1.Screening and preliminary validation of new protein markers in maternal serum for early diagnosis of Down syndrome
Ying JIANG ; Bo ZHANC ; Wei LI ; Yaojin LI ; Mei DONG
Chinese Journal of Laboratory Medicine 2012;35(4):328-332
Objective To assess the clinical application values of the protcin markers associated with Down syndrome (DS) in maternal serum which were screened and identified.Methods Seven maternal serum samples with DS fetus ( DS group) and 7 maternal serum samples with normal fetus ( control group) in the second trimester were separated by two-dimensional gel electrophoresis (2-DE).The differentinal expression profile of proteome in maternal serum from DS group was established.The differentially expressed proteins were screened by mass spectrometry (MS) and some proteins were verified by Western blotting (WB).Results Twenty-nine proteins were discovered to be differentially expressed by more than 1.5 folds in maternal serum from DS group,among which 19 proteins were up-regulated and 10 proteins were downregulated.Eight proteins displayed 2 or more folds changes in maternal serum from DS group were identified by MS and possibly matched with 12 proteins in Ameracan National Center of Biotchnology Information (NCBI) protein sequence database,such as dGTPase and Beta2-Glycoprotein Ⅰ (β2-GPI),etc.The resuhs of WB showed that the mean a values of dGTPase and β2-GPI were 21 567.0 ± 3009.4 and 22 097.0 ±3958.9 in the DS group,3957.7 ± 250.9 and 1799.7 ± 105.5 in the control group respectively,which presented that the expression of dGTPase and β2-GPI significantly increased in DS group (t'dGTPase =- 17.66,t'β2-GPI =- 14.83,P <0.0001 ).Conclusions 2-DE and MS are effective methods for preliminary identification of protein markers associated with DS in maternal serum.dGTPase and β2-GPI verified by WB laid a solid fundation for further screening new biologic markers for early diaglosis of DS.
2.Effects of transplantation of adrenomedullin gene modified bone marrow mesenchymal stem cells on cardiac function in rats with heart failure
Mei-ling, ZHANC ; Li-li, LI ; He-fei, LI ; Hui-bo, CHEN ; Yu-mei, LIU ; Yao, ZHANG
Chinese Journal of Endemiology 2012;31(6):625-630
Objective To investigate the effects of transplantation of bone marrow mesenchymal stem cells (BMSCs) transfected with adrenomedullin (ADM) on cardiac function in heart failure rats and the mechanism.Methods BMSCs were isolated from femur and tibia marrow of 10 rats,20 days old,body weight 30-50 g,and in vitro cultured.The third passage of BMSCs were tuansfected with adenovirus containing ADM and labeled with green fluorescent protein(GFP).Before transplantation,BMSCs were labeled with 4',6-diamidino-2-phenylindole (DAPI).Eighty healthy male Wistar rats weighted 180-200 g were randomly divided into 2 groups according to body weight:control group (n =10) was injected with normal saline (NS); diffuse myocardial injury heart failure rat model(n =70) was established by subcutaneous injection of isoproterenol (ISO,170 mg/kg) every day for 4 consecutive days.Four weeks after administration of ISO,heart function was assessed by echocardiography,the 39 rats with left ventricle ejection fraction(LVEF) < 70% of global heart failure model were randomly divided into three groups in accordance with the level of heart function:untransfected group,transfected group and NS group.DAPI labeled untransfected BMSCs suspension,ADM gene transfected BMSC suspensions (3 × 106/150 μl) and equal volume of NS were injected into the left ventricular anterior wall in 4 places in each goup.Control group received thoracotomy only.Four weeks after transplantation,rats were examined by ultrasound echocardiography,then were sacrificed and left ventricular were dissected.The myocardium was stained with Massons trichrome to analyze myocardial tissue fibrosis.The transplanted cells were observed by fluorescence microscopy and matrix metalloproteinase-2 (MMP-2) expression of myocardial tissue was detected in each group by Western blotting.Results After in vitro culture for three days,the BMSCs began to grow adherently,tended to be fused about 10 days,in the fusiform shape.Four weeks after transplantation,ultrasound echocardiography results showed that rat cardiac left ventricular end systolic diameter (LVDs),LVEF,and left ventricular cardiac fractional shortening (LVFS) were different between groups,and the difference were statistically significant(F =5.838,32.983,51.714,P < 0.05 or P < 0.01).Compared with the control group[(86.50 ± 1.54)%,(50.66 ± 1.87)%],the LVEF and LVFS of NS groups[(56.67 ± 6.86)%,(26.27 ± 4.01)%],the transfected group[(79.40 ± 1.70)%,(43.48 ±2.15)%] and untransfected group[(69.24 ± 7.30)%,(34.59 ± 5.13)%] were significantly lower(all P < 0.05);compared with the NS group,the LVEF and LVFS of the transfected groups and the untransfected group were significantly increased(all P < 0.05) ; compared with the untransfected group,the LVEF and LVFS of the transfected group were increased (all P < 0.05).Compared with the control group [(3.16 ± 0.22)mm],the LVDs of the NS group[(5.35 ± 1.57)mm] was significantly increased (P < 0.01); compared with the NS group,the LVDs of the transfected group and the untransfected group[(3.95 ± 0.55),(4.24 ± 0.92)mm] were significantly decreased (all P < 0.05).There was no statistically significant difference between the control group,the transfected group and the untransfected group in LVDs (P > 0.05).It can clearly be seen that there was GFP and DAPI labeled transplanted cells under a fluorescence microscope in the myocardial tissue transplanted area.There was significant difference in myocardial fibrosis area and the myocardial tissue protein expression of MMP-2 between groups(F =533.75,32.777,all P < 0.01).The area ratio of the NS group[(15.200 ± 0.356)%,0.584 ± 0.013],the transfected group[(8.530 ± 0.573)%,0.386 ± 0.017] and the untransfected group [(10.670 ± 0.369)%,0.438 ± 0.015] and the MMP-2 protein expression were significantly higher than that of the control group[(1.070 ± 0.113)%,0.319 ±0.013,all P < 0.01)]; compared with the NS group,the two index of the transfected group and the untransfected group were decreased (all P < 0.05).Compared with the untransfected group,the two index of the transfected group was decreased (all P < 0.05).Conclusion Transplantation of ADM gene transfected BMSCs can improve heart function of rats with heart failure significantly and reduce myocardial fibrosis.