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-beta1 (TGF-beta1) 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.

Increasing evidence suggests that angiotensin II (Ang II) may act as a growth factor for the heart. However, direct effects of Ang II on mammalian cardiac cells (myocytes and nonmyocytes), independent of secondary hemodynamic and neurohumoral effects, have not been well characterized. Therefore, we analyzed the molecular phenotype of cultured cardiac cells from neonatal rats in response to Ang II. In addition, we examined the effects of selective Ang II receptor subtype antagonists in mediating the biological effects of Ang II. In myocyte culture, Ang II caused an increase in protein synthesis without changing the rate of DNA synthesis. In contrast, Ang II induced increases in protein synthesis, DNA synthesis, and cell number in nonmyocyte cultures (mostly cardiac fibroblasts). The Ang II-induced hypertrophic response of myocytes and mitogenic response of fibroblasts were mediated primarily by the AT1 receptor. Ang II caused a rapid induction of many immediate-early genes (c-fos, c-jun, jun B, Egr-1, and c-myc) in myocyte and nonmyocyte cultures. Ang II induced "late" markers for cardiac hypertrophy, skeletal alpha-actin and atrial natriuretic factor expression, within 6 hours in myocytes. Ang II also caused upregulation of the angiotensinogen gene and transforming growth factor-beta 1 gene within 6 hours. Induction of immediate-early genes, late genes, and growth factor genes by Ang II was fully blocked by an AT1 receptor antagonist but not by an AT2 receptor antagonist. These results indicate that: (1) Ang II causes hypertrophy of cardiac myocytes and mitogenesis of cardiac fibroblasts, (2) the phenotypic changes of cardiac cells in response to Ang II in vitro closely mimic those of growth factor response in vitro and of load-induced hypertrophy in vivo, (3) all biological effects of Ang II examined here are mediated primarily by the AT1 receptor subtype, and (4) Ang II may initiate a positive-feedback regulation of cardiac hypertrophic response by inducing the angiotensinogen gene and transforming growth factor-beta 1 gene.

Hypertrophy is a fundamental adaptive process employed by postmitotic cardiac and skeletal muscle in response to mechanical load. How muscle cells convert mechanical stimuli into growth signals has been a long-standing question. Using an in vitro model of load (stretch)-induced cardiac hypertrophy, we demonstrate that mechanical stretch causes release of angiotensin II (Ang II) from cardiac myocytes and that Ang II acts as an initial mediator of the stretch-induced hypertrophic response. The results not only provide direct evidence for the autocrine mechanism in load-induced growth of cardiac muscle cells, but also define the pathophysiological role of the local (cardiac) renin-angiotensin system.

The PTEN/PI3K signaling pathway regulates a vast array of fundamental cellular responses. We show that cardiomyocyte-specific inactivation of tumor suppressor PTEN results in hypertrophy, and unexpectedly, a dramatic decrease in cardiac contractility. Analysis of double-mutant mice revealed that the cardiac hypertrophy and the contractility defects could be genetically uncoupled. PI3Kalpha mediates the alteration in cell size while PI3Kgamma acts as a negative regulator of cardiac contractility. Mechanistically, PI3Kgamma inhibits cAMP production and hypercontractility can be reverted by blocking cAMP function. These data show that PTEN has an important in vivo role in cardiomyocyte hypertrophy and GPCR signaling and identify a function for the PTEN-PI3Kgamma pathway in the modulation of heart muscle contractility.

External load plays a critical role in determining muscle mass and its phenotype in cardiac myocytes. Cardiac myocytes have the ability to sense mechanical stretch and convert it into intracellular growth signals, which lead to hypertrophy. Mechanical stretch of cardiac myocytes in vitro causes activation of multiple second messenger systems that are very similar to growth factor-induced cell signaling systems. Stretch of neonatal rat cardiac myocytes stimulates a rapid secretion of angiotensin II which, together with other growth factors, mediates stretch-induced hypertrophic responses in vitro. In this review, various cell signaling mechanisms initiated by mechanical stress on cardiac myocytes are summarized with emphasis on potential mechanosensing mechanisms and the relationship between mechanical loading and the cardiac renin-angiotensin system.

Phosphoinositide 3-kinase (PI3K) has been shown to regulate cell and organ size in Drosophila, but the role of PI3K in vertebrates in vivo is not well understood. To examine the role of PI3K in intact mammalian tissue, we have created and characterized transgenic mice expressing constitutively active or dominantnegative mutants of PI3K in the heart. Cardiacspeci®c expression of constitutively active PI3K resulted in mice with larger hearts, while dominantnegative PI3K resulted in mice with smaller hearts. The increase or decrease in heart size was associated with comparable increase or decrease in myocyte size. Cardiomyopathic changes, such as myocyte necrosis, apoptosis, interstitial ®brosis or contractile dysfunction, were not observed in either of the transgenic mice. Thus, the PI3K pathway is necessary and suf®-cient to promote organ growth in mammals.

An unresolved question in cardiac biology is whether distinct signaling pathways are responsible for the development of pathological and physiological cardiac hypertrophy in the adult. Physiological hypertrophy is characterized by a normal organization of cardiac structure and normal or enhanced cardiac function, whereas pathological hypertrophy is associated with an altered pattern of cardiac gene expression, fibrosis, cardiac dysfunction, and increased morbidity and mortality. The elucidation of signaling cascades that play distinct roles in these two forms of hypertrophy will be critical for the development of more effective strategies to treat heart failure. We examined the role of the p110␣ isoform of phosphoinositide 3-kinase (PI3K) for the induction of pathological hypertrophy (pressure overload-induced) and physiological hypertrophy (exercise-induced) by using transgenic mice expressing a dominant negative (dn) PI3K(p110␣) mutant specifically in the heart. dnPI3K transgenic mice displayed significant hypertrophy in response to pressure overload but not exercise training. dnPI3K transgenic mice also showed significant dilation and cardiac dysfunction in response to pressure overload. Thus, PI3K(p110␣) appears to play a critical role for the induction of physiological cardiac growth but not pathological growth. PI3K(p110␣) also appears essential for maintaining contractile function in response to pathological stimuli.

One of the least-understood areas in biology is the determination of the size of animals and their organs. In Drosophila, components of the insulin receptor phosphoinositide 3-kinase (PI3K) pathway determine body, organ, and cell size. Several biochemical studies have suggested that Akt/protein kinase B is one of the important downstream targets of PI3K. To examine the role of Akt in the regulation of organ size in mammals, we have generated and characterized transgenic mice expressing constitutively active Akt (caAkt) or kinasedeficient Akt (kdAkt) specifically in the heart. The heart weight of caAkt transgenic mice was increased 2.0-fold compared with that of nontransgenic mice. The increase in heart size was associated with a comparable increase in myocyte cell size in caAkt mice. The kdAkt mutant protein attenuated the constitutively active PI3K-induced overgrowth of the heart, and the caAkt mutant protein circumvented cardiac growth retardation induced by a kinase-deficient PI3K mutant protein. Rapamycin attenuated caAkt-induced overgrowth of the heart, suggesting that the mammalian target of rapamycin (mTOR) or effectors of mTOR mediated caAktinduced heart growth. In conclusion, Akt is sufficient to induce a marked increase in heart size and is likely to be one of the effectors of the PI3K pathway in mediating heart growth.