SERIAL CULTURING OF HUMAN BRONCHIAL EPITHELIAL-CELLS DERIVED FROM BIOPSIES
SERIAL CULTURING OF HUMAN BRONCHIAL EPITHELIAL-CELLS DERIVED FROM BIOPSIES
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DOI：
10.1007/bf02633985
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发表时间：
1993-05-01
影响因子：
2.1
通讯作者：
PONEC, M
中科院分区：
文献类型：
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作者：
DEJONG, PM;VANSTERKENBURG, MAJA;PONEC, M
在本研究中，我们描述了一系列培养人支气管上皮细胞来源于活检获得纤维支气管镜。使用外植体或上皮细胞悬液结合饲养层技术，从少量材料（2 mm活检钳）开始细胞培养。无论是否使用外植体或解离细胞，细胞增殖速率和传代次数（最多8次传代）均相似。为了调节分化的程度，支气管上皮细胞在浸没、低钙（0.06 mM）（增殖）、正常钙（1.6 mM）（分化增强）条件下或在空气-液体界面处培养。根据细胞形态、细胞角蛋白表达和纤毛活性评估支气管上皮细胞培养物的特征。在浸没条件下培养的细胞形成由最多三层杯形小立方细胞组成的多层，其外观类似于体内的基底细胞。在暴露于空气的培养物中，形成的多层由三至六层组成，表现出鳞状化生。在培养的支气管上皮细胞中的细胞角蛋白的档案是相似的，在淹没和空气暴露的文化和在体内发现的档案相媲美。除了细胞角蛋白，波形蛋白共表达的一部分传代细胞。在原代培养物中观察到纤毛活性，无论培养物是从外植体还是从解离的细胞建立的。这种活性在传代培养时丧失，并且通过延长培养期不能恢复。与浸没培养相反，尽管出现鳞状化生，但在气液界面培养时，细胞显示纤毛重现。人支气管上皮细胞培养物可以是控制支气管上皮细胞功能调节机制的代表性模型。
In the present study we describe the establishment of serial cultures of human bronchial epithelial cells derived from biopsies obtained by fiberoptic bronchoscopy. The cell cultures were initiated from small amounts of material (2 mm forceps biopsies) using either explants or epithelial cell suspensions in combination with a feeder-layer technique. The rate of cell proliferation and the number of passages (up to 8 passages) achieved were similar, irrespective of whether the explants or dissociated cells were used. To modulate the extent of differentiation, the bronchial epithelial cells were cultured either under submerged, low calcium (0.06 mM) (proliferating), normal calcium (1.6 mM) (differentiation enhancing) conditions, or at the air-liquid interface. Characterization of the bronchial epithelial cell cultures was assessed on the basis of cell morphology, cytokeratin expression, and ciliary activity. The cells cultured under submerged conditions formed a multilayer consisting of maximally three layers of polygonal-shaped, small cuboidal cells, an appearance resembling the basal cells in vivo. In the air-exposed cultures, the formed multilayer consisted of three to six layers exhibiting squamous metaplasia. The cytokeratin profile in cultured bronchial epithelial cells was similar in submerged and air-exposed cultures and comparable with the profile found in vivo. In addition to cytokeratins, vimentin was co-expressed in a fraction of the subcultured cells. The ciliary activity was observed in primary culture, irrespective of whether the culture had been established from explants or from dissociated cells. This activity was lost upon subculturing and it was not regained by prolongation of the culture period. In contrast to submerged cultures and despite the squamous metaplasia appearance, the cells showed a reappearance of cilia when cultured at the air-liquid interface. Human bronchial epithelial cell cultures can be a representative model for controlling the mechanisms of regulation of bronchial epithelial cell function.