Tissue inhibitor of metalloproteinase-4 instigates apoptosis in transformed cardiac fibroblasts

Abstract

Tumor cells become malignant, in part, because of their activation of matrix metalloproteinases (MMPs) and inactivation of tissue inhibitor of metalloproteinases (TIMPs). Myocardial tumors are rarely malignant. This raises the possibility that the MMPs and TIMPs are differentially regulated in the heart compared to other tissues. Therefore, we hypothesized that a tissue specific tumor suppressor exists in the heart. To test this hypothesis we prepared cardiac tissue extracts from normal (n = 4), ischemic cardiomypathic (ICM) [n = 5], and dilated cardiomyopathic (DCM) [n = 8] human heart end-stage explants. The level of cardiospecific TIMP-4 was determined by SDS-PAGE and Western-blot analysis. The results suggested reduced levels of TIMP-4 in ICM and DCM as compared to normal heart. TIMP-4 was purified by reverse phase HPLC and gelatin-sepharose affinity chromatography. Collagenase inhibitory activity of chromatographic peaks was determined using fluorescein-conjugated collagen as substrate and fluorescence spectroscopy. The activity of TIMP-4 (27 kDa) was characterized by reverse zymography. The role of TIMP-4 in cardiac fibroblast cell migration was examined using Boyden chamber analysis. The results suggested that TIMP-4 inhibited cardiac fibroblast cells migration and collagen gel invasion. To test whether TIMP-4 induces apoptosis, we cultured cardiac normal and polyomavirus transformed fibroblast cells in the presence and absence of TIMP-4. The number of cells were measured and DNA laddering was determined. The results suggested that TIMP-4 controlled normal cardiac fibroblast transformation and induced apoptosis in transformed cells. Cardiospecific TIMP-4 plays a significant role in regulating the normal cell phenotype. The reduced levels of TIMP-4 elicit cellular transformation and may lead to adverse extracellular matrix degradation (remodeling), cardiac hypertrophy and failure. This study suggests a possible protective role of TIMP-4 in other organs which are susceptible to malignancy.

Myocardial Matrix Remodeling and the Matrix Metalloproteinases: Influence on Cardiac Form and Function

“…It also appears that TIMP-4 can bind to pro-MMP-2, but whether this can yield an activational complex with MT1-MMP remains unknown (26). In addition, studies have demonstrated that TIMPs exhibit growth factor-like properties and participate in apoptosis (32,73,145,248,307,451). For example, TIMP-1 can suppress programmed cell death in a number of cell culture systems through the induction of antiapoptotic genes (32,144,145,307).…”

“…For example, TIMP-1 can suppress programmed cell death in a number of cell culture systems through the induction of antiapoptotic genes (32,144,145,307). TIMP-1 and TIMP-2 can induce cell growth in a number of cell types including fibroblasts, whereas TIMP-4 may actually suppress cell growth (24,248,451,517). These antiapoptotic/growth properties of TIMPs appear to be independent of any direct MMP inhibitory effects and therefore suggest the presence of a TIMP/receptor interaction (144,145,248).…”

Matrix Metalloproteinases: Drug Targets for Myocardial Infarction

“…MMP-2 secreted by myofbroblasts in vitro, and could be further induced by IL-1 , TNF , TGF , mechanical load and oxidative stress [33]. Increased TIMP-2 is associated with increased fibroblast collagen synthesis, while TIMP-4 inhibits cell growth and triggers apoptosis of differentiated myofibroblasts [39]. TIMP-3 facilitates programmed cell death of both normal and injured myofibroblasts to prevent excessive myocardial fibrosis [34].…”

Tissue Inhibitor of Metalloproteinase-3 Induces a Fas-associated Death Domain-dependent Type II Apoptotic Pathway

“…Mitsiades et al (53) have described the induction of Fas-mediated apoptosis in Ewing's sarcoma cell lines by synthetic metalloproteinase inhibitors. Additionally, TIMP-4 has recently been shown to promote apoptosis of transformed cardiac fibroblasts (54).…”