Kay E P, Lee M S, Seong G J, Lee Y G
Department of Ophthalmology, Yonsei University College of Medicine, Seoul, Korea.
Curr Eye Res. 1998 Mar;17(3):286-93. doi: 10.1076/ceyr.17.3.286.5212.
Although transforming growth factor-betas (TGF-beta s) inhibit epithelial cell proliferation, these same substances stimulate cell proliferation of fibroblasts. In order to elucidate the mechanism of stimulatory activity of TGF-beta on fibroblast, the present study was performed to determine whether TGF-beta might be an indirect mitogen acting through induction of an endogenous growth factor(s) that then acts as the direct mitogen in an autocrine manner in corneal stromal fibroblasts (CSFs).
Cell proliferation was determined either by counting cell numbers or by analyzing the incorporation of [3H]-thymidine into DNA. The synthesis of TGF-beta, TGF-beta receptors, FGF-2 and p27 was analyzed by immunoprecipitation and immunoblotting.
TGF-beta 1, TGF-beta 2, and TGF-beta 3 significantly stimulated cell proliferation of CSFs in a dose-dependent manner. The medium conditioned by CSFs and subsequently activated by acid-inhibited cell proliferation of corneal endothelial cells by 40%. When the acid-activated media conditioned by CSFs were immunoprecipitated with either combined anti-TGF-beta 1 and TGF-beta 2 antibodies or anti-TGF-beta 3 antibody, all three TGF-beta s, with an apparent molecular size of 25 kDa, were detected, whereas CSFs produced an 80-kDa latent form of TGF-beta 1. These cells can also express TGF-beta type II receptor and betaglycan. Interestingly, CSFs produced and secreted 18-kDa FGF-2, the synthesis of which is further stimulated by either TGF-beta 1 or TGF-beta 3, while both the neutralizing antibody to FGF-2 and the FGF-2 specific antisense oligonucleotide primers significantly inhibited the stimulatory activities of TGF-beta 1 in CSFs. The expression of p27, a negative regulator in cell cycle, was not altered by TGF-beta.
These findings indicate that CSFs produce both TGF-beta s and FGF-2 and that FGF-2 appears to be a direct stimulator for TGF-beta-mediated cell proliferation in CSFs.
尽管转化生长因子-β（TGF-β）抑制上皮细胞增殖，但这些物质却能刺激成纤维细胞的增殖。为阐明TGF-β对成纤维细胞的刺激活性机制，本研究旨在确定TGF-β是否可能是一种间接促有丝分裂原，它通过诱导内源性生长因子起作用，然后该生长因子以自分泌方式作为角膜基质成纤维细胞（CSF）中的直接促有丝分裂原。
通过细胞计数或分析[3H] - 胸腺嘧啶核苷掺入DNA来测定细胞增殖。通过免疫沉淀和免疫印迹分析TGF-β、TGF-β受体、FGF-2和p27的合成。
TGF-β1、TGF-β2和TGF-β3以剂量依赖性方式显著刺激CSF的细胞增殖。CSF条件培养基经酸激活后，角膜内皮细胞的增殖被抑制了40%。当用抗TGF-β1和TGF-β2联合抗体或抗TGF-β3抗体对CSF酸激活的条件培养基进行免疫沉淀时，检测到所有三种TGF-β，其表观分子大小为25 kDa，而CSF产生80 kDa的潜伏形式的TGF-β1。这些细胞还可表达TGF-β II型受体和β聚糖。有趣的是，CSF产生并分泌18 kDa的FGF-2，TGF-β1或TGF-β3可进一步刺激其合成，而FGF-2的中和抗体和FGF-2特异性反义寡核苷酸引物均显著抑制TGF-β1对CSF的刺激活性。细胞周期负调节因子p27的表达未被TGF-β改变。
这些发现表明CSF可产生TGF-β和FGF-2，并且FGF-2似乎是CSF中TGF-β介导的细胞增殖的直接刺激因子。
