Nguyen K D, Hoang A T, Lee D A
Jules Stein Eye Institute, Los Angeles, CA 90024-7004.
Invest Ophthalmol Vis Sci. 1994 Jun;35(7):3064-70.
Gamma-interferon (gamma-IFN) has been shown to be a potent inhibitor of collagenous protein production independent of its effects on noncollagenous protein production and cell proliferation in vitro. To understand further the processes controlling tissue fibrosis and the potential use of gamma-IFN as an antifibrotic treatment after glaucoma filtering surgery, the in vitro effects of recombinant gamma-IFN on procollagen mRNA production were studied.
Subconfluent human Tenon's capsule fibroblast cultures were exposed to 10, 50, 500, and 1000 U/ml of human recombinant gamma-IFN for 48 hours and to 500 U/ml for 12, 24, and 72 hours. After the incubation period, polyA+ mRNAs were isolated by oligo (dT) cellulose columns, separated according to size by electrophoresis through a denaturing agarose gel, and transferred to an activated nylon membrane for Northern blot analysis. The levels of type III (alpha 1) procollagen, type I (alpha 1) procollagen, and fibronectin (noncollagenous protein) mRNA were determined by hybridization with radiolabeled cDNA probes specific for these components followed by autoradiography.
Densitometric analysis showed gamma-IFN selectively inhibited type III and type I procollagen mRNA synthesis from 24% (10 U/ml) to 99% (1000 U/ml) while leaving fibronectin mRNA synthesis unaffected. The degree of inhibition was also time dependent; more inhibition occurred with increasing incubation time.
These results indicate that gamma-IFN is able to regulate collagen synthesis at the transcriptional level and that its inhibition is relatively specific. Gamma-interferon's specific inhibitory effects may offer advantages over current therapies in modulating the fibrotic response after glaucoma filtering surgery.
γ-干扰素（γ-IFN）已被证明是胶原性蛋白产生的有效抑制剂，在体外其作用独立于对非胶原性蛋白产生和细胞增殖的影响。为了进一步了解控制组织纤维化的过程以及γ-IFN作为青光眼滤过术后抗纤维化治疗的潜在用途，研究了重组γ-IFN对前胶原mRNA产生的体外作用。
将未长满的人Tenon囊成纤维细胞培养物分别用10、50、500和1000 U/ml的人重组γ-IFN处理48小时，以及用500 U/ml处理12、24和72小时。孵育期结束后，通过寡聚（dT）纤维素柱分离polyA + mRNA，通过变性琼脂糖凝胶电泳按大小分离，并转移到活化的尼龙膜上进行Northern印迹分析。通过与针对这些成分的放射性标记cDNA探针杂交，然后进行放射自显影，测定III型（α1）前胶原、I型（α1）前胶原和纤连蛋白（非胶原性蛋白）mRNA的水平。
光密度分析显示，γ-IFN选择性抑制III型和I型前胶原mRNA合成，抑制率从24%（10 U/ml）到99%（1000 U/ml），而纤连蛋白mRNA合成未受影响。抑制程度也呈时间依赖性；随着孵育时间延长，抑制作用增强。
这些结果表明，γ-IFN能够在转录水平调节胶原合成，且其抑制作用相对具有特异性。γ-干扰素的特异性抑制作用可能在调节青光眼滤过术后的纤维化反应方面比目前的治疗方法具有优势。
