ABSTRACT
In women with preeclampsia (PE), endothelial cell (EC) dysfunction can lead to altered secretion of paracrine factors that induce peripheral vasoconstriction and proteinuria. This study examined the hypothesis that PE sera may directly or indirectly, through human umbilical vein ECs (HUVECs), stimulate phospholipase C-gamma1-1,4,5-trisphosphate (PLC-gamma1-IP3) signaling, thereby increasing protein kinase C-alpha (PKC-alpha) activity, collagen I expression and intracellular Ca2+ concentrations ([Ca2+]i) in human umbilical artery smooth muscle cells (HUASMCs). HUASMCs and HUVECs were cocultured with normal or PE sera before PLC-gamma1 silencing. Increased PLC-gamma1 and IP3 receptor (IP3R) phosphorylation was observed in cocultured HUASMCs stimulated with PE sera (P<0.05). In addition, PE serum significantly increased HUASMC viability and reduced their apoptosis (P<0.05); these effects were abrogated with PLC-gamma1 silencing. Compared with normal sera, PE sera increased [Ca2+]i in cocultured HUASMCs (P<0.05), which was inhibited by PLC-gamma1 and IP3R silencing. Finally, PE sera-induced PKC-alpha activity and collagen I expression was inhibited by PLC-gamma1 small interfering RNA (siRNA) (P<0.05). These results suggest that vasoactive substances in the PE serum may induce deposition in the extracellular matrix through the activation of PLC-gamma1, which may in turn result in thickening and hardening of the placental vascular wall, placental blood supply shortage, fetal hypoxia-ischemia and intrauterine growth retardation or intrauterine fetal death. PE sera increased [Ca2+]i and induced PKC-alpha activation and collagen I expression in cocultured HUASMCs via the PLC-gamma1 pathway.
Subject(s)
Adult , Female , Humans , Pregnancy , Young Adult , Apoptosis , Calcium/metabolism , Cell Line , Cell Survival , Cells, Cultured , Coculture Techniques , Collagen Type I/analysis , Human Umbilical Vein Endothelial Cells , Muscle, Smooth, Vascular/cytology , Phospholipase C gamma/genetics , Pre-Eclampsia/blood , Protein Kinase C-alpha/metabolism , RNA Interference , Signal TransductionABSTRACT
Objective To investigate the effects of preeclampsia serum on activity and invasive ability of cultured cytotrophoblasts of first trimester of pregnancy. Methods Cytotrophoblasts of normal 6 to 8-week pregnancy were cultured by tissue explants adherent method,and were incubated with serum of women with normal pregnancy(normal group)and preeclampsia(preeclampsia group),respectively for 24 h.The activity of cytotrophoblasts was examined by CCK-8,invasive ability was determined by Transwell invasion assay,and expression of MMP-2,MMP-9 and PAI-1 mRNA was detected by reverse transcription-polymerase chain reaction(RT-PCR). Results The activity and invasive ability of cytotrophoblasts in preeclampsia group were lower than those in normal group(P<0.05,P<0.01).Compared with normal group,the expression of MMP-2 and MMP-9 mRNA of cytotrophoblasts was significantly lower(P<0.01),and the expression of PAI-1 mRNA was significantly higher(P<0.01).In both groups,the expression of MMP-2 mRNA was negatively related to that of PAI-1 mRNA(r=-0.985,P<0.01;r=-0.933,P<0.05),while there was no correlationship between the expression of MMP-9 mRNA and that of PAI-1 mRNA. Conclusion The preeclampsia serum may affect the invasive ability of cytotrophoblasts by regulating the expression of MMP-2,MMP-9 and PAI-1.
ABSTRACT
Objective To obtain the optimal extraction process of Prunella vulgaris L.Methods With the dependent variables of ethanol concentration,reflux time and solvent fold,and dependent variable of the amount of baicalin,Central Composite Design/Response Surface Method was applied to optimize the reflux extraction process.Predictions were carried out by comparing the observed and predicted values.Results The optimum conditions of extraction process were 45% ethanol,2 hours for reflux,4 fold sovent.Regression coefficients of binomial fitting complex model was 0.9759.Bias between observed values and predicted values was-3.86%.Conclusion The optimum model is simple,precise and highly predictive.
ABSTRACT
Objective To establish a HPLC method for determining the content of paeoniflorin in Dahuang Zhechong Tablets. Methods Stationary phase was C18 column (4.6 mm?250 mm, 5 ?m), mobile phase was acetonitrice-0.1% phosphoric (14∶86), detection wavelength was 230 nm, flow rate was 1.0 mL/min and column temperature was 30 ℃. Results There was good linear relationship between the concentration of paeoniflorin and area integral value in the range of 0.10～0.90 ?g, r =0.999 7. The average recovery was 99.98% with RSD=0.81% (n =6). Conclusion This method is simple, accurate, with good reproducibility and high precision, and can be used to control the quality of Dahuang Zhechong Tablets effectively.
ABSTRACT
Objective To study the effects of ligustrazine(LTZ) on the preeclampyic umbilical serum induced-expression of precollagen I,III in cultured human umbilical artery smooth muscle cell(HUSMC). Methods The cultured HUSMC was treated with LTZ for 30 min, and then incubated with medium containing 20% serum either from women with preeclampsia or normal pregnant women for 48h. The cell activity was determined by MTT, the cell cycle was analyzed by flow cytomerty, and RT-PCR analysis was used to dectect the expression of precollagen I,III. Results The cell proliferation, percentage of S and G2/M phases, and expression of precollagen I obviously increased in HUSMC incubated with preeclampsia umbilical serum compared with normal pregnant women one(P
ABSTRACT
Objective To study the effect of protein kinase C-nuclear factor-kappa B (PKC-NF-?B) signal transduction pathway on the umbilical sera-induced human umbilical artery smooth muscle cell (HUASMC) proliferation and the expression of precollagen Ⅰ, and Ⅲ mRNA in preeclamptic women. Methods HUASMC from normal pregnancy were cultured and generated. After 30 minutes of incubation with or without Polymyxin B (PMB), the umbilical serum from the preeclamptic women was added. Two hours later, the expression of Ⅰ-?B and NF-?B were detected by Western blot. After 48 h, the activity of HUASMC was evaluated by MTT, the cell cycle by flow cytomerty and the expression of precollagenⅠ,ⅢⅢ and bcl-3 mRNA by reverse transcription-polymerase chain reaction(RT-PCR). Results Ⅰ-?B level (0.3765?0.0321) and the percentage of G_0+G_1 phase cells (62.53?2.85)% in preeclamptic group were significantly lower than those in control group [(0.7857? 0.0296),(78.66?3.25)%, P