de Carvalho D F, Silva K L, de Oliveria D A, Villa-Verde D M, Coelho H S, Silva L C, Nasciutti L E
Departamento de Histologia e Embriologia, Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Cidade Universitária-Ilha do Fundão, Brazil.
Biol Cell. 2000 Aug;92(5):351-62. doi: 10.1016/s0248-4900(00)01076-5.
GH3B6 cells, a rat pituitary tumor cell line, synthesize and secrete large amounts of prolactin (PRL) in vitro. In the present work, we evaluated the capacity of these cells to express extracellular matrix (ECM) components and receptors in vitro. The expression of laminin (LN), fibronectin (FN) and type IV collagen (CIV) was investigated by immunofluorescence assays. In comparison to PRL distribution, where around 50-70% of the cells contained PRL concentrated in the Golgi region, a variable immunolabeling for the three ECM components could be observed in the majority of GH3B6 cells. Importantly, this pattern was not modified when cells were cultured in the presence of 30 nM thyroliberin (TRH). The expression of the ECM receptors: alpha5beta1 (FN receptor), alpha6beta1 (LN receptor) and CD44 (hyaluronic acid receptor) could be demonstrated by cytofluorometric analysis. Using biochemical procedures, we analyzed the synthesis and secretion of glycosaminoglycans (GAGs). The cells synthesized and secreted mainly heparan sulfate (75%) with a minor amount of chondroitin sulfate/dermatan sulfate. In an attempt to evaluate the individual contribution of the ECM components to influence cell morphology and PRL distribution in vitro, GH3B6 cells were cultivated separately on LN, FN and CIV substrates. Under all conditions, it was possible to observe an increase of cell adherence to the substrate, accompanied with changes of cellular morphology, characterized by the appearance of cytoplasmatic processes. However, no changes on PRL distribution could be observed. Our results suggest that endocrine tumor cell lines are involved in synthesis of ECM components and receptors.
GH3B6细胞是一种大鼠垂体瘤细胞系，在体外能合成并分泌大量催乳素（PRL）。在本研究中，我们评估了这些细胞在体外表达细胞外基质（ECM）成分及受体的能力。通过免疫荧光测定法研究了层粘连蛋白（LN）、纤连蛋白（FN）和IV型胶原（CIV）的表达。与PRL的分布情况相比，约50 - 70%的细胞中PRL集中在高尔基体区域，在大多数GH3B6细胞中可观察到三种ECM成分的免疫标记情况各不相同。重要的是，当细胞在30 nM促甲状腺素释放激素（TRH）存在的情况下培养时，这种模式并未改变。通过细胞荧光分析可证实ECM受体：α5β1（FN受体）、α6β1（LN受体）和CD44（透明质酸受体）的表达。我们采用生化方法分析了糖胺聚糖（GAGs）的合成与分泌。这些细胞主要合成并分泌硫酸乙酰肝素（75%），少量分泌硫酸软骨素/硫酸皮肤素。为了评估ECM成分对体外细胞形态和PRL分布的单独影响，将GH3B6细胞分别培养在LN、FN和CIV基质上。在所有条件下，都能观察到细胞对基质的黏附增加，同时伴随着细胞形态的改变，其特征为出现细胞质突起。然而，未观察到PRL分布有变化。我们的结果表明，内分泌肿瘤细胞系参与了ECM成分及受体的合成。
