No abstract

The epicardium makes essential cellular and paracrine contributions to the growth of the fetal myocardium and the formation of the coronary vasculature. However, whether the epicardium has similar roles postnatally in the normal and injured heart remains enigmatic. Here, we have investigated this question using genetic fate-mapping approaches in mice. In uninjured postnatal heart, epicardial cells were quiescent. Myocardial infarction increased epicardial cell proliferation and stimulated formation of epicardium-derived cells (EPDCs), which remained in a thickened layer on the surface of the heart. EPDCs did not adopt cardiomyocyte or coronary EC fates, but rather differentiated into mesenchymal cells expressing fibroblast and smooth muscle cell markers. In vitro and in vivo assays demonstrated that EPDCs secreted paracrine factors that strongly promoted angiogenesis. In a myocardial infarction model, EPDC-conditioned medium reduced infarct size and improved heart function. Our findings indicate that epicardium modulates the cardiac injury response by conditioning the subepicardial environment, potentially offering a new therapeutic strategy for cardiac protection.

Detection of matrix metalloproteinase (MMP) activities in the urine from patients with a variety of cancers has been closely correlated to disease status. Among these activities, the presence of a group of high molecular weight (HMW) MMPs independently serves as a multivariate predictor of the metastatic phenotype (1). The identity of these HMW MMP activities has remained unknown despite their novelty and their potentially important applications in non-invasive cancer diagnosis and/or prognosis. Here, we report the identification of one of these HMW urinary MMPs of ϳ125-kDa as being a complex of gelatinase B (MMP-9) and neutrophil gelatinase-associated lipocalin (NGAL). Multiple biochemical approaches verified this identity. Analysis using substrate gel electrophoresis demonstrated that the 125-kDa urinary MMP activity co-migrates with purified human neutrophil MMP-9⅐NGAL complex. The 125-kDa urinary MMP-9⅐NGAL complex was recognized by a purified antibody against human NGAL as well as by a monospecific antihuman MMP-9 antibody. Furthermore, these same two antibodies were independently capable of specifically immunoprecipitating the 125-kDa urinary MMP activity in a dose-dependent manner. In addition, the complex of MMP-9⅐NGAL could be reconstituted in vitro by mixing MMP-9 and NGAL in gelatinase buffers with pH values in the range of urine and in normal urine as well. Finally, the biochemical consequences of the NGAL and MMP-9 interaction were investigated both in vitro using recombinant human NGAL and MMP-9 and in cell culture by overexpressing NGAL in human breast carcinoma cells. Our data demonstrate that NGAL is capable of protecting MMP-9 from degradation in a dose-dependent manner and thereby preserving MMP-9 enzymatic activity. In summary, this study identifies the 125-kDa urinary gelatinase as being a complex of MMP-9 and NGAL and provides evidence that NGAL modulates MMP-9 activity by protecting it from degradation. Matrix metalloproteinases (MMPs)1 are a family of endopeptidases whose activities depend on metal ions, such as Zn 2ϩ and Ca 2ϩ . Collectively, MMPs are capable of degrading all of the molecular components of extracellular matrix, the barrier separating the tumor cells from their normal surrounding tissues, which is disassembled as part of the metastatic process (2). MMPs have been shown to play critical roles in a variety of biological as well as pathological processes, especially in tumor cell invasion and metastasis, and the overproduction of MMPs by tumor cells or surrounding stromal cells has been correlated with the metastatic phenotype (3).We have recently reported that intact and biologically active MMPs can be detected in the urine of cancer patients and are independent predictors of disease status (1). These urine samples were obtained from patients with a variety of cancers including prostate, renal, bladder, and breast carcinomas. The MMP activities detected in these urine samples included MMP-9 (gelatinase B, type IV collagenase, EC3.4.24.35) and MMP-2 (gelatinase A, type IV co...