Oishi K, Itoh Y, Isshiki Y, Kai C, Takeda Y, Yamaura K, Takano-Ohmuro H, Uchida M K
Department of Pharmacology, Meiji Pharmaceutical University, Tokyo 204-8588, Japan.
Am J Physiol Cell Physiol. 2000 Nov;279(5):C1432-42. doi: 10.1152/ajpcell.2000.279.5.C1432.
String-shaped reconstituted smooth muscle (SM) fibers were prepared in rectangular wells by thermal gelation of a mixed solution of collagen and cultured SM cells derived from guinea pig stomach. The cells in the fiber exhibited an elongated spindle shape and were aligned along the long axis. The fiber contracted in response to KCl (140 mM), norepinephrine (NE; 10(-7) M), epinephrine (10(-7) M), phenylephrine (10(-6) M), serotonin (10(-6) M), and histamine (10(-5) M), but not acetylcholine (10(-5) M). Phentolamine (10(-7) M) produced a parallel rightward shift of the NE dose-response curve. Moreover, NE-induced contraction was partially inhibited by nifedipine and completely abolished by the intracellular Ca(2+) chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester, the myosin light chain kinase inhibitor ML-9, the Rho kinase inhibitor Y-27632, and papaverine. A [(3)H]quinuclidinyl benzilate binding study revealed that the loss of response to acetylcholine was due to the loss of muscarinic receptor expression during culture. The expression of contractile proteins in the fibers was similar to that in cultured SM cells. These results suggest that, although the fiber is not a model for fully differentiated SM, contractile mechanisms are maintained.
通过对胶原蛋白与源自豚鼠胃的培养平滑肌（SM）细胞的混合溶液进行热凝胶化处理，在矩形孔中制备出丝状重构平滑肌纤维。纤维中的细胞呈现出细长的纺锤形，并沿长轴排列。该纤维对氯化钾（140 mM）、去甲肾上腺素（NE；10⁻⁷ M）、肾上腺素（10⁻⁷ M）、去氧肾上腺素（10⁻⁶ M）、5-羟色胺（10⁻⁶ M）和组胺（10⁻⁵ M）产生收缩反应，但对乙酰胆碱（10⁻⁵ M）无反应。酚妥拉明（10⁻⁷ M）使NE剂量-反应曲线平行右移。此外，硝苯地平部分抑制NE诱导的收缩，而细胞内Ca²⁺螯合剂1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸乙酰甲酯、肌球蛋白轻链激酶抑制剂ML-9、Rho激酶抑制剂Y-27632和罂粟碱则完全消除该收缩。[³H]奎宁环基苯甲酸酯结合研究表明，对乙酰胆碱反应的丧失是由于培养过程中毒蕈碱受体表达的丧失。纤维中收缩蛋白的表达与培养的SM细胞中的相似。这些结果表明，尽管该纤维不是完全分化的SM模型，但收缩机制得以维持。
