Kay E P, Lee H K, Park K S, Lee S C
Department of Ophthalmology, Yonsei University College of Medicine, Seoul, Korea.
Invest Ophthalmol Vis Sci. 1998 Mar;39(3):481-6.
To understand the mechanism of fibrosis after filtering surgery for glaucoma, the effect of transforming growth factor-beta (TGF-beta) was studied in subconjunctival fibroblasts (SCFs). TGF-beta, universal inhibitor of cell proliferation, stimulates the cell proliferation of fibroblasts. SCFs were evaluated for their production of TGF-beta and fibroblast growth factor 2 (FGF-2) to determine whether TGF-beta may be an indirect mitogen acting through the induction of an endogenous growth factor, or factors, that then acts as the direct mitogen in an autocrine manner.
Cell proliferation was determined either by counting cell numbers or by analyzing the incorporation of [3H]thymidine into DNA. The synthesis of TGF-beta and FGF-2 was analyzed by immunoprecipitation and immunoblotting.
TGF-beta 1, TGF-beta 2, and TGF-beta 3 stimulated the cell proliferation of SCFs in a dose-dependent manner. The media conditioned by SCFs, which were subsequently activated by acid, stimulated cell proliferation of corneal stromal fibroblasts. When the acid-activated media conditioned by SCFs were immunoprecipitated, respectively, either with anti-TGF-beta 1 and TGF-beta 2 antibodies or with anti-TGF-beta 3 antibody, TGF-beta s, with an apparent molecular size of 25 kDa, were detected, whereas SCFs produced an 80-kDa latent form of TGF-beta 1. Interestingly, SCFs produced and secreted an 18-kDa extracellular isoform of FGF-2, the synthesis of which is further stimulated by TGF-beta 1 and TGF-beta 3, respectively, whereas the neutralizing antibody to FGF-2 and the FGF-2-specific antisense oligonucleotide primers inhibited the stimulatory activities of TGF-beta 1 in SCFs.
These findings indicate that SCFs produce TGF-beta and FGF-2 and that FGF-2 seems to be the direct stimulator of TGF-beta-mediated cell proliferation in SCFs.
为了解青光眼滤过性手术后纤维化的机制，研究了转化生长因子-β（TGF-β）在结膜下成纤维细胞（SCFs）中的作用。TGF-β作为细胞增殖的通用抑制剂，却能刺激成纤维细胞的增殖。对SCFs产生TGF-β和成纤维细胞生长因子2（FGF-2）的情况进行评估，以确定TGF-β是否可能是通过诱导内源性生长因子或多种因子而间接发挥促有丝分裂作用，这些因子随后以自分泌方式作为直接促有丝分裂原发挥作用。
通过细胞计数或分析[3H]胸腺嘧啶核苷掺入DNA的情况来测定细胞增殖。通过免疫沉淀和免疫印迹分析TGF-β和FGF-2的合成。
TGF-β1、TGF-β2和TGF-β3以剂量依赖性方式刺激SCFs的细胞增殖。经酸激活的SCFs条件培养基能刺激角膜基质成纤维细胞的增殖。当分别用抗TGF-β1和TGF-β2抗体或抗TGF-β3抗体对经酸激活的SCFs条件培养基进行免疫沉淀时，检测到表观分子大小为25 kDa的TGF-βs，而SCFs产生一种80 kDa的潜伏形式的TGF-β1。有趣的是，SCFs产生并分泌一种18 kDa的细胞外FGF-2异构体，其合成分别受到TGF-β1和TGF-β3的进一步刺激，而FGF-2的中和抗体和FGF-2特异性反义寡核苷酸引物抑制了TGF-β1对SCFs的刺激活性。
这些发现表明SCFs产生TGF-β和FGF-2，并且FGF-2似乎是TGF-β介导的SCFs细胞增殖的直接刺激因子。
