OBJECTIVE:To study tanshinone ⅡA inhibiting the proliferation and migration induced by homocysteine(Hcy)of rat aortic vascular smooth muscle cells(VSMCs)and its signal pathway. METHODS:VSMCs were selected for the following ex-periments. In order to validate TanshinoneⅡA inhibiting the proliferation and migration induced by Hcy of VSMCs,VSMCs were di-vided into control group,Hcy group(1 000μmol/L),TanshinoneⅡA low-dose,medium-dose and high-dose groups(5,10,20 μg/L). In mechanism study test,VSMCs were divided into control group,TanshinoneⅡA group(20 μg/L),rapamycin group(20 nmol/L), MHY1485 group(10μmol/L),TanshinoneⅡA+rapamycin group(TanshinoneⅡA,20 μg/L+rapamycin,20 nmol/L),TanshinoneⅡA+MHY1485 group (TanshinoneⅡA,20 μg/L+MHY1485,10 μmol/L). In validation test of P70S6K and p-P70S6K pathway expres-sion,rapamycin and MHY1485 were used for inhibitory test and activation test,respectively. The proliferation of VSMCs was de-termined by ELIASA,and Transwell chambers and wound healing test were employed to test the migratory ability of VSMCs. West-ern blotting were used to investigate the expressions of P21,P27,MMP-2,MMP-9,P70S6K and p-P70S6K in VSMCs. RE-SULTS:In validation test,compared with control group,24,48 h absorbance,migration area,the number of VSMCs penetration and the expression of MMP-2,MMP-9 and p-P70S6K increased significantly,while the expression of P21 and P27 decreased sig-nificantly(P<0.01). Compared with Hcy group,48 h absorbance of TanshinoneⅡA low-dose group,24,48 h absorbance and the expression of MMP-2 of TanshinoneⅡA medium-dose and high-dose groups,migration area,the number of VSMCs penetration, the expression of MMP-9 and p-P70S6K in low-dose,medium-dose and high-dose groups all decreased significantly;the expres-sion of P21 and P27 increased significantly in TanshinoneⅡA medium-dose and high-dose groups(P<0.01);there was no statisti-rapamycin group,while 24,48 h absorbance increased significantly in MHY1485 group(P<0.01). Compared with TanshinoneⅡA group,24,48 h absorbance decreased significantly in TanshinoneⅡA+rapamycin group,while 12,24,48 h absorbance,migration area and the number of VSMCs penetration increased significantly in TanshinoneⅡA+MHY1485 group (P<0.01). In inhibitory test,compared with control group,the expression of p-P70S6K decreased significantly in TanshinoneⅡA group (P<0.01);com-pared with TanshinoneⅡA group,the expression of p-P70S6K decreased significantly in rapamycin group and TanshinoneⅡA+rapa-mycin group(P<0.01);in activation test,the expression of p-P70S6K decreased significantly in TanshinoneⅡA group(P<0.01), compared with TanshinoneⅡA group,the expression of p-P70S6K increased significantly in MHY1485,TanshinoneⅡA+MHY1485 group (P<0.01). CONCLUSIONS:TanshinoneⅡA can inhibit the proliferation and migration of VSMCs by suppressing mTOR/P70S6K signal pathway.

Objective To investigate the expression and clinical significance of matrix metalloproteinase(MMP-9)and tension protein homologous gene in gingival cancer tissue.Methods 50 cases in Guyuan people's hospital from June 2013to December 2016 were selected to study the expression of matrix metalloproteinase(MMP-9)and tension protein homologous gene in gingival cancer tissue.Results The positive rate of PTEN in gingival cancer group was 54.00%, the positive rate of PTEN was 94.00% in normal tissue, normal control group positive rate of PTEN was significantly higher than that of gingival cancer group(P<0.05);the positive rate of MMP-9 in gingival cancer group was 76.00%, the positive rate of control group was 20.00%, gingival cancer group was significantly higher than the normal control group(P<0.05).The expression of MMP-9 and PTEN in in gingival cancer tissue was unrealted with gender, age, and tissue differentiation, and associated with lymph node metastasis and tissue infiltration(P<0.05).Conclusion PTEN expression is low in gingival cancer tissue low, MMP-9 expression is high in gingival cancer tissues.PTEN and MMP-9 are closely related to the incidence and development of gingival cancer.Therefore, MMP-9 positivity is a risk factor for gingival cancer, while PTEN positivity is a protective factor for gingival cancer.

Objective To study the ultrastructural changes of gunshot wounds at the brain stem and the cause of the death. Method A case died of the head injury and two cases died of gunshot wounds of the brain stem were studied. Brain stem tissues were taken 25 min after death. Tissue blocks from the tegmentum of the midbrain and the pons as well as the gray matter of the medulla oblongata in the deep portion of the base of the fourth ventricle were taken and were examined with ultramicroscopy and scanning electron microscopy. Results (1) The gunshot wound injuries inside the brain including burn and injuries caused by punch and pressure; (2) Although the primary gunshot wounds were located at the midbrain, the pathological changes extended to the whole brain stem; (3)The injuries of the axons and the nerve cells were more severe and extensive than those of the glia fibers. (4)The changes of the burn included condensation and degeneration of the myelin sheath and axonal cytoplasm, especially the mitochondria as well as edema of the myelin sheath and the axonal cytoplasm. The punch and pressure injuries included the torsion, deformation, fusion, compression, breach, torn and displacement of the axons with loss of cytoplasm as well as the deviation of mitochondria, micro ?neuro - fibrils and micro - tubules in the axons. Conclusion The severe and extensive injuries of the axons of nerve fibers in the brain stem might play an important role in the cause of death of gunshot.

Objective To construct and purify the recombinant protein of platelet-activating factor acetylhydrolase (PAF-AH) isoform I , and study the enzyme activity by different substrates. Methods The (3 subunit of PAF-AH isoform I was cloned and expressed in E. coli. Exogenously expressed recombinant protein was purified to SDS-PAGE homogeneity, and its activity was identified by arylesterase detection. Phenylacetate, 1-O-hexadecyl-2-deoxy-2-thioacetyl-sn-glycero-3-phosphocholine ( 2-Thio PAF) and l-myristoy1-2-( 4-nitrophenylsuccinyl) phosphatidylcholine (the latter two were commercial plasma PAF-AH substrates) were used for the substrate identification. The plasma type PAF-AH was served as positive control. Results Recombinant protein of β subunit of PAF-AH isoform I was successfully constructed and expressed in E. coli after purification. Compared with positive control, the recombinant protein could hydrolyze phenylacetate and 2-Thio PAF, but could not hydrolyze l-myristoyl-2-( 4-nitrophenylsuccinyl) phosphatidylcholine. Conclusion Recombinant protein of β subunit of PAF-AH isoform I can be successfully constructed. There are differences in the substrate specification to the two commercial PAF substrates for PAF-AH isoform I and plasma type PAF-AH, which provides a quick method to differentiate PAF-AH isoform I from plasma type PAF-AH.

Objective To observe the effect of heat needle treatment on the inflammatory reaction in the damaged tissues of rats with the third lumbar transverse process syndrome. Methods Ninety Sprague-Dawley rats were randomly divided into 6 groups, namely normal group, model group, drug group, heat needle group, conventional acupuncture group and warming acup-moxibustion group, 15 rats in each group. The rat model of the third lumbar syndrome was established by embedding gelatin sponge. Two weeks later, the rats in various groups were given corresponding treatment respectively. Before treatment and on the 7th, 14th and 28th day of treatment, the blood samples were collected from jugular vein, and then the serum inflammatory cytokines of tumor necrosis factor alpha (TNF-α), interleukin (IL)-1β and IL-6 were detected by enzyme-linked immunosorbent assay (ELISA). On the 28th day of treatment, all of the rats were sacrificed, the expression levels of TNF-α, IL-1βand IL-6 mRNA in the muscle tissue were detected by reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), and the pathological changes in local muscle tissue were observed by HE staining method. Results The serum contents of the inflammatory cytokines of TNF-α, IL-1β and IL-6 in the model group and treatment groups after modeling were significantly higher than those in the normal group (P < 0.05). After treatment, the contents of TNF-α, IL-1β and IL-6 in various treatment groups were decreased significantly as compared with those in the model group, and the decrease was most obvious in the heat needle group. The expression levels of TNF-α, IL-1β and IL-6 mRNA in muscle tissues of drug group, heat needle group, conventional acupuncture group and warming acup-moxibustion group were lower than those of the model group (P < 0.05), and the heat needle group had the lowest levels. HE staining results showed that heat needle group had milder inflammatory reaction and better muscle fiber structure than the model group. Conclusion Heat needle treatment can improve the local inflammatory response and promote the repair of tissue damage through decreasing the expression levels of inflammatory cytokines of TNF-α, IL-1βand IL-6.

Objective:Select a suitable macroporous resin for the purification technic of total saponins from Panacis Japonici Rhizoma and determine the parameter of purification technic. Methods:Made the content of total saponins as the index, used static adsorption test and combined the adsorption kinetic parameters to select the type of macroporous resin. By using dynamic adsorption experiment to investigate the technical parameters of the purified macroporous resin extracted from Panacis Japonici Rhizoma. Then the preparation technic of the total saponins of Panacis Japonici Rhizoma was determined. Results:The D101 macroporous resin could absorpt and desorpt total saponins of Panacis Japonici Rhizoma effectively. The optimal purification parameters were as follow: the loading mass concentration was 0.1 g/ml (based on crude drug), and the loading volume was 100 ml (which means the loading volume of resin per ml was equivalent to 3.3 grams of crude drug). During the elution process, distilled water (3 BV) and 20% ethanol (3 BV) were used to remove impurity, and then 70% ethanol elution (6 BV) was used to enrich the total saponins. The flow rate of loading and elution was 0.5 ml/min. The transfer rate of total saponins could reache 85.6%. Conclusion:The D101 macroporous resin can effectively enrich and purify the total saponins of Panacis Japonici Rhizoma, which provides the scientific basis for the development and utilization of Panacis Japonici Rhizoma.

Abstract: Objective　To understand the epidemiological characteristics of the migratory bird population with hypertension in Hainan and to provide a basis for the health management of the migratory bird population. Methods　One hundred and eighty migratory birds with hypertension who had lived in in the Mangrove Bay community of Chengmai, Hainan for 3 months or more were selected as the study population. Demographic data, including gender, age, ethnicity, occupation, medication use, family history of hypertension, hometown living environment, Hainan living environment (presence of water area and greenbelt around the residence, type of water area and greenbelt), smoking and alcohol consumption, and height and weight were collected by questionnaire and physical examination to investigate the epidemiological characteristics of the study subjects, and unconditional dichotomous logistic regression analysis was used to identify factors affecting the reduction of blood pressure among the hypertensive migratory bird population before and after taking medication in Hainan. Results　A total of 180 study subjects were included, including 77 (42.8%) males and 103 (57.2%) females, with 78 (43.3%) subjects aged ≤65. The hypertension levels were: 87 (48.3%) had normal high blood pressure, 56 (31.1%) had grade 1 hypertension and 37 (20.6%) had grade 2 hypertension. The higher the blood pressure classification, the lower the percentage of migratory individuals whose blood pressure dropped during their stay in Hainan. Age, hypertension classification, dietary habits, living environment, body mass index (BMI), and alcohol consumption were associated with a decrease in blood pressure during Hainan among the study participants who took hypertension medication regularly. Salty diet (OR=2.778, 95%CI:1.070-7.213, P=0.036) and alcohol consumption (OR=2.686, 95%CI:1.042-6.925, P=0.041) were unfavorable factors for lower diastolic blood pressure before taking medication; overweight (OR=3.487, 95%CI:1.306-9.310, P=0.013) was an unfavorable factor in the reduction of diastolic blood pressure after taking medication. Conclusion　The blood pressure reduction is more significant in migratory individuals in migratory individuals under 65 years old, with a light diet, no alcohol consumption and normal BMI. Environmental factors, age, dietary habits and lifestyle are the main causes of blood pressure, which can provide a reference basis for the health management of the migratory birds with hypertension in Hainan.

To obtain the pure and soluble P51 antigen of HIV-1 strain CN54, we transformed the Escherichia. coli strain BL21 codonplus-RIL with recombinant plasmid pTHioHisA51 which carries a gene encoding the Polymerase (Pol) P51 antigen of HIV-1 CN54 formerly, and induced protein expression by IPTG. We purified the recombinant protein with Chelating Sepharose FF-Ni and DEAE-Sepharose FF column chromatography, then renatured the recombinant protein by dialyzation. Purified protein was identified by Western blotting. We labeled and coated antigen P51 in a dual-antigen sandwich system, and tested it with serum samples from HIV-infected individuals. The results showed that P51 was expressed as inclusion body, and represented about 50% of total cellular protein. After purification and renaturation, the purity of P51 was up to 95%. Western blotting and sandwich ELISA demonstrated that recombinant P51 had good anti-HIV antibody specificity and sensitivity. The results suggested that recombinant HIV-1 P51 can be prepared as diagnostic reagent, and provides valuable support for HIV-1 detection and vaccine research.
Escherichia coli ; genetics ; metabolism ; HIV Antibodies ; blood ; immunology ; HIV Infections ; immunology ; virology ; HIV Reverse Transcriptase ; biosynthesis ; genetics ; immunology ; HIV-1 ; classification ; immunology ; Humans ; Protein Renaturation ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Sensitivity and Specificity

Clinically, hepatocellular carcinoma (HCC) is one of the most common malignant tumors with a high rate of morbidity and mortality. Hepatitis B virus infection is the main cause of hepatocellular carcinoma in China and it is a serious threat to people's health. Antiviral drugs such as nucleos(t)ide analogues and interferon can inhibit viral replication and liver fibrosis progression and reduce the occurrence of hepatitis B-related HCC. This article reviews the effects of different antiviral therapy on the occurrence of hepatitis B-related hepatocellular carcinoma in recent years.

ObjectiveTo investigate the effect of entecavir (ETV) alone or combined with interferon (IFN) on the risk of hepatocellular carcinoma (HCC) in patients with chronic hepatitis B (CHB). MethodsA retrospective analysis was performed for 409 patients with CHB who were admitted to Beijing Ditan Hospital from January 2008 to December 2014, and according to their antiviral therapy, they were divided into ETV+IFN group with 169 patients (IFN treatment for ≥6 months) and ETV group with 240 patients (ETV treatment for ≥12 months). The patients were followed up to June 2019, and the development of HCC was the outcome event. The Mann-Whitney U test was used for comparison of continuous variables between two groups, and the chi-square test was used for comparison of categorical variables between two groups. The propensity score matching (PSM) method was used to eliminate baseline differences between groups, the Kaplan-Meier method and the log-rank test were used to compare the incidence rate of HCC between groups, and the Cox proportional-hazards regression model analysis was used to investigate the risk factors for the development of HCC. ResultsThe median follow-up time in this study was 5.4 years (IQR: 4.9-7.9). There was no significant difference in the cumulative incidence rate of HCC between the two groups before and after PSM (before PSM: 1.2% vs 2.8%, χ2=1.423, P=0.233; after PSM: 1.7% vs 4.1%, χ2=1.676, P=0.195), and the subgroup analysis also showed no significant difference in the cumulative incidence rate of HCC between the two groups in the non-high risk population (1.3% vs 1.5%, χ2=0.335, P=0.563). The Cox proportional-hazards regression model showed that age was an independent risk factor for HCC (hazard ratio=1.107, 95% confidence interval: 1.005-1.219, P=0.038). ConclusionFor CHB patients without a high risk of cancer, compared with ETV monotherapy, ETV combined with IFN for at least 6 months does not significantly reduce the risk of HCC.