Inhibition of resvertrol on osteoblast-like trans-differentiation of vascular smooth muscle cells / 中草药

Objective: To investigate the inhibition of resvertrol (Res) on angiosteosis, and to observe the effects of Res at different concentration on osteogenic conversion of human umbilical artery vascular smooth muscle cells (hVSMCs) induced by elevated calcium (Ca2+) and phosphate (P) culture. Methods: The hVSMCs were cultured in vitro, and identified by morphology and α-smooth muscle actin (α-SMA) antibody. High calcium (2.5 mmol/L) and high phosphate (3.0 mmol/L) culture was used to induce the osteogenic differentiation of hVSMCs. The calcification was confirmed by Alizarin red staining. Cells were divided into five groups such as normal control (NC, normal concentration of Ca2+ and P), positive control (PC, high Ca2+ and high P), and three Res (5, 10, and 20 μmol/L) groups. Calcium deposition in cells was measured by Arsenazo III after 12 d. Real-time PCR and Western blotting were used to detect mRNA and protein expression of BAP, BMP2, and OPN. Results: Compared to NC group, calcium deposition in the cells dramatically increased by five times in PC group, the protein expression of α-SMA decreased, and the mRNA and protein expression of BAP, BMP-2, and OPN obviously increased (P < 0.05). Res at different concentration could reduce the cellular Ca2+. The levels of BAP, BMP2, and OPN were highest in PC group than NC and three Res groups. Furthermore, the expression of BMP2 and OPN decreased with increasing Res concentration. Conclusion: Res prevents the process of the osteogenic differentiation of hVSMCs induced by high calcium and high phosphate. The degree of osteogenic differentiation is falling with increasing Res concentration.

Inhibition of rapamycin effect on proliferation of HUASMCs / 中国药学杂志

OBJECTIVE: To test effects of rapamycin on proliferation of cultured human umbilical arterial smooth muscle cells (HUASMCs) in vitro and the expression of interleukin-6 level.

Human umbilical artery smooth muscle exhibits a 2-APB-sensitive capacitative contractile response evoked by vasoactive substances and expresses mRNAs for STIM, Orai and TRPC channels

After depletion of intracellular Ca2+ stores the capacitative response triggers an extracellular Ca2+ influx through store-operated channels (SOCs) which refills these stores. Our objective was to explore if human umbilical artery smooth muscle presented this response and if it was involved in the mechanism of serotonin- and histamine-induced contractions. Intracellular Ca2+ depletion by a Ca2+-free extracellular solution followed by Ca2+ readdition produced a contraction in artery rings which was inhibited by the blocker of Orai and TRPC channels 2-aminoethoxydiphenyl borate (2-APB), suggesting a capacitative response. In presence of 2-APB the magnitude of a second paired contraction by serotonin or histamine was significantly less than a first one, likely because 2-APB inhibited store refilling by capacitative Ca2+ entry. 2-APB inhibition of sarcoplasmic reticulum Ca2+ release was excluded because this blocker did not affect serotonin force development in a Ca2+-free solution. The PCR technique showed the presence of mRNAs for STIM proteins (1 and 2), for Orai proteins (1, 2 and 3) and for TRPC channels (subtypes 1, 3, 4 and 6) in the smooth muscle of the human umbilical artery. Hence, this artery presents a capacitative contractile response triggered by stimulation with physiological vasoconstrictors and expresses mRNAs for proteins and channels previously identified as SOCs.

The establishment and identification of human umbilical artery endothelial cells-smooth muscle cells co-culture model / 复旦学报（医学版）

Objective To establish a human umbilical artery EC-SMC co-culture model, and mimic the morphological and functional characteristics of human arterial wall, for further reseach of the pathological mechanism and therapy of atherosclerosis and imflammatory damage. Methods We secceeded in the primary culture of human umbilical artery endothelial cells (HUAEC) and human umbilical artery smooth muscle cells (HUASMC) by collagenase perfusion digestion and tissue planting, respectively. HUASMCs were incubated in a medium with ascorbic acid at the concentration greater than 50 μg/mL to produce collagen, which was considered as the extracellular matrix for ECs. Then HUAECs were seeded directly upon HUASMCs in a saturate density for sufficient direct physical interaction between ECs and SMCs. The morphological characteristic of EC-SMC co-culture was identified by immunofluorescence staining, and the function of EC-SMC co-culture was identified by Dil-Ac-LDL uptake test. Results The morphological identification showed that the entire surface of HUASMCs was covered by a confluent monolayer confluent monolayer, which indicated that the model had simulated the morphological characteristic of human arterial wall. The results of Dil-Ac-LDL uptake test showed that there was a fluorescent signal in HUAECs. Compared with EC monoculture, the Dil-Ac-LDL uptake of HUAECs was increased significantly in the co-culture system. All the reseach results indicated that there was an interaction between HUAECs and HUASMCs in the co-culture system. Conclusions In the present study, human umbilical artery EC-SMC co-culture model was constructed successfully, which could mimic the morphological characteristic and basic functions of human arterial wall.

Pravastatin inhibits ossific calcification of human umbilical artery vascular smooth muscle cells induced by tumor necrosis factor α / 中华肾脏病杂志

ObjectiveTo investigate the effects of pravastation intervention on tumor necrosis factor (TNF)-α-indueed ossifie calcification in human umbilical artery smooth muscle cells (hUASMCs). MethodshUASMCs were cultured by tissue explant in vitro, hUASMC were treated with TNF-α 50 μg/L and pravastatin of three different concentrations. The calcium deposition was determined by O-cresolphthalein eomplexone method. The mRNA expression of BAP and OPN was determined by real time-PCR. The protein expression of BAP, OPN and BMP-2 was determined by Western blotting. ResultsPravastatin inhibited the proliferation of hUASMC (r=-0.946, P＜0.01) and decreased the cell calcium deposition (r=-0.973, P＜0.01) in a dosedependent manner. Pravastatin down-regulated the expression of BAP, OPN and BMP-2 induced by TNF-α in a dose-dependent manner (mRNA, r=-0.972, P＜0.01;BAP protein, r=-0.820, P＜0.01;OPN protein, r=-0.972, P＜0.01;BMP-2 protein, r=-0.928, P＜0.01). ConclusionPravastatin can inhibit the proliferation of hUASMC, decrease the cell calcium deposition and inhibit the ossifie calcification of hUASMC induced by TNF-α.

Effects of Bupivacaine and Magnesium on Serotonin-induced Vasocontraction in an Isolated Human Umbilical Artery / 대한마취과학회지

BACKGROUND: Serotonin is found in the blood of the umbilical cord at birth in concentrations sufficiently high to affect vascular tone. Serotonin has been suggested to be involved in the pathogenesis of preeclampsia. Magnesium sulfate (MgSO4) is used to treat convulsions and hypertension in patients with preeclamptic toxemia. Bupivacaine is used in the epidural anesthesia for a cesarean section. The effects of magnesium and bupivacaine on serotonin-induced vasocontraction in a human umbilical artery was investigated. METHODS: Experiments were performed on 52 human umbilical arteries. The rings were suspended in an organ bath to record isometric mechanical activity. The concentration-contraction responses to bupivacaine, magnesium and serotonin were measured respectively. Vessels were pretreated with bupivacaine (10(-5) M) or magnesium (2 mM or 6 mM), and then serotonin (10(-9) M - 10(-6) M) was added cumulatively. Data analysis was assessed by an unpaired t test, one-way ANOVA and a Kruskal-Wallis test. RESULTS: Bupivacaine induced a contraction of umbilical arterial rings, and showed a maximal contraction (51.8 +/- 6.1%) at a concentration of 43nM. Magnesium induced relaxation of the umbilical artery in a concentration dependent manner. Pretreatment with bupivacaine (10(-5) M) potentiated significantly the concentration response to serotonin (P < 0.05). Pretreatment with MgSO4 (2 mM or 6 mM) significantly suppressed the contractile response to serotonin (P < 0.05). CONCLUSIONS: Bupivacaine, magnesium and serotonin are vasoactive on human umbilical arteries. Magnesium exerts a strong relaxant effect on serotonin induced vasocontraction in the human umbilical artery. Potentiation of serotonin induced vasoconstriction by bupivacaine may play a significant role in the reduction of umbilicoplacental blood flow.

Inhibitory Effects of Ligustrazine on the Expression of Precollagen I,III Induced by Umbilical Serum of Preeclampsia in Cultured Human Umbilical Artery Smooth Muscle Cell / 中国医师杂志

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