ABSTRACT
Objective:To investigate the effect of Danggui Buxuetang（DGBX）on the functional activity of rat endothelial progenitor cells（EPCs）exposed to different luminar shear stress （SS）. Method:EPCs isolated from rat bone marrow were incubated on a parallel plate flow chamber at a steady SS of 0， 0.12， 1.2， 2.4 Pa for 6 h，then the cells exposed to different SS were randomly divided into 8 groups： control group （perfused with serum free medium），simvastatin group（0.1 μmol·L<sup>-1 </sup>simvastatin），3 DGBX groups（low，medium，high-dose DGBX）and 3 inhibitor groups（3 DGBX groups with LY294002）. After 12 h，the samples were collected for the detection of cell proliferation ，migration，tubule formation ，the secretion of nitric oxide （NO） ，and the expressions of endothelial nitric oxide synthase（eNOS） mRNA and protein kinase B（Akt），respectively. Result:Compared with the control group，simvastatin and DGBX（high-dose）could both promote the functional activities and NO secretion，and up-regulate the expressions of eNOS mRNA and Akt protein in EPCs exposed to different SS（<italic>P</italic><0.05），while DGBX（mid-dose）could do these only at 0 Pa. However，LY294002 could inhibit all effects of DGBX on EPCs. Conclusion:SS seems to play an important role in the effect of DGBX on EPCs，and DGBX could promote the functional activity of EPCs exposed to SS by up-regulating the expressions of NO/eNOS/Akt.
ABSTRACT
Objective:To observe protective effect of Danggui Buxuetang (DBT) on oxidative stress injury of mouse bone marrow-derived endothelial progenitor cells (EPCs) against induced by hydrogen peroxide (H2O2). Method:Monocytes from bone marrow of mice were obtained by density gradient centrifugation, and EPCs were obtained by specific culture medium. The experiment was divided into blank group，model group，DBT group (100，200，400 mg·L-1). Methyl thiazolyl tetrazolium（MTT） assay was used to determine the survival rate of EPCs and establish the cell injury model induced by H2O2. MTT，transwell chamber，matrigel and superoxide fluorescent anion probe (DHE) were used to detect the proliferation，migration，in vitro angiogenesis and ROS level，detection of autophagy by Western blot. Result:Compared with blank group，the proliferation ability，migration ability，the number of lumens and the length of tubule branches of EPCs in the model group were significantly reduced (P<0.01)，the level of intracellular reactive oxygen species （ROS） was significantly increased (P<0.01)，the expression of p62，the light chain microtubule associated protein 1 protein light chain 3 Ⅱ type （LC3-Ⅱ） protein of microtubule associated protein 1，was significantly increased (P<0.01). Compared with model group，DBT group increased the ability of cell proliferation and migration （P<0.01）. In addition，DBT increased the expression of LC3-Ⅱ protein in a concentration dependent manner (P<0.01)，and decreased the expression of p62 protein (P<0.01). Conclusion:DBT can improve the autophagy level of EPCs under oxidative stress, promote the proliferation, migration and angiogenesis of injured EPCs, and protect the biological function of EPCs under oxidative stress.
ABSTRACT
Objective: To investigate the protective effect of Danggui Buxue Tang(DGBX)with Angelicae Sinensis Radix(AS) and Astragali Radix(AR)at different radios on impaired functional activity of endothelial cells(ECs)exposed to low-fluid shear stress (FSS). Method: Low FSS was loaded by a parallel plate flow chamber,and ECs were divided into normal FSS group,low FSS group(each preincubated with M199 medium for 2 h),simvastatin group(preincubated with 0.1 μmol·L-1 simvastatin for 2 h),and 3 DGBX groups(preincubated with 3 g·L-1 AS and AR at 1:1,1:3,1:5 for 2 h,respectively). Then,the normal group was exposed to 1.2 Pa FSS,while the rest groups were all exposed to low FSS. At time points of 30,60, 360 min,the proliferation was detected by methyl thiazoly tetrazolium(MTT),the secretion of nitric oxide(NO) was detected by nitrase reduction test,and the mRNA and protein expressions of endothelial nitric oxide synthase(eNOS) were detected by Real-time fluorescence guantitative polymerase chain reaction(Real-time PCR) and Western blot,respectively. Result: Compared with the normal group, the secretion of NO and the expression of eNOS in ECs were both increased significantly at 60 min (PConclusion: DGBX could protect the functional activity of ECs exposed to low FSS.
ABSTRACT
<p><b>OBJECTIVE</b>To explore VEGF siRNA's effect on the immature fetal retinal microvascular endothelial cells in vitro.</p><p><b>METHOD</b>The fresh retinal micrangium was primarily cultured to obtain microvascular endothelial cells. CoCl2 was used to simulate oxygen-deficient conditions. siRNA directed against human VEGF was designed and chemically synthesized. There were 3 groups in our experiment: VEGF siRNA group, hypoxia control group, and negative siRNA control group. The fetal retinal micrangium vascular endothelial cells were transfected by using liposome. The expression levels of VEGF mRNA and protein were evaluated by RT-PCR and Western blotting 24, 48, 72 h after transfection, cell proliferation was evaluated by MTT method.</p><p><b>RESULT</b>The expression levels of VEGF mRNA decreased by 21.05%, 79.67%, and 90.48% 24 h, 48 h, and 72 h after transfection as compared to those in hypoxia control group, the expression level of VEGF protein had decreased by 14.58%, 66.97%, and 81.61% as compared to those in hypoxia control group. The siRNA could decrease cell proliferation under hypoxia too, the multiplication rate after 12, 24, 48, and 72 h decreased by 15.0%, 42.9%, 78.3% and 65.9%.</p><p><b>CONCLUSION</b>VEGF siRNA could down-regulate the expression of VEGF in immature fetal retinal microvascular endothelial cells and suppressed cell proliferation. Application of siRNA to inhibit expression of VEGF may be a hopeful way to prevent and cure ROP.</p>