Michael Zeisberg

Fibroblasts in cancer

Tumours are known as wounds that do not heal — this implies that cells that are involved in angiogenesis and the response to injury, such as endothelial cells and fibroblasts, have a prominent role in the progression, growth and spread of cancers. Fibroblasts are associated with cancer cells at all stages of cancer progression, and their structural and functional contributions to this process are beginning to emerge. Their production of growth factors, chemokines and extracellular matrix facilitates the angiogenic recruitment of endothelial cells and pericytes. Fibroblasts are therefore a key determinant in the malignant progression of cancer and represent an important target for cancer therapies.

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.

BMP-7 counteracts TGF-β1–induced epithelial-to-mesenchymal transition and reverses chronic renal injury

Bone morphogenic protein (BMP)-7 is a 35-kDa homodimeric protein and a member of the transforming growth factor (TGF)-β superfamily. BMP-7 expression is highest in the kidney, and its genetic deletion in mice leads to severe impairment of eye, skeletal and kidney development. Here we report that BMP-7 reverses TGF-β1–induced epithelial-to-mesenchymal transition (EMT) by reinduction of E-cadherin, a key epithelial cell adhesion molecule. Additionally, we provide molecular evidence for Smad-dependent reversal of TGF-β1–induced EMT by BMP-7 in renal tubular epithelial cells and mammary ductal epithelial cells. In the kidney, EMT-induced accumulation of myofibroblasts and subsequent tubular atrophy are considered key determinants of renal fibrosis during chronic renal injury. We therefore tested the potential of BMP-7 to reverse TGF-β1–induced de novo EMT in a mouse model of chronic renal injury. Our results show that systemic administration of recombinant human BMP-7 leads to repair of severely damaged renal tubular epithelial cells, in association with reversal of chronic renal injury. Collectively, these results provide evidence of cross talk between BMP-7 and TGF-β1 in the regulation of EMT in health and disease.

Fibroblasts Derive from Hepatocytes in Liver Fibrosis via Epithelial to Mesenchymal Transition

Activated fibroblasts are key contributors to the fibrotic extracellular matrix accumulation during liver fibrosis. The origin of such fibroblasts is still debated, although several studies point to stellate cells as the principal source. The role of adult hepatocytes as contributors to the accumulation of fibroblasts in the fibrotic liver is yet undetermined. Here, we provide evidence that the pro-fibrotic growth factor, TGF-β1, induces adult mouse hepatocytes to undergo phenotypic and functional changes typical of epithelial to mesenchymal transition (EMT). We perform lineage-tracing experiments using AlbCre. R26RstoplacZ double transgenic mice to demonstrate that hepatocytes which undergo EMT contribute substantially to the population of FSP1-positive fibroblasts in CCL4-induced liver fibrosis. Furthermore, we demonstrate that bone morphogenic protein-7 (BMP7), a member of the TGFβ superfamily, which is known to antagonize TGFβ signaling, significantly inhibits progression of liver fibrosis in these mice. BMP7 treatment abolishes EMT-derived fibroblasts, suggesting that the therapeutic effect of BMP7 was at least partially due to the inhibition of EMT. These results provide direct evidence for the functional involvement of adult hepatocytes in the accumulation of activated fibroblasts in the fibrotic liver. Furthermore, our findings suggest that EMT is a promising therapeutic target for the attenuation of liver fibrosis.

Fibroblasts in Kidney Fibrosis Emerge via Endothelial-to-Mesenchymal Transition

Fibroblasts are key mediators of fibrosis in the kidney and other organs, but their origin during fibrosis is still not completely clear. Activated fibroblasts likely arise from resident quiescent fibroblasts via epithelial-to-mesenchymal transition and from the bone marrow. Here, we demonstrate that endothelial cells also contribute to the emergence of fibroblasts during kidney fibrosis via the process of endothelial-to-mesenchymal transition (EndMT). We examined the contribution of EndMT to renal fibrosis in three mouse models of chronic kidney disease: (1) Unilateral ureteral obstructive nephropathy, (2) streptozotocin-induced diabetic nephropathy, and (3) a model of Alport renal disease. Approximately 30 to 50% of fibroblasts coexpressed the endothelial marker CD31 and markers of fibroblasts and myofibroblasts such as fibroblast specific protein-1 and alpha-smooth muscle actin. Endothelial lineage tracing using Tie2-Cre;R26R-stop-EYFP transgenic mice further confirmed the presence of EndMT-derived fibroblasts. Collectively, our results demonstrate that EndMT contributes to the accumulation of activated fibroblasts and myofibroblasts in kidney fibrosis and suggest that targeting EndMT might have therapeutic potential.

Discovery of Endothelial to Mesenchymal Transition as a Source for Carcinoma-Associated Fibroblasts

Activated fibroblasts are associated with many different tumors. Myofibroblasts, activated fibroblasts, and perivascular mesenchymal cells such as pericytes play a role in cancer progression. Many studies suggest that myofibroblasts facilitate tumor growth and cancer progression. The source for myofibroblasts and other activated fibroblasts within the tumors is still debated. Although de novo activation of quiescent fibroblasts into alpha-smooth muscle actin (alpha SMA)-positive myofibroblasts is one likely source, epithelial to mesenchymal transition and bone marrow recruitment are also evolving as possible mechanisms for the emergence of a heterogeneous population of carcinoma-associated fibroblasts. Here, we show that transforming growth factor-beta1 could induce proliferating endothelial cells to undergo a phenotypic conversion into fibroblast-like cells. Such endothelial to mesenchymal transition (EndMT) is associated with the emergence of mesenchymal marker fibroblast-specific protein-1 (FSP1) and down-regulation of CD31/PECAM. Additionally, we show EndMT in tumors using the B16F10 melanoma model and the Rip-Tag2 spontaneous pancreatic carcinoma model. Crossing Tie2-Cre mice with R26Rosa-lox-Stop-lox-LacZ mice allows for irreversible tagging of endothelial cells. We provide unequivocal evidence for EndMT at the invasive front of the tumors in these transgenic mice. Collectively, our results show that EndMT is a unique mechanism for the accumulation of carcinoma-associated fibroblasts and suggest that antiangiogenic treatment of tumors may have a direct effect in decreasing activated fibroblasts that likely facilitate cancer progression.

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.

Physiological levels of tumstatin, a fragment of collagen IV α3 chain, are generated by MMP-9 proteolysis and suppress angiogenesis via αVβ3 integrin

We demonstrate a physiological role for tumstatin, a cleavage fragment of the α3 chain of type IV collagen (Col IVα3), which is present in the circulation. Mice with a genetic deletion of Col IVα3 show accelerated tumor growth associated with enhanced pathological angiogenesis, while angiogenesis associated with development and tissue repair are unaffected. Supplementing Col IVα3-deficient mice with recombinant tumstatin to a normal physiological concentration abolishes the increased rate of tumor growth. The suppressive effects of tumstatin require αVβ3 integrin expressed on pathological, but not on physiological, angiogenic blood vessels. Mice deficient in matrix metalloproteinase-9, which cleaves tumstatin efficiently from Col IVα3, have decreased circulating tumstatin and accelerated growth of tumor. These results indicate that MMP-generated fragments of basement membrane collagen can have endogenous function as integrin-mediated suppressors of pathologic angiogenesis and tumor growth.

The role of epithelial-to-mesenchymal transition in renal fibrosis

Epithelial-to-mesenchymal transition (EMT) involving injured epithelial cells plays an important role in the progression of fibrosis in the kidney. Tubular epithelial cells can acquire a mesenchymal phenotype, and enhanced migratory capacity enabling them to transit from the renal tubular microenvironment into the interstitial space and escape potential apoptotic cell death. EMT is a major contributor to the pathogenesis of renal fibrosis, as it leads to a substantial increase in the number of myofibroblasts, leading to tubular atrophy. However, recent findings suggest that EMT involving tubular epithelial cell is a reversible process, potentially determined by the surviving cells to facilitate the repopulation of injured tubules with new functional epithelia. Major regulators of renal epithelial cell plasticity in the kidney are two multifunctional growth factors, bone morphogenic protein-7 (BMP-7) and transforming growth factor β1 (TGF-β1). While TGF-β1 is a well-established inducer of EMT involving renal tubular epithelial cells, BMP-7 reverses EMT by directly counteracting TGF-β-induced Smad-dependent cell signaling in renal tubular epithelial cells. Such antagonism results in the repair of injured kidneys, suggesting that modulation of epithelial cell plasticity has therapeutic advantages.