Yoshino K, Tseng S C, Pflugfelder S C
Department of Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Florida 33136, USA.
Exp Cell Res. 1995 Sep;220(1):138-51. doi: 10.1006/excr.1995.1300.
Human lacrimal gland biopsies were serially incubated in chelating and enzymatic solutions. Viable single cells were evaluated for outgrowth in growth factor enriched medium using the following culture substrates: Matrigel, Type I collagen gel with or without fibroblasts, and plastic. Each epithelial outgrowth was characterized morphologically and immunohistochemically, and their growth and viability were examined by BrdU labeling and a quantitative cell viability assay. Synthesized proteins were evaluated by ELISA, SDS-PAGE, and 14C-labeled amino acid incorporation. Lacrimal gland epithelial cells plated on Matrigel formed clusters with central cavities that contained lactoferrin, mimicking acinar complexes in vivo. Cells plated on collagen gel or collagen gel containing fibroblasts formed islands or a monolayer, and lactoferrin was detected in incomplete cavities of epithelia on the latter substrate. Epithelial cells plated on plastic formed a monolayer and cellular expression of lactoferrin was weak and sporadic. Cellular release of the lactoferrin measured by ELISA supported the results of immunohistochemistry. Cells grown on plastic had the highest proliferative rate, whereas those grown on Matrigel showed the lowest proliferative rate. These results indicate that different substrates modulate lacrimal gland epithelial cell morphology, proliferative rate, and production of the tear protein lactoferrin. Matrigel promotes acinar differentiation to a greater extent than collagen gel and plastic. Incorporation of fibroblasts in collagen gel substrate promotes significant effects on growth and differentiation.
人泪腺活检组织在螯合剂和酶溶液中进行连续孵育。使用以下培养底物评估在富含生长因子的培养基中生长的活单细胞：基质胶、含或不含成纤维细胞的I型胶原凝胶以及塑料。对每个上皮细胞生长物进行形态学和免疫组织化学表征，并通过BrdU标记和定量细胞活力测定来检查它们的生长和活力。通过ELISA、SDS-PAGE和14C标记氨基酸掺入来评估合成蛋白。接种在基质胶上的泪腺上皮细胞形成带有中央腔的簇，中央腔内含有乳铁蛋白，类似于体内的腺泡复合体。接种在胶原凝胶或含成纤维细胞的胶原凝胶上的细胞形成岛状或单层，并且在后者底物上上皮细胞的不完全腔中检测到乳铁蛋白。接种在塑料上的上皮细胞形成单层，乳铁蛋白的细胞表达较弱且呈散发性。通过ELISA测量的乳铁蛋白细胞释放支持免疫组织化学结果。在塑料上生长的细胞增殖率最高，而在基质胶上生长的细胞增殖率最低。这些结果表明，不同的底物可调节泪腺上皮细胞的形态、增殖率以及泪液蛋白乳铁蛋白的产生。与胶原凝胶和塑料相比，基质胶在更大程度上促进腺泡分化。在胶原凝胶底物中加入成纤维细胞对生长和分化有显著影响。
