Objective To investigate the effects of radix salvia miltiorrhizae on the apoptosis of hepatoeytes during cold preservation and reperfusion injury in rat donor liver. Methods Forty male SD rats were divided into experimental group, control group and sham operation group. The rat model of liver transplantation was established according to the Kamada method. The grafts were preserved in lactated Ringer's solution with radix salvia miltiorrhizae in experimental group and in lactated Ringer's solution in control group for 5 hours, then they were transplanted orthotopically. Six hours after transplantation, the recipients were sacrificed, and the serum ALT and AST were detected by automatic biochemistry analyzer, the hepatocyte apoptosis by TUNEL, the expression of Bcl-2 and FasL protein by flow cytometry. The histopathological changes of the liver grafts were observed under light microscope. Results The levels of ALT and AST in the experimental group were significantly lower than those in the control group after reperfusion. Compared with that in the control group, the apoptosis index of the hepatoeyte was signifieandy decreased in the experimental group ( F = 133.802, P <0.05 ), while the level of Bel-2 protein expression was increased ( F = 91.063, P < 0.01 ). No statistical difference upon FasL protein expression was detected between the 2 groups( F = 1.329, P >0.05). The histopathological injury of the liver grafts in the experimental group was significantly slighter than that in the control group. Conclusions Radix salvia mihiorrhizae inhibits the apoptosis of hepatocytes by increasing the Bcl-2 protein expression during cold preservation and reperfusion injury, so it has a protective effect on the liver graft against ischemia reperfusion injury.

Objective: To investigate the late identification of newly reported HIV/AIDS cases in Wuxing District of Huzhou City, Zhejiang Province from 2012 to 2021, and analyze the influencing factors, so as to provide the basis for improving AIDS prevention and control measures and promoting early identification of HIV/AIDS cases. Methods: Data pertaining to demographics, CD4+T lymphocyte counts, transmission routes and identification routes among newly reported HIV/AIDS cases in Wuxing District from 2012 to 2021 were collected through the HIV/AIDS Comprehensive Control System of Chinese Disease Prevention and Control Information System. The proportions of late identification of cases with different characteristics were descriptively analyzed, and factors affecting late identification were identified using a multivariable logistic regression model. Results: Totally 627 HIV/AIDS cases were reported in Wuxing District from 2012 to 2021, including 555 males (88.52%), and had a mean age of (40.30±15.98) years. There were a total of 212 cases with late identification, and the annual average proportion was 33.81%. The proportions of late identification ranged from 51.85% in 2012 to 28.00% in 2021, with no significant changing trend (χ2trend=1.152, P=0.283). Multivariable logistic regression analysis showed that higher proportion of late identification was more likely seen in the registered cases in Wuxing District (OR=1.651, 95%CI: 1.140-2.393), cases at ages of 40 years and older at diagnosis (OR=1.719, 95%CI: 1.068-2.766), and cases identified by medical institutions (OR=1.809, 95%CI: 1.136-2.881). Conclusion HIV/AIDS cases registered in Wuxing District, aged 40 years and older, and identified by medical institutions have higher proportions of late identification in Wuxing District from 2012 to 2021.

Adult ; Antiphospholipid Syndrome ; complications ; Humans ; Male ; Myocardial Infarction ; complications

Objective To construct eukaryotic expression plasmid pEE14.1-dsFv?pr+,and detect the expression of the recombined gene in eukaryotic CHO-K1 cells.Methods The cationic DNA fragment was cloned into the 3' of VH gene by overlapping extension PCR,and the 6?His tab was inserted to the 3' of VL and human IFN-? gene by the same way.The above mentioned recombinant VH and VL genes were inserted into a pCI-GPI vector first,and then cloned into the pEE14.1 vector to construct the recombinant plasmid pEE14.1-dsFv?pr+.Finally,the recombinant plasmid was transfected into the CHO-K1 cells by LipofectamineTM 2000,and the expression was detected by RT-PCR,ELISA and Western blotting.Results The enzyme digestion and sequencing analysis showed that the recombinant plasmid was successfully constructed.RT-PCR showed that only the cells with transfected plasmid can generate the specific 1700bp fragment.ELISA analysis showed that the production of IFN-?expressed in the supernatant of transfected cells was about 1.1ng/ml.Also,Western blotting could reveal the characteristic band of HBsAg dsFv?pr+ protein.Conclusion The antibody targeting to human IFN-?genes has been successfully expressed in a single open reading frame.Changing the electricity of the antibody may provide the necessary condition for the study of the a new type of anti-HBV drug in nanoscale in the future.

Objective To construct the prokaryotic expression plasmid of human prostate stem cell antigen (PSCA),to induce the expression of GST-PSCA fusion protein in E. coli BL21,and to identify the purified recombinant fusion protein. Methods The fragment of PSCA gene was amplified by PCR,and then cloned into the pGEM-T Easy vector. The transitional plasmid Teasy-PSCA was identified by DNA sequencing. The PSCA gene was digested from the plasmid Teasy-PSCA by restrictive enzyme BamH I and Sal I,and then inserted into the pET42a vector which contains a glutathione s-transterase (GST) tag. Following the double restriction enzyme digestion,the recombinant plasmid pET42a-PSCA was obtained and transformed into E. coli BL21 (DE3). The expression of GST-PSCA fusion protein was induced with IPTG. The recombinant fusion protein was purified by passing over a Glutathione Sepharose 4B column,and was identified by SDS-PAGE and Western blotting. Results The length of amplified PSCA gene fragment was consistent with that expected,and the sequence was correct as exemplified by the PSCA gene reported in GenBank. The result of enzyme digestion indicated that the prokaryotic expression plasmid pET42a-PSCA was successfully constructed. After transformation with pET42a-PSCA and induction with IPTG,the recombinant target protein of about 43kD was obtained. The GST-PSCA fusion protein was correctly identified by SDS-PAGE and Western blotting. Conclusions The prokaryotic expression plasmid of human PSCA gene has been successfully constructed. The GST-PSCA fusion protein may express and be purified in E. coli BL21,and it lays a foundation for further study on the anti-prostate cancer gene vaccine.

Objective To evaluate of the clinical effect of triple arthrodesis and bone grafting combined with plat screw internal fixation for treatment of necrosis of talus. Methods The clinical data of 24 patients with necrosis of talus who were treated by triple arthrodesis and bone grafting combined with plat screw internal fixation from April 2011 to January 2015 in Xiangya Second Hospital of Central South Univer-sity were retrospectively analyzed. And the clinical effect were measured by AOFAS Ankle Hind-foot Scale System. Results The 24 patients were followed up for 26 months averagely (12~36 months). According to AOFAS Ankle Hind-foot Scale System, there were 4 cases of mod-erate and 20 cases of bad before treatment, and there were 18 cases of excellent, 4 cases of good, and 2 cases of moderate after the treat-ment. Conclusion It can achieve good clinical results in treatment of necrosis of talus through triple arthrodesis and bone grafting combined with plat screw internal fixation.

Objective To investigate the influence of tissue-specific growth hormone receptor (GHR)deficiency in type 1 diabetes in the mice at the gene level using pancreaticβcells combined with streptozotocin (STZ)-induced type 1 diabetes model.Methods The experiment was divided into four groups:knockout mice group (LLc knockout group), using the homozygotes (LLc:LL+Cre) producted by pancreaticβ cell-specific expressed recombinant enzyme mice (RIP-Cre)and Cre-LoxP system modified GHR mice (Floxed,LL);LL control group, containing Floxed GHR allele homozygous mice (LL);LLc STZ group and LL STZ group (STZ was used for inducing type 1 diabetes model mice). The mice with feeding glucose≥25 mmol · L-1 were considered to be successful models.The Glucose Tolerance Test (GTT),pancreas tissue HE staining and immunohistochemistry were performed in the mice.Results The blood glucose of the mice in LL STZ group and LLc STZ group and LLc STZ group were increased after inj ection of STZ and the models achieved the diagnostic criteria for diabetes 1 6 d later.The results of GTT showed that compared with LLc control group and LLc knockout group, the blood glucose levels of the mice in LL STZ and LLc STZ groups were increased (P<0.05).There was no significant change of morphology and structure of islets between LL control group and LLc knockout group detected by HE staining. The immunohistochemistry results showed that the insulin level of the mice in LL STZ group was significantly reduced compared with LL control group;the insulin level of the mice in LLc STZ group was reduced compared with LLc control group.Conclusion Pancreaticβcell GHR gene knockout has no effect on the blood glucose and the function ofβcells in the mice with STZ-induced type 1 diabetes.

Objective To explore the possibility that rat bone mesenchymal cells (BMSCs) can differentiate into hepatocytes under the affection of fetal liver filtrate. Methods PAS and green indigo dye were used to detect glycogen and differential level of hepatocytes, respectively. The concentration of ALT, AST, ALP in the culture supernatant were served as markers of hepaocyte function. Results Fourteen days after induced by the fetal liver filtrate, BMSCs changed their shapes into polygon, oval or round. Some of BMSCs were positive for AFP and ALB at 7 days after induction, then the number of positive cells increased, and most of BMSCs expressed AFP and ALB till 21days. The PAS reaction and indocyanine green(ICG) intaking also appeared at 7days. Enzyme in supernatant such as ALT, AST, ALP were fristly detected at 7days and peaked at 14days,then the level declined. Conclusion The fetal rat liver filtrate was able to induce BMSCs into cells with function and characteristics of hepatocytes.

BACKGROUND: Vascular endothelial growth factor family and its receptor play an important role in the process of angiogenesis and neovascularization. Recently, the effect of vascular endothelial growth factor family on blood has been paid much attention. OBJECTIVE: To observe the expression of vascular endothelial growth factor A (VEGFA), vascular endothelial growth factor C (VEGFC) and their receptors as well as angiopoietin-1 (ang-1),angiopoietin-2(ang-2) and their receptors in the blood island of yolk sac and aorta-gonadalmesonephros (AGM) region at embryonic 3 to 12 weeks, so as to investigate the effect of VEGF and angiopoietin family in the process of haematogenesis. DESIGN: Single sample observation. SETTING: Weifang Medical College, Experimental Center of Morphology and Staff Room of Anatomy. MATERIALS: This experiment was carried out at Weifang Medical College from February 2002 to August 2004. Specimens were collected from 47 pregnant women's 3-to-12-week abortive embryos. Informed consent was obtained from the pregnant women. METHODS: Specimens were performed successive sections. Two sections were drawn out every other 10 sections. Hematoxylin-eosin staining and immunohistochemical staining were performed. Polyclonal anti-vascular endothelial growth factor C, fms like protein tyrosine kinase(PTK), flt-4,ang-1, ang-2,Tie-2 antibody, monoclonal anti-vascular endothelial growth factor A and fetal liver kinase 1 (flk1) antibody were used for incubation overnight at 4 ℃ .Goat anti-mouse IgG and SABC solution were used separately for 2 hours. 3,3'2 diaminoazobenzidine (DAB) was used to develop. Serum of normal rabbit or mouse were used separately to replace primary antibody as negative control. They were observed and taken photographs under optical microscope. MAIN OUTCOME MEASURES: ①Blank expression of VEGFA and VEGFC and their receptors in the blood island of yolk sac and AGM region at embryonic 3 to 12 weeks. ②Blank expression of ang-1 and ang-2 and their receptors in the blood island of yolk sac and AGM region at embryonic 3 to 12 weeks. RESULTS: ①On the 21st to 25th day , vascular endothelial cells and hematopoietic cells in blood island of yolk sac strongly expressed VEGFA and its receptor fms like PTK and flk1, VEGFC and its receptor flk4, ang2 and its receptor Tie-2 protein, weakly expressed ang-1 protein. ②From the fourth week of development, dorsal aorta and mesonephros expressed above-mentioned various factors and their receptors . Immune positive reactants gathered in the large and round hematopoietic cells with nucleus. The cell quantity reached the peak at week 7. After week 8, the number of positive cells was significantly decreased, and almost all the blood cells were immune negative reaction at week 12. ③Gonad mainly expressed VEGFA , fms like PTK, flt-4,ang-1 ,ang-2 and Tie-2 protein at weeks 6 to 8 , but did not express VEGFC and flkl. ④ The expression of above-mentioned factors were not detected in the vascular endothelial cells in AGM region. CONCLUSION: Hematopoietic cells and endothelial cells of blood island of yolk sac of human embryo as well as dorsal aorta and mesonephros coexpresses various factors related to angiogenesis and haematogenesis. Haematogenesis of human embryos occurs at the fourth week.

Objective: To investigate the expression of ATP-Binding Cassette Proteins including P-gp (P-glycoprotein), MRP1 (multidrug resistance associated protein 1) and BCRP (breast cancer resistance protein) in hepatocellular carcinoma and its relationship with pathological features. Methods: The expression of P-gp/MDR1 (multidrug resistance gene 1), MRP1 and BCRP in hepatocellular carcinoma was examined by RT-PCR and immunohistochemistry in 34 cases of hepatocellular carcinoma and 19 cases of paraneoplastic hepatic tissues. Results: The expression of MDR1, MRP1 and BCRP mRNA (messenger ribonucleic acid) was 1.15±0.24, 0.64±0.33, and 1.07±0.32 in hepatocellular carcinoma and 0.36±0.14, 0.19±0.06, and 0.31±0.09 in paraneoplastic hepatic tissues. The expression of MDR1, MRP1 and BCRP mRNA was 1.38±0.26, 0.73±0.35, and 1.34±0.21 in poorly differentiated hepatocellular carcinoma and 0.74±0.32, 0.30±0.11, and 0.45±0.13 in well differentiated hepatic tissues. The immunohistochemical positive substance was detected in the plasma membrane and cytoplasm. The positive rates of P-gp, MRP1 and BCRP were 82.35%, 58.82%, and 79.41% in hepatocellular carcinoma and 42.11%, 26.32%, and 36.84% in paraneoplastic hepatic tissues, respectively. The positive rates of P-gp, MRP1 and BCRP were 100.00%, 81.25%, and 100.00% in poorly differentiated hepatocellular carcinoma and 66.67%, 38.89%, and 61.11% in well differentiated hepatic tissues. The expression of three indicies in hepatocellular carcinoma was higher than that in paraneoplastic hepatic tissues (P<0.05). The expression of P-gp/MDR1, MRP1 and BCRP in poorly differentiated hepatocellular carcinoma was higher than that in well differentiated hepatic tissues (P<0.05). No correlation was found among the three indices. Conclusion: Intrinsic multidrug resistance exsists in hepatocellular carcinoma, with various mechanisms. The multidrug resistance of HCC (hepatic cell carcinoma) is related to P-gp/MDR1, MRP1 and BCRP. MRP1 and BCRP may be targets for reversing multidrug resistance.