Objective To explore the capacity of hematopoietic cells from human umbilical cord blood (UCB) induced by hematopoietic growth fators (HGFS) to differentiate committedly to granulocyte in vitro . Methods Suspension culture of isolated mononuclear cells from fresh UCB samples were conducted for 5 weeks in IMDM medium containing UCB serum and HGFS. The expansion folds of nucleated cells, the changes of CD34+antigen by immunofluorometry, and the colony forming unit granulocyte macrophage(CFU GM) by progenitor clonogenic assay were observed. Results Culture of mononuclear cells with the combination of granulocyte colony stimulating factor(G CSF), granulocyte macrophage colony stimulating factor (GM CSF), stem cell factor(SCF), interluekin 3(IL 3) and interleukin 6(IL 6) for 3 weeks markedly increased the total count of nucleated cells and CFU GM, being 58.3 and 12.2 folds of the pre cultured, and the cultured cells were mainly neutrophils. However, CD34+ cells in nucleated cells declined from 0.97% to 0.05%. According to the culture condition, a single portion of fresh UCB could increase nucleated cells by (5.1?2.3) fold and CFU GM by (2.2?0.95) fold. Conclusion Hematopoietic cells of UCB may be induced to differentiate committedely to granulocyte lineage by this culture system in vitro to result in the increase of granulocytes and CFU GM in single portion of cord blood.

Objective To study the value of MR perfusion-weighted imaging(PWI)after administration of acetazolamide(ACZ)in evaluating the cerebrovascular capacity.Methods MR perfusion-weighted imaging were performed before and after ACZ administeredorally in 7 cases with cerebral infarction.The values of rCBV and rMTT were analysed and the cerebrovascular capacity were evaluated.Results The prolonged rMTT was found in rMTT map before and after ACZ in all cases.Only 5 subjects were abnormal at rCBV map visually.rMTT was increased at the abnormal region on rMTT map.The abnormal regions at rCBV maps included both rCBV increased or decreased.The abnormal region was larger at rMTT map than that at rCBV map.Conclusion MR perfusion-weighted imaging with ACZ test has significant value in assessing the cerebrovascular capacity.

Acute Disease ; Adolescent ; Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; Toluene ; poisoning ; Xylenes ; poisoning ; Young Adult

Objective To explore the expression and significance of hedgehog signal molecules (Ptch, Smo and Gli1 ) in pancreatic cancer. Methods Two hundred SD rats were randomly divided into DMBA group ( group A, n = 90), cyclopamine intervening group ( group B, n = 90) and control group ( group C, n = 20).For group A and B, DMBA was directly implanted into the parenchyma of the pancreas to establish the model of pancreatic cancer. The rats in group B were treated with 6.25 ml/kg cyclopamine and DMSO solution intraperitoneally daily. All rats were sacrificed four months later to observe the pancreatic tissue pathologic changes, and immunohistochemistry SP was used to detect the expression of Ptch, Smo, Gli1 protein in pancreatic cancer and normal pancreatic tissue. Results The prevalence rate of pancreatic cancer in group A was 57.5％ (46/80), the maximum size of the tumor was 0.5 ～ ＞2 cm; the prevalence rate of pancreatic cancer in group B was 17.1％ ( 14/82), the maximum size of the tumor was 0.5 ～ 2.0 cm, and the difference between the two group was statistically significant (P ＜0.05). The positive expression rate of Ptch, Smo and Gli1 protein was 82.6％, 73.9％ and 65.2％ in DMBA group, and was 50.0％, 42.9％ and 28.6％ in cyclopamine group, and the difference between the two group was statistically significant ( P ＜ 0.05 ). Ptch,Smo and Gli1 protein was expressed in normal pancreatic tissue. Conclusions Direct implantation of DMBA in the parenchyma of rat pancreas can induce pancreatic cancer with a high incidence in a short time.Hedgehog signal protein expression is significantly increased, cyclopamine can inhibit the occurrence and progression of pancreatic cancer by inhibiting Hedgehog messenger expression.

Objective To investigate the effects of angiotensin Ⅱ (Ang Ⅱ ) and losartan on the expression of small ubiquitin-related modifier(SUMO) protein (SUMO1 ,SUMO2/3) in cultured rat glomerular mesangial cells(GMCs) .Methods In vitro cul-tured HBZY-1 rat GMCs were divided into 5 groups:normal control group(NC group) ,different concentrations of Ang Ⅱinterven-tion groups(A1 ,A2 ,A3 groups) and losartan treatment group(MT group) .The expression of SUMO1 and SUMO2/3 protein and mRNA of each group was measured by Western blot and RT-PCR .Results Compared with the NC group ,the expression of SU-MO1 and SUMO2/3 protein and mRNA in the Ang Ⅱintervention groups and the losartan treatment group was increased signifi-cantly (P<0 .01);Compared with the Ang Ⅱintervention groups ,the expression of SUMO1 and SUMO2/3 protein and mRNA in the losartan treatment group had no statistically significant difference .Conclusion Ang Ⅱ up-regulates the expression of SUMO protein in rGMCs by a dose-dependent manner in certain range ,this effect is not blocked by losartan ,Ang Ⅱ may be involved in the pathogenesis of diabetic nephropathy .

BACKGROUND: Some researches suggest that placenta is regarded as a new source of mesemchymal stem cells (MSCs). Adherent cells derived from placenta tissue have similar morphological characteristics and surface markers to MSCs; meanwhile, they can differentiate into osteoblasts and nerve cells.OBJECTIVE: To find a new source and way for MSCs separation.DESIGN: Observational study.SETTING: The Third People's Hospital of Wuxi.MATERIALS: Placenta was sourced from cesarean in our department of obstetrics and gynecology and provided confirmed consent from the relatives and ethics committee. The main reagents contained culture medium, sABC kit, DAB staining kit, caprine-anti-rat FITC, recombinant human basic fibroblast growth factor (rh-bFGF), rabbit-anti-human glial fibriliary acidic protein (GFAP).METHODS: The experiment was carried out in the Laboratory of Cells & Molecular Biology, the Third People's Hospital of Wuxi from May 2005 to August 2006. Placenta entity tissue was digested, adherent cells were cultured, morphological characteristics were observed, and growth curves of placenta-derived multipotent cells (PDMCs) in various generations were drawn. On the 1st and 7th days, supernatant was derived from cells in primary culture to measure content of β-glycerophosphate disodium (β-HCG) with chemiluminescence technique. In addition, expression of surface antigen and differentiating potency were detected at the same time. At 24 hours (at phase of nerve cells) and 2 weeks (at phase of osteoblasts) after inducible differentiation, cultured cells were dealt with routine immunocytochemical stain, and then they were observed under routine microscope or fluorescence microscope.MAIN OUTCOME MEASURES: ① Morphological indexes and growth curves of PDMCs; ② measurement of β-HCG in supernatant of PDMCs and expression of antigen of PDMCs with flow cytometer; ③ immunohistochemical analysis of PDMCs during and after inducible differentiation.RESULTS: ① Primary culture of PDMCs: After digestion of placenta tissue, a few of adherent cells were obtained and gradually formed thin and flat monolayer cells two weeks later. The monolayer cells grew like whirlpool or cluster. With the increase of cell density, soma was slenderer and slenderer and like fibroblast. ② Growth curves of PDMCs: Growth latency ranged from 2 to 8 days after cell inoculation. During this period, adherence was observed gradually but not obviously amplified. Eight days later, cells entered log growth phase. During this period, proliferation was active and cell process surrounding extended under phase contrast microscope. A lot of MSCs were observed in division phase between two nuclei. In addition, density was increased and cells connected to each other. Within 11-14 days, growth curve gradually entered platform phase. MSCs covered the bottom of bottle, cells slowly expanded, and primary culture stopped. ③ Measurement of β-HCG: Expression of β-HCG was not detected in supernatant at two time points. ④ Characteristics of surface antigen of PDMCs: PDMCs could express CD29, CD44 and CD105, but not CD34, CD45,CD19 and CD106. ⑤ Inducible differentiation of PDMCs: At 24 hours after inducing to nerve cells, form of PDMCs obviously changed, soma rebounded, refraction of nucleus was partially reinforced, structure was similar to dendrite and axis-cylinder, and positive neuro-specific enolase and GFAP were observed after staining.CONCLUSION:Placenta tissue contains PDMCs whose morphological function is similar to MSCs; in addition, placenta is regarded as an effective source of MSCs.

BACKGROUND: Multipotential mesenchymal stem cells (MSCs) can be differentiated into bone, fat, cartilage, muscle and endothelial cell at the specific conditions, so it draws great attentions of the related study.OBJ ECTTVE: To establish the method of in vitro culture and expansion of human umbilical blood-derived MSCs and investigate their influencing factors.DESTGN: Randomized and controlled trials.SETTING: Department of Tissue Engineering and Cell Biology in Wuxi Third People's Hospital.MATERTALS: Seventy cases of human umbilical cord blood (HUCB) of healthy neonates (selected from Parturient Room, Department of Gynecology and Obstetrics, Wuxi Third People's Hospital, with the informed consents of the puerpera and their relatives), 40 mL each; dulbecco's minimal essential medium (DMEM) of low glucose (LG) at the dose of 50 g/L, fetal bovine serum (FBS), mycillin and trypsin (all were from Gibco), FITC-labeled mice anti-human CD29, CD105 and CD106 (Ancell) monoclonal fluorescent antibody, PE-labeled mice anti-human CD34, CD44, CD45, CD19, HLA-DR (Immunotech)monoclonal fluorescent antibody, Ficoll separation medium (Pharmacia).METHODS: The experiment was carried out in Department of Tissue Engineering and Cell Biology in Wuxi Third neonatal cord blood were anticoagulated by 25 u/mL heparin, and 60 cases of isolated HUCB mononuclear cell were precipitated and suspended by cell culture fluid (50 g/L DMEM-LG + 50 g/L FBS + 10 g/L mycillin), while other 10 cases were purified with DMEM of high glucose (100 g/L) to produce adherent layer. Once the cells reached fluences of FBS coating on adhesive rats of MSCs: Some of the purified HUCB MSCs at logarithmic phase were growth curve of MSCs: The growth of cells were observed by phase contrast microscope (OLYMPUS CK40), and were detected by EPICS-ALTRA flow cytometry.FBS coating and without FBS coating at the different time points, and their adhesive rates.RESULTS: All 70 cases of HUCB MSCs were adopted in this study, however, 10 samples failed to obtain the ideal MSCs cord blood MSCs after culture. During 2 weeks of culture, primary cells began to double for 3-4 days and reached conand CD105, but did not express antigens CD34, CD45, CD106 and HLA-DR, which was identical to surface labeling of those without FBS coating (P＜0.01).CONCLUSrON: MSCs in HUCB can be cultured and expanded in vitro, and are regarded as an alternative source of MSCs for experimental and clinical applications. Moreover, high glucose can depress the growth and proliferation of MSCs in HUCB.

Objective To detect the expression of NF‐κB ,MCP‐1 in the endometrial carcinoma and their significance .Methods The expression of NF‐κB ,MCP‐1 in 52 cases of patients with endometrial carcinoma selected from January 2003 to December 2013 , and in 16 cases of endometrial atypical hyperplasia and 19 cases of proliferative endometrial were detected by immunohistochemistry assay .The relationship and correlation between clinical pathological characteristics was analyzed .Results The expressions of NF‐κB ,MCP‐1 proteins were significantly higher in the endometrial carcinoma than those of the endometrial atypical hyperplasia and proliferative endometrial (P<0 .05) .The expressions of NF‐κB and MCP‐1 were related with histological grade and myometrial in‐vasion (P<0 .05) ,but independent of clinical stage and lymph node metastasis (P>0 .05) .The expressions of these two proteins were positively correlated(r=0 .895 ,P<0 .05) .Conclusion NF‐κB ,MCP‐1 may participate in the progress of endometrial carcino‐ma .

OBJECTIVE: To investigate the anti-tumor effect of bufalin and its regulation on Bcl-2 and Bax proteins in orthotopically transplanted tumor of human hepatocellular carcinoma in nude mice. METHODS: Orthotopically transplanted tumor of human hepatocellular carcinoma was established in nude mice. The mice were randomly divided into five groups: high-dose bufalin-treated group (1.5 mg/kg), medium-dose bufalin-treated group (1 mg/kg), low-dose bufalin-treated group (0.5 mg/kg), adriamycin-treated group (8.0 mg/kg), and normal saline-treated group. After 25 days, mice were sacrificed. The tumor volume was measured, and the pathological changes of tumor tissues were detected by HE staining to observe the tumor necrosis degree. Cell morphological changes were also observed by an electron microscopy. Label index of tumor cell apoptosis was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), and the expressions of Bcl-2 and Bax proteins were determined by immunohistochemical method. RESULTS: The tumor volume in the bufalin-treated groups was shrunk significantly compared with that in the normal saline-treated group (P<0.01). The survival time of the bufalin-treated groups was prolonged compared with that of the adriamycin-treated group and the normal saline-treated group P<0.05. Apoptotic characteristics could be seen in tumor tissues of the bufalin-treated groups. The label index of tumor cell apoptosis in the bufalin-treated groups (5.87+/-2.13, 8.86+/-2.96 and 10.60+/-3.42 in low-, medium- and high-dose groups respectively) was higher than that in the adriamycin-treated group (3.28+/-0.98) (P<0.05, P<0.01). The expression of Bax was up-regulated, while no changes were detected as to Bcl-2 protein in tumors of the bufalin-treated groups. CONCLUSION: Bufalin has significant anti-tumor effect on the orthotopically transplanted tumor of human hepatocellular carcinoma in nude mice. Its effect might be related to up-regulation of Bax protein and inducement of the tumor cell apoptosis.

Aim To investigate the protective effect and mechanism of madecassoside from hydrocotyle sibthorpioides (MHS)on learning and memory impair-ment induced by D-galactose (D-gal)in mice.Meth-ods Totally 75 SPF Kunming male mice were divided into normal control group,model group,low-,middle-and high-doses of MHS treated groups.The dementia models were induced with D-gal.The learning and memory functions were tested by Morris water maze, the level of Aβ1 -42 and its related proteins in the hip-pocampus was determined by Western blot,and the ex-pression of Aβ-related genes were determined by RT-PCR.Results Compared with model group,MHS markedly decreased the content of Aβ1 -42 ,inhibited the expression of APP,BACE1 and CatB,but promo-ted the expression of NEP and IDE.In addition,AHS significantly increased the expression of plasticity-relat-ed proteins including PSD-95,p-NMDAR1,p-CaMKII, p-PKACβ,PKCγ,p-CREB and BDNF.Conclusions MHS could remarkably ameliorate the learning and memory impairment induced by D-gal in mice,which may be due to its ability to inhibit the Aβgeneration and deposition and promote synaptic plasticity related protein expression.