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AIM: To study the effect of calcium sensing receptor (CaSR) on icariin (ICA) induced mouse embryonic stem cells ( mESCs) to differentiate into cardiomyocytes in vitro.METHODS:mESCs were cultured to embry-oid bodies ( EBs) by direct suspension method and the differentiation of EBs into cardiomyocytes was induced by ICA.The expression of cardiac specific proteinsα-actinin and cardiac troponin-I ( cTnI) was analyzed by Western blot and immuno-fluorescence.The differentiation rate was determined by flow cytometry.The ultrastructure of the derived cardiomyocytes was further characterized by transmission electron microscopy.The expression of cardiac-specific transcription factors Nkx2.5 and GATA-4,as well as CaSR was detected by Western blot.RESULTS: After induction with ICA, the positive characteristics of myocardial cells appeared in the EBs cultured for 2 d.The expression of cardiac-specific sarcomeric pro-tein actinin (α-actinin) and cTnI showed an overall upward trend by Western blot in different phases of ICA induced differ-entiation.The expression of CaSR, Nkx2.5 and GATA-4 was the highest at an early stage of ICA-induced differentiation. Neomycin (an activator of CaSR) up-regulated CaSR, NKx2.5 and GATA-4 expression in the EBs at early stage of ICA-in-duced differentiation, all of which were reversed by NPS2390 ( an inhibitor of CaSR) .CONCLUSION:CaSR is function-ally expressed in mESC-derived cardiomyocytes, and activation of CaSR is involved in the differentiation of mESCs into car-diomyocytes by facilitating the expression of NKx2.5 and GATA-4.
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Objective To study the protective effects of choline on myocardial ischemia rat heart and its potential mecha -nisms .Methods Ischemia hypoxia environment was simulated with low value of pH (pH 6 .8) and lack of oxygen .Calcium currents were recorded by whole cell patch under the voltage clamp configuration .The alternations in[Ca2 + ]induced by KCl was detected by laser scanning confocal microscope in ventricular myocytes ,then disccuss the effects of choline on calcium and calcium store in cells . Results The normalized peak currents of ICa-L in ventricular myocytes were larger in pH 6 .8 group than those in pH 7 .4 group , which can be attenuated by choline .The(Ca2 + )i induced by KCl in ventricular myocytes were significantly increased in pH 6 .8 Ty-rode solution and its increasing can be suppressed by choline .4-DAMP can inhibit the suppressing effect of choline .Conclusion The possible mechanism of M 3 receptor involved in the protection of ischemic myocardium by inhibiting myocardial cells in ICa-L ,in-hibiting intracellular calcium overload .
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Objective To observe the vasodilation effect of extract of Jasminum samba (EJs), a kind of traditional Chinese medicine, on ex vivo rat thoracic aortic rings, and to investigate its mechanism. Methods On ex vivo aortic ring perfusion device, influence of EJs on contraction of the aorta induced by phenylephrine (PE) or potassium chloride (KCl) was observed. Influence of N-nitro-L-arginine-methylester ( L-NAME ), barium chloride ( BaCl2 ), glibenclamide ( Gli ) on vasodilating effect of EJs (0. 5, 1, 2, 4, 8 g·L-1 ) was detected. Effect of EJs on the contraction of calcium chloride (CaCl2 ) and PE in Ca2+-free medium was detected. [ Ca2+ ] i in vascular smooth muscle cells was determined by using laser scanning confocal microscope (LSCM). Results In blood vessels with intact endothelium, EJs concentration-dependently decreased PE- or KCl-induced vasoconstriction, the maximum dilating effect being (105. 0±3. 2)% and (78. 0±6. 5)% , respectively; L-NAME affected the vasodilatory effect of EJs on thoracic aorta rings ( P<0. 01), the maximum dilatory effect being (58. 0 ± 6. 9)% . BaCl2 and Gli had significant influence on vasodilation of EJs, and the contraction was obviously attenuated (P<0. 01), the maximum dilatory effect being (37. 0±5. 2)% and (78. 0±10. 0)% , respectively. EJs significantly inhibited contracting effect of PE on thoracic aorta rings in Ca2+-free medium (P<0. 01). The maximum contraction effect was (70. 0±6. 3)% . EJs inhibited CaCl2-induced vasoconstriction (0. 5-8 mmol·L-1 ), and vasoconstriction was decreased by (65. 0±3. 2)% . LSCM recorded that Fmax / F0 of 4 and 8 g·L-1 EJs was (2. 0±0. 2) and (1. 5±0. 2), respectively. Conclusion EJs exerted a dose-dependent vasodilating effect on rat isolated aorta rings. The mechanism might be related to promoting NO release, activating K+channels and decreasing the concentration of cytoplasmic Ca2+.
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Purpose To observe the functional expression of calcium sensing receptor ( CaSR) in human gastric cancer SGC-7901 cell line, the effect of CaSR on intracellular calcium, cell proliferation and migration of SCG-7901. Methods The expression and distribu-tion of CaSR were detected by Western blotting and immunofluorescence observation in SGC-7901. The intracellular concentration of free calcium ( [ Ca2+] i ) was determined by confocal laser scanning microscopy. MTT, flow cytometry and scratch test were used to an-alyze the impact of CaSR the proliferation and the migration capabilities of SGC-7901 cell. Results CaSR protein was expressed in SGC-7901. Extracellular calcium or calindol significantly increased the expression of [Ca2+]i, CaSR and E-cadherin;In addition, the migration capabilities were decreased. Conclusion CaSR is expressed in SGC-7901. The activation of CaSR induces the expression of E-cadherin, and decreases migration ability.
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AIM:To observe the functional expression of calcium-sensing receptor (CaSR) in the mouse em-bryonic stem cells (mESCs).METHODS:The expression and distribution of CaSR were detected by Western blotting and immunofluorescence observation in 129 mouse ES-D3 cells.The intracellular concentration of free calcium ([Ca2+]i) was determined by confocal laser scanning microscopy .The cell viability was analyzed by MTT assay and flow cytometry .RE-SULTS:CaSR protein was expressed in mESCs .Extracellular calcium or neomycin significantly increased the expression of CaSR and [Ca2+]i.Neomycin increased the cell viability , up-regulated the protein expression of p-ERK2.These effects of neomycin were inhibited by NPS2390.CONCLUSION:CaSR is expressed in mESCs .The activation of CaSR is involved in the proliferation of mESCs .