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BACKGROUND:To improve the survival rate of transplanted hematopoietic stem cells, dynamic monitoring of plasma content of human cytomegalovirus (HCMV) and rapid screening of early active HCMV infection in hematopoietic stem celltranplantation recipients, thus to guide the clinical medication, is preferred. OBJECTIVE:To evaluate the usefulness of fluorescence quantitative PCR assay for early detection of HCMV activation after hematopoietic stem celltransplantation. METHODS:Real-time fluorescence quantitative PCR assay was applied for HCMV monitoring in 656 blood samples from 41 hematopoietic stem celltranplantation recipients and 60 control blood samples. RESULTS AND CONCLUSION:In 656 blood samples, 96 samples were positive, and the HCMV DNA copies ranged from 5.0×102 to 1.0×107 IU/mL. Timely initiation of therapy resulted in the rapid clearance of DNA-viraemia but it recurrenced in short time by drug-resistent. Two cases from 12 postive recipients died from HCMV infection. The quantitative detection of HCMV DNA by real-time PCR is a rapid method for monitoring HCMV infection and the early diagnosis in patients after hematopoietic stem celltransplantation.
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BACKGROUND:Mesenchymal stem cells(MSCs) have immunoregulation,can reduce the occurrence rate of graft versus host disease following transplantation,and treat autoimmune disease. OBJECTIVE:To observe the immunoregulation of human umbilical cord(UC) -MSCs on bone marrow T-lymphocyte of patients with aplastic anemia. DESIGN,TIME AND SETTING:A randomized grouping design,in vitro cytological controlled study. Cases were obtained from the Department of Hematology,First Hospital and Second Hospital Affiliated to Nanchang University. Experiments were conducted at the Institute of Hematology,Second Hospital Affiliated to Nanchang University from September 2007 to November 2008. PARTICIPANTS/MATERIALS:A total of 22 patients with aplastic anemia were enrolled at the Department of Hematology,First Hospital and Second Hospital Affiliated to Nanchang University. There were 6 cases of severe aplastic anemia(3 males and 3 females) ,with a median age of 38(16-68) years;16 cases of chronic aplastic anemia(9 males and 7 females) ,with a median age of 41(25-59) years. Umbilical cord was obtained from healthy full-term fetus by normal uterine-incision delivery for isolation,culture and determination of human UC-MSCs. METHODS:MSCs were extracted from human umbilical cord. T-lymphocytes were isolated from bone marrow of patients with aplastic anemia by density gradient centrifugation. MSCs were cocultured with T-lymphocytes. In the control group,T-lymphocytes were incubated at 1?105/well. In the UC-MSCs group,human UC-MSCs were incubated at various densities of 1?105/well,0.5?105/well,0.1?105/well and 0.05?105/well. In the experimental group,T-lymphocytes were cocultured with UC-MSCs at 1:1,1:0.5,1:0.1,1:0.05. MAIN OUTCOME MEASURES:Inhibitory rate of human UC-MSCs at various concentrations on T-lymphocytes of aplastic anemia patients was measured by MTT assay. CD4+/CD8+ changes of the T cells were analyzed by flow cytometry. Expression of the two marrow hematopoietic factors(interleukin-2 and ?-interferon) in the supernatant was evaluated by ELISA. RESULTS:The second passage of MSCs had typical morphology of fibroblast,and highly expressed CD166,CD29,CD54,but lowly expressed CD13,CD34,CD45. MSCs could be induced into adipocytes. UC-MSCs showed inhibitory effect on lymphocyte proliferation of aplastic anemia patients when UC-MSCs and lymphocytes mixed at the ratio of 1:1,0.5:1,0.1:1,and their inhibitory rate were(56.2?12.1) %,(43.7?10.4) %,(28.6?8.9) %. The effect was the strongest at the ratio of 1:1. UC-MSCs adjusted the proportion of CD4+ and CD8+,inhibited the secretion of interleukin-2 and ?-interferon by T cells of aplastic anemia patients. CONCLUSION:Human UC-MSCs can mediate an immunoregulation effect on T lymphocytes of aplastic anemia patients in vitro and the inhibitory effects are dependent on the amount of UC-MSCs.
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OBJECTIVE:To investigate the use of antibacterials and to evaluate the distribution and drugs resistance of pathogenic bacteria causing hospital nosocomial infection between 2005 and 2007 in our hospital. METHODS:The antibiotic resistance and the data of drug use from 2005 to 2007 in our hospital were analyzed statistically. RESULTS:Over the 3 years,the production rate of ESBLs of Escherichia.coli ascended year by year. The proportion of methicillin resistant staphylococcus aureus (MRSA) in staphylococcus aureus were 79.4%,66.3% and 73.9% respectively over the 3 years. The DDDs of antimicrobials showed an year-on-year increase,with third-generation cephalosporins and fluoroquinolones topping the first two places. CONCLUSION:Antibacterial drugs should be used rationally in clinical practice so as to reduce antibiotic resistance.
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OBJECTIVE:To optimize the extraction of effective ingredients and purification for crude glycyrrhizic acid and liquiritin from Radix et Rhizoma Glycyrrhizae with different methods. METHODS:The contents and extraction percentages of glycyrrhizic acid and liquiritin were used as indexes to optimize the extraction solvent. Then the extraction process was optimized with orthogonal experiment taking the concentration of ammonium ethanol,the amount of solvent and the extracting times as indexes. Crude glycyrrhizic acid was purified with the method of recrystallization and crude liquiritin was purified with organic solvent method.RESULTS:The optimal extracting solvent was ammonium ethanol. The optimum extraction conditions were as follows:extracting the solvent 4 times(2 h/time) by adding 5 times volume of 60% ethanol(containing 0.3% ammonia water). The purity of glycyrrhizic acid was over 85% with total extraction rate at 43%,and content of liquiritin was above 15% with its total extraction rate at 67%. CONCLUSION:The method is simple and feasible and from which glycyrrhizic acid and liquiritin was been obtained,Radix et Rhizoma Glycyrrhizae can broaden the scope of the application.
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Objective To establish a method of culturing mesenchymal stem cells from umbilical cord blood. Methods The mononuclear cell fraction of umbilical cord blood was isolated with the aid of a Ficoll/Hypaque gradient, then MSC were cultivated in serum containing medium or serum-free medium. The serum containing medium (SC) consisted of LG-DMEM supplemented with 10% fetal calf serum and 10-6M hydrocortisone, with a final cell concentration of 1?106 cells/ml. The serum-free medium (SF) was supplemented with 1?10 -6M hydrocortisone and 40ng/ml fibronectin with a final cell concentration of 1?106 cells/ml. After a week, 50% of the media were renewed. Before reaching the monolayer phase cells were trypsinised and re-cultured under similar conditions for the another 3 weeks. Cells were counted at the beginning and the end of the culturing period. Phenotypes were identifiad by PAS, NAE, ALP staining. Identification of surface markers and cell cycle analysis of MSCs were performed with flow cytometry. Results Adherent cells appeared 24h after plating of mononuclear cells. The cells formed adherent heterogeneous cell populations after 4-7 days in culture, and they consisted of round and spindle-like cells. In the primary passage of culture, the cells proliferated slowly and became confluent in 14-20 days. When subcultured, the heterogeneous cell populations became homogeneous assuming flat and fibroblast-like shape. Though colonies in the medium containing serum formed earlier than those in the serum-free medium, passage times and cell morphology were the same. The total cell numbers in SF group were lower than those of SC group. The percentage of cells at G0-G1 cell cycle was higher in SF group than that of SC group with significant difference. By flow cytometry analysis, cells of two groups were negative for CD34, CD13 and CD45, but strongly positive for specific surface markers such as CD166, CD29, CD105 and CD54. Conclusion Serum-free medium was superior to medium containing serum for culturing MSCs.