Hasegawa M, Kobayashi M, Oyamada H, Kamishima T, Yoshida C, Ohata H, Maruyama I, Momose K, Gomi Y
University of Showa, School of Pharmaceutical Sciences, Tokyo, Japan.
Nihon Heikatsukin Gakkai Zasshi. 1987 Feb;23(1):35-46. doi: 10.1540/jsmr1965.23.35.
To prepare single smooth muscle cells from the taenia coli of guinea pig, the application of papain to the enzymatic solution was examined under two conditions: 1) the isolation in a modified Tyrode solution (containing 0.18 mM Ca2+: 0.18 mM Ca2+-Tyrode solution) and 2) the isolation in a high-K+ Tyrode solution (Na+ was replaced by K+, and Ca2+ was not added: high-K+ Tyrode solution). The presence of papain during collagenase digestion reduced contamination of broken cells and cell debris. In the case of the high-K+ Tyrode solution, papain increased the yield of single cells significantly. The cells were contracted in a dose-dependent manner by Ca2+ in the high-K+ Tyrode solution and by carbachol in 0.18 mM Ca2+-Tyrode solution; furthermore, the contractions were antagonized by verapamil and atropine, respectively. Treatment with papain did not affect cell sensitivity to the stimulants. Therefore, our results suggest that the addition of papain is useful for the isolation of single cells to investigate the physiological and pharmacological characteristics of smooth muscle.
为了从豚鼠结肠带制备单个平滑肌细胞，在两种条件下研究了在酶溶液中加入木瓜蛋白酶的情况：1）在改良的台氏液（含0.18 mM Ca2+：0.18 mM Ca2+ - 台氏液）中分离；2）在高钾台氏液（Na+被K+取代且不添加Ca2+：高钾台氏液）中分离。在胶原酶消化过程中加入木瓜蛋白酶可减少破碎细胞和细胞碎片的污染。在高钾台氏液的情况下，木瓜蛋白酶显著提高了单细胞的产量。在高钾台氏液中，细胞对Ca2+呈剂量依赖性收缩，在0.18 mM Ca2+ - 台氏液中对卡巴胆碱呈剂量依赖性收缩；此外，这些收缩分别被维拉帕米和阿托品拮抗。用木瓜蛋白酶处理不影响细胞对刺激物的敏感性。因此，我们的结果表明，添加木瓜蛋白酶有助于分离单个细胞以研究平滑肌的生理和药理特性。
