were divided into control group (0 μmol·L-1 AdipoRon) and 20, 40, 60, 80,and 100 μmol·L-1 AdipoRon groups. The proliferation rates of the cells in various groups at 24, 48,and 72 h were detected by CCK-8 assay.The U251 and U87 MG cells were divided into control group (0 μmol·L-1 AdipoRon) and 40, 60, and 80 μmol·L-1 AdipoRon groups.The clone formation rates of the cells in various groups were detected by clone formation assay, the scratch healing rates of the cells in various groups were detected by scratch healing experiment, the apoptotic rates and the percentages of the cells at different cell cycles were detected by flow cytometry, and the expression levels of AMP-dependent protein kinase (AMPK) and phosphorylated AMPK (p-AMPK) proteins in the cells in various groups were detected by Western blotting method.The U251 cells were divided into shNC control group(given no treatment) and shNC group(given 40 μmol·L-1 Adiporon), AdipoR1 knockdown group (given 40 μmol·L-1 Adiporon, knockdown AdipoR1), AdipoR2 knockdown group (given 40 μmol·L-1 Adiporon, knockdown AdipoR2) and AdipoR1+AdipoR2 co-knockdown group (given 40 μmol·L-1 Adiporon, knockdown AdipoR1 and AdipoR2). The proliferation rates of the U251 cells in various groups were detected by CCK-8 assay, and the expression levels of AdiopR1 and AdiopR2 mRNA in the U251 cells were detected by real-time fluorescence quantitative PCR (RT-qPCR)method.

Results

Compared with control group, the proliferation rates of the U251 and U87 MG cells in different concentrations of AdipoRon groups were decreased at 24, 48 and 72 h (P<0.05 or P<0.01). Compared with control group, the clone formation rates of the U251 and U87 MG cells in 40, 60,and 80 μmol·L-1 AdipoRon groups were decreased (P<0.01). Compared with control group, the scratch healing rates of the U251 and U87 MG cells in 40, 60,and 80 μmol·L-1 AdipoRon groups were decreased after 48 h treatment (P<0.01). Compared with control group, the apoptotic rates of the U251 cells in 60 and 80 μmol·L-1 AdipoRon groups and the U87 MG cells in 80 μmol·L-1 AdipoRon group were increased (P<0.01); the percentages of the U251 and U87 MG cells at G0/G1 phase in 40, 60, and 80 μmol·L-1 AdipoRon groups were increased (P<0.01). Compared with control group, the expression levels of p-AMPK protein in the U251 cells in 60 and 80 μmol·L-1 Adiporon groups were increased (P<0.01),and the expression levels of p-AMPK protein in the U87 MG cells in 40, 60, and 80 μmol·L-1 Adiporon group were increased (P<0.05 or P<0.01). Compared with shNC control group, the expression level of AdipoR1 mRNA in the U251 cells in AdipoR1 knockdown group was decreased (P<0.01), and the expression level of AdipoR2 mRNA in the U251 cells in AdipoR2 knockdown group was decreased (P<0.01). Compared with shNC control group, after treated with of 40 μmol·L-1 Adiporon, the proliferation rates of the U251 cells in AdipoR1 knockdown group, AdipoR2 knockdown group, and AdipoR1+AdipoR2 co-knockdown group were increased (P<0.05 or P<0.01).

Conclusion

Adiporon can inhibit the proliferation, migration and apoptosis of the glioma cells and block the cell cycle at G0/G1 phase, and its mechanism may be that adiporon interacts with the adiponectin receptors AdipoR1 and AdipoR2 to promote the AMPK phosphorylation, thus affecting the biological behaviors of the glioma cells.

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Inductive effect of peiminine on apoptosis of lung cancer A549 cells and its mechanism
Ming xing YANG,Wen DONG,Ji LI
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  711-717.  DOI: 10.13481/j.1671-587X.20220320
Abstract ( 372 )   HTML ( 4 )   PDF (1118KB) ( 351 )  
Objective

To discuss the effect of different concentrations of peiminine (PMI) on the apoptosis of the human lung cancer A549 cells, and to clarify its possible molecular mechanism.

Methods

The A549 cells were treated with different concentrations (0.025, 0.050, 0.100, 0.200 and 0.400 mmol·L-1) of PMI for 24, 48 and 72 h,respectively. The proliferation activity of the A549 cells was detected by MTT method, and the half inhibitory concentration (IC50) was calculated.The A549 cells were treated with different concentrations (0.050,0.100,0.200 mmol·L-1)of PMI or 0.200 mmol·L-1 PMI combined with p38 MAPK inhibitor SB203580 for 48 h. The cells were divided into blank control group, different concentrations(0.050, 0.100 and 0.200 mmol·L-1) of PMI groups and combination group (0.200 mmol·L-1 PMI + 20 μmol·L-1 SB203580).The apoptotic rates of the A549 cells in various groups were detected by Annexin Ⅴ-FITC/PI method; the expression levels of apoptosis-related proteins B-cell lymphoma-2 (Bcl-2),Bcl-2 associated X protein(Bax),and cleaved-caspase-3 and mitogen-activated protein kinases (p38MAPK)/p53 signaling pathway related proteins phosphorylaed p38MAPK (p-p38 MAPK) (Thr180/Thr182), p-p53 (ser15), p53, PUMA, and murine double minute 2(MDM2) in the A549 cells in various groups were detected by Western blotting method the localization of p53 protein of the A549 cells in various groups was observed by immunofluorescence assay.

Results

Compared with control group, the survival rates of the A549 cells in 0.10 and 0.20 mmol·L-1 PMI groups were decreased (P<0.05), the apoptotic rates were increased (P<0.05), and the expression levels of Bax, cleaved-caspase-3, p-p38 MAPK (Thr180/Thr182), p-p53 (ser15), p53 and PUMA proteins were increased (P<0.05),and the expression levels of Bcl-2 and MDM2 proteins were decreased (P<0.05).Compared with 0.20 mmol·L-1 PMI group, the survival rate of the A549 cells in combination group was increased (P<0.05), the apoptotic rate was decreased (P<0.05), the expression levels of Bax, cleaved-caspase-3, p-p38MAPK (Thr180/Thr182), p-p53 (ser15), p53, and PUMA proteins were decreased (P<0.05), while the expression levels of Bcl-2 and MDM2 proteins were increased (P<0.05).The immunofluorescence results showed that the phenomenon of p53 protein transferring from cytoplasm to nucleus could be seen, and the p53 protein was expressed in both the cytoplasm and nucleus.

Conclusion

Peiminine can induce the apoptosis of the A549 cells, and its molecular mechanism may be related to the p38 MAPK/ P53 signaling pathway mediated apoptosis.

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Inhibitory effect of miR-124-3p on proliferation and invasion of oral squamous cell carcinoma cells and its mechanism
Zhuangzhi WU,Xiaoning HE,Siqi CHEN
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  718-727.  DOI: 10.13481/j.1671-587X.20220321
Abstract ( 331 )   HTML ( 0 )   PDF (1659KB) ( 274 )  
Objective

To observe the effect of microRNA-124-3p (miR-124-3p) on the proliferation and invasion of oral squamous cell carcinoma (OSCC) HSC2 and KON cells, and to explore its possible mechanism.

Methods

The expression levels of miR-124-3p and brain-derived nerve growth factor (BDNF) mRNA in the OSCC tissue and OSCC cells were detected by real-time fluorescence quantitative PCR (RT-qPCR) method, and the relationship between the expression of miR-124-3p and the survival rate of the OSCC patients was analyzed;the volumes and masses of tumor of the nude mice in various groups were analyzed;the expression levels of BDNF protein in HSC2 and KON cells were detected by Western blotting method;the HSC2 and KON cells were divided into miR-124-3p group(transfected miR-124-3p),miR-NC group(transfected with miR-NC),inhibitor-NC(transfected with inhibitor-NC),miR-124-3p inhibitor group(transfected with miR-124-3p inhibitor),siBDNF group(transfected with siBDNF),BDNF group(transfected with BDNF),and miR-124-3p+BDNF group(transfected with miR-124-3+BDNF).The survival rates of the HSC2 and KON cells were detected by CCK-8 method,and the invasion rates of HSC2 and KON cells in various groups were detected by Transwell experiment; the targeting relationship between miR-124-3p and BDNF was also detected by double luciferase reporter gene experiment, and the targeting relationship between miR-124-3p and BDNF was detected by RNA immunoprecipitation experiment.

Results

The expressions of miR-124-3p in OSCC tissue and HSC2 and KON cells were low, and the expression of BDNF mRNA was high;the expression of miR-124-3p was correlated with the survival rate of the OSCC patients(P<0.05). Compared with miR-NC group, the survival rates and the invasion rates of the HSC2 and KON cells in miR-124-3p group were decreased (P<0.05), and the tumor volumes and mass of the nude mice were decreased (P<0.05). Compared with inhibitor-NC group, the survival rates and the invasion rates of the HSC2 and KON cells in siBDNF group were decreased (P<0.05). Compared with inhibitor-NC group, the survival rates and the invasion rates of the HSC2 and KON cells in BDNF group were increased (P<0.05).BDNF was a potential target of miR-124-3p. Compared with miR-124-3p group, the survival rates and the invasion rates of the HSC2 and KON cells in miR-124-3p+BDNF group were increased (P<0.05).

Conclusion

miR-124-3p can inhibit the proliferation and invasion of the OSCC cells, and its mechanism may be related to the targeted binding of BDNF.

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Effect of proteasome inhibitor lactacystin on oxidative damage of dopaminergic neurons in substantia nigra of rats
Qi PAN,Zizhen WANG,Fuyue YE,Wei XING,Jiahan LIN,Jianyue LU,Kun YANG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  728-733.  DOI: 10.13481/j.1671-587X.20220322
Abstract ( 194 )   HTML ( 0 )   PDF (500KB) ( 76 )  
Objective

To observe the effect of proteasome inhibitor lactacystin on the death and oxidative damage of dopaminergic neurons in substantia nigra of the rats, and to explore the effect of oxidative damage on the death of dopaminergic neurons induced by proteasome inhibitor.

Methods

Seventy SD rats were randomly divided into lactacystin group and normal saline group,and there were 35 rats in each group. The proteasome inhibitor lactacystin was injected into the right substantia nigra of the rats in lactacystin group, and the rats in control group were injected with the same volume of normal saline. The shuttle distance, the number of lower limb standing, and the total running time of the rats in two groups were detected by open field experiment; the forelimb asymmetry indexes of the rats in two groups were detected by forelimb asymmetry experiment; apomorphine induced rotation experiment was used to test the rotation behavior of the rats in two groups. The levels of hydroxyl free radical, reduced glutathione(GSH), malondialdehyde(MDA), protein carbonyl and 8-hydroxydeoxyguanosine(8-OHdG) in substantia nigra of the rats in two groups were detected by microplate method; the damage of tyrosine hydroxylase(TH) immunopositive neurons in substantia nigra of the rats in various groups was detected by immunohistochemistry.

Results

Compared with normal saline group, the shuttle distance, the number of standing, and the total running time of the rats in lactacystin group were significantly decreased(P<0.05). The forelimb asymmetry index of the rats in lactacystin group was significantly higher than that in normal saline group (P<0.05).After injected with apomorphine, the rats in lactacystin group showed constant rotation to the healthy side.After administrated for 7 and 21 d, compared with normal saline group, the levels of hydroxyl radical, MDA, protein carbonyl, and 8-OHdG in right substantia nigra of the rats in lactacystin group were increased significantly (P<0.05),while the levels of GSH were decreased significantly (P<0.05).After administrated for 21 d, the TH immunopositive neurons in right substantia nigra of the rats in lactacystin group were significantly lost. Compared with normal saline group, the survival rate of TH immunopositive neurons in substantia nigra of the rats in lactacystin group was significantly decreased (P<0.05).

Conclusion

Injection of proteasome inhibitor lactosin can induce the oxidative damage of dopaminergic neurons in substantia nigra of the rats, and eventually lead to the death of neurons.

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Effects of miR-107 on regulation of immune escape and taxol resistance of ovarian cancer cells
Qinghui WANG,Bo LI,Chuancui HU,Mingchao NIE,Xiaomei ZHENG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  734-743.  DOI: 10.13481/j.1671-587X.20220323
Abstract ( 351 )   HTML ( 1 )   PDF (989KB) ( 108 )  

Objective: To investigate the effect of microRNA-107(miR-107)on the taxol (TAX)-resistance in the ovarian cancer cells by regulating immune escape through the mitogen-activated protein 3 kinase 7 (MAP3K7),and to charify its mechanism.

Methods

The 293T cells were divided into miR-NC group(transfected with miR-NC) and miR-107 mimics group(transfected with miR-107 mimics).The ovarian cancer SKOV3 cells were treated with concentration gradient increasing method to obtain the TAX resistant SKOV3/TAX cells. The SKOV3/TAX cells were randomly divided into blank control group, TAX (given 4.0 μmol·L-1 TAX) group, TAX+miR-107 mimics (given 4.0 μmol·L-1 TAX and transfected with miR-107 mimics) group,TAX+miR-107 mimics+pcDNA-MAP3K7 (given 4.0 μ mol·L-1 TAX and transfected with miR-107 mimics and pcDNA-MAP3K7), and co-cultured with the human peripheral blood lymphocytes HPBL, respectively. Then they were divided into HPBL group, HPBL+SKOV3/TAX group, HPBL+ SKOV3/TAX+TAX group, HPBL + SKOV3/TAX+TAX+miR-107 mimics group and HPBL+ SKOV3/TAX+TAX+miR-107 mimics+pcDNA-MAP3K7 group. The expression levels of miR-107 in the cells in various groups were detected by real-time fluorescence quantitative PCR (RT-qPCR)method, the targeting relationship between miR-107 and MAP3K7 was verified by double luciferase reporter gene experiment, the expression levels of MAP3K7 and FAS proteins in the cells in various groups were detected by Western blotting method, the survival rates of the cells in various groups were detected by CCK-8 method, and the apoptotic rates of the cells in various groups were detected by flow cytometry.

Results

Compared with the human ovarian epithelial cells HOSEpic, the expression level of miR-107 in the ovarian cancer cells was decreased (P<0.05). There was a targeted binding site between 3′-UTR of MAP3K7 and miR-107. The double luciferase reporter gene experiment results showed that compared with miR-NC group, the luciferase activity of wild-type MAP3K7 in miR-107 over-expression group was decreased (P<0.01). The Western blotting results showed that the expression level of MAP3K7 protein in the cells in miR-107 over-expression group was significantly lower than that in miR-NC group (P<0.01). The RT-qPCR results showed that compared with blank control group, the expression level of miR-107 mRNA in the SKOV3/TAX cells in TAX group was increased(P<0.01); compared with TAX group, the expression level of miR-107 mRNA in the SKOV3/TAX cells in miR-107 MICs+TAX group was increased(P<0.05). Compared with blank control group, the expression levels of MAP3K7 and FAS proteins in the SKOV3 / TAX cells in TAX group were decreased (P<0.01); compared with TAX group, the expression levels of MAP3K7 and FAS proteins in the cells in TAX+miR-107 mimics SKOV3/TAX group were decreased (P<0.05 or P<0.01); compared with TAX+miR-107 mimics group, the expression levels of MAP3K7 and FAS proteins in the SKOV3/TAX cells in TAX+miR-107 mimics+pcDNA-MAP3K7 group were increased(P<0.05 or P<0.01).Compared with HPBL+SKOV3/TAX group, the expression level of FAS protein and apoptotic rate of the HPBL cells in HPBL+SKOV3/TAX+TAX group were decreased (P<0.05); compared with HPBL+SKOV3/TAX+TAX group, the expression level of FAS protein and the apoptotic rate in the HPBL cells in HPBL+SKOV3/TAX+TAX + miR-107 mimics group were decreased(P<0.05); compared with HPBL+SKOV3/TAX+TAX+miR-107 mimics group, the expression level of FAS protein in the HPBL cells in HPBL+SKOV3/TAX+TAX+miR-107 mimics+pcDNA-MAP3K7 group was decreased (P<0.01), and the apoptotic rate was decreased (P<0.01). Compared with blank control group, the survival rate of the SKOV3/TAX cells in TAX group was decreased (P<0.05);compared with TAX group,the survival rate of the SKOV3/TAX cells in TAX+miR-107 mimics group was significantly decreased(P<0.05). Compared with TAX + miR-107 mimics group, the survival rate of the SKOV3/TAX cells in TAX+miR-107 mimics+pcDNA-MAP3K7 group was increased (P<0.05). The apoptotic rate of the SKOV3/TAX cells in TAX group was significantly higher than that in blank control group (P<0.05), the apoptotic rate of the SKOV3/TAX cells in TAX+miR-107 group was significantly higher than that in TAX group (P<0.05), and the apoptotic rate of the cells in TAX+miR-107 mimics+pcDNA-MAP3K7 group was lower than that in TAX+miR-107 mimics group (P<0.05).

Conclusion

MiR-107 is low-expressed in the SKOV3/TAX cells, and over-expression of miR-107 could inhibit the immune escape of the SKOV3/TAX cells and alleviate the TAX resistance of the ovarian cancer cells by inhibiting the expression of MAP3K7.

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Improvement effect of p38 MAPK inhibitor on chronic obstructive pulmonary disease injury in mice through inhibiting cell pyrotosis mediated by NLRP3 pathway
Ming LI,Qiuting WANG,Shan CHEN,Huifang SHI
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  744-754.  DOI: 10.13481/j.1671-587X.20220324
Abstract ( 960 )   HTML ( 12 )   PDF (1689KB) ( 388 )  
Objective

To investigate the effect of p38 mitogen-activated protein kinase (p38 MAPK) inhibitor SB303580 (SB) on the progression of chronic obstructive pulmonary disease (COPD) in the mice, and to elucidate its possible mechanism.

Methods

A total of 48 C57BL/6 mice were divided into control group,SB group, COPD group, and COPD+SB group, and there were 12 mice in each group. The mice in COPD group and COPD+SB group were given cigarette smoke and lipopolysaccharide (LPS) to establish the COPD models. The airway resistance after inhalation of methacholine (Mch) was observed to evaluate the lung function of the mice in various groups, and the pathomophology of lung tissue of the mice in various groups was observed by HE staining. The total number of white blood cells, neutrophils, macrophages and lymphocytes, the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β), and interleukin-18 (IL-18) in pulmonary alveolar lavage fluid (BALF) of the mice in various groups were detected by cell counting and ELISA method.The mouse macrophage RAW264.7 cells were stimulated by cigaritte smoke extract (CSE) in vitro, and the cells were divided into control group, SB group, CSE group and SB+CSE group. Cell immunofluorescence staining was used to observe the expressions of NOD-like receptor protein 3(NLRP3) and cleaved caspase-3 in the macrophages in various groups, and flow cytometry was used to detect the pyroptotic rates and early apoptotic rates of the cells in various groups,and Western blotting method was used to detect the expression levels of phosphorylated p-38(p-p-38),pyroptosis-related and nuclear factor κB(NF-κB) signaling pathway related poteins in the macrophages in various groups.

Results

The COPD mouse models were successfully established.Compared with control group, the airway resistance of the mice in COPD group and COPD+SB group, the total number of white blood cells, neutrophils, macrophages and lymphocytes in BALF, the levels of TNF-α, IL-6, IL-1β ,IL-18 in BALF, and the expression levels of p-p-38, NLRP3,cysteine protease 1(caspase-1),apoptosis-associated spotted protein(ASC),NF-κB p65,and Toll-like receptor 4 (TLR4) proteins in lung tissue of the mice were increased (P<0.05 or P<0.01);the lung tissue was obviously damaged, the airway epithelial cells were exfoliated, and some adjacent alveoli fused to bullae.Compared with control group, the airway resistance of the mice in SB group, the total number of white blood cells, neutrophils, macrophages and lymphocytes in BALF, the levels of TNF-α, IL-6, IL-1β ,and IL-18 in BALF, and the expression levels of NLRP3, caspase-1, ASC, NF-κB p65, and TLR4 proteins in lung tissue had no significant differences(P>0.05); the lung tissue was normal, but the expression level of p-p-38 in lung tissue was significantly decreased (P<0.05). Compared with COPD group, the airway resistance of the mice in COPD+SB group, the total number of white blood cells, neutrophils, macrophages and lymphocytes in BALF, the levels of TNF-α, IL-6, IL-1β, and IL-18 in BALF, and the expression levels of NLRP3, caspase-1, ASC, NF-κB p65,and TLR4 proteins were decreased(P<0.05);the pathological injury of lung tissue of the mice in COPD+SB group was alleviated(P<0.05). In vitro experiments, compared with control group, the expression amounts of NLRP3 and cleaved caspase-3, pyroptotic rates and early apoptotic rates of the cells, the expression levels of p-p-38, ASC, NF-κB p65, TLR4, and cleaved caspas-3 proteins in the macrophages in CSE group and CSE+SB group were increased (P<0.05 or P<0.01);the pyroptotic rate and early apoptotic rate of the cells, and expression levels of NLRP3, cleaved caspase-3, ASC, NF-κB p65, TLR4, and cleaved caspase-3 proteins in the macrophages in SB group had no significant differences(P>0.05), and the expression level of p-p-38 in SB group was decreased (P<0.05). Compared with CSE group, the early apoptotic rate of the macrophages and the expression level of cleaved caspase-3 in the cells in CSE+SB group were increased (P<0.05),and the pyroptotic rate, the expression levels of p-p-38, NLRP3, ASC, NF-κB p65,TLR4, and cleaved caspase-3 proteins were decreased (P<0.05).

Conclusion

SB may improve the lung injury and inflammatory response of COPD by inhibiting the pyroptosis of macrophages mediated by the NLRP3 pathway.

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Research in clinical medicine
Bioinformatics analysis on screening of colon cancer core genes and independent prognostic factors
Qian LIU,Guoping QI,Huayi YU,Yuyang DAI,Wenbin LU,Jianhua JIN
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  755-765.  DOI: 10.13481/j.1671-587X.20220325
Abstract ( 623 )   HTML ( 4 )   PDF (1898KB) ( 163 )  
Objective

To mine the gene expression data of colon cancer in Gene Expression Omnibus (GEO) database and The Cancer Genome Atlas (TCGA) database,and to explore the potential core genes and the independent prognostic factors of colon cancer.

Methods

TCGA database was used to retrieve the gene expression levels and clinical feature data in colon cancer, and the GEO database was used to retrieve the core gene expression levels in colon cancer and adjacent tissues. Weighted correlation network analysis(WGCNA), Wenn analysis, protein-protein interaction(PPI) network construction and Hub gene screening, Gene Ontology(GO)enrichment analysis and Kyoto Encylopaedia of Genes and Genomes (KEGG) enrichment analysis, immune cell subtype analysis on pan oncogene,pan cancer heat map and correlation analysis of core gene, correlation analysis on core gene and tumor microenvironment and risk regression model analysis on core gene were performed. The expression levels of NKD1 and AXIN2 in the colon cancer tissue and adjacent tissue were detected by real-time fluorescence quantitative PCR (RT-qPCR),Western blotting and immunohistochemistry to further confirm the screening results.

Results

The TCGA data analysis results showed that 1 208 genes were highly expressed, 252 genes were highly expressed in the GSE37182 chip. The intersection of 4 groups of differential analysis data was taken, 13 common genes were obtained by Venn analysis. The PPI network was constructed, and 7 genes (ARID3A,SALL4,NKD1,KND2,AXIN2,CA9,and TACSTD2) were found to be the potential core genes of colon cancer. The GO analysis results showed that these 7 genes were mainly involved in positive/negative regulation of Wnt signaling pathway, basolateral plasma membrane or bound to ubiquitin protein ligases. The KEGG enrichment analysis results showed that these 7 genes were mainly involved in the Hippo signaling pathway and Wnt signaling pathway.The pan-cancer gene analysis results showed that these 7 core genes were significantly highly expressed in the colon cancer tissue, and the expression levels of NKD1 and AXIN2 genes in the colon cancer tissue had a high positive correlation (r=0.69). High expression of TACSTD2 gene was shown in the C1,C2,C3, and C6 immune cell subtypes.The correlation analysis results between the expression of core genes and the tumor microenvironment showed that the expression levels of NKD2,AXIN2,ARID3A,and NKD1 genes in the colon cancer tissue were negatively correlated with the comprehensive score(P<0.01). The survival prognostic analysis results of colon cancer patients showed that only AXIN2 was a significant independent prognostic factor (P<0.05), and it was a low-risk independent prognostic factor in the colon cancer patients. The RT-qPCR,Western blotting,and immunohistochemistry detection results showed that NKD1 was highly expressed in both the colon cancer tissue and colon cancer cells, and the expression levels of NKD1 and AXIN2 were significantly positively correlated(P<0.01).

Conclusion

Seven core genes are screened by the bioinformatics screening of colon cancer database, among which AXIN2 is an independent prognostic factor for the colon cancer patients, and the expression level of NKD1 in the colon cancer tissue is positively correlated with the expression level of AXIN2.

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Effect of preoperative glycemic level on infection-related complications of diabetic patients after flexible ureteroscopic lithotripsy
Yongxin ZHAI,Huaifeng TA,Yi ZHANG,Qijia SUN,Ming ZHANG,Shuqiang FENG,Ranwei LI
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  766-772.  DOI: 10.13481/j.1671-587X.20220326
Abstract ( 313 )   HTML ( 0 )   PDF (520KB) ( 94 )  
Objective

To observe the influencing factors of infection-related complications of the patients with type 2 diabetes mellitus (T2DM) after flexible ureteroscopic lithotripsy (FURL),and to explore the relationship between the preoperative glycemic level and postoperative infection-related complications of the T2DM patients, and to provide the basis for the early prevention and treatment of infection-related complications of the T2MD patients after FULR.

Methods

A total of 83 patients diagnosed as urolithiasis with concurrent T2DM and received FURL were retrospectively analyzed. They were divided into infection group and non-infection group according to whether or not infection occurred after surgery. Univariate and multivariate Logistic regression analysis were used to examine the independent risk factors for the postoperative infections. Receiver operating characteristic (ROC) curve was used to define the well-controlled level of preoperative glycemic;based on the optimal cut-off value, the patients were divided into good glycemic control group (glycemic level≤7.7 mmol·L-1) and poor glycemic control group (glycemic level>7.7 mmol·L-1).The incidence frequency of postoperative infection-related complications and avarage postoperative hospital stay of the patients in two groups were compared.

Results

The univariate Logistic regression analysis results showed that the preoperative glycemic level, the stone diameter≥2 cm, preoperative urine routine leukocyte count ≥10/HP,the positive preoperative urine culture,and the operation time≥90 min were the independent influencing factors for the postoperative infection-related complications of the T2DM patients.The multivariate Logistic regression analysis reesults showed that the preoperative glycemic level,the operation time ≥90 min,the stone diameter≥2 cm,and positive preoperative urine culture were the independent influencing factors for the postoperative infection-related complications of the T2DM patients.The ROC curve analysis results showed that if the optimal cut-off value of the preoperative glycemic level was 7.7 mmol·L-1, the sensitivity to predict the postoperative infection-related events of the patients was 70.37%, and the specificity was 96.43%.The incidence frequency of postoperative infection-related complications and the average postoperative hospital stay of the patients in good glycemic control group were lower than those in poor glycemic control group (P<0.05).

Conclusion

The preoperative glycemic level, positive preoperative urine culture, operative time≥90 min and stone diameter≥2 cm are the independent influencing factors for the infection-related complications of the T2DM patients after FURL. When the preoperative glycemic level≤7.7 mmol·L-1 is used as the standard for good glycemic control, the incidence of postoperative infection can be effectively reduced and the postoperative hospital stay can be shortened.

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Inhibitory effect of silencing expression of DNMT3a on progression of psoriasis like cell cycle and cell proliferation and its mechanism
Yanbin PAN,Jiaguang SU,Meile TAN,Meng YANG,Wenfei TAN,Yuxiu HUANG,Shihao MENG,Yaohui HUANG,Jianqiang LIANG,Xuefang SU,Zichan HUANG,Jianmin LI
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  773-782.  DOI: 10.13481/j.1671-587X.20220327
Abstract ( 438 )   HTML ( 12 )   PDF (1760KB) ( 278 )  
Objective

To investigate the inhibitory effect of silencing the expression of DNA methyltransferase 3A (DNMT3a) on the progression of psoriasis like HaCaT cell cycle and cell proliferation,and to clarify its possible mechanism.

Methods

The skin tissues of 15 patients with psoriasis vulgaris (psoriasis group) and 15 healthy controls (control group) were collected. Real-time fluorescence quantitative PCR(RT-qPCR) method was used to detect the expression levels of DNMT3a mRNA in skin tissue of the subjects in two groups. The Si-DNMT3a silenced by DNMT3a was transfected into the HaCaT cells by siRNA transfection technology. RT-qPCR and Western blotting methods were used to detect the transfection efficiency of si-DNMT3a. The HaCaT cells were induced by M5 method and the psoriasis like cell model was established. The cells were divided into control group, si-DNMT3a group, si-NC group, and HaCaT group. The proliferation rates of the cells in various groups were detected by CCK-8 method, the percentages of the cells at different cell cycles in various groups were detected by flow cytometry, and the apoptotic rates of the cells in various groups were detected by Annexin Ⅴ-FITC test. MSP test was used to detect the methylation levels of LATS1 gene promoter in skin tissue of the subjects in psoriasis group and control group. The expression levels of large tumor suppressor gene 1(LATS1), Yes associated protein(YAP1), cyclin D1, CDK6, and cleaved caspase-3 proteins in the HaCaT cells in various groups were detected by Western blotting method.

Results

Compared with control group, the expression level of DNMT3a mRNA in skin tissue of the patients in psoriasis group was significantly increased (P<0.01). Compared with HaCaT group,the expression levels of DNMT3a mRNA and protein in the HaCaT cells in si-NC group were decreased (P<0.05). Compared with HaCaT group, the proliferation rates of the HaCaT cells and the percentages of cells at S phase in control group, si -NC group,and si- DNMT3a group were increased (P<0.05), and the percentages of cells at G1 phase and the apoptotic rates were significantly decreased (P<0.05);compared with control group and si-NC group, the proliferation rates of the HaCaT cells and the percentage of cells at S phase in si-DNMT3a group were significantly decreased(P<0.05),and the apoptotic rate and the percentage of cells at G1 phase were increased (P<0.05). The MSP results showed that the methylation level of LATS1 promoter in skin tissue of the patients in psoriasis group was higher than that in control group (P<0.01). The Western blotting results showed that compared with HaCaT group, the expression levels of YAP1, Cyclin D1, and CDK6 proteins in the HaCaT cells in control group, si-NC group,and si-DNMT3a groups were increased (P<0.05), while the expression levels of LATS1 and cleaved caspase-3 proteins were decreased (P<0.05); compared with control group and si-NC group, the expression levels of YAP1, cyclin D1, and CDK6 proteins in the HaCaT cells in si- DNMT3a group were decreased (P<0.05), and the expression levels of LATS1 and cleaved caspase-3 proteins were increased (P<0.05).

Conclusion

Silencing the DNMT3a expression may down regulate the expression of YAP1 by reducing the methylation level of LATS1, so as to inhibit the progression of psoriasis like HaCaT cell cycle and cell proliferation and induce the apoptosis.

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Analysis on causes for early death and its risk factors of patients with multiple myeloma in era of novel drugs
Shan GAO,Yutong WANG,Minqiu LU,Lei SHI,Bin CHU,Yuehua DING,Mengzhen WANG,Li BAO
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  783-789.  DOI: 10.13481/j.1671-587X.20220328
Abstract ( 460 )   HTML ( 2 )   PDF (486KB) ( 92 )  
Objective

To explore the causes and risk factors of early death in the patients with multiple myeloma (MM) in the era of novel drugs, and to provide the basis for the early identification of the high-risk patiens and guiding the treatment.

Methods

The clinical data of the patients with newly diagnosed multiple myeloma (NDMM) treated with proteasome inhibitor or immunomodulator-based chemotherapy were retrospectively analyzed. According to whether the overall survival (OS) was more than 24 months, they were divided into early death group (OS < 24 months) and control group (OS ≥ 24 months). The high-risk factors of the patients in two groups were statistically analyzed, including age, International Staging System(ISS) stage, Revised Intenational Staging System (R-ISS) stage, anemia, renal insufficiency, hypercalcemia, increasing of lactate dehydrogenase(LDH) level, bone marrow plasma cell ratio, extramedullary plasmacytoma, detection results of fluorescence in situ hybridization(FISH),and efficacy.Kaplan Meier method was used to draw the survival curves for survival analysis, Logistic regression analysis was used for stepwise screening for the risk factors, and the meaningful clinical indicators were screened, and multivariate analysis was used to explore the high-risk factors of early death.

Results

A total of 237 patients with NDMM were included, including 53 cases in early death group and 184 cases in control group. The median OS of the patients in early death group was 16 months, but the median OS of the patients in control group had not been reached by the end of follow-up. There was significant difference in OS of the patients between two groups (P<0.01). The causes for death of the patients in early death group included disease progression death (77.4%) and non-disease progression death (22.6%). The non-disease progression death included 4 cases of pneumonia, 3 cases of acute myocardial infarction, 1 case of acute cerebral infarction and 4 cases of ominous causes of death due to automatic abandonment of treatment. The univariate analysis results showed that the percentages of elderly patients (>65 years old), ISS stage Ⅲ, R-ISS stage Ⅲ, anemia, increasing of LDH level,bone marrow plasma cells ratio> 60%, extramedullary plasma cell tumor, 17p deficiency detected by FISH and failure to reach partial response(PR) in early death group were higher than those in control group (P<0.05).The multivariate regression analysis results showed that old age (P=0.007), R-ISS stage Ⅲ (P=0.003), extramedullary plasmacytoma (P=0.008) and failure to reach PR (P<0.01) were the independent risk factors for early death of the NDMM patients in the era of novel drugs.

Conclusion

The cause for early death in the era of novel drugs is disease progression; age> 65 years old, R-ISS stage Ⅲ, extramedullary plasmacytoma and the best curative effect not reaching PR are still the independent risk factors for early death of the NDMM patients. The intensive treatment of the NDMM patients with the above characteristics may improve the prognosis of the patients.

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Clinical medicine
Treatment of advanced rectal cancer with rectovaginal fistula through bevacizumab combined with FOLFIRI regimen: A case report and literature review
Junjie HOU,Xuguang MI,Xiaonan LI,Xiaonan LI,Ying YANG,Xianzhuo JIANG,Ying ZHOU,Zhiqiang NI,Ningyi JIN,Yanqiu FANG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  790-795.  DOI: 10.13481/j.1671-587X.20220329
Abstract ( 470 )   HTML ( 3 )   PDF (683KB) ( 156 )  
Objective

To investigate the efficacy of bevacizumab combined with chemotherapy in the treatment of the rectal cancer patients with rectovaginal fistula,and to provide the reference basis for the treatment of the disease.

Methods

The clinical data of one patient with advanced rectal cancer with rectovaginal fistula treated with bevacizumab combined with FOLFIRI were collected,and the characteristics and process of diagnosis and treatment were analyzed; the relevant literatures were summarized at the same time.

Results

A 64-year-old female patient received transabdominal rectal cancer surgery (Dixon surgery) in People’s Hospital of Jilin Province,and the postoperative pathological results showed moderately differentiated rectal adenocarcinoma,invasion of posterior wall of vagina and regional lymph node metastasis (pT4bN2M0, Stage ⅢC,Dukes C);the immunohistochemistry results showed MHS2(+),MLH1(+),PMS2(+),MSH6(+),S-100(-),and BRAFV600E(-),indicating proficiency of mismatch repair(pMMR);the patient had N- and K-RAS gene mutations.The pelvic MRI results at 4 weeks after surgery revealed the postoperative rectovaginal fistula before pre-adjuvant chemotherapy.The chemotherapy was postponed, and the disease continued to progress 5 months after surgery, and the palliative first-line and second-line chemotherapy were perfermed under compulsion.However, the disease continued to progress, and the third-line chemotherapy of bevalizumab combined with FOLFIRI regimen was performed and did not aggravate the rectovaginal fistula, and promoted the fistula healing,while significantly improved the quality of life of the patient.

Conclusion

Bevacizumab combined with chemotherapy is a feasible treatment option for the rectal cancer patients with rectovaginal fistula caused by the tumor progression and will not benefit from the cetuximab therapy.

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Pulmonary imaging changes as first manifestation of angioimmunoblastic T-cell lymphoma: A case report and literature review
Qian LI,Jingyi YUAN,Jiaqi ZHOU,Min ZHAO,Ke WANG
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  796-800.  DOI: 10.13481/j.1671-587X.20220330
Abstract ( 359 )   HTML ( 4 )   PDF (546KB) ( 150 )  
Objective

To explore the clinical characteristics, diagnostic process and treatment of angioimmunoblastic T-cell lymphoma (AITL), and to improve clinicians’ understandings of the disease.

Methods

The clinical data, imaging findings, bronchoscope and pathological findings of one patient with AITL were collected, the above data were analyzed, and the relevant literatures were reviewed.

Results

A 66-year-old female patient was admitted with fever accompanying with cough and expectoration for 15 d. The physical examination results showed that there was a red rash scattered on all four limbs, no abnormality of the thorax, and the trachea was in the center. Dullness of the left lower lung was found by percussion, and the breath sounds of the left lower lung were weakened, without other obvious positive signs.The chest CT results showed the multiple patchy high-density shadows in the lower lobe of both lungs, especially in the left lower lung, and the mediastinum and bilateral axillary lymph nodes were enlarged. The possibility of double pneumonia was considered initially, and the space-occupying lesions were excluded. The results of bronchoscopic lung biopsy (TBLB),mediastinal lymph node transbronchial needle aspiration biopsy (TBNA), and exfoliated pleural cells all excluded pulmonary malignancy. After anti-infection treatment and experimental anti-tuberculosis treatment, the patient still had repeated fever. The chest CT results showed that the high-density shadow range of the left lower lung was increased and consolidated. The comprehensive laboratory examination and imaging examination results showed no abnormalities, and the specific cause of the patient’s fever was unclear. The physical examination of the patient showed the multiple light red rashes on the trunk and limbs and the bilateral supraclavicular lymph node enlargement. Biopsy of right supraclavicular lymph node was performed, and AITL was finally reported pathologically.

Conclusion

AITL should be considered in the patients with unexplained fever with pulmonary imaging changes, rash and lymph node enlargement, if systemic anti-infective therapy does not respond well.

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Methodology
Establishment of molecular biological identification method for origin of true and false bear bile powder species and its evaluation
Yuan ZHAO,Huijian JIA,Shunjia SONG,Qi LI,Xuechao SHAO,Jinxia AI,Liyuan SUN
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  801-808.  DOI: 10.13481/j.1671-587X.20220331
Abstract ( 1354 )   HTML ( 5 )   PDF (1712KB) ( 219 )  
Objective

To establish and evaluate a rapid and specific method for identifying the biological origin of bear bile powder based on the multiplex polymerase chain reaction (PCR), and to provide the basis for the identifying of artificial mixed adulterated samples.

Methods

Using porcine, bovine and bear mitochondrial cytochrome b genes,SDS-proteinase K cleavage(SDS-PK) was used to extract the bile DNA, Primer Premier 5.0 software was used to design the species-specific primers that could amplify the different sizes of DNA fragments without cross reaction. After the specific amplification, the amplified bands were cloned and sequenced, and compared with the registered sequences of GenBank database. The multiplex PCR was established and optimized, and the specificity and sensitivity were evaluated,and the method was used to identify 27 simulated mixed adulterated samples.

Results

The established method for DNA extraction from pig, bovine,and bear could be completed within 2 h, and the DNA purities were 2.04, 1.69, and 1.70, respectively, and the concentrations were 158.63, 189.34,and 148.55 mg·L-1. The species-specific primers were designed to amplify the pig, bovine, and bear. The sequencing results showed that the homology between the samples and the registered sequences from GenBank database was more than 99%. The primers of each species in the PCR system had strong specificity and no non-specific amplification. In triple PCR system,the primers did not interfere with each other;the detection sensitivity was high, and it could be successfully detected when the DNA concentration of any of the three target samples reduced to 1 mg·L-1. The test results of 27 artificially simulated bear bile powder adulterated samples with different species and different proportions were consistent with the preset mixing conditions. The blind random sampling was repeated for 5 times, and the results were stable.

Conclusion

The method that can simultaneously identify the pig, bovine, and bear bile powder through a single experiment is successfully established, which is simple, sensitive and efficient, and can be applied to the identification of bear bile powder and its common animal orgin adulteration.

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Preparation and evaluation of sodium alginate-locust bean gum interpenetrating polymer network hydrogel drug sustained-release system
Da LI,Yanhong JIA,Zebing ZHANG,Xiufeng HAO
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  809-817.  DOI: 10.13481/j.1671-587X.20220332
Abstract ( 481 )   HTML ( 4 )   PDF (2037KB) ( 92 )  
Objective

To investigate the drug loading and release properties of different ratios of sodium alginate (SA)-carob gum (LBG) interpenetrating polymer network(IPN) hydrogels, and to evaluate the persistent killing effect of IPN hydrogels coated with cisplatin (DDP) on the oral squamous cell carcinoma CAL-27 cells and its mechanism.

Methods

Different ratios of SA and LBG were dissolved in the deionized water.The IPN hydrogels of different formulations were prepared by ion gel method with glutaraldehyde (GA) and calcium chloride(CaCl2) as the cross-linking agents. The physicochemical structures of the hydrogels were characterized with scanning electron microscope (SEM) and Fourier transform infrared spectroscopy (FTIR). The entrapment efficiency(EE) of the hydrogels was determined by ultraviolet spectrophotometry and the drug release curves were drawn. The particle sizes and drug EE of the hydrogel beads under different ratios of LBG and GA were shown by three-dimensional histogram. Combined with the EE and drug release curve, the optimized formula was selected to wrap DDP to prepare DDP hydrogel beads. The CAL-27 cells were divided into control group, DDP solution group and DDP hydrogel beads group. The survival rates and apoptotic state of the CAL-27 cells in various groups at different time points were observed by MTS and AO/PI fluorescence staining.

Results

The SEM results showed that the surface of the dried gel beads was rough or even uneven and wrinkled. The three-dimensional histogram results showed that the particle size was increased with the increasing of LBG level and was decreased with the increasing of GA level. With the increasing of LBG and GA levels, the EE of the hydrogel was increased. The FTIR results showed that DDP was encapsulated successfully by the IPN hydrogel. When the ratio of SA and LBG was 2.25∶0.75, the level of GA was 3%, and the concentration of CaCl2 was 3%, the EE of hydrogel was 76.8%, and the drug sustained release time was 48 h.The MTS assay results showed that after treated for 6 and 12 h, the survival rates of the CAL-27 cells in DDP hydrogel beads group were slightly higher than those in DDP solution group (P<0.05). At 24 and 48 h, the survival rates of the CAL-27 cells in DDP hydrogel beads group were lower than those in DDP solution group (P<0.05). The cells in control group grew well from 6 to 48 h, and the proliferated cells were dyed green. At 6 and 12 h, the number of CAL-27 cells in DDP solution group and DDP hydrogel beads group was less than that in control group, and the number of survival cells in DDP hydrogel beads group was slightly more than that in DDP solution group. After 24 h, the growth rate of the CAL-27 cells in DDP hydrogel beads group became slower, and the number of survival cells in DDP hydrogel beads group was less than that in DDP solution group at 24 and 48 h.

Conclusion

DDP-IPN hydrogel with sustained-release DDP is successfully prepared and optimized, which has a sustained killing effect on the oral squamous cell carcinoma CAL-27 cells, and it is expected to develop into a new drug sustained-release system.

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Review
Research progress in effect of hard tissue factors on morphology of soft tissue in corrective orthodontic treatment
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  818-824.  DOI: 10.13481/j.1671-587X.20220333
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Progress research in role of TLRs signaling pathway and Cross- talk of TLRs in inflammatory diseases
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  825-831.  DOI: 10.13481/j.1671-587X.20220334
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Research progress in application of injectable platelet rich fibrin in treatment of oral diseases
Journal of Jilin University(Medicine Edition). 2022, 48 (3):  832-838.  DOI: 10.13481/j.1671-587X.20220335
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