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The role of PI3K and Hippo signaling in chronic pancreatitis (CP) pathogenesis is unclear. Therefore, we assessed the involvement of these pathways in CP by examining the PI3K and Hippo signaling components PTEN and SAV1, respectively. We observed significant decreases in pancreatic PTEN and SAV1 levels in 2 murine CP models: repeated cerulein injection and pancreatic ductal ligation. Additionally, pancreas-specific deletion of Pten and Sav1 (DKO) induced CP in mice. Pancreatic connective tissue growth factor (CTGF) was markedly upregulated in both CP models and DKO mice, and pancreatic CCAAT/ enhancer-binding protein-[alpha] (CEBPA) expression was downregulated in the CP models. Interestingly, in pancreatic acinar cells (PACs), CEBPA knockdown reduced PTEN and SAV1 and increased CTGF levels in vitro. Furthermore, CEBPA knockdown in PACs induced acinar-to-ductal metaplasia and activation of cocultured macrophages and pancreatic stellate cells. These results were mitigated by CTGF inhibition. CP in DKO mice was also ameliorated by Ctgf gene deletion, and cerulein-induced CP was alleviated by antibody-mediated CTGF neutralization. Finally, we observed significantly decreased PTEN, SAV1, and CEBPA and increased CTGF levels in human CP tissues compared with nonpancreatitis tissues. Taken together, our results indicate that dysregulation of PI3K and Hippo signaling induces CP via CTGF upregulation., Introduction Chronic pancreatitis (CP) is characterized by continuous inflammatory destruction of the pancreas and replacement with fibrotic tissues, leading to the permanent loss of pancreatic function (1). CP is caused [...]

Although peptides are safe and useful as therapeutics, they are often easily degraded or metabolized. Dampening the clearance system for peptide ligands is a promising strategy for increasing the efficacy of peptide therapies. Natriuretic peptide receptor B (NPR-B) and its naturally occurring ligand, C-type natriuretic peptide (CNP), are potent stimulators of endochondral bone growth, and activating the CNP/NPR-B system is expected to be a powerful strategy for treating impaired skeletal growth. CNP is cleared by natriuretic peptide clearance receptor (NPR- C); therefore, we investigated the effect of reducing the rate of CNP clearance on skeletal growth by limiting the interaction between CNP and NPR-C. Specifically, we generated transgenic mice with increased circulating levels of osteocrin (OSTN) protein, a natural NPR-C ligand without natriuretic activity, and observed a dose-dependent skeletal overgrowth phenotype in these animals. Skeletal overgrowth in OSTN-transgenic mice was diminished in either CNP- or NPR-C-depleted backgrounds, confirming that CNP and NPR-C are indispensable for the bone growth-stimulating effect of OSTN. Interestingly, double-transgenic mice of CNP and OSTN had even higher levels of circulating CNP and additional increases in bone length, as compared with mice with elevated CNP alone. Together, these results support OSTN administration as an adjuvant agent for CNP therapy and provide a potential therapeutic approach for diseases with impaired skeletal growth., Introduction Most bones in mammals are formed through endochondral ossification, in that cartilaginous anlage is formed first and is subsequently replaced by calcified bone tissue (1). Although many signaling pathways, [...]

Kake T, Kitamura H, Adachi Y, Yoshioka T, Watanabe T, Matsushita H, Fujii T, Kondo E, Tachibe T, Kawase Y, Jishage K, Yasoda A, Mukoyama M, Nakao K. Chronically elevated plasma C-type natriuretic peptide level stimulates skeletal growth in transgenic mice. Am J Physiol Endocrinol Metab 297: E1339-E1348, 2009. First published October 6, 2009; doi: 10.1152/ajpendo.00272.2009.--C-type natriuretic peptide (CNP) plays a critical role in endochondral ossification through guanylyl cyclase-B (GC-B), a natriuretic peptide receptor subtype. Cartilage-specific overexpression of CNP enhances skeletal growth and rescues the dwarfism in a transgenic achondroplasia model with constitutive active mutation of fibroblast growth factor receptor-3. For future clinical application, the efficacy of CNP administration on skeletal growth must be evaluated. Due to the high clearance of CNP, maintaining a high concentration is technically difficult. However, to model high blood CNP concentration, we established a liver-targeted CNP-overexpressing transgenic mouse (SAP-CNP tgm). SAP-CNP tgm exhibited skeletal overgrowth in proportion to the blood CNP concentration and revealed phenotypes of systemic stimulation of cartilage bones, including limbs, paws, costal bones, spine, and skull. Furthermore, in SAP-CNP tgm, the size of the foramen magnum, the insufficient formation of which results in cervico-medullary compression in achondroplasia, also showed significant increase. CNP primarily activates GC-B, but under high concentrations it cross-reacts with guanylyl cyclase-A (GC-A), a natriuretic peptide receptor subtype of atrial natriuretic peptides (ANP) and brain natriuretic peptides (BNP). Although activation of GC-A could alter cardiovascular homeostasis, leading to hypotension and heart weight reduction, the skeletal overgrowth phenotype in the line of SAP-CNP tgm with mild overexpression of CNP did not accompany decrease of systolic blood pressure or heart weight. These results suggest that CNP administration stimulates skeletal growth without adverse cardiovascular effect, and thus CNP could be a promising remedy targeting achondroplasia. blood level elevation doi: 10.1152/ajpendo.00272.2009

CCN1 (cysteine-rich protein 61; Cyr61) is an extracellular matrix-associated signaling molecule that functions in cell migration, adhesion, and differentiation. We previously reported that CCN1 is induced at podocytes in rat anti-Thy-1 glomerulonephritis, a well-known model of reversible glomerular injury, but its expression and significance in the human kidney remain totally unknown (Sawai K, Mori K, Mukoyama M, Sugawara A, Suganami T, Koshikawa M, Yahata K, Makino H, Nagae T, Fujinaga Y, Yokoi H, Yoshioka T, Yoshimoto A, Tanaka I, Nakao K. J Am Soc Nephrol 14:1154-1163, 2003). Here we report that, in the human kidney, CCN1 expression was confined to podocytes in normal adult and embryonic glomeruli from the capillary loop stage. Podocyte CCNI expression was decreased in IgA nephropathy, diabetic nephropathy, and membranous nephropathy, whereas it remained unchanged in minimal change disease and focal segmental glomerulosclerosis. Downregulation of CCN1 was significantly greater in diseased kidneys with severe mesangial expansion. CCN1 protein was also localized in the thick ascending limb of Henle's loop, distal and proximal tubules, and collecting ducts, which was not altered in diseased kidneys. In vitro, recombinant CCN1 protein enhanced endothelial cell adhesion, whereas it prominently inhibited mesangial cell adhesion. CCN1 also completely suppressed mesangial cell migration, suggesting its role as a mesangial-repellent factor. In cultured podocytes, CCN1 markedly induced the expression of cyclin-dependent kinase inhibitor [p27.sup.Kip1] as well as synaptopodin in a dose-dependent manner and suppressed podocyte migration. These data indicate that CCN1 is expressed in podocytes, can act on glomerular cells to modulate glomerular remodeling, and is downregulated in diseased kidneys, suggesting that impairment of CCN1 expression in podocytes may contribute to the progression of glomerular disease with mesangial expansion. podocyte; glomerular visceral epithelial cell; cysteine-rich protein 61; mesangial cell; synaptopodin

Connective tissue growth factor (CTGF) is one of the candidate factors mediating downstream events of transforming, growth factor-[beta] (TGF-[beta]), but its role in fibrogenic properties of TGF-[beta] and in tubulointerstitial fibrosis has not yet been clarified. Using unilateral ureteral obstruction (UUO) in rats, we analyzed gene expression of TGF-[beta]1, CTGF, and fibronectin. We further investigated the effect of blockade of endogenous CTGF on TGF-[beta]-induced fibronectin expression in cultured rat renal fibroblasts by antisense oligodeoxynucleotide (ODN) treatment. After UUO, CTGF mRNA expression in the obstructed kidney was significantly upregulated subsequent to TGF-[beta]1, followed by marked induction of fibronectin mRNA. By in situ hybridization, CTGF mRNA was detected mainly in the interstitial fibrotic areas and tubular epithelial cells as well as in parietal glomerular epithelial cells in the obstructed kidney. The interstitial cells expressing CTGF mRNA were also positive for [alpha]-smooth muscle actin. CTGF antisense ODN transfected into cultured renal fibroblasts significantly attenuated TGF-[beta]-stimulated upregulation of fibronectin mRNA and protein compared with control ODN transfection, together with inhibited synthesis of type I collagen. With the use of a reporter assay, rat fibronectin promoter activity was increased by 2.5-fold with stimulation by TGF-[beta]1, and this increase was abolished with antisense CTGF treatment. Thus CTGF plays a crucial role in fibronectin synthesis induced by TGF-[beta], suggesting that CTGF blockade could be a possible therapeutic target against tubulointerstitial fibrosis. transforming growth factor-[beta]; in situ hybridization; obstructive nephropathy; antisense oligonucleotide; reporter assay

Atrial natriuretic polypeptide (ANP) is a hormone secreted by the atrial tissue of the heart as a result of an increase in blood pressure. This hormone regulates blood pressure, blood volume, and cardiac output, or the amount of blood pumped by the ventricles per minute. ANP also stimulates the elimination of salt in the urine, and these actions contribute to a reduction in the workload on the heart. The expression and distribution of ANP in the ventricles was assessed in tissue obtained at autopsies of 27 children with Kawasaki disease, an acute fever-associated illness. This disease affects children under five years of age and may be associated with the development of aneurysms in the coronary arteries, which supply the heart. Patients with Kawasaki disease often die of myocarditis, or inflammation of the heart muscle; coronary thrombosis, the formation of blood clots; rupture of coronary aneurysms; or coronary heart disease. In this study, 14 children died in the acute stage of the disease, including 3 of myocarditis and 11 of coronary heart disease. Evidence of a heart attack was detected in three of the children who died of coronary heart disease. Seven of the 14 children with acute-stage deaths developed heart failure 2 to 22 days before death. The remaining 13 children developed coronary artery aneurysms and died during the recovery period. Three children with granulations, or granular tissue, and congestive heart failure had detectable amounts of ANP in the heart cells surrounding the granulations. These three patients died within eight days after a heart attack. ANP was detected near sites of fiber-like tissue formation in 10 patients with a previous heart attack, including four with a history of congestive heart failure. These findings suggest that ANP was secreted into the ventricle more than a week after a heart attack in children with Kawasaki disease who died during recovery. In addition, a history of congestive heart failure was associated with an increase in the expression of ANP in patients with Kawasaki disease and a previous heart attack. (Consumer Summary produced by Reliance Medical Information, Inc.)

Atrial natriuretic peptide (ANP) is a protein hormone released by the atria of the heart in response to an increase in blood pressure. ANP affects blood pressure and volume and cardiac output, the amount of blood emptied from the ventricle per minute, and increases the elimination of salt from the body; these actions serve to decrease the workload on the heart. Although ANP is normally produced, stored and released from the atria, the upper receiving chambers of the heart, studies have shown that it is also present in the enlarged or failing ventricle, a pumping chamber. ANP levels were shown to be increased in the blood and ventricular tissues of patients with myocardial infarction, the deterioration of heart tissue due to cessation of blood supply. An antibody, or protein factor, that specifically binds to human ANP was used to detect ANP in the ventricles of five patients with myocardial infarction and five normal subjects. ANP was present in the ventricular tissue of patients with myocardial infarction. When infarction had affected both ventricles, ANP was detected in both ventricles. In a case of ventricular aneurysm, in which a portion of the ventricle wall is abnormally dilated, ANP was found in the affected area. The hormone was not observed in the ventricles of normal subjects. The presence of ANP in the ventricle in disease conditions such as myocardial infarction may serve as one of the body's compensatory mechanisms, or methods for coping with decreased heart function under abnormal conditions. (Consumer Summary produced by Reliance Medical Information, Inc.)

To elucidate the significance of oxidative stress in the modulation of endothelial functions, we examined the effects of [H.sub.2][O.sub.2] on the expression of two endothelium-derived vasoactive peptides, endothelin (ET) and adrenomedullin (Am), and their interaction. [H.sub.2][O.sub.2] dose dependently suppressed ET secretion and ET-1 mRNA expression in bovine carotid endothelial cells (ECs). Menadion sodium bisulfate, a redox cycling drug, also decreased ET secretion in a dose-dependent manner. Catalase, a [H.sub.2][O.sub.2] reductase, and dl-[Alpha]-tocopherol (vitamin E) significantly inhibited [H.sub.2][O.sub.2]-induced suppression of ET secretion. Downregulation of ET-1 mRNA under oxidative stress was regulated at the transcriptional level. In contrast, [H.sub.2][O.sub.2] increased Am secretion (and its mRNA expression) accompanied by the augmentation of cAMP production. Am, as well as 8-bromo-cAMP and forskolin decreased ET secretion in a dose-dependent fashion. Furthermore, an anti-Am monoclonal antibody that we developed abolished [H.sub.2][O.sub.2]-induced suppression of ET secretion at 6-24 h after the addition of [H.sub.2][O.sub.2]. [H.sub.2][O.sub.2] increased the intracellular [Ca.sup.2+] concentration ([[[Ca.sup.2+]].sub.i]). Moreover, treatment with ionomycin, a [Ca.sup.2+] ionophore, and thapsigargin, an inhibitor of endoplasmic reticulum ATPase, decreased ET secretion dose dependently for 3 h. These results suggest that the production of ET was decreased via activation of the Am-cAMP pathway and by the elevation of [[[Ca.sup.2+]].sub.i] under oxidative stress. These findings elucidate the coordinate expression of two local vascular hormones, ET and Am, under oxidative stress, which may protect against vascular diseases. intracellular [Ca.sup.2+]; hydrogen peroxide; cAMP; nitric oxide; C-type natriuretic peptide

Cardiac fibrosis, defined as a proliferation of interstitial fibroblasts and biosynthesis of extracellular matrix components in the ventricles of the heart, is a consequence of remodeling processes initiated by pathologic events associated with a variety of cardiovascular disorders, which leads to abnormal myocardial stiffness and, ultimately, ventricular dysfunction. Brain natriuretic peptide (BNP) is a cardiac hormone produced primarily by ventricular myocytes, and its plasma concentrations are markedly elevated in patients with congestive heart failure and acute myocardial infarction. However, its precise functional significance has been undefined. In this paper, we report the generation of mice with targeted disruption of BNP ([Nppb.sup.-/-] mice). We observed multifocal fibrotic lesions in the ventricles from [Nppb.sup.-/-] mice. No signs of systemic hypertension and ventricular hypertrophy are noted in [Nppb.sup.-/-] mice. In response to ventricular pressure overload, focal fibrotic lesions are increased in size and number in [Nppb.sup.-/-] mice, whereas no focal fibrotic changes are found in wild-type littermates ([Nppb.sup.+/+] mice). This study establishes BNP as a cardiomyocytederived antifibrotic factor in vivo and provides evidence for its role as a local regulator of ventricular remodeling.