ABSTRACT
ObjectiveThe purpose of this study was to observe the correlation of mTOR and VEGF gene with nephropathy indicators in diabetic rats. Methods Forty-eight male Sprague-Dawley(SD)rats were divided into diabetes mellitus group(DM=28)and control group(NDM=20). Diabetic models were produced by injection of streptozotocin. In the courses of 12,16,20 and 24 weeks,the histology scores(HS)and glomerular basement membrane(GBM)thickness were collected. The protein and mRNA expressions of the gene of mTOR,VEGF and VEGFR2 were observed by immunohistochemistry and real-time quantitative polymerase chain reaction (RT-Q-PCR)by SYBR Green. And the standardized cycle of threshold(SCt)was regarded as the indicators of the mRNA expression. Results HS and GBM thickness were significantly higher in DM rats than those in NDM rats,especially in DM rats of the courses of 20 and 24 weeks(P < 0.01). IHC scores of VEGF and VEGFR2 were higher in total DM rats and were positively correlated with each other. There were positive correlations between HS with VEGF and VEGFR2 in total DM rats(P < 0.05). SCts of VEGF and VEGFR2 were significantly higher and were positively correlated with each other in total DM rats(P < 0.01). SCt of VEGF and GBM thickness showed positive correlation in total DM rats. SCt of VEGF was highest in the course of 12w DM rats. SCt of VEGFR2 gradually decreased following by the diabetic course,and was lowest in the course of 24w. There were no significantly differences in IHC scores and SCt of mTOR between DM and NDM rats. But the IHC scores of mTOR,VEGF and VEGFR2 were positively correlated with each other and with HS in total DM rats(P < 0.05). Conclusion HS and GBM thickness were higher in diabetic rats,especially in the course of 24w,which could reflect the injury of nephropathy. The protein and mRNA of VEGF and VEGFR2 were high expressed in kidney of DM rats and increased with the increasing of diabetic course. The mRNA expression of VEGF was positively correlated with GBM thickness of in diabetic nephropathy(DN). The protein expressions of mTOR,VEGF and VEGFR2 might have synergistic effects in DN of DM rats. But the results could not exclude the influences of different courses,sample size and experimental conditions.
ABSTRACT
Objective To purify BMSCs by a simple Colony-Forming system and identify BMSCs in vitro.Methods BMSCs were planted at low-density ( 10/cm2 ) in the early stage of isolation,additional cells were scraped until a colony formed,secondly seeded those Colony-Forming cells at low-density.This process was repeat again when passaged cells formed another colony.This so called Colony-Forming selection was repeated several times until highly purified cells were obtained.Biological characters of BMSCs were observed.The surface antigens of BMSCs were identified by flow cytometry.The multipotentiality of BMSCs was assayed for differentiation into either osteoblasts or adipocytes.Results Homogeneous cells were obtained after BMSCs were passaged twice to thrice by Colony-Forming system.These BMSCs highly expressed positive surface antigens of BMSCs ( CD29 in 98.8％,CD90 in 98.4％ ) meanwhile seldom expressed negative surface antigens of BMSCs (CD31 in 2.6％,CD45 in 3％ ).BMSCs efficiently differentiated into osteoblasts and adipocytes.Conclusions Colony-Forming system is a simple and effective way to get highly purified BMSCs in a short time.
ABSTRACT
Objective To investigate the expression of matrix metalloproteinase 2 (MMP2) and MMP9 in the breast cancer and its relationship with tumor invasive potentiality,node metastasis and prognostic significance.Methods 130 cases of breast cancer and 10 cases of breast benign disease were studied by immunohistochemical staining with MMP2,MMP9 monoclonal antibodies.Results The positive expression rate of MMP2 and MMP9 were 83.08％ (108/130) and 71.54％ (93/130) in the breast cancers,which was significantly higher than those in breast benign disease ( P ＜0.01 ).The expressions of MMP2 and MMP9 obviously increased with tumor sizes and clinical staging( P ＜0.05,P ＜ 0.01 ).Over expression of MMP2 and MMP9 in the breast cancers with axillary lymph node metastases were significantly higher than those without lymph node metastases ( P ＜ 0.01 ).Conclusions The expression of MMP2 and MMP9was closely correlated to lymph node metastasis,and may act as markers for lymph node metastasis and prognostic in breast cancer.