Eric G. Neilson

Epithelial-mesenchymal transition and its implications for fibrosis

Epithelial to mesenchymal transition (EMT) is a central mechanism for diversifying the cells found in complex tissues. This dynamic process helps organize the formation of the body plan, and while EMT is well studied in the context of embryonic development, it also plays a role in the genesis of fibroblasts during organ fibrosis in adult tissues. Emerging evidence from studies of renal fibrosis suggests that more than a third of all disease-related fibroblasts originate from tubular epithelia at the site of injury. This review highlights recent advances in the process of EMT signaling in health and disease and how it may be attenuated or reversed by selective cytokines and growth factors.

Stromal fibroblasts in cancer initiation and progression

It is widely accepted that the development of carcinoma — the most common form of human cancer — is due to the accumulation of somatic mutations in epithelial cells. The behaviour of carcinomas is also influenced by the tumour microenvironment, which includes extracellular matrix, blood vasculature, inflammatory cells and fibroblasts. Recent studies reveal that fibroblasts have a more profound influence on the development and progression of carcinomas than was previously appreciated. These new findings have important therapeutic implications.

Evidence that fibroblasts derive from epithelium during tissue fibrosis

Interstitial fibroblasts are principal effector cells of organ fibrosis in kidneys, lungs, and liver. While some view fibroblasts in adult tissues as nothing more than primitive mesenchymal cells surviving embryologic development, they differ from mesenchymal cells in their unique expression of fibroblast-specific protein-1 (FSP1). This difference raises questions about their origin. Using bone marrow chimeras and transgenic reporter mice, we show here that interstitial kidney fibroblasts derive from two sources. A small number of FSP1(+), CD34(-) fibroblasts migrate to normal interstitial spaces from bone marrow. More surprisingly, however, FSP1(+) fibroblasts also arise in large numbers by local epithelial-mesenchymal transition (EMT) during renal fibrogenesis. Both populations of fibroblasts express collagen type I and expand by cell division during tissue fibrosis. Our findings suggest that a substantial number of organ fibroblasts appear through a novel reversal in the direction of epithelial cell fate. As a general mechanism, this change in fate highlights the potential plasticity of differentiated cells in adult tissues under pathologic conditions.

Endothelial-to-mesenchymal transition contributes to cardiac fibrosis

Cardiac fibrosis, associated with a decreased extent of microvasculature and with disruption of normal myocardial structures, results from excessive deposition of extracellular matrix, which is mediated by the recruitment of fibroblasts. The source of these fibroblasts is unclear and specific anti-fibrotic therapies are not currently available. Here we show that cardiac fibrosis is associated with the emergence of fibroblasts originating from endothelial cells, suggesting an endothelial-mesenchymal transition (EndMT) similar to events that occur during formation of the atrioventricular cushion in the embryonic heart. Transforming growth factor-β1 (TGF-β1) induced endothelial cells to undergo EndMT, whereas bone morphogenic protein 7 (BMP-7) preserved the endothelial phenotype. The systemic administration of recombinant human BMP-7 (rhBMP-7) significantly inhibited EndMT and the progression of cardiac fibrosis in mouse models of pressure overload and chronic allograft rejection. Our findings show that EndMT contributes to the progression of cardiac fibrosis and that rhBMP-7 can be used to inhibit EndMT and to intervene in the progression of chronic heart disease associated with fibrosis.

Biomarkers for epithelial-mesenchymal transitions.

Somatic cells that change from one mature phenotype to another exhibit the property of plasticity. It is increasingly clear that epithelial and endothelial cells enjoy some of this plasticity, which is easily demonstrated by studying the process of epithelial-mesenchymal transition (EMT). Published reports from the literature typically rely on ad hoc criteria for determining EMT events; consequently, there is some uncertainty as to whether the same process occurs under different experimental conditions. As we discuss in this Personal Perspective, we believe that context and various changes in plasticity biomarkers can help identify at least three types of EMT and that using a collection of criteria for EMT increases the likelihood that everyone is studying the same phenomenon - namely, the transition of epithelial and endothelial cells to a motile phenotype.

Heart repair by reprogramming non-myocytes with cardiac transcription factors

The adult mammalian heart possesses little regenerative potential following injury. Fibrosis due to activation of cardiac fibroblasts impedes cardiac regeneration and contributes to loss of contractile function, pathological remodelling and susceptibility to arrhythmias. Cardiac fibroblasts account for a majority of cells in the heart and represent a potential cellular source for restoration of cardiac function following injury through phenotypic reprogramming to a myocardial cell fate. Here we show that four transcription factors, GATA4, HAND2, MEF2C and TBX5, can cooperatively reprogram adult mouse tail-tip and cardiac fibroblasts into beating cardiac-like myocytes in vitro. Forced expression of these factors in dividing non-cardiomyocytes in mice reprograms these cells into functional cardiac-like myocytes, improves cardiac function and reduces adverse ventricular remodelling following myocardial infarction. Our results suggest a strategy for cardiac repair through reprogramming fibroblasts resident in the heart with cardiogenic transcription factors or other molecules.

Mechanisms of Tubulointerstitial Fibrosis

Purpose of reviewTubulointerstitial fibrosis is the final common pathway to end-stage renal disease. Understanding the mechanisms of tubulointerstitial fibrosis is essential in establishing novel therapeutic strategies for the prevention or arrest of progressive kidney diseases. The present review focuses on a newly proposed mechanism of tubulointerstitial fibrosis, one that emphasizes the roles of epithelial-mesenchymal transition and cellular activation. Recent findingsAmong the cells that accumulate in the renal interstitium, fibroblasts are the principal effectors mediating tubulointerstitial fibrosis. By contrast, the phagocytosis of extracellular matrix and apoptotic cells by macrophages may actually exert a beneficial effect. Interstitial fibroblasts are more heterogeneous than expected, and during renal fibrosis new fibroblasts are derived mainly through epithelial-mesenchymal transition. The intracellular signaling pathways leading to initiation of epithelial-mesenchymal transition remain largely unknown, though recent studies have identified β-catenin and Smad3 activation of lymphoid enhancer factor, integrin-linked kinase, and small GTPases and mitogen-activated protein kinases as key components. Transforming growth factor-β is believed to be a critical fibrogenic factor, but recent studies have also focused on transforming growth factor-β independent pathways as mechanisms of tubulointerstitial fibrosis. As the mechanisms underlying tubulointerstitial fibrosis leading to epithelial-mesenchymal transition have been identified, so have cytokines that efficiently antagonize renal fibrosis, particularly bone morphogenic protein-7 and hepatocyte growth factor. SummaryIn combination with traditional angiotensin converting enzyme inhibitors, newly identified cytokines may eventually form the basis for new therapeutic strategies aimed at inhibiting the progression of renal disease.The pathologic paradigm for renal progression is advancing tubulointerstitial fibrosis. Whereas mechanisms underlying fibrogenesis have grown in scope and understanding in recent decades, effective human treatment to directly halt or even reverse fibrosis remains elusive. Here, we examine key features mediating the molecular and cellular basis of tubulointerstitial fibrosis and highlight new insights that may lead to novel therapies. How to prevent chronic kidney disease from progressing to renal failure awaits even deeper biochemical understanding.

Early role of Fsp1 in epithelial-mesenchymal transformation

A seamless plasticity exists among cells shifting between epithelial and mesenchymal phenotypes during early development and again later, in adult tissues, following wound repair or organ remodeling in response to injury. Fsp1, a gene encoding a fibroblast-specific protein associated with mesenchymal cell morphology and motility, is expressed during epithelial-mesenchymal transformations (EMT) in vivo. In the current study, we identified several cytokines that induce Fsp1 in cultured epithelial cells. A combination of these factors, however, was most efficacious at completing the process of EMT. The optimal combination identified were two of the cytokines classically associated with fibrosis, i.e., transforming growth factor-β1 (TGF-β1) and epidermal growth factor (EGF). To confirm that it was the induction of Fsp1 by these cytokines mediating EMT, we used antisense oligomers to block Fsp1 production and subsequently measured cell motility and markers of EMT phenotype. The antisense oligomers suppressed Fsp1 expression and epithelial transformation; therefore, we conclude that the appearance of Fsp1 is an important early event in the pathway toward EMT.

Angiotensin II as a renal growth factor.

Angiotensin II (ANG II) can have multiple effects on the kidney, including influences on the regulation of glomerular hemodynamics and tubular transport as well as consequences for the glomerular processing of macromolecules. The recognized suppressive effects of angiotensin-converting enzyme inhibitors on compensatory renal growth, even in the absence of hemodynamic effects, and the well-documented proliferative effect of ANG II on vascular smooth muscle cells have provided the background for the recent intensive interest in this peptide as a renal cytokine. Diverse cell types along the nephron express a variety of ANG II receptors. These receptors and their putative signal transduction pathways have been best characterized in mesangial and proximal tubular cells. Culture experiments provide convincing evidence that ANG II can be a phenotypic influence on these cell types. The growth responses and the associated signal transduction pathways, however, are different in mesangial and proximal tubular cells. These ANG II-mediated responses are also associated with an increase in the synthesis of distinct collagen subtypes, potentially linking the growth stimulatory effects of ANG II to the irreversible changes of glomerulosclerosis and tubulointerstitial fibrosis observed in chronic renal failure. Preventing the intrarenal actions of ANG II with angiotensin-converting enzyme inhibitors or with the new, orally active, selective ANG II receptor antagonists may provide a rational therapeutic approach to attenuate the progression of a variety of kidney diseases.

Loss of TGF- β type II receptor in fibroblasts promotes mammary carcinoma growth and invasion through upregulation of TGF- α -, MSP- and HGF-mediated signaling networks

Stromal fibroblasts regulate epithelial cell behavior through direct and indirect cell–cell interactions. To clarify the role of TGF-β signaling in stromal fibroblasts during mammary development and tumorigenesis, we conditionally knocked out the TGF-β type II receptor gene in mouse mammary fibroblasts (Tgfbr2fspKO). Tgfbr2fspKO mice exhibit defective mammary ductal development, characterized in part by increased ductal epithelial cell turnover associated with an increase in stromal fibroblast abundance. Tgfbr2fspKO mammary fibroblasts transplanted with mammary carcinoma cells promote growth and invasion, which is associated with increased activating phosphorylation of the receptors: erbB1, erbB2, RON, and c-Met. Furthermore, the increased receptor phosphorylation correlates with increased secretion of the cognate ligands by Tgfbr2fspKO fibroblasts. Treatment of tumor cells with fibroblast-conditioned medium leads to increased tumor cell proliferation and motility, which are blocked by addition of pharmacologic inhibitors of TGF-α signaling or neutralizing antibodies to macrophage-stimulating protein (MSP), HGF, or c-Met. These studies characterize a significant role for stromal TGF-β signaling in mammary tissue homeostasis and mammary tumor progression via regulation of TGF-α, MSP, and HGF signaling pathways.