Animals ; Gene Expression Regulation ; drug effects ; Gene Targeting ; methods ; Humans ; Oligonucleotides, Antisense ; genetics ; pharmacology ; RNA Interference ; RNA, Small Interfering ; genetics ; Transfection

Objective To explore the effects of oxidative stress in porcine iliac endothelial cells(PIECs) induced by high glucose. Methods After being intervened by high glucose for some time, dihydroethidium (DHE) or dihydrorhodamine 123 (DHR123) was used as a reactive oxygen species(ROS) capture. The mean fluorescent intensity(MFI) of above probes which were the products of intracellular oxidation was detected by fluorescent microscopy and flow cytometry, and the level of ROS was thus measured. With lucigenin as chemiluminescence agent, luminescence changes were detected by chemiluminescence analyzer after addition of NADPH to observe the effect of high glucose on activity of NADPH oxidase(NOX). Results Intracellular MFI was markedly elevated with the concentration of high glucose and time of exposure to high glucose. It was revealed by flow cytometry that the NOX activity was significantly activated compared with normal medium treated PIECs(P

To investigate the effects of icariin, an effective extract from traditional Chinese medicine Epimedium pubescens with the function of tonifying kidney, in promoting osteogenesis of mesenchymal stem cell line C3H10T1/2, and to explore the underlying mechanism.

Objective: To observe the therapeutic effect of intravenous thrombolysis of recombinant streptokinase (r SK) on acute myocardial infarction (AMI) and its safety. Methods: Twenty patients with AMI received r SK for thrombolysis.The reopening rate of infarct related artery,side effect and the fatality rate in hospital were observed. Results: The reopening rate of infarct related artery was 75%. The incidence of slight hemorrhage, fever, low blood pressure,which could be corrected in short time, was 5%, 15%, 15%, respectively. The fatality rate 5 weeks after AMI was 10%. Conclusion: The therapeutic effect of r SK in the thrombolytic therapy of AMI is definite and the safety is fine.

Objective To explore the effect of advanced glycation end products(AGEs) on the oxidative stress in porcine vascular endothelial cells(PIECs). Methods After being intervened by AGEs for some time,cell viability was detected by MTT.2′,7′-dichlorofluorescein diacetate(DCFH-DA) was used as a reactive oxygen species(ROS) capture agent.The fluorescent intensity of 2′,7′-dichlorofluorescein(DCF),which was the product of cellular oxidation of DCFH-DA,was detected by flow cytometry,and the level of ROS was thus measured. Results Viability of PIECs was inhibited by AGEs in a dose-and time-dependent fashion(P

BACKGROUND: Adipose tissue-derived mesenchymal stem cells (ADMSCs) have the multilineage differentiation potential, and are relatively easier to be obtained, thus they have attracted more and more attention as a new seed cell for cell engineering.OBJECTIVE: To observe the in vitro culture conditions of ADMSCs isolated from rat's subcutaneous adipose tissue, and identify them using immunohistochemical staining.DESIGN: An animal experiment.SETTING; Department of Cardiology, the General Hospital of Chinese PLA.MATERIALS: One healthy male Wistar rat of clean degree, 4 months old, weighing 200 g, was used. DMEM, fetal bovine serum were from GIBCO; Monoclone antibodies of rabbit-anti-rat CD13, CD34, CD44, CD45, CD105, D-related human leucocyte antigen (HLA-DR), factor-Ⅷ, vov Willebrand factor (VWF), Myosin, SABC kits and DAB staining kit from Wuhan Booster Biological Engineering, Co.,Ltd; Adeno-associated virus encoding green fluorescent protein from Vector Gene Technology Company Limited (Beijing).METHODS: The experiments were carried out in the Department of Internal Medicine, the General Hospital of Chinese PLA in October 2006. ① Cell isolation and culture: 0.3 g adipose tissue was cut from subcutaneous adipose tissue of Wistar rat's groin under aseptic condition, then minced and digested before culture, DMEM was changed at 2-3 days after plenty of fusiform-shap ed attached cells were observed under microscope, and the cell growth was observed. The cell concentration was adjusted to 2×107 L-1, then seeded into 96-well plate, and 100 μL for each well. From the second day, 3 wells were randomly selected every day, the cells were released with tripsin, and counted with blood cell counting chamber under inverted microscope. ② Cell viability assay: ADMSCs of passages 3 to 8 were added to DMSO freeze medium, and thawed after 2-4 weeks, and the cell viability was assessed by trypan blue dye exclusion. ③Immunohistochemical staining and identification: 2 ×107 L -1 cells were seeded to culture plate, then the immunohistochemical (SABC method) identification and Oil red O staining were performed to determine the cell surface antibodies of CD13, CD34, CD44, CD45, CD105, HLA-DR, factor-Ⅷ, HLA-DR and VWF. ④Lineage-specific differentiation and identification: The ADMSCs were plated on multi-well chamber and induced with lineage-specific media supplementation at least two weeks and identified by histologic/immunohistochemical assay of Oil red O for adipogenisis, alkaline phosphatase (ALP) stain for osteogenisis and Myosin monoclonal antibody for myogenisis. ⑤Transfected adenovirus carried green fluorescence protein (AD-GFP) medium: The fourth generation of ADMSCs were seeded on 96-well plate, 3 000 cells for each well, serum-free DMEM was changed after 24 hours, and added by AD-GFP at the same time, then transfected with different multiplicity of infection (MOI) of 1∶50, 1∶100, 1∶150 and 1∶200respectively, and then the transfection was observed.MAIN OUTCOME MEASURES: ① Results of cell isolation and culture; ② Cell viability after freezing and thawing; ③Results of immunohistochemical staining and identification; ④ Results of lineage-specific differentiation and identification;⑤ Results of transfected adenovirus carried AD-GFP.RESULTS: ① About 3.6×105 attached cells were obtained from 0.3 g subcutaneous adipose tissue, and these cells could be subcultured for passages in vitro with stable population doubling time. ② The cells were thawed after freezing for 2-3 weeks, and the trypan blue staining showed that the cell viability was above 90%. ③ The immunocytochemical staining showed that CD13, CD44, CD105 were positive and CD45, factor-Ⅷ, HLA-DR and VWF negative in different generations. ④ From the second generation, a few Oil red O positively stained cells were observed, which were obviously increased after prolonging the refreshing. After lineage-specific differentiation, the cells were all positive by Oil red O staining, ALP staining and Myosin immunohistochemical staining. ⑤ 72 hours after transfection, it was observed under fluorescence microscope that most cells were green fluorescence when the MOI value was 1∶200, the transfection was successful, and it was generally determined that the transfection rate was above 90%.CONCLUSION: A large number of ADMSCs with multilineage differentiation potential can be easily obtained from rat adipose tissue, osteoblast, myoblasts, they can be expanded in large quantity and stored in vitro for long time, AD-GFP were also successfully transfected.

Objective To investigate the protective effects of erythropoietin (EPO) on myocardium injury after sepsis in rats in order to clarify the mechanisms.Methods The rat models of sepsis were produced by cecal ligation and perforation method (CLP).Ninty-six healthy SD rats were randomly (random number) divided into 3 groups:the sham operation group (Sham group,n =32),the sepsis group (CLP group,n =32),and the EPO treatment group (EPO group,n =32) treated with EPO 1000 IU/kg intraperitoneal injection after the CLP.The observation intervals were set at 3 h,6 h,12 h and 24 h after the surgery.The cardiac hemodynamics of the CLP group were measured.Plasma levels of inflammatory factors and myocardial enzyme indicators were determined by enzyme-linked immunosorbent assay (ELISA) ; Membrane potential levels of chondriosome of myocardial cells,cell apoptosis rates and expressions of nuclear factor-κB p65 of myocardium tissue were detected by flow cytometer; Then the pathological change of myocardium with HE staining was observed under light microscopy.Results (1) Compared with the CLP group,left ventricle systolic pressure (LVSP),left ventricle diastolic end pressure (LVEDP),the maximum rate of left ventricle rise and fall (+ dp/dtmax and-dp/dtmax) in the EPO group improved (P ＜0.05,P＜0.01); (2) Compared with the Sham operation group,plasma levels of tumor necrosis factoralpha (TNF-α),interleukin-6 (IL-6),C-reactive protein (CRP),cardiac troponin-I (cTNI),creatine kinase (CK),lactate dehydrogenase (LDH) and glutamine-oxaloacetic transaminase (GOT) in the CLP group increased at each interval (P ＜ 0.05),and those biomarkers in the EPO group were lower than those in the CLP group (P ＜ 0.05) ; On the contrary,plasma level of anti-inflammatory factor IL-10 in EPO group was higher than that in the CLP group (P ＜ 0.01) ; (3) Compared with the Sham operation group,the cell apoptosis rates in the CLP group increased significantly (P ＜ 0.01),and it decreased obviously in the EPO group (P ＜ 0.01); (4) Compared with the Sham group,membrane potential levels of chondriosome of myocardial cell in the CLP group decreased (P ＜0.01),while it increased in the EPO group in comparison with the CLP group (P ＜ 0.01) ; (5) Pathological changes of myocardium after the CLP could be lessened by the EPO treatment.Conclusions EPO may increase membrane potential levels of chondriosome and decrease the apoptosis rates of myocardial cell in rats after sepsis,and it may reduced the production of inflammatory factors to protect the myocardial cell by down-regulating NF-κB p65.Both increased membrane potential level of chonriosome and decreased inflammatory factor may implicate in myocardium protection thereby improving cardiac function after sepsis.

The true meanings of the terms of traditional Chinese medicine (TCM) need to be analyzed on a logical basis. It is not suitable to use a new term to interpret an old term of TCM, or arbitrarily specify the special term of TCM corresponding to some substances of modern medicine. In philosophy of language, language has a logical structure, which reflects the structure of the world, that is to say, language is the picture of the world in a logical sense. Using this idea, the authors collected the ancient literature on "kidney essence", and extracted each necessary condition for "kidney essence". All necessary conditions formed a sufficient condition to define the term "kidney essence". It is expected that this example can show the effectiveness of philosophy of language in analysis of the terms of TCM.

Objective To explore the effects of high glucose on oxidative stress of human peritoneal mesothelial cells(HPMCs).Methods HPMCs were cultured in vitro and identified by immunohistochemistry, and cells of second generation were selected. After HPMCs were treated by glucose with different concentrations for some time, MTT method was employed to detect the cell viability. 2’,7’-dichlorofluorescein diacetate (DCFH-DA) was used as a reactive oxygen species (ROS) capture. The cell viability of HPMCs and ROS level were analysed after being intervened by glucose with different concentrations and for different time. Results Viability of HPMCs was significantly inhibited in a dose-and time-dependent manner by high glucose(P