生物活性转化生长因子-β1（TGFβ1）稳定转染与外源性TGFβ1处理对小鼠皮肤鳞状细胞癌CH72细胞中92-kDa IV型胶原酶表达的相反作用。

Opposite effect of stable transfection of bioactive transforming growth factor-beta 1 (TGF beta 1) versus exogenous TGF beta 1 treatment on expression of 92-kDa type IV collagenase in mouse skin squamous cell carcinoma CH72 cells.

作者信息

Rundhaug J E, Park J, Pavone A, Opdenakker G, Fischer S M

机构信息

University of Texas M. D. Anderson Cancer Center, Science Park-Research Division, Smithville, Texas 78957, USA.

出版信息

Mol Carcinog. 1997 Jun;19(2):122-36. doi: 10.1002/(sici)1098-2744(199707)19:2<122::aid-mc7>3.0.co;2-h.

DOI:10.1002/(sici)1098-2744(199707)19:2<122::aid-mc7>3.0.co;2-h

PMID:9210959

Abstract

We have previously shown that transforming growth factor-beta 1 (TGF beta 1) mRNA is consistently overexpressed in squamous cell carcinomas relative to normal mouse skin. Here we show that 92-kDa type IV collagenase (matrix metalloproteinase) (MMP-9) mRNA was likewise progressively overexpressed during mouse skin carcinogenesis. To determine if overexpression of MMP-9 and TGF beta 1 are linked, we stably transfected a bioactive TGF beta 1 into a mouse skin squamous cell carcinoma cell line (CH72), which resulted in about twofold to three-fold higher levels of secreted active TGF beta 1. Active TGF beta 1-transfected cells grew only slightly, but not significantly, more slowly in vitro and in vivo than vector-only transfectants. Two clones overexpressing active TGF beta 1 secreted much reduced levels of MMP-9 activity, as determined by zymogram analyses. However, treatment of these clones with 40 pM exogenous TGF beta 1 for 48 h enhanced secretion of MMP-9 activity. Constitutive mRNA expression of MMP-9 was reduced twofold to 70-fold in five untreated active TGF beta 1-transfected clones relative to the other transfectants. In contrast, treatment with 40 pM exogenous TGF beta 1 induced MMP-9 mRNA expression in a time-dependent fashion, from twofold to fourfold after 4 h to a maximum of 12- to 19-fold after 24-48 h. Induction of MMP-9 mRNA was dose dependent at TGF beta 1 concentrations of 4-400 pM. Thus, stable transfection of bioactive TGF beta 1 downregulated whereas exogenous TGF beta 1 treatment upregulated MMP-9 activity and expression. Treatment of transfectants with a neutralizing TGF beta 1 antibody slightly downregulated constitutive MMP-9 mRNA (20-30%) but completely blocked induction by exogenous TGF beta 1. Thus, the effect of TGF beta 1 transfection was not due to secreted TGF beta 1 but may have been a secondary effect.

摘要

我们之前已经表明，相对于正常小鼠皮肤，转化生长因子-β1（TGFβ1）mRNA在鳞状细胞癌中持续过度表达。在此我们表明，92-kDa IV型胶原酶（基质金属蛋白酶）（MMP-9）mRNA在小鼠皮肤致癌过程中同样逐渐过度表达。为了确定MMP-9和TGFβ1的过度表达是否相关联，我们将具有生物活性的TGFβ1稳定转染到小鼠皮肤鳞状细胞癌细胞系（CH72）中，这导致分泌的活性TGFβ1水平提高了约两倍至三倍。活性TGFβ1转染的细胞在体外和体内生长仅略微变慢，但并不显著，比仅转染载体的细胞慢。通过酶谱分析确定，两个过度表达活性TGFβ1的克隆分泌的MMP-9活性水平大幅降低。然而，用40 pM外源性TGFβ1处理这些克隆48小时可增强MMP-9活性的分泌。相对于其他转染细胞，在五个未处理的活性TGFβ1转染克隆中，MMP-9的组成型mRNA表达降低了两倍至70倍。相反，用40 pM外源性TGFβ1处理以时间依赖性方式诱导MMP-9 mRNA表达，4小时后增加两倍至四倍，24 - 48小时后最高增加12至19倍。在TGFβ1浓度为4 - 400 pM时，MMP-9 mRNA的诱导呈剂量依赖性。因此，生物活性TGFβ1的稳定转染下调了MMP-9的活性和表达，而外源性TGFβ1处理则上调了其活性和表达。用中和性TGFβ1抗体处理转染细胞略微下调了组成型MMP-9 mRNA（20 - 30%），但完全阻断了外源性TGFβ1的诱导作用。因此，TGFβ1转染的作用并非由于分泌的TGFβ1，而可能是一种继发性效应。