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Objective To investigate the effect of reducing OPTN gene expression on the sensitivity of cells to anticancer drugs by establishing the human SKOV3 cell line stably knocking down OPTN protein with RNA interference technique and human SKOV3 cell line stably knockout OPTN protein with CRISPR/Cas9 gene editing technique. Methods The sequences of shRNA and sgRNA were designed for OPTN gene and ligated with lentiviral vector plasmid pGPU6_GFP_Neo and knockout plasmid PX459 vector to produce recombinant vector. The recombinant plasmid vector and packaging plasmids(psPAX2 and pMD2.G) were co-transfected into 293 T cells for virus packaging. The supernatant of virus was collected to infect SKOV3 cells. SKOV3 cell line with stable knockdown and stable knockout of OPTN gene was obtained.RT-PCR was used to detect the changes of OPTN gene expression in knockdown group. PCR was used to detect the changes of genomic DNA in the knockout group. The expression of OPTN protein was detected by Western blot. The effect of DDP on the proliferation of ovarian cancer cells was detected by MTT. The expression of mRNA of caspase3, Bax and Bcl-2 was detected by qRT-PCR. The expression of caspase3, Bax and Bcl-2 proteins was detected by Western blot. Results Lentiviral vector and gene knockout vector were constructed successfully. In SKOV3 cells,the expression of OPTN protein in SKOV3 cells decreased(knockout cell line) or knocked out completely(knockout cell line).Ovarian cancer cells(SKOV3) with OPTN gene knockdown and knockout were less sensitive to DDP. Compared with the control group, the inhibition rate of DDP on cells was lower(P<0.05,P<0.01), and the effect of gene knockout cells was better. The expression of apoptosis-related genes Caspase3 and Bax mRNA and protein decreased, while the expression of Bcl-2 increased in SKOV3 cells with stable knockdown and knockout of OPTN protein by qRT-PCR and Western blot(P<0.05,P<0.01). Conclusion Reducing or removing the expression of OPTN gene in ovarian cancer can reduce the sensitivity of ovarian cancer cells to the anticancer drug DDP, which may be achieved by affecting apoptosis-related pathways.

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Objective To investigate the inhibitory effect of breviscapine(Bre) on pyroptosis and apoptosis of neuronal cells in chronic cerebral ischemia(CCI) rats by regulating the activation of NLRP3 inflammasome. Methods Male SD rats were randomly divided into sham group, 2 VO group, 2 VO+breviscapine(2.5, 5, 10 mg/kg) groups. The model of CCI was established by permanently ligating the bilateral common carotid arteries(2 VO) in rats. The learning and memory ability of rats were detected by Morris water maze test. The pathological changes and apoptosis of brain tissue were detected by HE and Tunel staining respectively. NLRP3, Caspase 1, interleukin(IL)-1β,IL-6 and cleaved Caspase-3 proteins expression levels were detected by Western blot. Results Compared with the sham group, the spatial learning and memory ability was disordered, the latency was significantly prolonged, the target quadrant dwell time and the number of crossing target quadrants significantly reduced in the 2 VO group rats(P<0.01). The apoptosis, degeneration and necrosis have been increased in cortical and hippocampal neurons in the 2 VO group rats. Also, the NLRP3 inflammasome in the hippocampus was activated, the expressions of Caspase 1, IL-6 and IL-1β protein were up-regulated, and Caspase-3 protein was activated, which further promoted neuronal apoptosis in the 2 VO group rats. Compared with the 2 VO group, Bre significantly reduced the latency of the platform found, increased the target quadrant residence time and the number of crossing target quadrants in rats. Bre reduced apoptosisand the pathological damage of cortical and hippocampal neurons, increased the number of neuronal cells, and improved neuronal cell morphology and alleviated nuclear pyknosis in CCI rats. In addition, Bre significantly inhibited the activation of NLRP3 inflammasome in hippocampus, down-regulated the expression of Caspase 1, IL-6 and IL-1β, inhibited neuronal apoptosis and the activation of Caspase-3 proteinand. Conclusion Bre can significantly improve the cognitive function of CCI rats, and reduce the pathological damage of ischemic neurons. The mechanism may be related to inhibiting the activation of NLRP3 and pyroptosis pathway.

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Objective To study the effect of long non-coding RNA family with sequence similarity 83 member H antisense RNA 1(lncRNA FAM83 H-AS1) on cell cycle, proliferation and metastasis of gastric cancer cells, and explore its possible mechanisms. Methods qRT-PCR was used to detect the expression level of FAM83 H-AS1 in gastric cancer cell lines MKN-45, SGC-7901, MGC-803 and human immortalized normal gastric epithelial cell line GES-1. The highest expression of gastric cancer cell line was selected for FAM83 H-AS1 siRNA infection, which was divided into si-NC group and si-FAM83 H-AS1 group. The expression of FAM83 H-AS1 in each group was detected by qRT-PCR. The effect of FAM83 H-AS1 on cell cycle was detected by flow cytometry. The effect of FAM83 H-AS1 on cell proliferation was detected by MTS and plate cloning assay. The effect of FAM83 H-AS1 on cell metastasis ability was detected by Transwell assay. Western blot was used to detect the effect of FAM83 H-AS1 on the expression of cyclinD1 and cyclin dependent kinase 4(CDK4) and epithelial-mesenchymal transition(EMT) marker proteins. Results The results of qRT-PCR showed that the expression level of FAM83 H-AS1 in gastric cancer cell lines was higher than that in human immortalized normal gastric epithelial cell line GES-1(P<0.05), and the gastric cancer cell line MKN-45 with the highest FAM83 H-AS1 expression level was selected to transfect FAM83 H-AS1 siRNA. qRT-PCR results showed that the expression of FAM83 H-AS1 decreased in si-FAM83 H-AS1 cells(P<0.05), the proliferation and metastasis ability and the expression of cyclinD1 and CDK4 proteins in si-FAM83 H-AS1 group MKN-45 cells decreased, the expression of EMT marker protein E-cadherin protein increased, and the expression of N-cadherin and Vinmentin protein decreased. Conclusion Interference with the expression of FAM83 H-AS1 inhibits the proliferation and metastasis of gastric cancer cells. FAM83 H-AS1 may be a novel target for the treatment of gastric cancer.

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Objectives To investigate the effect and mechanism of ginsenoside Rb1 on neural stem cell injury induced by lidocaine. Methods Primary neural cells were labeled with anti-Nestin antibody and anti-CD133 antibody. Neural stem cells were sorted out by using a flow cytometer, and their characteristics were examined by RT-qPCR and immunofluorescence staining. The isolated neural stem cells were cultured with a medium containing lidocaine or ginsenoside Rb1, and the effects of lidocaine or Rb1 on the neural stem cell JNK signaling pathway and expression of apoptosis-related molecules were detected by RT-qPCR and Western blot. Neural stem cells were co-cultured with lidocaine and Rb1, and the effect of Rb1 on lidocaine induced neural stem cell apoptosis were examined. Results Through two-step screening, spinal cord neural stem cells with higher purity were obtained. The stem cells expressed SOX2, a marker gene of neural stem cells, and could differentiate into related neural cells. Lidocaine treatment could activate the JNK signaling pathway and induce apoptosis of neural stem cells, while addition of ginsenoside Rb1 inhibited the activation of the JNK signaling pathway and partially restored the apoptosis of neural stem cells caused by lidocaine treatment. Conclusion Ginsenoside Rb1 inhibits lidocaine treatment-activated JNK signaling pathway and restores spinal cord neural stem cell apoptosis induced by lidocaine treatment.

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Objective To explore the impact of the status of human epidermal growth factor receptor 2 gene(HER2) gene on the metabolism of gastric cancer with computational gene sets enrichment analysis(GSEA). Methods The expression datasets of gastric cancer were downloaded from the cancer genome atlas(TCGA),gene expression omnibus(GEO) and ArrayExpress. Phenotype labels of Her2 high and low expression were made, which were applied in the GSEA. The meaningful metabolic genesets were found at false discovery rate(FDR) q value<25% and/or nominalPvalue<0.01. Heatmap was made to compare the meaningful gene sets in different datasets. Results Among the 10 datasets including TCGA STAD and GSE66229, peroxisome, N-glycan and pyrimidine metabolisms were significantly affected at FDR q val<25% and nomPval<0.01, glycosylphospatidylinositol GPI anchor biosynthesis, glycerolphosphalipid metabolism and sphingolipid metabolism atPvalue<0.01. Conclusion Multiple metabolic gene sets are affected by Her2 status with evidences in multiple datasets.

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Objective To develop an octreotide(Oct) functionalized polymeric nanosized contrast agent for targeted magnetic resonance(MR) imaging of tumor. Methods Reversible addition-fragmentation chain transfer(RAFT) polymerization had been employed to synthesize branched copolymer nanoparticles composing n-butyl methacrylate and ethylene glycol dimethacrylate copolymer segments as the core, and oligoethylene glycol methacrylate together with segment of 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid as the shell respectively, followed by the modification of Oct as corona. These polymeric nanoparticles were chelated with gadolinium ions to obtain a novel tumor-targeted branched copolymer nanosized MR contrast agent [P(DO3 A-Gd)-Oct]. The morphology of the products was analyzed by nuclear magnetic resonance(NMR), dynamic light scattering(DLS) and transmission electron microscope(TEM). The stability, hemolysis rate, cytotoxicity andin vivosafety of P(DO3 A-Gd)-OCT were preliminarily tested. Magnetic resonance relaxation rates were measured using a clinical 3.0 T magnetic resonance imager andin vivomagnetic resonance imaging was performed in H22 hepatoma bearing mice. Results According to the DLS and TEM results, the hydrodynamic diameter of P(DO3 A-Gd)-Oct was about 20 nm. After the exposure to P(DO3 A-Gd)-Oct, the hemolysis rate was only 0.34 %, and no obvious cyto-toxicity was observed. Compared with normal mice without any treatment, there was no obvious change of the pathological structures of major organs or the major blood biochemical indexes after the intravenous(i.v.) injection of P(DO3 A-Gd)-Oct. The relaxivity of P(DO3 A-Gd)-Oct(r1) was 8.33 mM-1s-1. After 60 min and 120 min of the i.v. injection, there were obvious increase of the signal in the tumor region in T1 weighted images of H22 tumor bearing mice, which was attributed to the increased concentration of gadolinium ions in tumor tissues compared with the non-targeted control. Conclusion P(DO3 A-Gd)-Oct exhibits bio-compatibility and excellent MR contrast ability, which can contribute to the enhancement of the sensitivity and accuracy in the diagnosis of malignant tumors such as liver tumor.

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Objective To explore the effects of sinomenine(SIN) on proliferation, apoptosis, invasion and migration ability of gastric cancer cell line SGC-7901 and its mechanism. Methods Normal human gastric mucosal epithelial cells GES-1 and human gastric cancer cells SGC-7901 were normally cultured, and they were enrolled as blank control group(Control). They were treated with 0.5, 1.0 and 2.5 mmol/L SIN for 48 h. The positive control(50 μmol/L celecoxib) was set up. Cell growth was detected by CCK-8 method. The apoptosis was detected by flow cytometry. Cell invasion was detected by Transwell chamber. Cell migration was detected by scratch test.Western blot was applied to detect the expression of Bax,Bcl-2,Cleaved caspase3,E-cadherin,N-cadherin and Vimentin. The targeted relationship between miR-33 a-5 p and lactate dehydrogenase A( LDHA) was analyzed by online prediction software,which was confirmed by double luciferase report. The miR-33 a-5 p mimics was transfected into SGC-7901 cells. The expression of miR-33 a-5 p and LDHA mRNA in cells was detected by RT-qPCR. The LDHA overexpression vector was recombined and constructed to transfected into SGC-7901 cells. The cells were treated with 0. 5,1. 0 and 2. 5 mmol/L SIN for 48 h. The cell proliferation,apoptosis,invasion and migration were detected. Results Compared with Control group,SIN could increase apoptosis rate of SGC-7901 cells,the expression of Bax,Cleaved caspase-3,E-cadherin and miR-33 a-5 p,while decreased survival rate,invasion number,migration rate,the expression of Bcl-2,N-cadherin,Vimentin and LDHA mRNA( P<0. 05). LDHA was the predicted target spot of miR-33 a-5 p. SIN could up-regulate the expression of miR-33 a-5 p,and down-regulate the expression of LDHA mRNA( P<0. 05). Compared with Control group,after LDHA overexpression,apoptosis rate of SGC-7901 decreased,while the expression of LDHA protein,survival rate,invasion number and migration rate increased( P<0. 05). After SIN treatment,apoptosis rate of SGC-7901 increased,while the expression of LDHA protein,survival rate,invasion number and migration rate decreased( P<0. 05). Compared with pcDNA-LDHA group,after SIN intervention,apoptosis rate of SGC-7901 increased,while survival rate,invasion number and migration rate decreased( P<0. 05). Conclusion SIN can down-regulate LDHA level by targeting miR-33 a-5 p,mediate cell proliferation,apoptosis,invasion and migration for inhibiting gastric cancer cell line SGC-7901.

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Objective To investigate the mechanism of propofol alleviating nerve function damage and oxidative stress in rats undergoing cerebral ischemia reperfusion(CI/R) by inhibiting the activation of extracellular signal regulated kinase 1/2(ERK12) and nuclear transcription factor kappa(BNF-κB p65). Methods A total of 75 SD rats were divided into sham operation group, model group, low-dose propofol group, high-dose propofol group and positive control group by random grouping method.The scores of nerve function deficits and postural reflex were compared among all groups. The cerebral infarction rate, cerebral water-containing rate and cerebral index in each group were calculated by 2,3,5-triphenyl tetrazolium chloride(TTC) method. The expression of brain-derived neurotrophic factor(BDNF) and nerve growth factor(NGF) in prefrontal lobe of the rats in each group was detected by RT-PCR and Western blot. The phosphorylation situations of ERK1/2 and NF-κB p65 were detected by Wetern blot. The pathological changes of neurons at hippocampus areas in each group were observed by HE staining. The levels of serum oxidative stress markers [malondialdehyde(MDA), superoxide dismutase(SOD), lactate dehydrogenase(LDH)] were detected by the kits. Results HE staining results showed that the cell distribution was scattered in model group, and there were many necrotic cells. After propofol treatment, the cell distribution in hippocampus areas was relatively even, and necrotic cells were reduced. Compared with sham operation group, scores of nerve function and postural reflex, cerebral infarction rate, cerebral water-containing rate, cerebral index, expression levels of BDNF, NGF, phosphorylated-ERK1/2(p-ERK1/2) and phosphorylated-p65 protein(p-p65), levels of MDA and LDH increased in model group(P<0.05), there was no change in the expression of total ERK1/2 or p65 protein(P>0.05), SOD level decreased(P<0.05). Compared with model group, SOD level increased in low-dose propofol group, high-dose propofol group and positive control group(P<0.05), there was no change in the expression of total ERK1/2 or p65 protein(P>0.05), scores of nerve function and postural reflex, cerebral infarction rate, cerebral water-containing rate, cerebral index, expression levels of BDNF, NGF, p-ERK1/2 and p-p65 protein, MDA and LDH decreased(P<0.05). Conclusion Propofol can protect against cerebral damage caused by ischemia-reperfusion through inhibiting phosphorylation of ERK12 and NF-κB p65 and reducing oxidative stress.

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Objective To explore the mechanism of modified Tuoli Xiaodu powder in the treatment of radiation proctitis based on TLR4/NF-κB signaling pathway. Methods Forty-eight SD rats were randomly divided into 6 groups, except for the control group, all the other groups were induced to make the rat model of radiation proctitis. After 3 d of normal feeding, the rats began enema, the control group and model group were enema with 2 ml 0.9% normal saline, and the positive control group were treated with 2 ml montmorillonite powder(0.2 g/ml) combined with dexamethasone(1 mg/ml) enema, modified Tuoli Xiaodu powder 2 ml enema with low(1 mg/ml), medium(2 mg/ml) and high dose(4 mg/ml) was performed once a day for 10 d. Then rats were killed, the diameter of rectum was measured, the histopathological changes of rectum were observed by HE staining, the levels of serum interferon-γ(IFN-γ), tumor necrosis factor-α(TNF-α), interleukin-4(IL-4), IL-10, epidermal growth factor(EGF) and IgA, IgM and IgG were measured by ELISA, the expressions of TLR4, Myd88, NF-κB p65, IκBα in rectum were detected by Western blot, and the expressions of TLR4, Myd88, NF-κB p65, IL-6 and IL-10 mRNA in rectum were detected by qRT-PCR. Results High dose of modified Tuoli Xiaodu powder could significantly improve the pathological changes of rectal tissue and reduce the score of rectal inflammation(P<0.05). Compared with the model group, the high dose of modified Tuoli Xiaodu powder could significantly increase the levels of serum EGF, IL-4, IL-10, IgA, IgM and IgG(P<0.05). The expression of IFN-γ, TNF-α, TLR4, Myd88, NF-κBp65, IκBα and TLR4, Myd88, NF-κBp65 and IL-6 mRNA significantly decreased in rats(P<0.01). Conclusion The high dose of modified Tuoli Xiaodu powder can significantly improve the pathological changes of rectum in rats with radiation proctitis and play a significant role in the expression of TLR4/NF-κB signaling pathway related factors and inflammatory cytokines.

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Objective To explore the pathogenesis of skeletal myopathy in patients with heart failure by observing the effects of dexmedetomidine on the proliferation and differentiation of mouse myoblast C2 C12. Methods Immunohistochemistry was used to analyze the expression characteristics of adrenergic receptors in C2 C12 cells.C2 C12 cells were divided into 6 groups: the control group and dexmedetomidine 5 concentration gradient groups(5, 10, 20, 40, 80 mmol/L). CCK-8 method was used to detect the effect of dexmedetomidine on C2 C12 cells proliferation. Differentiation medium was used to induce the differentiation of C2 C12 cells, and C2 C12 cell were dosed with single or continuous single administration. After 6 d of cell differentiation, the expression levels of myosin heavy chain(MYH), myocyte enhancer factor 2 C(MEF2 C) and myogenin(MyoG) in different groups were detected by immunofluorescence, and the number of myotubes containing different nuclei of C2 C12 cells was quantitatively analyzed.C2 C12 cells were divided into two groups: the control group and dexmedetomidine group(20 mmol/L). Then C2 C12 cells were dosed with continuous single administration and the expression levels of MYH, MEF2 C and MyoG of differentiated C2 C12 cells on the 2 nd, 4 th and 6 th day in different groups were detected by immunofluorescence. Results C2 C12 cells exhibited the characteristics of α2-adrenergic receptor expression. CCK-8 assay showed that dexmedetomidine inhibited the proliferation of C2 C12 cells in a concentration-dependent manner(P<0.05).The effect of single administration of dexmetomidine on the differentiation of C2 C12 cells was less significant than the effect of the continuous single administration of dexmetomidine, and with the increase of concentration in the continuous single administration of dexmetomidine groups, the expression levels of MYH, MEF2 C, MyoG gradually decreased, the number of C2 C12 cells differentiated into myotubes with multiple cell nuclei gradually decreased(P<0.05). On the 2 nd, 4 th and 6 th day after differentiation of C2 C12 cells, the expression levels of MYH, MEF2 C, MyoG in the 20 mmol/L group were all lower than those in the control group. Conclusion α2 adrenergic receptors exists in mouse myoblast C2 C12. Dexmetomide may inhibit the proliferation and differentiation of C2 C12 cells by binding to α2 adrenergic receptors, which is closely related to the decrease of the MEF2 C and MyoG expression levels.

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Objective To study the difference between the high temperature requirement serine proteinase A(HtrA) gene-deficient Streptococcus strains and high-virulent strains of Streptococcus mutans(S.mutans) in the formation of biofilm in vitro on non-stressful environment.Methods Taking the high virulence strain of HtrA screened from high caries-sensitive deciduous teeth obtained in the early stage and the high cariogenic HtrA gene defectivestrain and the international standard strain UA159 as the research objects, and the growth curve of them was evaluated by spectrophotometer turbidimetry. The biofilm model of the three strains was constructed in vitro,the morphology and structure of the biofilm were observed under scanning electron microscope( SEM),and the amount of biofilm formation was evaluated by crystal purple staining. Results Compared with HtrA gene-deficient strains,the bacterial concentration( OD600 value) of HtrA high virulent strains and UA159 standard strains was higher than that of HtrA gene deficient strain in each stage of S. mutans proliferation in the three groups,and the differences were statistically significant( P<0. 05). Under scanning electron microscope,the density of HtrA high virulence strain was the highest,followed by UA159 standard strain,and the density of HtrA deficient strain was the lowest.Semi-quantitative determination of biofilm crystal purple for 4,8,12,24 h,there was significant difference in OD600 value between HtrA high virulence strain and HtrA gene deficient strain( P<0. 001). Conclusion The formation of S. mutans biofilm is affected by HtrA,and the deletion of HtrA gene reduces bacterial adhesion and slows biofilm formation.

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Objective To investigate the interleukin-1β(IL-1β) induced apoptosis of thyroid cells in Hashimoto thyroiditis(HT) and its potential mechanism. Methods ① Factor associated suicide(Fas) mRNA expression in FRTL-5 cells were detected by real-time PCR under different concentration and reacting time of IL-1β.②The control group, IL-1β group, IL-1β+caspase8 inhibitor(Z-IETD-FMK) group were set up, and the apoptosis rate of each group was analysed by flow cytometry. Results ① The result of real-time PCR showed that there was a dose-time dependent relationship between IL-1β and Fas mRNA expression levels of FRTL-5 cells(P<0.05); ②Flow cytometry results showed that compared with the control group, the apoptosis rate of thyroid cells increased after IL-1β induction(P<0.05) and the increase of IL-1β-induced thyroid cell apoptosis was inhibited after combined with Z-IETD-FMK(P<0.05). Conclusion IL-1β may induce apoptosis of thyroid cells through Fas and caspase 8 pathways.

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Objective To explore the effect of GANT 61 on collagen induced arthritis(CIA) mice. Methods BALB/C mice were randomly divided into three groups [Control group, collagen induced arthritis(CIA) group, GANT61 group].CIA group and GANT61 group were induced arthritis with bovine type Ⅱ collagen. GANT61 group was injected with GANT61 on 0, 3, 6, 9, 12, 15, 18, 21 d after second immunization. Meanwhile the condition of joints was observed and the clinical score was made. On 39 th day after primary immunization, mice were executed to collect peripheral blood and serum for detecting the change of neutrophils and cytokines. Joints and synovial tissues were collected for pathological examination and synovioblast culture. Cell proliferation was detected by flow cytometry and CCK-8 assay. Apoptosis was detected by flow cytometry. Results Data showed that GANT61 could reduce joint swellingand clinical score of arthritis,the infiltration of inflammatory cells can also be reduced.Neutrophils and inflammation cytokines of interferon-gamma(IFN-γ), interleukin-6(IL-6), tumor necrosis factor-α(TNF-α). GANT61 could remarkably inhibit cell proliferation and induce cell apoptosis. Conclusion GANT61 can alleviate the pathological progress of arthritis in Balb/c mice by inhibiting the proliferation of synovial fibroblasts.

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Objective To evaluate the effects of mouse bone marrow mast cell derived exosomes on the proliferation, migration and invasion of hepatocellular carcinoma Hepa1-6 cells. Methods Mouse bone marrow mast cells were cultured by using IL-3 and SCF cytokinesin vitro. The cells were identified by toluidine blue staining, immunofluorescence and flow cytometry. Exosomes were isolated by ultracentrifugation from supernatants, and the morphology, diameter and marker protein of exosomes were detected by transmission electron microscopy, particle size analysis and Western blot. Hepa1-6 cells were treated with different concentrations of exosomes(5, 10, 20 μg/ml). CCK-8 test, Edu test, scratch test and transwell test were used to detect the proliferation, migration and invasion of hepatocellular carcinoma cells. Results The exosomes of bone marrow mast cells were round or oval cup shape, with a particle size of 30～150 nm, and expressed exosomal marker proteins CD63, Alix, and TSG101.The results of CCK-8 and Edu showed that exosomes derived from mast cells promoted the proliferation of Hepa1-6 cells, and the effect is concentration-dependent. Cell scarification results showed that exosomes derived from mast cells promoted the migration of Hepa1-6 cells. Transwell results showed that exosomes derived from mast cells enhanced the invasion ability of Hepa1-6 cells. Conclusion Bone marrow mast cell derived exosomes can promote the proliferation, migration and invasion of hepatocellular carcinoma Hepa1-6 cells.

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Objective To investigate the depressive-like behavior and expression of hippocampal brain-derived neurotrophic factor(BDNF) in different sexual cycles of female mice and the effect of estrogen deprivation and supplementation on depressive-like behaviors and BDNF expression, and to elucidate the role of estrogen in the pathogenesis and therapy of depression. Methods Female C57 mice were screened for proestrus and diestrus by using vaginal smear staining. The forced swim was used to evaluate the depressive-like behavior of these two groups of female mice. Real-time PCR and Western blot were used to detect the expression levels of total BDNF mRNA and protein in the hippocampus of these two groups. Sucrose preference test was used to examine the depressive-like behaviors of female mice after OVX mediated estrogen deprivation under basal and chronicunpredictable stress condition. The effect of estrogen deprivation and supplementation on depression-like behaviors and hippocampal total BDNF mRNA expression of female mice were detected by sucrose preference test, forced swim test and Real-time PCR under stress condition respectively. Results The forced swim test results showed that the immobility time of diestrus was significantly more than that of proestrus(P<0.05). Real-time PCR and Western blot results showed that the total mRNA and protein expression levels of hippocampal BDNF in the diestrus were significantly lower than those in the proestrus(allP<0.05). The sucrose preference results showed that under the basic condition, the sucrose preference of OVX mice was not significantly changed compared with the sham operation control group(P>0.05), under stress condition, the sucrose preference of OVX mice was significantly reduced(P<0.05). Sucrose preference test and forced swim test results showed that estrogen supplementation could significantly reverse the decreased sucrose preference and increased immobility time induced by estrogen deprivation under the stress condition. Real-time PCR results showed that total BDNF mRNA level in the hippocampus of female OVX mice was significantly reduced(P<0.05) compared to the sham operation group, However, the total mRNA level of hippocampal BDNF was significantly restored after exogenous estrogen supplementation(P<0.05). Conclusion Sexual cycles of female mice can affect depression-like behavior and hippocampal total BDNF mRNA expression significantly, estrogen deprivation can enhance the susceptibility to the CUS induced depression-like behaviors, and estrogen supplementation can reverse the depression-like behavior and reduce hippocampal BDNF expression.

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Objective To evaluate the expression pattern of SP142-PD-L1 in poorly differentiated gastric adenocarcinoma(PDGA) and investigate the clinical implication of programmed death-ligand 1(PD-L1) expression in PDGA classification.Methods The study comprised tumor tissues from 163 PDGA patients treated with surgically resection, which were analyzed for PD-L1 expression in tumor cells(TCs) as well as tumor associated immune cells(ICs) by immunohistochemistry. Tumor infiltrating lymphocytes(TILs) were determined by morphological and immunohistochemical evaluation. PDGA was classified into different subsets based on either PD-L1 expression or PD-L1 expression in combination with the feature of TILs.Results PD-L1 expression was observed in TCs and ICs,with positive rate of 36. 81% and 99. 39% respectively. High expression of PD-L1 in TCs was significantly correlated with histological subtype of tumor,status of lymphovascular and perineural involvement. High expression of PD-L1 in ICs was closely associated with several clinicopathological characteristics,including tumor size,histological subtype,pTNM stage,status of lymphovascular and perineural involvement,tumor budding as well as CD3+ TILs. There were significant associations of different PDGA subsets defined by either PD-L1 expression or PD-L1 expression in combination with CD3 + TILs with different clinicopathological features respectively. Conclusion The clinicopathological significance of PD-L1 expression in TC is different from that in IC. Combined evaluation ofPD-L1 expression level and comprehensive analysis of PD-L1 expression and TIL characteristics can be used for PDGA classification with prognosticsignificance,which has potential implication in the clinical diagnosis and treatment of PDGA.

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Objective To investigate the efficacy of endoscopic retrograde cholangiopancreatography(ERCP) in the treatment of benign bile duct lesions with cholangitis, and to analyze the influencing factors of postoperative recurrent cholangitis. Methods Eighty-six patients with benign bile duct benign lesions and cholangitis were retrospectively selected. All patients received ERCP. The patients′ laboratory examination index and improvement of main symptoms and complication rate were counted 2 weeks after operation. The recurrence rate of cholangitis was counted 1 year after follow-up, and the influencing factors of postoperative recurrent cholangitis were analyzed. Results In 2 weeks after surgery, the improvement of total bilirubin(TB), alanine aminotransferase(ALT), leucopenia and abdominal pain relief accounted for 75.58%, 81.40%, 87.21% and 88.37% respectively. There were 10 cases of complications(11.6%), including 3 cases of pancreatitis(3.5%), 3 cases of biliary tract infection(3.5%), 3 cases of hyperamylasemia(3.5%), and 1 case of hypoxemia(1.2%); no death and perforation, gastrointestinal bleeding occurred. There were 18 patients(20.93%) with recurrent cholangitis after operation. Multivariate logistic analysis showed that age>65 years, diverticulum, operation time>45 min and with gallbladder stone were recurrent bile ducts in patients with benign bile duct lesions and cholangitis. Risk factors for inflammation(P<0.05). Conclusion ERCP is effective in the treatment of benign lesions of the bile duct with cholangitis. The complications are less. The patients with age>65 years old, diverticulum and operation time>45 min, and with gallbladder stone are more likely to relapse cholangitis after surgery.

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Objective To explore the expression and clinical significance of interleukin-19(IL-19) in different types of chronic rhinosinusitis with nasal polyps(CRSwNP). Methods 42 CRSwNP patients were selected to collect nasal polyps during the operation: 20 patients with eosinophilic chronic rhinosinusitis with nasal polyps(ECRS), 22 patients with non-eosinophilic chronic rhinosinusitis with nasal polyps(non-ECRS). 20 patients in the control group were selected to collect mucosa of uncinate process during the operation. The percentage of eosinophils in tissues was measured by hematoxylin-eosin staining, and the expression of IL-19 was measured by qRT-PCR and immunohistochemistry staining. Results IL-19 was mainly expressed in epithelial and submucosal glands of polyps in ECRS. The mRNA and protein expression of IL-19 was significantly higher in ECRS than in non-ECRS,and higher than in control(P<0.05,P<0.05). In ECRS group, the mRNA level of IL-19 was positively correlated with CT Score, endoscopic score and percentage of eosinophils(P<0.05,P<0.05,P<0.05), but not with VAS score(P>0.05). Conclusion Compared with non-ECRS and normal nasal mucosa, the expression of IL-19 in ECRS is higher, and is positively correlated with clinical severity. IL-19 may aggravate clinical symptoms by promoting the aggregation of eosinophils.

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Objective To explore the distribution characteristics of single nucleotide polymorphisms(SNPs) of rs619586 and rs7927113 at metastasis-associated lung adenocarcinoma transcript 1(MALAT1) in Guangxi healthy population, and to further compare the distribution differences of polymorphisms among different populations. Methods The polymorphisms of MALAT1 were genotyped by the SNPscan technology in 195 samples from Guangxi, and statistical methods were used to compare the differences in genotype and allele frequency between Guangxi population and other populations from haplotypemap(HapMap-CHB, HapMap-JPT, HapMap-CEU and HapMap-YRI). Results The genotypes of rs619586 included AA(78.5%) and AG(21.5%) in Guangxi population, and the frequencies of A and G allele were 89.2% and 10.8% respectively. Rs7927113 had GG(97.4%) and AG(2.6%) genotypes, and the frequencies of allele were 98.7% and 1.3% for G and A respectively. There was no statistically significant difference in polymorphism of two loci between male and female samples(P>0.05). The genetic polymorphisms of rs619586 in Guangxi population were significantly different from HapMap-JPT, HapMap-CEU and HapMap-YRI(P<0.05). The differences in polymorphism of rs7927113 between Guangxi population and HapMap-YRI were significant(P<0.05). Conclusion The few mutations in MALAT1 rs619586 and rs7927113 in the Guangxi population differ from those of other regions in varying degrees, which lay the groundwork for the genetic analysis of local diseases related to MALAT1 gene.

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Objective To investigate the prenatal ultrasound diagnosis of abnormal invasive placenta by measuring the peak flow velocity of systolic blood flow under placenta. Methods A total of 133 pregnant women(≥28 weeks of gestation) who received placenta previa were enrolled, including complete placenta previa and anterior placenta with ≤2 cm distance between the placental edge and internal os. The US company VOLUSON E8 color Doppler ultrasound was used to examine the absence of the vocal cord area behind the placenta, the thickness of the posterior placental muscle layer<1 mm, the flow of the placental cavity ≥10 cm/s, and the peak systolic velocity(PSV). Ultrasound characteristics such as PSV were used to predict the accuracy of abnormal invasive placenta using receiver operating characteristic(ROC) curves. Results Compared with non-invasive placenta patients, the average gestational age of patients with invasive placenta decreased, the number of previous cesarean section increased, and PSV increased(P<0.05). There was no significant difference in the resistance index(RI) between patients with invasive placenta and non-invasive placenta(P>0.05). The area of PSV under the ROC curve predicting abnormal invasive placenta was 0.910(P<0.001, 95%CI: 0.873～0.962) while the area of RI under the ROC curve predicting abnormal invasive placenta was 0.562(P=0.336, 95%CI: 0.461～0.663). When the blood flow PSV under the placenta of the anterior wall of the uterus was ≥41 cm/s, the sensitivity of the abnormally invasive placenta was 89.47%, the specificity was 84.21%, the positive predictive value was 80.95%, and the negative predictive value was 91.43%. Conclusion The measurement of peak flow velocity during systolic blood flow in placenta may be a new marker for ultrasound diagnosis of abnormal invasive placenta, with high sensitivity and specificity.

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Objective To investigate the spontaneous activity of neurons in the resting state of patients exposed to car accident trauma. Methods 17 patients with posttraumatic stress disorder(PTSD group) and 23 patients without PTSD(non-PTSD group) after car accident trauma exposure were included in the study. The differences in fractional low-frequency amplitude(fALFF) between the PTSD and non-PTSD groups at 1 week and 2 months after the traumatic event were compared horizontally,and the PTSD and non-PTSD groups were compared longitudinally before and after fALFF at 1 week and 2 months after the traumatic event. Results Compared with the non-PTSD group,fALFF in the PTSD group decreased in the left posterior central gyrus after traumatic event at 1 week, fALFF in the PTSD group decreased in the left medial frontal gyrus at 2 months(P<0.01,GRF correction), and no abnormality was found in the residual brain areas. In the PTSD group, compared with the first week, fALFF significantly increased in the right medial temporal gyrus and left occipital lobe at 2 months after the traumatic event. In the non PTSD group, compared with the first week,fALFF decreased in the left cerebellum and left posterior cingulate gyrus and increased in the right precuneus at 2 months after the traumatic event(P<0.01, GRF correction), and no abnormality was found in the residual brain areas. Conclusion fALFF changes in the brain of patients with PTSD may be related to the pathophysiological mechanism of PTSD, which is helpful for early diagnosis and intervention of PTSD.

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Objective To investigate the effects of sevoflurane preconditioning on liver ischemia-reperfusion injury(LIRI) and plasma neutrophil nuclear transcription factor B(NF-κB) activity in Tibetan patients undergoing hepatic hydatid surgery. Methods 80 patients undergoing hepatic hydatid surgery were enrolled and were randomly divided into Tibetan generation living hydatid patient control group(group ZC), Tibetan generation living hydatid patient sevoflurane preconditioning group(group ZS), Han immigrant living hydatid patient control group(group HC) and Han immigrant living hydatid patient sevoflurane preconditioning group(group HS) by random number table method, 20 cases in each group. Group ZC and HC were not given any inhaled anesthetics. Group ZS and HS were given sevoflurane inhalation anesthesia for anesthesia-induced intubation. Before anesthesia induction(T0), immediate after porta hepatis occlusion(T1), immediate after porta hepatis opening(T2), at 1 h after ischemia/reperfusion(T3), at 6 h after ischemia/reperfusion(T4) and at 24 h after ischemia/reperfusion(T5), the levels of serum Alanine transaminase(ALT), aspartate transaminase(AST), lactic dehydrogenase(LDH),lactic acid, tumor necrosis factor α(TNF-α), interleukin6(IL-6) and neutrophil NF-κB activity at any time point were detected. Malondialdehyde(MDA)and superoxide dismutase(SOD) levels in normal liver tissues were detected at T3. Pathological morphology of liver tissues was observed by HE staining under light microscope. Results Compared with group ZC, the levels of ALT, AST, LDH, lactic acid, TNF-α and IL-6, and NF-κB activity decreased in group ZS at T1～T5(P<0.05), while SOD activity of liver tissue increased at T3(P<0.05), and MDA level of liver tissue decreased(P<0.05). Compared with group HC, the levels of ALT, AST, LDH, lactic acid, TNF-α and IL-6, and NF-κB activity decreased in group HS at T1～T5(P<0.05), while SOD activity of liver tissue increased at T3(P<0.05), and MDA level of liver tissue decreased(P<0.05). HE staining showed that pathological changes of liver tissue in group ZS and HS were alleviated. Conclusion Sevoflurane may alleviate LIRI in Tibetan patients undergoing hepatic hydatid surgery by inhibiting neutrophils NF-κB activity and the release of inflammatory factors.

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Objective To investigate the expression of vaccinia-related kinase 1(VRK1) in diffuse large B cell lymphoma(DLBCL) and its relationship with clinicopathology and prognosis, and its effect on the apoptosis of DLBCL cells. Methods RT PCR was used to detect the mRNA expression of VRK1 in 98 cases of DLBCL tissues and 62 cases of reactive hyperplasia lymphoid(RHL) lymph nodes. Western blot was performed on normal B cell immortalized cell lines HMy2.CIR and DLBCL cell lines including SU-DHL-4, OCI-Lyl9, OCI-Ly3, VAL, SU-DHL-2 and SU-DHL-6 to detect VRK1 protein expression, and VRK1 expression was analyzed based on DLBCL gene chip expression data in visualization software GEPIA and TCGA database. The relationship between VRK1 expression and clinicopathological features of patients with DLBCL was analyzed. The prognosis of patients with high VRK1 expression and low VRK1 expression was compared. RT PCR was used to detect the expression of DNA repair pathway-related proteins XRCC3 and CDC20 mRNA expression in DLBCL tissues, and to compare the co-expression relationship among VRK1, XRCC3 and CDC20. The silencing of VRK1-stable DLBCL cell line was constructed, and the proliferation and apoptosis ability of cells were detected by soft agar colony formation assay and Annexin V-FITC/PI double labeling assay. Results VRK1 was highly expressed in both DLBCL tissues and cells. VRK1 expression was associated with Ann Arbor staging and immunophenotyping in patients with DLBCL. The 5-year overall survival rate of patients with high VRK1 expression was lower than that of VRK1 low expression group. There was positive co-expression among VRK1, XRCC3 and CDC20. Silencing expression of VRK1 inhibited the proliferation of OCI-Ly3 cells and promoted the apoptosis of OCI-Ly3 cells. Conclusion Silencing VRK1 shows strong anti-tumor potential and has certain clinical significance. It provides new ideas and theoretical basis for future clinical diagnosis, prognosis and molecular targeted therapy of DLBCL based on VRK1.

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Objective To observe the change of quantitative parameters of dynamic enhanced magnetic resonance(DCE-MRI) during neoadjuvant chemotherapy(NAC) in breast cancer, and to investigate the feasibility of predicting the pathological efficacy of breast cancer with NAC.Methods Eighty patients with breast cancer received NAC were retrospectively analyzed. All patients underwent surgery after chemotherapy were divided into pathological complete response(pCR) group and non-pCR group. Statistical analysiswas used to compare the changes and differenceof DCE-MRI quantitative parameters before and after NACbetween pCR and non-pCR group. Pearson correlation analysis was used to analyze the correlation between the changes of DCE-MRI parametersand Δd before and after NAC treatment. The receiver operating characteristic curve(ROC) was used to analyze the diagnostic efficacy of DCE-MRI quantitative parameters. Results The 34 cases of 80 cases with invasive carcinoma were pCR and 46 cases of that were non-pCR. There was no significant difference in d value between p CR group and non-pCR group before NAC( P = 0. 320). The perfusion transfer coefficient( Ktrans) and reflux rate( Kep) values of p CR group before NAC were statistically lower than those of non-pCR group( P<0. 05). The values of d,Ktrans,Kepand Vpafter NAC in p CR group were statistically lower than those before NAC( P<0. 05),and the values of d and Ktransafter NAC in non-pCR group were lower than those before NAC( P<0. 05). The ΔKtransand ΔKepvalues between pCR group and non-pCR group were statistically different( ΔKtrans: P = 0. 022,ΔKep: P = 0. 016). Correlation analysis showed that there was a positive correlation between ΔKtransand Δd after NAC( r = 0. 730,P<0. 001). The areas under the ROC of Ktrans,ΔKtrans和 Kepwere 0. 822,0. 747 and 0. 705 respectively. The sensitivity and specificity of the diagnosis were 84. 000% and 81. 200%,84. 000% and 74. 910%,76. 000% and 79. 800% respectively.Conclusion Ktrans,ΔKtransand Kephave higher diagnostic efficacy in predicting the pathological response of NAC in locally advanced breast cancer.

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Objective To explore the effect of systemic lupus erythematosus(SLE) exacerbations on maternal and fetal outcomes. Methods The datas of 152 pregnant women who diagnosed with SLE were reviewed, 2 cases with labor induction were excluded because of fetal malformation. According to SLE disease activity index, the 150 patients were divided into two groups:activity group(≥5 points) with 41 cases and inactivity group(<5 points) with 109 cases. The pregnancy complications(preeclampsia, oligohydramnios and intrapartum hemorrhage) and pregnancy outcomes [days of pregnancy, birth weight, birth length, the size of the placenta, preterm delivery rate, cesarean section rate, fetal growth restriction rate(FGR), neonatal asphyxia] between the two groups were compared. Two independent samplest-test was used for the comparison of measurement datas, and chi-square test for qualitative variables. Results The incidence of preeclampsia in the activity group was significantly higher than that in the inactivity group(P<0.001). And the birth weight in the activity group was significantly lower than that in the inactivity group(P<0.01). The birth length in the activity group was smaller compared with that in the inactivity group(P<0.05). Moreover the incidence of neonatal asphyxia in the activity group was higher than that in the inactivity group(P<0.05). However, there were no statistically significant difference between oligohydramnios, bleeding volume during delivery, days of pregnancy, placental size, preterm birth, cesarean delivery, and FGR in the two groups. Conclusion SLE activity is an influential factor in the occurrence of preeclampsia. It can effect birth weight and birth length, and it is more likely to cause neonatal asphyxia. Therefore pregnant women with active SLE need to be jointly treated by rheumatologist and obstetricians. Close follow-up, timely adjustment of drugs and disease control within a stable range can greatly improve their pregnancy outcomes.

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Rat spleen cell suspension was prepared by common method. The expression levels of helper T cells(CD3+CD4+), cytotoxic T cells(CD3+CD8+), and B cells(CD45 R+) subsets tryptophan 2, 3-dioxygenase(TDO2) in rat spleen were detected by Cytomics FC500(FC500) and ImageStreamXMark Ⅱ(Mark Ⅱ) flow cytometry. The correlation and consistency of the results were observed respectively. The expression of TDO2in CD3+CD4+T cells, CD3+CD8+T cells and CD45 R+B cells were measured by two kinds of flow cytometry,and the correlation were well(R2>0.8). Bland-Altman method was used to analyze TDO2expression in CD3+CD4+T cells, CD3+CD8+T cells and CD45 R+B cells. The bias were-1.3%,-8.2% and 7.4%, respectively.TDO2were expressed in T and B cells of rat spleen. These two flow cytometries had good correlation and consistency in testing resualts. FC500 is easy and fast to operate. Mark-Ⅱ combines the high-throughput analysis of traditional flow cytometry with cell mor-phology analysis, which is highly accurate and can obtain specific cell pictures.

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To compare the clinical outcomes and safety of minimally invasive incisionviavideo-assisted thoracoscopic surgery(VATS) and traditional median full sternotomy incision for aortic valve replacement(AVR). There were 63 patients underwent AVRviaminimally invasive incision as Mini-AVR group including 52 cases through partial upper mini-sternotomy and 11 cases through right anterior minithoracotomy. 135 patients were undergone traditional median full sternotomy AVR as control group(Full-AVR group). The preoperative general data, intraoperative and postoperative indicators and prognosis were compared between the two groups. The clinical effects were also compared between the two minimally invasive incisions. The proportion of New York heart association(NYHA) grade IV in the Mini-AVR group was less than that in the Full-AVR group and the difference was statistically significant(P<0.01). The cardiopulmonary by pass and aortic cross-clamp time in Mini-AVR group were significantly longer than that in Full-AVR group(P<0.01). However, the duration of ICU stay time and post-operative hospital stay, incision length, volume of drainage on the first day postoperative, chest tube duration, and volume of red blood cell transfusion in Mini-AVR group were significantly lower than those in Full-AVR group(P<0.01). The cardiopulmonary bypass(CPB) time and aortic cross-clamp time of right anterior minithoracotomy were the longest in three methods for AVR, with statistically significant difference(P<0.01). The Mini-AVRviaVATS is safe and effective, among which partial upper mini-sternotomy is preferred and right anterior minithoracotomy is the second choice.

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To investigate the color Doppler ultrasound detection application in hypertensive disorders of pregnancy( HDP) pregnant outcome by analyzing the correlations of some parameters. 80 cases of late pregnancy HDP pregnant women were selected and analyzed after grouping. The changes of left uterine artery( UtA),fetal umbilical artery( UA) resistance index( RI),pulsatile index( PI) and ratio of systolic peak velocity to diastolic peak velocity( S/D) in the very low birth weight and asphyxia groups were significant( P<0. 05). Comprehensive monitoring of uterine artery,fetal umbilical artery and middle cerebral artery( MCA) has practical clinical value in evaluating HDP pregnancy outcome.

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Direct fluorescence labeling and multiparameter flow cytometry were used to detect different T lymphocyte subsets such as total T lymphocytes, helper T lymphocytes, cytotoxic T lymphocytes, regulatory T cells(Treg cells) and activated effector T cells(Teff cells) in patients with secondary myeloid leukemia retrospectively. By comparing secondary acute myeloid leukemia(S-AML), primary acute myeloid leukemia(P-AML) and healthy control group, the differences in Treg and Treg/Teff among the three groups were found to be statistically significant(P<0.05). There was an imbalance in the immune status of patients with acute myeloid leukemia. Moreover, the immune imbalance was even more pronounced in s-aml patients, which might provide new ideas for the prevention and treatment of S-AML in the future.

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To explore the risk factors of recurrence of idiopathic generalized epilepsy after drug withdrawal. 96 patients were divided into relapse group and non relapse group within one year after drug withdrawal,The relevant factors were statistically analyzed. Female,abnormal electroencephalogram( EEG) for the first time,abnormal EEG after discontinuation,seizure frequency>5 times/year,withdrawal time≤6 months,combined medication were all related to risk factor for relapse after drug withdrawal( P<0. 05). Female,abnormal EEG for the first time,abnormal EEG after discontinuation,seizure frequency>5 times/year,withdrawal time≤6 months,combined medication( P<0. 05) were independent risk factors for relapse. The risk of relapse after withdrawal of antiepileptic drugs is affected by many factors,so it needs to be evaluated before withdrawal of antiepileptic drugs to reduce relapse.

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To explore the clinical characteristics of smear negative pulmonary tuberculosis and the diagnostic value of multi-color nested real-time fluorescence quantitative nucleic acid amplification(Gene Xpert-MTB/RIF). The clinical data of 400 patients were analyzed retrospectively, and the value of Gene Xpert-MTB/RIF in the diagnosis of smear negative pulmonary tuberculosis was discussed. The results showed that the incidence of fever, cough/expectoration and neutrophil increase in smear negative pulmonary tuberculosis was significantly lower, the proportion of diabetes mellitus and chronic obstructive pulmonary disease was small, and the incidence of pleurisy and pulmonary fibrosis in imaging was higher than that in smear positive pulmonary tuberculosis. Gene Xpert-MTB/RIF technology has a better diagnostic value than traditional inspection methods, which is worthy of clinical promotion.