Kaartinen L, Nettesheim P, Adler K B, Randell S H
Department of Pharmacology and Toxicology, College of Veterinary Medicine, Helsinki, Finland.
In Vitro Cell Dev Biol Anim. 1993 Jun;29A(6):481-92.
In vitro culture conditions enabling rat tracheal epithelial (RTE) cells to differentiate to mucociliary, mucous, or squamous phenotypes are described. Medium composition for rapid cell growth to confluence in membrane insert cultures was determined, and the effects of major modifiers of differentiation were tested. Retinoic acid (RA), collagen gel substratum, and an air-liquid interface at the level of the cell layer were required for expression of a mucociliary phenotype which most closely approximated the morphology of the tracheal epithelium in vivo. Large quantities of high molecular weight, hyaluronidase-resistant glycoconjugates, most likely mucin glycoproteins, were produced in the presence of RA when the cells were grown with or without a collagen gel and in submerged as well as in interface cultures. However, extensive ciliagenesis was dependent on the simultaneous presence of RA, collagen gel, and an air-liquid interface. When RA was omitted from the media, the cells became stratified squamous and developed a cornified apical layer in air-liquid interface cultures. This phenotype was accompanied by loss of transglutaminase (TGase) type II and keratin 18 and expression of the squamous markers TGase type I and keratin 13. The ability to modulate RTE cell phenotypes in culture will facilitate future studies investigating molecular regulation of tracheal cell proliferation, differentiation, and function.
本文描述了能够使大鼠气管上皮（RTE）细胞分化为黏液纤毛、黏液或鳞状表型的体外培养条件。确定了在膜插入培养中使细胞快速生长至汇合的培养基成分，并测试了主要分化调节剂的作用。要表达最接近体内气管上皮形态的黏液纤毛表型，需要视黄酸（RA）、胶原凝胶基质以及细胞层水平的气液界面。当细胞在有或没有胶原凝胶的情况下生长，以及在浸没培养和界面培养中时，在RA存在的情况下会产生大量高分子量、抗透明质酸酶的糖缀合物，很可能是黏蛋白糖蛋白。然而，广泛的纤毛形成依赖于RA、胶原凝胶和气液界面的同时存在。当培养基中不含RA时，细胞在气液界面培养中会变成分层的鳞状，并形成角质化的顶端层。这种表型伴随着转谷氨酰胺酶（TGase）II型和角蛋白18的丧失以及鳞状标志物TGase I型和角蛋白13的表达。在培养中调节RTE细胞表型的能力将有助于未来研究气管细胞增殖、分化和功能的分子调控。
