To explore the interventional effect of salidroside （SAL） on acute asthmatic airway inflammation in the mice， and to clarity its mechanism.

Fifty clean BALB/c female mice were randomly divided into control group， ovalbumin （OVA） group， low dose of SAL group， high dose of SAL group and dexamethasone group， and there were 10 mice in each group. The mouse asthma model was established by intraperitoneal injection of 200 μL mixed solution （containing normal saline， 10 mg OVA， and 1 mg aluminum hydroxide） on the 1st， 7th and 14th days. On the 21st day， the sensitized mice were placed in the experimental glass box. In the box，the mice were stimulated with 0.1 g OVA+10 mL physiological saline pulverization for 30 min， once a day， lasted for 7 d.^{The mice in low and high doses of SAL groups were intraperitoneally injected with 30－60 mg·kg－1 SAL treatment solution 1 h before each challenge， and the mice in dexamethasone group were intraperitoneally injected with 2 mg·kg－1 dexamethasone treatment solution， while the mice in control group were replaced with the same dose of saline. The enhanced pause（Penh） values of the mice in various groups were detected and the airway reactivities of the mice were evaluated； the morphology of lung tissue of the mice in various groups was observed by HE staining； the numbers of neutrophils， eosinophils and lymphocytes in alveolar lavage fluid （BALF） of the mice in various groups were calculated by direct counting method； ELISA method was used to detect the levels of interleukin 1β （IL-1β）， interleukin 4 （IL-4）， interleukin 13 （IL-13），interleukin 17A （IL-17A）， interleukin 18 （IL-18） ，and interleukin 33 （IL-33） in BALF of the mice in various groups； the expression level of NOD-like receptor family protein 3 （NLPR3），apoptosis-associated speck-like protein containing a CARD （ASC），Caspase-1， interleukin-1β precursor （pro-IL-1β） and IL-1β proteins in lung tissue of the mice in various groups were determined by Western blotting method.}

Compared with control group， the Penh value of the mice in OVA group was increased （P<0.05）； compared with OVA group， the Penh value of the mice in high dose of SAL group was increased （P<0.05）. Compared with control group， the airway smooth muscle of the mice in OVA group was thickened and a large amount of inflammatory cell infiltration was seen； compared with OVA group，the number of inflammatory cells around the airway of the mice in high dose of SAL group was decreased （P<0.05）. Compared with control group， the numbers of neutrophils， eosinophils and lymphocytes in BALF of the mice in OVA group were increased （P<0.05）.The levels of IL-1β， IL-4， IL-13， IL-17A， IL- 18， and IL-33 in BALF of the mice in OVA group were significantly increased （P<0.05）；compared with OVA group， the numbers of neutrophils， eosinophils and lymphocytes in BALF of the mice in high dose of SAL group were decreased （P<0.05）， the levels IL -1β， IL-4， IL-13， IL-17A， IL-18 ，and IL-33 in BALF of the mice in OVA group were decreased （P<0.05）. Compared with control group， the expression levels of NLPR3， ASC， Caspase-1， pro-IL-1β ，and IL-1β proteins in lung tissue of the mice in OVA group were significantly increased （P<0.05）； compared with OVA group， the expression levels of NLPR3， ASC， Caspase-1，pro-IL-1β ，and IL-1β proteins in lung tissue of the mice in high dose of SAL group were decreased （P<0.05）.

SAL can reduce the airway inflammation in the mice with acute asthma， and its mechanism may be related to down-regulation the expression of NLRP3 inflammasome.

To investigate the improvement effect of Sanjiao acupuncture on the neurological function and stromal cell-derived factor-1α/chemokine receptor 4 （SDF-1α/CXCR4） pathway of the rapid aging model （SAMP8）mice， and to explore the mechanism of Sanjiao acupuncture in the SAMP8 mice.

The experiment was divided into model group （SAMP8 mice）， non-acupuncture group （SAMP8 mice + non-acupuncture）， acupuncture group （SAMP8 mice + acupuncture）， acupuncture + AMD3100 group （SAMP8 mice + acupuncture + AMD3100 treatment），and control group （SAMR1 mice），and there were 10 mice in each group. Morris water maze test was used to detect the learning and memory function indexes of the mice in various groups； enzyme-linked immunosorbent assay （ELISA） was used to detect the levels of amyloid β （Aβ） 1-42 in hippocampus tissue of the mice in various groups； choline acetyltransferase （chAT） and acetylcholinesterase （AchE） kits were used to detect the activities of chAT and AchE in hippocampus tissue of the mice in various groups； Western blotting method was used to detect the expression levels of SDF-1α and CXCR4 proteins in hippocampus tissue of the mice in various groups.

Compared with control group， the escape latency of the mice in model group was prolonged（P<0.05）， the level of Aβ1-42 in hippocampus tissue of the mice was increased （P<0.05）， the number of crossing the platform， the ratio of swimming distance in the original quadrant to total distance， the ratio of swimming time in the original quadrant to total time， the activities of chAT and AchE in hippocampus tissue，and the levels of SDF-1α and CXCR4 proteins in hippocampus tissue of the mice were decreased （P<0.05）； compared with model group， the escape latency of the mice in acupuncture group was shortened（P<0.05）， the level of Aβ1-42 in hippocampus tissue of the mice was decreased （P<0.05）， the number of crossing the platform， the ratio of swimming distance in the original quadrant to total distance， and the ratio of swimming time in the original quadrant to total time， the activity of chAT in hippocampus tissue，and the expression levels of SDF-1α and CXCR4 proteins in hippocampus tissue of the mice were increased （P<0.05）； compared with acupuncture group， the escape latency of the mice in acupuncture + AMD3100 group was prolonged（P<0.05）， the level of Aβ1-42 in hippocampus tissue of the mice was increased （P<0.05），the number of crossing the platform， the ratio of swimming distance in the original quadrant to total distance， and the ratio of swimming time in the original quadrant to total time， the activity of chAT in hippocampus tissue，and the expression levels of SDF-1α and CXCR4 proteins in hippocampus tissue of the mice were decreased（P<0.05）.

Sanjiao acupuncture may reduce the accumulation of Aβ1-42 by activating the SDF-1α/CXCR4 pathway， and improve the neurological function of the SAMP8 mice.

To predict the ubiquitination sites of human tumor necrosis factor receptor-associated factor 6（TRAF6） gene and construct the ubiquitination mutant plasmid， and to explore the effect of mutant plasmid on the relative luciferase activity of nuclear factor kappa-B（NF-κB）and activator protein-1（AP-1） in the human colorectal cancer HCT116 and SW480 cells.

UbPred， UbiSite， and BDM-PUB softwares were used to predict the ubiquitination sites of TRAF6 gene；the mutation primers were designed by CE Design V1.04 software， and the mutation kits were used for site-directed mutation； the mutated target fragment was amplified by PCR method；the amplified products were digested by Dpnl enzyme to remove the methylated template plasmids；the digested products were recombined under the catalysis of Exnase Ⅱ to obtain the recombinant plasmids；the recombinant plasmids were transformed into the competent cells of DH5α E. coli and the sequence was sequenced. The relative luciferase activities of NF-κB and AP-1 in the colorectal cancer HCT116 and SW480 cells were detected by dual-luciferase reporter gene system.

After DNA sequencing， the ubiquitination mutation site was successfully mutated， and the ubiquitination mutant plasmid was successfully constructed. Compared with TRAF6 wild-type gene strain， the relative luciferase activities of NF-κB and AP-1 in the colorectal cancer HCT116 and SW480 cells were decreased after transfected with 124mut， 319mut， and 331mut plasmids （P<0.05 or P<0.01）， and the relative luciferase activities of NF-κB and AP-1 in the colorectal cancer HCT116 and SW480 cells were the most significantly decreased after transfected with 124mut plasmid（P<0.01）.

The ubiquitination mutant plasmids are successfully constructed.The 124th amino acid of TRAF6 is the most important ubiquitination site， which may affect the activities of NF-κB and AP-1 factors in downstream signaling pathways.

To discuss the effect of over-expression of peroxiredoxin 6 （PRDX6） gene on the proliferation， invasion and migration of liver cancer cells， and to clarify its possible mechanism.

The HepG2 and Hep3B cells were established and divided into PRDX6 over-expression group（tansfected with over-expression vector PIRES-EGFP-PRDX6） and empty vector group（tansfected with control vector PIRES）.The expression levels of PRDX6 mRNA and protein in the liver cancer cells in two groups were detected by Real-time fluorescence quantitative（RT-qPCR） and Western blotting methods；the proliferation rates of the liver cancer cells in two groups were detected by MTT method in vitro；scratch assay and Transwell chamber assay were used to evaluate the wound healing rate and the invasion number of liver cancer cells in two groups； Western blotting method was used to detect the expression levels of PRDX6， matrix matalloproteinases-2（MMP-2）and matrix matalloproteinases-9（MMP-9）proteins in the liver cancer cells in two groups.

The RT-qPCR and Western blotting results showed that compared with empty vector group， the expression levels of PRDX6 mRNA and protein in PRDX6 over-expression group were significantly increased （P<0.05）.The MTT results showed that compared with empty vector group， the proliferation activities of the liver cancer cells in PRDX6 over-expression group were significantly increased （P<0.05）. The scratch assay and Transwell chamber assay results showed that compared with empty vector group， the wound healing rates and the number of invasion liver cancer cells in PRDX6 over-expression group were significantly increased （P<0.05）.The Western blotting results showed that that compared with empty vector group， the expression levels of MMP-2 and MMP-9 proteins in the liver cancer cells in PRDX6 over-expression groups were increased（P<0.05）.

PRDX6 could significantly promote the proliferation， invasion， and migration of liver cancer cells， and its mechanism may be related to its regulatory effect on the expression levels of MMP- 2 and MMP- 9 proteins.

To observe the effect of urolithin B （UB） on the proliferation， migration， invasion and apoptosis of human glioblastoma（GBM）U118 MG cells，and to discuss its mechanism.

The U118 MG cells were cultured in vitro and divided into control group（0 μmol·L^－1 UB） and different concentrations （20， 40， 80， 120， 160 and 200 μmol·L^－1） of UB groups. After cultured for 24， 48 and 72 h， the proliferation rates of U118 MG cells in various groups were detected by CCK-8 method.The U118 MG cells were divided into control group（0 μmol·L^－1 UB） and different concentrations （40， 80 and 120 μmol·L^－1） of UB groups，and clone formation experiment was used to detect the rates of clone formation of U118 MG cells in various groups； cell scratch healing test was used to detect the scratch healing rates of U118 MG cells in various groups； Transwell chamber assay was used to detect the percentages of invasion U118 MG cells in various groups； flow cytometry was used to detect the apoptotic rates of U118 MG cells and the percentages of U118 MG cells at different cell cycles in various groups； Western blotting method was used to detect the expression levels of Vimentin，epithelial adhesive protein（E-cadherin），B cell lymphoma-2（Bcl-2） ，and Bcl-2 related X protein（Bax） in the U118 MG cells in various groups.

The CCK-8 results showed that compared with control group， the proliferation rates of U118 MG cells in different concentrations of UB groups at 24， 48 and 72 h after treatment were decreased（P<0.01） in a dose and time-dependent manner. The clone formation experiment results showed that compared with control group， the rates of clone formation of U118 MG cells in 40， 80 and 120 μmol·L^－1 UB groups were decreased（P<0.01） in a dose-dependent manner.The cell scratch test results showed that compared with control group， the scratch healing rates of U118 MG cells in 40， 80 and 120 μmol·L^－1 UB groups at 24 and 48 h after treatment were significantly decreased（P<0.05 or P<0.01） in a dose-dependent manner.The Transwell chamber assay results showed that compared with control group， the percentages of invasion U118 MG cells in 40， 80 and 120 μmol·L^－1 UB groups were decreased significantly（P<0.05 or P<0.01） in a dose- dependent manner.The flow cytometry results showed that compared with control group， the apoptotic rates of U118 MG cells in 80 and 120 μmol·L^－1 UB groups were increased（P<0.05 or P<0.01） in a dose-dependent manner， and the percentages of U118 MG cells at G₂/M phase were increased （P<0.01）. The Western blotting results showed that compared with control group， the expression levels of Vimentin and Bcl-2 proteins in U118 MG cells in different concentrations of UB groups were decreased （P<0.05 or P<0.01），and the expression levels of E-cadherin and Bax proteins were increased （P<0.01）.

UB can inhibit the proliferation， migration and invasion of human GBM U118 MG cells， inhibit the expression of Bcl-2 protein， increase the expression of Bax protein， induce the apoptosis， and result in G₂/M phase arrest.

To inhibit and activate the phosphorylation of mammalian target protein of rapamycin （mTOR） by rapamycin （RAPA） and MHY1485，and to discuss the effect of level of phosphorylated mTOR（p-mTOR） on the proliferation， autophagy and differentiation of osteoblasts and its mechanism.

The mouse MC3T3-E1 osteoblasts were divided into control group， 20 nmol·L^－1 RAPA group， 60 nmol·L^－1 RAPA group， 160 nmol·L^－1 RAPA group and 2.0 μmol·L^－1 MHY1485 group. The percentages MC3T3E1 osteoblasts at different cell cycles in various groups were detected by flow cytometry；the expression levels of mTOR and downstream signals—IF4E-binding protein 1 （4E-BP1） and ribosome protein subunit 6 kinase 1 （S6K1），B cell lymphoma-2（Bcl-2），Bcl-2 related X protein（Bax），and Caspase-3，alkaline phosphatase （ALP）， Runt-related transcription factor 2 （Runx2），and osteoblast-related transcription factor Osterix were detected by Western blotting and Real-time fluorescence quantitative PCR（RT-qPCR） methods；the mineralization ability of MC3T3-E1 osteoblasts in various groups was observed by Alizarin red staining.

Compared with control group， the proliferation activities of MC3T3-E1 osteoblasts in 20 and 60 nmol·L^－1RAPA groups were significantly increased（P<0.05 or P<0.01）， the percentages of MC3T3-E1 osteoblasts at G₁ phase were increased（P<0.05 or P<0.01），the percentages of the MCET3-E1 osteoblasts at S phase were increased（P<0.05 or P<0.01），the expression levels of p-mTOR and its downstream pathway of p-4E-BP1 and p-S6K1， as well as p62，Bax，and Cleaved-Caspase-3 were decreased（P<0.05 or P<0.01），and the expression levels of LC3-Ⅱ， Bcl-2， ALP and Runx2 were increased （P<0.05 or P<0.01）.Compared with control group，the proliferation activity of MC3T3-E1 osteoblasts in 160 nmol·L^－1 RAPA group was significantly decreased（P<0.01），and the expression levels of p-mTOR，p-4E-BP1， and p-S6K1 were decreased （P<0.05 or P<0.01）；the proliferation activity of MC3T3-E1 osteoblasts in 2.0 μmol·L^－1 MHY1485 group was decreased（P<0.01），and the expression levels of p-mTOR，p-4E-BP1， and p-S6K1 were decreased（P<0.01）；however，the expression levels of Bax and Cleaved-Caspase-3 in both 160 nmol·L^－1 RAPA and 2.0 μmol·L^－1 MHY1485 groups were increased（P<0.05 or P<0.01），and the expression levels of Bcl-2， ALP， Runx2， and Osterix were decreased（P<0.05 or P<0.01）.Twelve days after treatment， the mineralized nodules were found in the MC3T3-E1 osteoblasts in 20 and 60 nmol·L^－1RAPA groups， and only a very small amount of calcium salt were found in the MC3T3-E1 osteoblasts in 160 nmol·L^－1 RAPA and 2.0 μmol·L^－1 MHY1485 groups.

Mild to moderate inhibition of mTOR phosphorylation can promote the proliferation， autophagy and differentiation of the MC3T3-E1 osteoblasts，while excessive inhibition and enhancement of mTOR phosphorylation have the opposite effect.

To observe the effect of miR-146b on the expression of intercellular adhesion molecule-1 （ICAM-1） in lung tissue of the rats with acute respiratory distress syndrome （ARDS）， and to clarify the molecular mechanism of miR-146b in the treatment of ARDS.

The ARDS rat models were established by injecting oleic acid into the tail vein， and the successfully modeled rats were divided into model group（only given oleic acid）， agomir negative control group and miR-146b agomir group， with 15 rats in each group； another 15 rats were selected as sham operation group（only given saline）. One hour before operation， the rats in miR-146b agomir group were given miR-146b agonist for intervention， and the rats in agomir negative control group were given miR-146b agonist negative control reagent for intervention.Twenty-four hours after successful modeling， the partial pressure of oxygen （PaO₂） and oxygenation index （OI） of the rats in various groups were detected by the blood gas system； the wet/dry weight ratios （W/D） of lung tissue of the rats in various groups were detected； the levels of interleukin-1β （IL-1β）， interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α） in bronchoalveolar lavage fluid （BALF） of the rats in various groups were detected by ELISA assay； the pathomorphology of lung tissue of the rats in various groups was observed by HE staining； the expression levels of miR-146b and ICAM-1 mRNA in lung tissue of the rats in various groups were detected by Real-time fluorescence quantitative PCR（RT-qPCR） method； the expression levels of ICAM-1 protein in lung tissue of the rats in various groups were detected by Western blotting method； the targeting relationship between miR-146b and ICAM-1 was detected by dual luciferase reporter system.

Compared with model group and agomir negative control group， the PaO₂ and OI， and the ratio of W/D of lung tissue of the rats in miR-146b agomir group were increased （P<0.01）， and the levels of IL-1β， IL-6， and TNF-α in BALF of the rats in miR-146b agomir group were increased （P<0.01）.The HE staining results showed that the structure of lung tissue of the rats in sham operation group was normal， there was no obvious inflammatory pathological changes， while the pathological changes such as alveolar hemorrhage， alveolar wall thickening and red blood cell exudation in the lung tissue were found in model group and agomir negative control group， and the above pathomorphological changes of lung tissue of the rats in miR-146b agomir group were alleviated. The expression levels of ICAM-1 mRNA and protein of the rats in miR-146b agomir group were significantly lower than those in model group and agomir negative control group （P<0.01）， while the expression level of miR-146b was significantly higher than those in model group and agomir negative control group （P<0.01）.The dual luciferase reporter system experiment results confirmed that the miR-146b could targetedly regulate the ICAM-1 gene.

MiR-146b can reduce the inflammation level of lung tissue of the ARDS rats， relieve the lung tissue function damage， and protect the lung tissue from ARDS by targeted down-regulation of the expression level of ICAM-1.

To observe the effect of isorhynchophylline （IRN） on TNF-α-induced the apoptosis and the release of inflammatory factors in the human bronchial epithelial （16HBE） cells， and to explore its possible mechanism.

After treated with different concentrations［0（control group），2.5， 5.0， 10.0， 20.0， 30.0 and 40.0 μmol·L^－1］ of IRN for 24 h， the 16HBE cells were cultured with 20 mg·L^－1 TNF-α for 18 h. The 16HBE cells were divided into control group， TNF-α group （20 mg·L^－1）， IRN group （20 μmol·L^－1）， TNF-α+IRN group （20 mg·L^－1 TNF-α and 20 μmol·L^－1 IRN）， TNF-α+IRN+miR-192-5p inhibitor group （20 mg·L^－1 TNF-α， 20 μmol·L^－1 IRN and 50 nmol·L^－1 miR-192- 5p inhibitor） and TNF-α+IRN+pcDNA3.1 CXCR5 group ［20 mg·L^－1 TNF-α， 20 μmol·L^－1 IRN and 2 μmol·L^－1 pcDNA3.1-C-X-C chomokine receptor type 5（CXCR5）］. CCK-8 method was used to detect the viabilities of 16HBE cells in various groups； Real-time fluorescence quantitative PCR（RT-qPCR） method was used to detect the expression levels of miR-192-5p and CXCR5 mRNA in the 16HBE cells in various groups；Western blotting method was used to detect the expression levels of interleukins-1β （IL-1β）and monocyte chemoattactant protein-1 （MCP-1），B cell lymphoma-2（Bcl-2）， Cleaved-Caspase 3， CXCR5， and phosphorylated p38（p-P38）， c-Jun N-terminal kinase （JNK）， c-Jun， and nuclear factor-κB （NF-κB） p65 proteins in the 16HBE cells in various groups； ELISA method was used to detect the levels of IL-1β and MCP-1 in supernatant of the 16HBE cells in various groups. TargetScan7.1 website was used to predict the target genes and the targeted binding association between miR-192-5p and CXCR5 was verified by dual-luciferase reporter gene assay.

Compared with control group， the viability of the 16HBE cells in TNF-α group was significantly decreased （P<0.05）；compared with control group， the viabilities of 16HBE cells in 5.0，10.0，20.0， and 30.0 μmol·L^-1 IRN groups were increased （P<0.05）. Compared with control group， the expression levels of miR-192-5p and Bcl-2 proteins in the 16HBE cells in TNF-α group were significantly decreased （P<0.05）， the expression levels of IL-1β， MCP-1，Cleaved-Caspase-3， p-p38， p-JNK， p-c-Jun and p-NF-κB p65 proteins， the apoptotic rate and the levels of IL-1β and MCP-1 in supernatant of the 16HBE cells were significantly increased （P<0.05）； compared with TNF-α group， the expression levels of miR-192-5p and Bcl-2 protein in the 16HBE cells in TNF-α+IRN group were significantly increased （P<0.05）， the the expression levels of IL-1β， MCP-1，Cleaved-Caspase-3， p-p38， p-JNK， p-c-Jun and p-NF-κB p65 proteins， the apoptotic rate and the levels of IL-1β and MCP-1 in supernatant of the 16HBE cells were significantly decreased （P<0.05）； compared with TNF-α+IRN+NC inhibitor group， the expression levels of miR-192-5p and Bcl-2 protein in the 16HBE cells in TNF-α+IRN+ miR-192-5p inhibitor group were significantly decreased （P<0.05），the expression levels of IL-1β， MCP-1，Cleaved-Caspase-3， p-p38， p-JNK， p-c-Jun and p-NF-κB p65 proteins，and the apoptotic rate and the levels of IL-1β and MCP-1 in supernatant of the 16HBE cells were significantly increased （P<0.05）. Compared with NC mimic and NC inhibitor groups， the expression levels of CXCR5 mRNA and protein in the 16HBE cells in miR-192-5p mimic group were significantly decreased （P<0.05）， and the expression levels of CXCR5 mRNA and protein in the 16HBE cells in miR-192-5p inhibitor group were significantly increased （P<0.05）. Compared with control group， the expression level of CXCR5 protein in the 16HBE cells in TNF-α group was significantly increased （P<0.05）， the cell viability was decreased （P<0.05）， and the apoptotic rate and the levels of IL-1β and MCP-1 in supernatant of the 16HBE cells were increased （P<0.05）；compared with TNF-α group， the expression level of CXCR5 protein in the 16HBE cells in TNF-α+IRN group was significantly decreased （P<0.05），the cell viability was increased （P<0.05），and the apoptotic rate and the levels of IL-1β and MCP-1 were significantly decreased （P<0.05）； compared with TNF-α+IRN+ pcDNA3.1 group， the expression level of CXCR5 protein in the 16HBE cells in TNF-α+IRN+ pcDNA3.1 CXCR5 group was significantly increased （P<0.05），the cell viability was decreased （P<0.05），and the apoptotic rate and the levels of IL-1β and MCP-1 in supernatant of the 16HBE cells were increased （P<0.05）.

IRN can reduce the TNF-α-induced apoptosis and release of inflammatory factors in the human bronchial epithelial cells， and its mechanism may be related to the miR-192-5p/MAPKs/NF-κB signal pathway.

To investigate the effect of centromere protein U （CENPU） on cisplatin resistance of ovarian cancer（OC） cells， and to analyze its mechanism.

The expression levels of CENPU protein in the OC cells （OVCAR3 and SKOV3 cells） and cisplatin resistant OC cells （OVCAR3/DDP and SKOV3/DDP cells） were detected by Western blotting method. The OVCAR3/DDP and SKOV3/DDP cells were divided into shControl group （transfected with shControl plasmid） and CENPU short hairpin RNA plasmid（shCENPU） group （transfected with shCENPU plasmid）.The cell viabilities in two groups after stimulated with 0－100 μmol·L^－1 cisplatin for 24 h were detected by CCK-8 assay； after stimulated with 20 μmol·L^－1 cisplatin for 24 h， the apoptotic rates of cells in two groups were detected by flow cytometry； the sphere formation efficiencies （SFE） of the cells in two groups were detected by tumor sphere formation assay； the expression levels of self-renewing related gene sex determining region Y （SRY）-related high-mobility-group （HMG）-box protein-2 （Sox2）， Nanog and octamer-binding transcription factor-4 （Oct4） mRNA in the cells in two groups were detected by Real-time fluorescence quantitative PCR（RT-qPCR） method； the expression levels of Wnt/β-catenin pathway related proteins wingless type MMTV integration site family member 1 （Wnt1）， cyclinD1， c-Myc， and β-catenin in the cells in two groups were detected by Western blotting method.

Compared with the OVCAR3 and SKOV3 cells， the expression levels of CENPU protein in the OVCAR3/DDP and SKOV3/DDP cells were significantly increased （P<0.05）. After treated with cisplatin， compared with shControl group， the viabilities of OVCAR3/DDP and SKOV3/DDP cells in shCENPU group were significantly decreased（P<0.05）， and the apoptotic rates were significantly increased （P<0.05）. Compared with shControl group， the SFE and the expression levels of Sox2， Nanog， and Oct4 mRNA， and the expression levels of Wnt1， cyclinD1， c-Myc， and β-catenin proteins in the OVCAR3/DDP and SKOV3/DDP cells in shCENPU group were significantly decreased（P<0.05）.

CENPU knockdown can inhibit the self-renewal，cisplatin resistance， and Wnt/β-catenin signaling activity in cisplatin resistant OC cells.

To investigate the effect of betulinic acid（BA） on the proliferation and apoptosis of the myeloma cells through the Janus kinase 2（JAK2）/signal transducers and activators of transcription 3（STAT3） signaling pathway and its mechanism.

The myeloma U266 cells were treated with different concentrations（0，20，40，60，80，100，120，and 160 μmol·L^－1） of BA as different concentrations of BA groups， at the same time，blank group （without BA and without cells） was set up，and the U266 cells without BA treatment were used as control group.The proliferation rates of the U266 cells in various groups were detected. The U266 cells were divided into control group and 20， 40，and 60 μmol·L^－1 BA groups. The proliferation rates of the cells in various groups were detected by CCK-8 method；the apoptotic rates of the cells in various groups were detected by Annexin Ⅴ-FITC/PI double staining method； the expression levels of c-myc oncogene（c-Myc），cell cycle proteins D1（cyclin D1），B cell lymphoma-2（Bcl-2），Bcl-2 associated X protein （Bax） and JAK2/STAT3 signaling pathway proteins in the cells in various groups were detected by Western blotting method.The nude mice were divided into control group（given 100 μL DMSO）and 20，30 and 40 mg·kg^－1 BA group（given 20，30，40 mg·kg^－1 BA）；the weights and volumes of tumor of the nude mice in various groups were observed.

Compared with control group， the proliferation rate of the myeloma U266 cells at 48 and 72 h after treatment，the expression levels of C-MYc， cyclin D1， Bcl-2， and JAK2/STAT3 signaling pathway proteins in the cells and the tumor weight of nude mice in 20，40， and 60 μmol·L^－1 BA groups were significantly decreased（P<0.05）， while the apoptotic rates and the expression levels of Bax protein were significantly increased （P<0.05）.Forty-eight and 72 h after treatment，compared with 20 μmol·L^－1 BA group，the proliferation rates of the myeloma U266 cells in 40 and 60 μmol·L^－1 BA groups， and the expression levels of c-MYc， cyclin D1， Bcl-2，and JAK2/STAT3 signaling pathway proteins in the cells in 40 and 60 μmol·L^－1 BA groups were significantly decreased（P<0.05）， while the apoptotic rates and the expression levels of Bax protein were significantly increased （P<0.05）；compared with 40 μmol·L^－1 BA group， the proliferation rates of the cells in 60 μmol·L^－1 BA group，the expression levels of c-MYc， cyclin D1， Bcl-2， and JAK2/STAT3 signaling pathway proteins in the cells in 60 μmol·L^－1 BA group were significantly decreased（P<0.05）， while the apoptotic rates and the expression level of Bax protein were significantly increased （P<0.05）.Compared with control group， the tumor weights and volumes of the nude mice in 20， 30，and 40 mg·kg^－1 BA groups were significantly decreased （P<0.05）. Compared with 20 mg·kg^－1 BA group， the tumor weight and volume of the nude mice in 30 and 40 mg·kg^－1BA groups were significantly decreased （P<0.05）.Compared with 30 mg·kg^－1 BA group， the tumor weight and volume in 40 mg·kg^－1 BA group were significantly decreased （P<0.05）.

BA can promote the apoptosis of myeloma cells and inhibit the proliferation of myeloma cells and the growth of transplanted tumor in the nude mice. The mechanism may be related to the inhibition of JAK2/STAT3 signaling pathway activation by BA.

To investigate the effects of baicalin （BAL）combined with metformin（DMBG） on the endocrine indicatros， inflammatory factors， endometrial condition and insulin resistance of the rats with polycystic ovary syndrome （PCOS） and their mechanisms.

A total of 32 rats were divided into control group， dehydroepiandrosterone（DHEA） group， DMBG group， and BAL + DMBG group，and there were 8 rats in each group.Except control group，the rats in other groups were given DHEA to establish the PCOS models.The blood of abdominal aorta of the rats was taken， and the levels of serum endocrine factorsluteinizing hormone（LH）， follicle-stimulating hormone（FSH）， testosterone（T），estradiol（E2）and tumor necrosis factor-α（TNF-α），C-reactive protein（CRP）， interleukin 6（IL-6）and interleukin 18（IL-18）were determined by enzyme-linked immunosorbent assay （ELISA）method； radioimmunoassay was used to detect the levels of fasting insulin（FINS）and fasting blood-glucose（FBG），and the insulin resistance index（HOMA-IR） was calculated；the area ratio of endometrial space to stroma and thickness of endometrium of the rats in various groups were detected；Western blotting method was used to detect the expression amounts of phosphorylated phosphatidylinositide 3-kinases （p-PI3K），phosphatidylinositide 3-kinases（PI3K）， phosphorylated protein kinase B （p-AKT） and protein kinase B （AKT） in ovary tissue of the rats in various groups.

Compared with control group， the serum levels of FSH and E2 of the rats in DHEA group were decreased（P<0.05），and the serum levels of LH，T， TNF-α， CRP， IL-6，and IL-18 were increased （P<0.05），the serum levels of LH，T，FBG，FINS and HOMA-IR were increased （P<0.05），and the area ratio of endometrial space to stroma and the thickness of endometrium were decreased（P<0.05）.Compared with DHEA group， the serum levels of FSH， E2， TNF-α， CRP， IL-6，and IL-18 of the rats in BAL+DMBG group and DMBG group were increased （P<0.05），the serum levels of LH，T，FBG，FINS， and HOMA-IR were decreased （P<0.05），and the area ratios of endometrial space to stroma and the thicknesses of endometrium were increased（P<0.05）.Compared with DMBG group， the serum levels of FSH， E2， TNF-α， CRP， IL-6，and IL-18 of the rats in BAL+DMBG were increased （P<0.05），the serum levels of LH，T，FBG，FINS， and HOMA-IR were decreased （P<0.05）， and the area ratio of endometrial space to stroma and the thickness of endometrium were increased（P<0.05）.

BAL combined with DMBG can improve insulin resistance and endometrium condition by up-regulating the expressions of PI3K / AKT signaling pathway proteins in ovarian tissue of the PCOS rats，correcting endocrine disorders，and inhibiting the chronic low-grade inflammatory reaction.

To investigate the effect of miR-106b targeting transforming growth factor-β receptor1（TGF-βR1）on the invasion and migration of colon cancer cells， and to clarify the targeting relationship between miR-106b and TGF-βR1 and its possible mechanism.

A total of 30 cases of colon cancer tissue and adjacent normal colon tissue，the colon cancer SW-480 cells and normal colon epithelial NCM460 cells were selected. The Real-time fluorescence quantitative PCR（RT-qPCR）method was used to detect the expression levels of miR-106b and TGF-βR1 mRNA in different kinds of tissues and cells.The colon cancer SW-480 cells were transfected and divided into empty vector group （transfected with miR-NC），miR-106b group （transfected with miR-106b-mimics），pGL3-TGF-βR1 group （transfected with pGL3-TGF-βR1）， and miR-106b-mimics+pGL3-TGF-βR1 group （transfected with miR-106b-mimics and pGL3-TGF-βR1）. The level of miR-106b in different kinds of tissues and cells，and the expression levels of TGF-βR1 mRNA in different kinds of cells were detected by RT-qPCR method；the targarc relationship between miR-106b and TGF-βR1 was predicted by bio-information analysis method.The luciferase activities of SW-480 cells in various groups were detected by luciferase reporter assay；the invasion abilities of the colon cancer SW-480 cells in various groups were detected by Transwell chamber test； the wound healing rates of the colon cancer SW-480 cells in various groups were detected by cell scratch test； the expression levels of TGF-βR1， phosphorylated Smad family member 2 （p-Smad2） and phosphorylated Smad family member 3（p-Smad3） proteins in colon cancer SW-480 cells in various groups were detected by Western blotting method.

The level of miR-106b in the colon cancer tissue and colon cancer SW-480 cells were lower than those in normal colon tissue and normal colon epithelial NCM460 cells （P<0.01）.The results of double luciferase reporter gene analysis showed that TGFβR1 was the target gene of miR-106b.Compared with empty vector group，the expression levels of TGF-βR1 mRNA and protein in the colon cancer SW-480 cells in miR-106b-mimics+pGL3-TGF-βR1 group were significantly decreased（P<0.01）；the number of invasion cells and wound healing rate in the colon cancer SW-480 cells were significantly increased （P<0.01），and the expression levels of p-Smad2 and p-Smad3 proteins in the colon cancer SW-480 cells were decreased （P<0.01）.Compared with miR-106b group，the expression levels of TGF-βR1 mRNA and protein in the SW-480 cells in miR-106b-minics+PGL3-TGF-βR1 group were significantly increased（P<0.01），the number of invasion cells and the scrath healing rate were decreased（P<0.01），and the expression levels of p-Smad2 and p-Smad3 proteins in the cells were significantly increased（P<0.01）.

Over-expression of miR-106b may promote the invasion and migration of the colon cancer cells expression by inhibiting the TGF-β / Smad pathway.

To investigate the effect of metformin on the proliferation of the non-small cell lung cancer A549 cells ，and to clarify its possible mechanism.

The A549 cells were divided into control group and different concentrations （1， 2， 4， 8， 16 and 32 mmol·L^－1） of metformin groups.MTT method was used to detect the proliferation rates of A549 cells in various groups after treated for 24， 48 and 72 h； clone formation experiment was used to detect the clone formation number of A549 cells in various groups； cell scratch test was used to detect the migration rates of A549 cells in various groups； RT-PCR method was used to detect the expression levels of phosphoinositide-3-kinase （PI3K）， protein kinase B （AKT）， and glycogen-synthase kinase-3β （GSK3β） mRNA in the A549 cells in various groups； Western blotting method was used to detect the expression levels of PI3K， PI3K（p-PI3K）， AKT， phosphorylated AKT（p-AKT）， GSK3β ，and phosphorylated GSK3β （p-GSK3β） proteins in the A549 cells in various groups.

Compared with control group， the proliferation rates of lung cancer A549 cells in different concentrations of metformin groups after treated for 24， 48 and 72 h were decreased （P<0.05）； compared with control group， the number of clone formation in the A549 cells in 2 and 8 mmol·L^－1 metformin groups was decreased （P<0.05）； compared with control group， the migration rates of the A549 cells in 2 and 8 mmol·L^－1 metformin groups were decreased （P<0.05）； compared with control group， the expression levels of PI3K， AKT and GSK3β mRNA in the A549 cells in 2 and 8 mmol·L^－1 metformin groups were significantly decreased （P<0.05）； compared with control group， the expression levels of PI3K， p-PI3K， p-AKT， and p-GSK3β proteins in the A549 cells in 2 and 8 mmol·L^－1 metformin groups were significantly decreased （P<0.05）.

Metformin can inhibit the proliferation and migration of lung cancer A549 cells， and its mechanism may be related to the inhibition of PI3K/AKT/GSK3β signaling pathway.

A total of 40 young rats were used to establish the RE models and then they were randomly divided into model group （n=8），low dose of Cornus officinalis polysaccharide group （n=9），high dose of Cornus officinalis polysaccharide group（n=10）， and positive control group （n=10）；sham operation group was set up at the same time（n=10）. The young rats in low and high doses of Cornus officinalis polysaccharide groups were given 0.14 and 0.56 g·kg^－1Cornus officinalis polysaccharide by gavage，the young rats in positive control group were given 154 mg·kg^－1 lamotrigine， and the young rats in control group and model group were given the same amount of normal saline. The epoleptic behavioral changes of the young rats in various groups were observed；the patholomorphology of hippocampus tissue of the young rats in various groups were observed by HE staining；the serum levels of tumor necrosis factor-α （TNF-α） and interleukin-1β （IL-1β） of the young rats in various groups were detected by ELISA method， and expression levels of nuclear factor-κB （NF-κB）， MDR1b， and MVP mRNA and proteins in hippocampus tissue of the young rats in various groups were detected by RT-qPCR and Western blotting methods.

Compared with sham operation group， the total number of seizures of the young rats in model group was increased（P<0.05），the total duration was prolonged（P<0.05）， the serum levels of TNF-α and IL-1β were increased（P<0.05），and the expression levels of NF-κB， MDR1b， and MVP mRNA and proteins in hippocampus tissue of the young rats were increased （P<0.05）. Compared with model group， the total number of seizures of the young rats in low and high doses of Cornus officinalis polysaccharide groups and positive control group was decreased（P<0.05），the total duration was shortened（P<0.05）， the serum levels of TNF-α and IL-1β were decreased （P<0.05）， and the expression levels of MDR1b， MVP mRNA and proteins in hippocampus tissue of the young rats were decreased （P<0.05）.The HE staining results showed that the morphology and structure of the cells in hippocampus area of the young rats in sham operation group were normal； the cells in hippocampus area of the young rats in model group were arranged disorderly， a large number of cells showed vacuole-like changes， and a large number of nuclear ruptures and nuclear pyknosis were seen； the cells in hippocampus area of the young rats in low and high doses of Cornus officinalis polysacharide groups and positive control group were arranged neatly and the cell outline was clear， with a small amount of vacuolar degeneration， nuclear rupture， and nuclear pyknosis.

Cornus officinalis polysaccharide can improve the seizure behavior of RE young rats， and its mechanism may be related to the inhibition of MDR1b and MVP mRNA and protein expressions.

To investigate the inhibitory effect of mini-chromosome maintenance protein 10（MCM10） silencing on the proliferation of breast cancer cells，and to elucidate its mechanism.

The human triple negative breast cancer （TMBC）MDA-MB-231 cells were divided into control group，MCM10 interference group 1 （shMCM10-1 group）and MCM10 interference group 2（shMCM10-2 group）. The lentivirus interference vectors of MCM10-1 group and MCM10-2 were constructed and infected into the MDA-MB231 cells in shMCM10-1 group and shMCM10-2 group，respectively. The MDA-MB-231 cells in control group were infected with the negative control virus. The expression levels of MCM10 in the MDA-MB-231 cells in various groups were measured by Real-time fluorescence quantitative PCR（RT-qPCR） and Western blotting methods； the proliferation of the MDA-MB-231 cells in various groups was monitored by Cellomics Count assay；the apoptotic rates of the MDA-MB-231 cells in various groups were detected by Flow cytometry， and the activities of Caspase3/7 in the MDA-MB-231 cells in various groups were measured by Caspase 3/7 activity detection kit.

The expression levels of MCM10 mRNA in the MDA-MB-231 cells in shMCM10-1 group and shMCM10-2 group were significantly lower than that in control group （P<0.01）， and the expression of MCM10 protein was not found.Compared with control group， the relative proliferation multiples of the MDA-MB-231 cells in shMCM10-1 group and shMCM10-2 group were significantly decreased （P<0.01），and the apoptotic rates of the MDA-MB-231 cells in shMCM10-1 group and shMCM10-2 group were significantly increased （P<0.01），and the activities of Caspase 3/7 in the MDA-MB-231 cells in shMCM10-1 group and shMCM10-2 group were significantly increased （P<0.01）.

MCM10 gene silencing can inhibit the proliferation of the MDA-MB-231 cells， and promote the apoptosis of the MDA-MB-231 cells and increase the activities of Caspase3/7 in the MDA-MB-231 cells.

To investigate the effect of exosomes carrying microRNA-196b-5p secreted by the bone marrow mesenchymal stem cells （MSCs） on the biological characteristics of the colon cancer cells.

The MSCs were isolated and cultured from bone marrow of the mice. The mi-196b-5p mimic and mi-196b-5p inhibitor were transfected into the MSCs. The culture medium was collected and the exosomes were separated. The morphology of exosomes was observed under transmission electron microscope.After identification of exosomes by expression of specific markers， the colon cancer CT26.WT cells of the mice were intervened with the exosomes. The colon cancer CT26.WT cells of the mice at logarithmic growth phase after resuscitation were divided into medium group， mimic-NC group， inhibitor-NC group，exosome group， mimic-exosome group and inhibitor-exosome group.The proliferation activities of the colon cancer CT26.WT cells in various groups were measured by MTT assay；the percentages of the colon cancer CT26.WT cells at different cell cycles in various groups were detected by flow cytometry；the apoptotic rates of the colon cancer CT26.WT cells in various groups were detected by flow cytometry； the number of migration and invasion colon cancer cells in various groups was evaluated by Transwell chamber assay.

Compared with exosome group， the proliferation activity， the number of migration cells and invasion cells of the colon cancer CT26.WT cells in mimic-exosome group were decreased （P<0.05）， and the percentage of colon cancer CT26.WT cells at G₁ phase and the apoptotic rate were increased（P<0.05）；the proliferation activity， the number of migration and invasion cells of colon cancer CT26.WT cells in inhibitor-exosome group were increased （P<0.05），and the apoptotic rate was decreased（P<0.05）.

The exosomes derived from MSCs carring a high level of miR-196b-5p can inhibit the proliferation， migration and invasion of the colon cancer CT26.WT cells， and induce the apoptosis.

The expression matrix of AML （GSE96535） was downloaded from NCBI Gene Expression Database （GEO）.The differentially expressed mRNA and long non-coding RNAs（lncRNAs） were screened using the edgeR package of R language. miRcode， miRDB， miRTarBase ，and TargetScan databases were used to predict the regulatory relationships between the differentially expressed lncRNA and miRNA， and the differentially expressed mRNA and miRNA. Cytoscape software was utilized for the construction of the ceRNA regulatory network of lncRNA-miRNA-mRNA.Gene Ontology（GO） and Kyoto Encyclopedia of Genes and Genomes（KEGG） were performed to conduct the enrichment analysis of the differentially expressed genes. The core gene， Hub gene， was screened by using CytoHubba plug-in. The GEPIA database was used to compare the expressions of the top 10 Hub genes in the AML patients and normal controls， and then the survival curve of Hub gene was plotted.

Compared with normal controls，a total of 1 105 mRNAs and 387 lncRNAs were differentially expressed in the AML patients. The constructed lncRNA-miRNA-mRNA ceRNA regulatory network consisted of 45 lncRNAs， 31 miRNAs and 89 mRNAs.The GO analysis results showed that the differentially expressed genes were mainly concentrated in the membrane microdomain and glutamate receptor binding， and regulated epithelial cell proliferation.The KEGG analysis results revealed that 19 pathways were enriched， including transcriptional regulation， proteoglycan regulation， and Ras signaling pathway. GATA binding protein 3（GATA3），WT1 transcription factor （WT1）， peroxisome proliferator activated receptor gamma （PPARG） and other top 10 Hub genes with the highest connectivity were screened by CytoHubba. The GEPIA database was performed to verify that GATA3， WT1， MYB proto-oncogene （MYB）， SRY-box transcription factor 4（SOX4） and E2F7 had the differential expression levels. Moreover， the survival rate of the patients in E2F7 low-expression group was higher than that in E2F7 high-expression group（P=0.028）.

The screened ceRNA network composed of lncRNA， miRNA and mRNA may promote the occurrence and development of AML.

To investigate the effect of miR-222-3p on the proliferation and apoptosis of thyroid cancer cells via regulating gene of phosphatase and tension homolog deleted on chromosome ten（PTEN），and to elucidate the role of miR-222-3p in ¹³¹I radiotherapy resistance of thyroid cancer cells and its mechanism.

The expression levels of miR-222-3p in the human thyroid cancer cells and human normal thyroid follicular epithelial Nth-ori 3-1 cells were determined by Real-time fluorescence quantitative PCR（RT-qPCR） method.The thyroid cancer K1 cells and ¹³¹I resistance of thyroid cancer K1R cells were used as the subjects and were divided into blank control group， ¹³¹I treatment group， ¹³¹I+miR-222-3p knockdown group， ¹³¹I+PTEN over-expression group，and ¹³¹I+PTEN over-expression+miR-222-3p over-expression group. The K1 cells and KIR cells in blank control group didn’t receive any treatment； both the K1 and K1R cells in ¹³¹I treatment group were treated with ¹³¹I at doses of 1 and 3 Gy； the cells in ¹³¹I+miR-222-3p knockdown group were transfected with miR-222-3p inhibitor prior to ¹³¹I treatment at dose of 1 and 3 Gy； the cells in ¹³¹I+PTEN over-expression group were transfected with pcDNA3.1-PTEN prior to ¹³¹I treatment with doses of 1 and 3 Gy； the cells in ¹³¹I+PTEN over-expression+miR-222-3p over-expression group were simultaneously transfected with pcDNA3.1-PTEN and miR-222-3p mimic prior to ¹³¹I treatment with doses of 1 and 3 Gy. Clony formation assay was used to detect the clony formation number of K1 and K1R cells；CCK-8 assay was used to detect the proliferation activities of K1 and K1R cells；Annexin Ⅴ-FITC / PI double staining method was used to detect the apoptotic rates of K1 and K1R cells；Western blotting method was used to detect the expression levels of PTEN protein in K1 and K1R cells； Double luciferase reporter gene assay was used to verify the targeting relationship between miR-222-3p and PTEN.

Compared with the Nth-ori 3-1 cells，the expression levels of miR-222-3p in thyroid carcinoma cells were increased （P<0.01）； the expression level of miR-222-3p in the thyroid cancer K1R cells was higher than that in thyroid cancer K1 cells （P<0.01）. Compared with ¹³¹I treatment group， the clony formation number of the K1 and K1R cells in ¹³¹I + miR-222-3p knockdown group was decreased（P<0.01）， the proliferation activities of K1 and K1R cells were decreased （P<0.01），and the apopotic rates were increased（P<0.01）. Compared with miR-NC-PTEN-WT group， the luciferase activity of HEK-293 cells in miR-222-3P-PTEN-WT group was decreased （P<0.01）. Compared with before knockdown， the expression level of PTEN protein in K1 and K1R cells were significantly increased after knockdown（P<0.01）. Compared with ¹³¹I + PTEN over-expression group， the colony formation number of K1 and K1R cells in ¹³¹I +PTEN over-expression+miR-222-3p over-expression group was increased （P<0.01）， the proliferation activities were increased （P<0.01）， and the apoptotic rate was decreased （P<0.01）.

Knockdown of miR-222-3p targeting PTEN and up-regulating its expression level can alleviate the ¹³¹I radiotherapy resistance of thyroid cancer cells.

To discuss the effect of C19ORF12 on the proliferation and chemo-sensitivity of the gastric cancer MKN45 cells，and to clarify its mechanism.

The gastric cancer MKN45 cells were divided into control group and C19ORF12 group. The proliferation activities of the gastric cancer MKN45 cells in two groups were detected by CCK-8 method； the number of colony formation of the gastric cancer MKN45 cells in two groups was detected by colony formation assay；the survival rates of the gastric cancer MKN45 cells after treated with chemotherapeutic drugs doxorubicin or hydrogen peroxide（H₂O₂） in two groups were detected by CCK-8 method； the apoptotic rates of the gastric cancer MKN45 cells in two groups were detected by flow cytometry； the subcellular localization of C19ORF12 in the gastric cancer MKN45 cells was detected by laser confocal microscope.The expression difference of C19ORF12 gene in Cancer Genome Atlas （TCGA） between normal gastric and gastric cancer tissues was analyzed by GEPIA.The effect of different expression level of C19ORF12 on the prognosis of gastric cancer patients was retrospectively analyzed by KM-plot.

Compared with control group， the proliferation activity and the number of colone formation of the gastric cancer MKN45 cells in C19ORF12 group were significantly increased （P<0.01）. Compared with control group， the survival rate of gastric cancer MKN45 cells in C19ORF12 group after treated with doxorubicin or H₂O₂ were increased（P<0.05 or P<0.01），and the apoptotic rates were decreased（P<0.01）. The C19ORF12 was mainly expressed in the mitochondrion.The bioinformatics analysis results showed that compared with normal gastric tissue， the level of C19ORF12 mRNA in gastric cancer tissue was increased（P<0.01）. Compared with the patients with low expression of C19ORF12， the overall survival time of gastric cancer patients with high expression of C19ORF12 was significantly decreased（P<0.01）.

C19ORF12 can promote the proliferation of gastric cancer cells and reduce the sensitivity of gastric cancer cells to chemotherapeutic drugs. C19ORF12 can be used as a potential target for tumor therapy.

To explore the effect of Carisolv chemomechanical method in treatment of deciduous tooth caries in the children， and to provide the evidence-based medical basis for the extensive application of this method in the clinical practice.

PubMed， CNKI， Medline， Wiley Online Library， Wanfang and Weipu databases were searched by computer from inception to October 31，2020.Meta-analysis was conducted on the complete randomized controlled trials that met the inclusion criteria.

A total of 11 completely randomized controlled trials involving 1 051 teeth of 645 patients were included. The painless rate of Carisolv chemomechanical method was significantly higher than other traditional methods（OR=4.84，95%CI：3.44－6.81，Z=9.04，P<0.01），and the incidence of postoperative complications of the patients treated with Carisolv chemomechanical method were lower than those of other traditional methods （RR=0.37，95%CI： 0.21－0.63，Z=3.60，P=0.0003）.The pediatric patients were more inclined to Carisolv chemomechanical method when choosing the retreatment method， but the difference in preference was not statistically significant（OR=16.99，95% CI：1.50－192.31，Z=2.29，P=0.02）.

The pediatric patients treated with Carisolv chemomechanical method are less likely to suffer from pain and complications， and this method is more likely to be accepted by the children. Carisolv is a suitable technique for the caries of children’s deciduous teeth.

To evaluate the effect of multiple-dose gonadotropin-releasing hormone agonist（GnRH-a） addition to luteal support in pregnancy outcomes and safety of the patients with first in-vitro fertilization-embryo implantation （IVF-ET） and their offsprings， and to guide the IVF-ET patients to use GnRH-a scientifically.

A total of 106 patients with the first IVF-ET were randomly assigned into control group （n=53） and multiple-dose GnRH-a group （n=53）. The patients in control group were given standard luteal support treatment， while the patients in multiple-dose GnRH-a were given standard luteal support plus GnRH-a at day 2， day 5 and day 8 after egg retrieval. The pregnancy outcomes including pregnancy rates， clinical pregnancy rates and live birth rates of the patients were compared between two groups， and the delivery outcomes included premature birth rate， low-birth weight birth rate and birth defect rate of the patients were compared between two groups. The serum progesterone levels of the patients in two groups were detected. The offsprings of the patients were followed up，and the Bayley Infant Development Scale （Bayley Ⅲ） and Child Behavior Checklist （CBCL） were used to evaluate the exercise， cognitive abilities， language and behavioral development of the children.

The pregnancy rate， clinical pregnancy rate， and live birth rates of the patients in multiple-dose GnRH-a group were statistically higher than those in control group （P<0.05）. There were no significant differences in the rates of premature birth， low-birth weight and birth defects of the patients between two groups （P>0.05）.On the 14th day after embryo implantation， the serum progesterone level of the patients in multiple-dose GnRH-a group was significantly higher than that in control group （P<0.05）.The subgroup analysis results showed that on the 14th day after embnyo implantation，the serum progesterone levels of the pregnancy and non-pregnant patients in multiple-dose GnRH-a group were significantly higher than those in control group， respectively （P<0.05）.There were no significant differences in Bayley Ⅲ score and CBCL score of the patients between two groups at 24th month （P>0.05）.

Addition of multiple-dose GnRH-a to standard lutal support treatment can achieve better pregnancy outcomes with sustained better safety.

To explore the application values of simplified Wells score， revised Geneva score，and D-dimer（DD） level in the diagnosis of malignant tumor complicated with pulmonary thromboembolism （PTE），and to provide the reference for PTE diagnosis in clinic.

The clinical data of 142 cases of malignant tumor patients suspected of PTE and underwent CT pulmonary angiography （CTPA） examination were analyzed retrospectively. The diagnosis of PTE was based on the results of CTPA， and 72 patients with malignant tumor complicated with PTE were regarded as PTE group and 70 patients with malignant tumor without PTE were regarded as non-PTE group.The risk factors of all the enrolled patients were investigated through the statistical analysis，the receiver operating characteristic curves（ROC curves） were generated and the Z test was applied to compare the area under the ROC curve （AUC） of the above scores， and their values in the dignosis of malignant tumor complicated with PTE were evaluated.

Adenocarcinoma，tumor metastasis，deep venous thrombosis （DVT） and positive DD were the independent risk factors for PTE in the patients with malignant tumor （P<0.05）.The sensitivities of simplified Wells score，revised Geneva score， and DD level were 87.5%，87.5%， and 97.2%，respectively；the negative predictive values were 74.3%， 70.0%， and 83.3%，respectively. The sensitivities of the two clinical score scales combined with DD level reached 100%，the misdiagnostic rate was lower，and the negative predictive value reached 100% which was of great value for exclusion. The AUC of simplified Wells score，revised Geneva score，DD level， revised Geneva score combined with DD level，and simplified Wells score combined with DD level were 0.762，0.687，0.649，0.741，and 0.786，respectively.The AUC of simplified Wells score combined with DD level was the greatest， which was larger than those of the other scales （P<0.05）.

Simplified Wells score combined with DD level shows a higher value in the diagnosis of malignant tumor complicated with PTE.

To investigate the effect of miR-30a-5p on 5-fluorouracil （5-FU） resistant of colorectal cancer（CRC） cells by targeting tripartite motif-containing protein 31 （TRIM31） gene， and to clarify its mechanism.

The CRC tissue of 27 patients with 5-FU chemotherapy-sensitive colorectal cancer （5-FU/S group） and 23 patients with 5-FU chemotherapy-resistant colorectal cancer （5-FU/R group） were collected. The expression levels of miR-30a-5p and TRIM31 mRNA in CRC tissue of the patients were detected by Real-time fluorescence quantitative PCR（RT-qPCR） method； the correlation between miR-30a-5p and TRIM31 mRNA expressions was analyzed by Pearson correlation analysis.The human CRC 5-FU resistant cells HT-29/5-FU cells and its parent HT-29 cells were cultured in vitro. The proliferation activities of two kinds of cells were detected by MTT assay， and the median inhibitory concentration （IC₅₀） and drug resistance index （RI） were calculated； the expression levels of miR-30a-5p and TRIM31 mRNA in two kinds of cells were detected by RT-qPCR method；the expression levels of TRIM31 protein in two kinds of cells were detected by Western blotting method. The HT-29/5-FU cells were further divided into blank control group， mimics NC group， miR-30a-5p mimics group， miR-30a-5p mimics+vector group and miR-30a-5p mimics+TRIM31 group. The cells in blank control group were not transfected， the cells in miR-30a-5p mimics group were transfected with miR-30a-5p mimics， the cells in miR-30a-5p mimics+vector group were transfected with miR-30a-5p mimics and negative control of vector， and the cells in miR-30a-5p mimics+TRIM31 group were transfected with miR-30a-5p mimics and TRIM31 over-expression vector. The expression levels of miR-30a-5p and TRIM31 mRNA in the HT-29/5-FU cells in various groups were detected by RT-qPCR method； the expression levels of TRIM31 protein in the HT-29/5-FU cells in various groups were detected by Western blotting method； the proliferation activities of the HT-29/5-FU cells in various groups were detected by MTT method； the apoptotic rates of the HT-29/5-FU cells in various groups were detected by flow cytometry；the targeted relationship between miR-30a-5p and TRIM31 was verified by dual-luciferase reporter gene system.

Compared with 5-FU/S group， the expression level of miR-30a-5p in cancer tissue of the patients in 5-FU/R group was decreased （P<0.01），and the expression level of TRIM31 mRNA was increased （P<0.01）. The expression levels of miR-30a-5p and TRIM31 mRNA were negatively correlated （R²=0.885，P<0.01）. Compared with HT-29 cells， the expression level of miR-30a-5p in the HT-29/5-FU cells was decreased （P<0.01），and the expression levels of TRIM31 mRNA and protein were increased （P<0.01）. The IC₅₀ values of HT-29/5-FU and HT-29 cells were （104.41±0.22） and （9.82±0.31） mg·L^－1， respectively， and the RI value was 12.42. Compared with blank control group or mimics NC group， the expression level of miR-30a-5p in the HT-29/5-FU cells in miR-30a-5p mimics group was increased （P<0.01）， the expression level of TRIM31 protein was decreased （P<0.01）， the proliferation activity of HT-29/5-FU cells was significantly decreased （P<0.01），and the apoptotic rate of HT-29/5-FU cells was significantly increased （P<0.01）. Compared with miR-30a-5p mimics group， the expression level of TRIM31 protein in the HT-29/5-FU cells in imiR-30a-5p mimics+TRIM31 group was increased （P<0.01）， the proliferation activity of HT-29/5-FU cells was significantly increased （P<0.01），and the apoptotic rate of HT-29/5-FU cells was significantly decreased （P<0.01）.The dual-luciferase reporter system results confirmed that TRIM31 was the target gene of miR -30a-5p.

Over-expression of miR-30a-5p enhances the sensitivity of the CRC drug-resistance cells to 5-FU by targeting the TRIM31 gene expression.

To detect the serum B vitamin levels in the breast cancer patients who were receiving different chemotherapy regimens， and to clarify the differences in serum B vitamin levels in the breast cancer patients with different chemotherapy regimens and their clinical significances.

The clinical data of breast cancer chemotherapy patients （case group，n=62） and healthy subjects （control group，n=48） were retrospectively analyzed.According to the serum levels of vitamin B12（VitB12），vitamin B9（VitB9） and vitamin B6（VitB6）， the breast cancer patients were divided into six subgroups：VitB12<0.025 μg·L^－1 group and VitB12≥0.025 μg·L^－1group，VitB9<3.93 μg·L^－1group and VitB9≥3.93 μg·L^－1 group， VitB6<1.955 μg·L^－1 group and VitB6≥1.955 μg·L^－1 group.The baseline characteristics and chemotherapy regimens of breast cancer patients in different subgroups were analyzed.The patients in case group were divided into AC-T group（n=21），EC-T group（n=22），TC group（n=11），and the other chemotherapy regimen group （n=8） according to the chemotherapy regimen. In the first chemotherapy cycle， in the early morning of the second day after infusion of all chemotherapeutics， the fasting serum was collected from the patients， and the serum B vitamin levels of the subjects in two groups were detected by liquid chromatography-tandem mass spectrometry.The levels of serum vitamin B in breast cancer patients with different chemotherapy regimens and the chemotherapy regimens of breast cancer patients with different serum B vitamins levels were compared and analyzed.

The level of serum VitB9 in the subjets in control group was significantly higher than that in case group （t=－3.343，P<0.05）.The serum levels of VitB12 and VitB6 in the patients in case group had no significant differences compared with control group（P>0.05）. There was a statistically significant difference in menopause or not between VitB12<0.025 μg·L^－1 group and VitB12≥0.025 μg·L^－1 group（t=7.127，P=0.008）.There were no statistically significant differences in age， body mass index（BMI）， TNM stage， tumor size， lymph node metastasis， estrogen receptor （ER）， progesterone receptor （PR），human epidermal growth factor receptor-2（Her-2），Ki-67， and pathological classification between VitB12<0.025 μg·L ^－1group and VitB12≥0.025 μg·L ^－1 group（P>0.05）；there was a statistically significant difference in lymphnode metastasis between VitB6<1.955 μg·L^－1 group and VitB6≥1.955 μg·L ^－1 group（t=6.057，P=0.014）；there were no statistically significant differences in age， BMI， menopause or not ， TNM stage， tumor size， ER， PR， Her-2， Ki-67， and pathological types between VitB6<1.955 μg·L^－1 group and VitB6≥1.955 μg·L^－1 group （P>0.05）.There were no significant differences in age， menopause or not， BMI， TNM stage， tumor size，lymphnode metastasis， ER，PR，Her-2，Ki-67， and pathological classification between VitB9<3.93 μg·L^－1 group and VitB9≥3.93 μg·L^－1 group（P>0.05）. Among the 62 breast cancer patients included， 11 cases were treated with TC chemotherapy regimen， 21 cases were treated with AC-T chemotherapy regimen，22 cases were treated with EC-T chemotherapy regimen， and 8 cases were treated with other chemotherapy regimens；there were no significant differences in the serum levels of VitB12， VitB9 and VitB6 among the breast cancer patients received different chemotherapy regimens（P>0.05）.

There are no significant differences in the serum vitamin B levels in the breast cancer patients received different chemotherapy regimens， but there is a significant difference in the serum VitB9 level in the breast cancer chemotherapy patients compared with normal controls.

A total of 22 patients diagnosed with EOC for the first time and underwent cytoreductive surgery were regarded as drug susceptibility group，and the patients agreed to take the miniPDX trial and signed the relevant consent forms. In addition， 41 EOC patients treated at the same time， whose basic conditions were matched with the patients in drug susceptibility group， were selected as control group. The tumor tissue of the patients in drug susceptibility group was used to get the cell suspension and encapsulated，and were implanted under the skin of nude mice to establish the miniPDX models. The mice were treated with different chemotherapeutic drugs for a week， then the capsules were taken out. The proliferation rates of the tumor cells in the capsules were determined by CellTiter-Glo fluorescent cell viability assay. The chemotherapy regiment with the lowest proliferation rate was selected to treat the patients.The patients in control group were given paclitaxel + carboplatin or paclitaxel + cisplatin chemotherapy regiment.The efficacies and the incidences of adverse reactions of the patients in two groups were evaluated. In addition， the tumor sections of the patients in two groups were stained with immunohistochemical assay of CK7， WT-1， p16， p53， Ki-67 and paired box protein-8（PAX-8），and the relationships between their positive results and the sensitivities of the patients to platinum-combined chemotherapy were analyzed.

Among the 22 patients in drug susceptibility group， 14 patients were complete remission （CR），and 4 patients were partial remission （PR），and the overall remission rate （ORR） was 81.8%（18/22）； among the 41 patients in control group，13 patients were CR， and 14 patients were PR， the ORR was 65.9% （27/41）； the ORR of patients in drug susceptibility group was higer than that in control group （P<0.05）. There were no significant differences in age， pathological type and stage of the patients between two groups （P>0.05）. There were no significant differences in the incidences of adverse reactions of the patients between two groups （P>0.05）. The results of immunohistochemistry indicated that the percentages of platinum-sensitive patients with p16， p53， Ki-67， and PAX-8 positive were significantly higher than the patients with p16， p53， Ki-67， and PAX-8 negative （P<0.05）， and there were no significant differences between the percentages of platinum-sensitive patients with CK7 and WT-1 positive and the patients with CK7 and WT-1 negative （P>0.05）.

To screen EOC chemotherapy regimens based on miniPDX animal model can improve the therapeutic efficacy of EOC patients； p16， p53， Ki-67， and PAX-8 positive expression can predict the platinum sensitivity of the EOC patients .

The clinical data of patients with extensive defects of posterior teeth restored by chair-side CAD / CAM all-ceramic restoration were retrospectively analyzed. A total of 107 teeth were selected， and the prostheses were all with edge-to-edge margins. The types of prostheses were counted. All the teeth were assessed by using “the modified USPHS criteria” immediately and 12 months after operation. The proportions and relative ratios of grade A of the restorations immediately and 12 months after operation were assessed by these indicator，such as restoration integrity， color match， restoration retention， tooth integrity， proximal contact， and periodontium health.

There were 32 endo-onlays and 75 endo-overlays. The comparation results of the A-grade proportion of each index during the observation period immediately and 12 months after operation showed that the restoration integrity of onlays was decreased from 100% to 96.88%， and the color matching of onlays was increased from 84.38% to 93.75%. The restoration retention of overlays was decreased from proportions of grade A of 100% to 98.67%， and the color matching of overlays was increased from 89.33% to 92.00%. The tooth integrities， proximal contact， and periodontium health of two kinds of restorations all remained 100%.Among the patients who failed to repair， 1 case was restoration exfoliation and 1 case was restoration fracture.

The endo-onlays and endo-overlays with edge to edge margin show good short-term clinical efficacies in restoring extensive posterior tooth defect， which is an ideal repair method.

To analyze the clinical manifestations， imagical and pathological features and treatment methods of one patient with small cell carcinoma （SCC） of gallbladder， and to improve the clinicians’understanding of SCC.

The clinical materials of one patient with gallbladder SCC were collected， and the clinical features， diagnosis and treatment methods of gallbladder SCC were discussed combined with the related literature review.

The male patient aged 51 years old was admitted to hospital because of “upper abdominal pain with radiating pain in the lower back for more than half a month”. The abdominal CT results indicated the gallbladder mass. The liver artery and vein angiography CT examination results showed that the gallbladder neoplasm invaded the common bile duct with common bile duct and intrahepatic bile duct dilatation， multiple liver metastasis and retroperitoneal lymph node metastasis in hilar region. The tumor-associated antigen CA125 level was 65.2 U·mL^－1 and neuron-specific enolase （NSE） level was 23.5 μg·L^－1， and both the levels were increased.The patient underwent ultrasound-guided puncture biopsy of liver space-occupying lesions and was pathologically diagnosed as metastatic SCC. Combined with the medical history and clinical data， gallbladder SCC with liver and retroperitoneal lymph node metastasis was diagnosed.After diagnosis， the patient received 2 cycles of etoposide oral chemotherapy， 4 cycles of nedaplatin + etoposide（EP） intravenous chemotherapy rather than the surgical treatment.The re-examination of abdominal CT results revealed that the lesion was significantly reduced. The patient was currently in stable condition and was under close follow-up.

The clinical manifestations of gallbladder SCC are poorly specific and prone to distant metastasis. Pathological diagnosis and immunohistochemistry are the golden standards for the diagnosis of gallbladder SCC. In this case， the EP regimen of gallbladder SCC is effective and the tumor volume is significantly reduced. The patients with gallbladder SCC should be followed up regularly during the treatment process.

To compare the results of three-dimensional transrectal ultrasound （3D-ERUS） and magnetic resonance imaging （MRI） in the diagnosis of T substages and circumferential resection margin of the patients with middle and lower rectal cancer before operation，and to explore the diagnosis value of 3D-ERUS in the preoperative assessment of T substages and circumferential resection margin in the patients with middle and lower rectal cancer.

The data of 94 patients with rectal cancer confirmed by pathology were collected，and all the patients were examined by 3D-ERUS and MRI before operation. Kappa test was used to compare the consistency between the results of 3D-ERUS and MRI in the diagnosis of T substages and the pathological results.The specificity， sensitivity， positive predictive value （PPV）， negative predictive value （NPV），precision，Recall and F1 were used to evaluate the results of 3D-ERUS and MRI in the diagnosis of T substages and circumferential resection margin in the patients with middle and lower rectal cancer. The differences between 3D-ERUS and MRI in the diagnosis of T substages and circumferential resection margin in the patients with middle and lower rectal cancer were compared.

The accuracies of 3D-ERUS and MRI and the pathological results in the diagnosis of T substages in the patients with middle and lower rectal cancer were 81.91%（Kappa=0.736，P<0.01）and 74.47%（Kappa=0.624，P<0.01），respectively，and there was no statistical difference in the diagnostic accuracy of T substages in the patients with middle and lower rectal cancer between the two methods（χ²=1.12，P=0.289）.The sensitivity， specificity， PPV， NPV， accuracy， recall and F1 diagnosed by 3D-ERUS in the rectal cancer patients at T_is， T₁ and T₂ stages were higher than those of MRI. The sensitivity， specificity， PPV， NPV， accuracies， recall and F1 diagnosed by MRI in the patients with rectal cancer at T₄ stage were higher than those of 3D-ERUS. The accuracies of 3D-ERUS and MRI in the diagnosis of circumferential resection margin were 88.30% and 84.04%，and there was no statistical difference in the diagnostic accuracy of circumferential resection margin in the patients with middle and lower rectal cancer between two methods（χ²=0.402，P=0.526）.

3D-ERUS and MRI have high accuracies in the evaluation of preoperative T substages and circumferential resection margin in the patients with middle and lower rectal cancer. 3D-ERUS has high clinical value in the evaluation of T substages of the patients with early rectal cancer.

To discuss the associations between cardiometabolic multimorbidity（CMM） and disability in the middle-aged and older Chinese adults ，and to provide the evidence for the risk assessment of disability of the middle-aged and older Chinese adults.

The participants aged over 45 years old were selected as the subjects based on the China Health and Retirement Longitudinal Survey from 2011 to 2015. The descriptive statistics were presented for the demographic characteristics.The generalized estimating equation model was used to assess the associations between the CMM and disability.

This study included 18 051 participants with activities of daily living （ADL） assessment and 18 541 with instrumental activities of daily living （IADL） assessment， respectively. After adjusted for covariates， CMM was associated with the higher risks for ADL （OR=2.18， 95%CI： 1.95－2.43） and IADL （OR=1.64， 95%CI： 1.50－1.80） disabilities. Compared with the participants without cardiometabolic disease （CMD）， the risks for ADL disability were 1.29， 2.13 ，and 3.41 times， and the risks for IADL disability were 1.17， 1.60 and 2.26 times in the participants with 1， 2， and 3 or more kinds of CMDs.In the participants with 2， 3， and 4 kinds of CMDs， the patterns with the highest risk for ADL disability were hypertension and stroke （OR=6.00， 95%CI： 4.27－8.44），hypertension， stroke and dyslipidemia （OR=11.43， 95%CI： 6.27－20.80），hypertension， heart disease， stroke and dyslipidemia （OR=9.44， 95%CI：4.19－21.20）；the patterns with the highest risk for IADL disability were hypertension and stroke （OR=5.76，95%CI：4.07－8.17）， hypertension， heart disease and stroke （OR=6.93， 95%CI：3.45－13.90）， and hypertension， heart disease， stroke and dyslipidemia （OR=8.56， 95%CI： 4.27－17.10）.

CMM is positively associated with the risk of disability in the middle-aged and older Chinese adults.Different counts and combinations of CMD may have differences in the influence degrees in the disability among the middle-aged and older Chinese adults.

To develop a double-labeled time-resolved immunofluorescence analysis （TRFIA） method for the determination of the levels of β-cross-linked C-terminal telopeptide of type Ⅰ collagen（β-CTX） and N-terminal middle molecular fragment of osteocalein（N-MID） of osteocalcin，and to evaluate its detection performance.

The 4E5和2B7 monoclonal antibodies（MAb）of β-CTX and N-MID antigens were constructed， and then coated as coating antibodies in 96-well plates， and the antibodies were labeled with europium ion（Eu³⁺） and samarium ion（Sm³⁺），respectively. A double antibody sandwich TRFIA method was established and prepared into a kit. The detection performance was evaluated by the sensitivity， accuracy （dilution recovery）， specificity， precision， stability， and clinical sample alignment of the kit.

The prepared double-labeled TRFIA Kit had a detection sensitivity of 0.025 μg·L^－1 for β-CTX and a linear range of 0.025－5.000 μg·L^－1；the detection sensitivity for N-MID was 0.5 μg·L^－1， and the linear range was 0.5－200.0 μg·L^－1； the average dilution recovery rate of β-CTX was 102.13%， and the average dilution recovery rate of N-MID was 103.02%；there were no obvious cross-reaction with other common bone test indicators and the specificity was high.The intra-assay coefficient of variation（CV） of β-CTX was 5.81%－7.82%， and the inter-assay CV was 5.97%－8.02%； the intra-assay CV of N-MID was 6.05%－8.32%， and the inter-assay CV was 6.14%－8.56%； the TRFIA Kit could be stored stably at 4 ℃ for half a year and stably stored at 37 ℃ for 7 d.

The established β-CTX and N-MID double-labeled TRFIA method has the advantages of high sensitivity， high specificity， high accuracy， convenience and speed， and is suitable for the detection of large-scale clinical samples.

To develop a scalable purification process for production of exosomes derived from bovine milk amenable to both laboratory preparation and industrial scale production，and to evaluate the molecular and cellular biological activities of bovine milk exosomes.

A variety of full fat or skim milk samples were taken from the market. The gold standard of exosome extraction， ultracentrifugation （UC）， was used as the control. A variety of chemical pretreatment methods were used to remove the milk protein，and tangential flow ultrafiltration （TFF） was used to further remove the residual milk protein； the primary purification and concentration of milk samples were realized；the bovine milk exosomes were purified by biosize exclusion chromatography （BE-SEC）；nano flow cytometry（NanoFCM） was used to detect the particle size， concentration and purity of the bovine milk exosomes； the morphology of bovine milk exosomes was observed under transmission electron microscope（TEM）；the sizes and purities of the bovine milk exosomes were verified by high performance liquid chromatography（HPLC）； the expressions of specific proteins of bovine milk exosomes were detected by Western blotting method； Gene Ontology （GO） enrichment was used to analyze the candidate target genes of differentially expressed microRNA（miRNA）； DAVID Bioinformatics Resources 6.8 and KOBAS software were used to detect the statistical enrichment of target candidate genes in Kyoto gene and genome encyclopedia （KEGG） pathway； CytoFlex flow cytometry was used to detect the uptake of bovine milk exosomes； flow cytometry was used to detect the apoptotic rate of Caco-2 cells after treated with bovine milk exosomes.

The process development study results showed that the precipitation pretreatment approach with sodium phosphate was superior to all other methods evaluated in regard to feasibility， product quality and reproducibility. TFF method could further remove the residual milk protein and achieved the purpose of liquid concentration， which was effectively linked to the downstream BE-SEC purification method. The exosomes obtained by the three-step process showed the typical disc-shaped or cup-shaped exosomes under TEM.The purity of the sample detected by NanoFCM method was high than that detected by UC，and its partical size was 30－150 nm.The Western blotting results showed that the exosome samples contained CD81， Alix，TSG101 and other exosome specific markers. The SEC-HPLC analysis results showed that the final products of different batches had a single homogeneous peak without free protein pollution. The GO and KEGG database analysis results showed that the milk exosomes were rich in immune related proteins and miRNA related to inflammatory response regulation signaling pathway. The exosomes obtained by the above preparation processes had the ability to penetrate the cell membrane efficiently， and the proportion of exosomes entering the cell within 4 h was 99%.

A three-step purification process is established by chemical precipitation pretreatment， TFF and BE-SEC.The purity and yield of bovine milk exosomes obtained by this method are better than those obtained by gold standard—UC.The physicochemical properties of the samples are in line with the standard of typical bovine milk exosomes， which could penetrate the cell membrane within a few hours， and are rich in miRNA related to immune and inflammatory response regulation.