| Background:Autologous transplantation of granulocyte colony-stimulating factor (G-CSF) mobilized peripheral blood mononuclear cells (M-PBMNCSs) has been shown to be effective in treating critical lower limb ischemia; however, the studies of the possible prognosis predictors after autologous M-PBMNCSs transplantation are inadequate.Objectives:To assess the possible factors affecting the results of M-PBMNCs therapy for lower limb ischemia.Method:We reviewed the clinical profiles of87patients with lower limb ischemia who were treated with M-PBMNCs therapy in our hospital between December2002and December2011, and we followed these patients. The variables studied included the following:the patient’s sex, patient’s age at first transplantation, Rutherford classification, vascular complications (including high blood pressure, ischemic cardiomyopathy, and cerebral vascular disease), history of smoking, leukocyte counts after mobilization, blood concentrations of hemoglobin(HGB), prothrombin time (PT), fibrinogen, liver enzymes (including alanine aminotransferase(ALT) and aspartate aminotransferase(AST)), creatinine(Cr), blood urea nitrogen(BUN), fasting blood glucose on first admission, and times of transplantation. The patients were divided into a good prognosis group and a poor prognosis group based on whether amputation happened. The significant differences of clinical variables between two groups were analyzed by Mann-Whitney test and Χ2test, and logistic regression analysis was used to study the variables representing the possible prognostic factors for amputation. Results and conclusion:Of the87patients,3patients died and1patient lost during the follow-up period. We analyzed83patients at last. The diseases included critical limb ischemia complicated by diabetes mellitus gangrene (35cases,42.2%), arteriosclerosis obliterans (31cases,37.3%), and thromboangiitis (17cases,20.5%). The mean age was62(30-87). The median follow-up time for the surviving patients was5years. The5-year amputation-free rate was72.2%and no adverse effects related to M-PBMNCS transplantation were observed. The significant prognostic factors possibly associated with poor angiogenesis were:fibrinogen>4g/L and fasting blood glucose>6mmol/L. Background:Vascular diseases are the most serious complications of diabetes mellitus (DM). Present studies showed that DM was associated with dysfunction of endothelial progenitor cells (EPCs), which might resulted in the increased risk of macro-and micro-vascular events in DM. EPCs were identified as bone marrow-derived endothelial precursor cells, which were essential in neovascularization. Oxidative stress played an important role in the mechanisms of EPCs dysfunction in DM. Heat shock factor-1(HSF-1), the main protector mechanism of cells, can protect cells from oxidative stress-induced toxic protein during regulating the gene expression of heat shock protein. However, studies about effects of high glucose on expression of HSF-1in EPCs under oxidative stress are limited.Objective:To explore the dysfunction of EPCs under high glucose and the role of high glucose in expression of HSF-1in EPCs under oxidative stress.Methods:Mononuclear cells were separated from human umbilical cord blood and the EPCs were cultured. The cultured cells were idetitified by flow cytometry. The cells were further characterized by Dil-acLDL uptaking by immunofluorescence; the EPCs were cultured in medium with glucose concentration of30mmol/L for5days. The proliferation of EPCs was determined by CCK-8, and the migration capacity was detected by transwell. After that, the PECs were cultured in the condition of oxidative stress caused by150μmol/L H2O2for6hours, and determined the changes of proliferation, migration and the expression of HSF-1by real time PCR before and after treatment. All results form EPCs from high glucose would be compared with EPCs cultured under medium without glucose (control group).Results:On day10of culture, flow cytometric analysis showed that the positive staining rate of cells for CD34, CD133and KDR were96.1%,97.6%and96.7%. Utilizing immunofluorescence, we observed that more than90%of cultured cells were positive for Dil-acLDL. Compared with that in the control group, the proliferation of EPCs cultured in high glucose medium was inhibited, with P value was0.025; the migration was also inhibited, but there was not statistic significance (P=0.421). However, after a short inducement by H2O2, the proliferation and migration was significantly decreased, the P values were0.016and0.024, while the control group didn’t show significant difference before and after treatment, the P values were separately0.353and0.273. Before the treatment of H2O2, the expression of HSF-1mRNA in EPCs of high glucose group was0.162, and the expression of EPCs in control group was0.377; there was no statistics difference between them; But after6h treatment of H2O2, expression of HSF-1mRNA in EPCs of both high glucose group and control group increased significantly, with P<0.05(P value of high glucose group:0.982; P value of high glucose group1.552). Furthermore, HSF-1mRNA expression was significantly higher than that of EPCs in high glucose group, with P=0.041.Conclusion:we concluded that high glucose could inhibited the proliferation of EPCs; and EPCs cultured in high glucose medium were more sensitive to oxidative stress. The expression of HSF-1in EPCs cultured in high glucose medium increased in oxidative stress, but the expression was significantly lower than that of control group. |
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