Effect of nitric oxide deficiency on the pulmonary PTHrP system.

Nitric oxide (NO) deficiency is common in pulmonary diseases, but its effect on pulmonary remodelling is still controversial. As pulmonary parathyroid hormone-related protein (PTHrP) expression is a key regulator of pulmonary fibrosis and development, the effect of chronic NO deficiency on the pulmonary PTHrP system and its relationship with oxidative stress was addressed. NO bioavailability in adult rats was reduced by systemic administration of L-NAME via tap water. To clarify the role of NO synthase (NOS)-3-derived NO on pulmonary expression of PTHrP, NOS-3-deficient mice were used. Captopril and hydralazine were used to reduce the hypertensive effect of L-NAME treatment and to interfere with the pulmonary renin-angiotensin system (RAS). Quantitative RT-PCR and immunoblot techniques were used to characterize the expression of key proteins involved in pulmonary remodelling. L-NAME administration significantly reduced pulmonary NO concentration and caused oxidative stress as characterized by increased pulmonary nitrite concentration and increased expression of NOX2, p47phox and p67phox. Furthermore, L-NAME induced the pulmonary expression of PTHrP and of its corresponding receptor, PTH-1R. Expression of PTHrP and PTH-1R correlated with the expression of two well-established PTHrP downstream targets, ADRP and PPARγ, suggesting an activation of the pulmonary PTHrP system by NO deficiency. Captopril reduced the expression of PTHrP, profibrotic markers and ornithine decarboxylase, but neither that of PTH-1R nor that of ADRP and PPARγ. All transcriptional changes were confirmed in NOS-3-deficient mice. In conclusion, NOS-3-derived NO suppresses pulmonary PTHrP and PTH-1R expression, thereby modifying pulmonary remodelling.

Effect of high fat diet on pulmonary expression of parathyroid hormone-related protein and its downstream targets.

AIMS: Parathyroid hormone-related protein (PTHrP) is involved in lung development and surfactant production. The latter one requires a paracrine interaction between type II alveolar cells and lipofibroblasts in which leptin triggers PTHrP-induced effects. Whether increased plasma leptin levels, as they occur in high fat diet, modify the expression of PTHrP remains unclear. Furthermore, the effect of high fat diet under conditions of forced pulmonary remodelling such as response to post myocardial infarction remains to be defined. MATERIALS AND METHODS: C57 bl/6 mice were randomized to either normal diet or high fat diet at an age of 6 weeks. Seven months later, the mice were euthanized and the lung was removed and frozen in fluid nitrogen until use. Samples were analyzed by real-time RT-PCR and western blot. Leptin deficient mice were used to investigate the effect of leptin on pulmonary expression of PTHrP more directly. A subgroup of mice with and without high fat diet underwent in vivo ischemia (45 min) and reperfusion (4 weeks). Finally, experiments were repeated with prolonged high-fat diet. KEY FINDINGS: High fat diet increased plasma leptin levels by 30.4% and the pulmonary mRNA expression of PTHrP (1,447-fold), PTH-1 receptor (4.21-fold), and PTHrP-downstream targets ADRP (7.54-fold) and PPARγ (5.27-fold). Pulmonary PTHrP expression was reduced in leptin deficient mice by 88% indicating leptin dependent regulation. High fat diet further improved changes in pulmonary adaptation caused by ischemia/reperfusion (1.48-fold increased PTH-1 receptor protein expression). These effects were lost during prolonged high fat diet. SIGNIFICANCE: This study established that physiological regulation of leptin plasma levels by high fat diet affects the pulmonary PTHrP expression and of PTHrP downstream targets. Modification of pulmonary expression of PTH-1 receptors by high fat diet after myocardial infarction suggests that the identified interaction may participate in the obesity paradox.

TGF-beta1 improves cardiac performance via up-regulation of laminin receptor 37/67 in adult ventricular cardiomyocytes.

TGF-beta1 plays an important role in cardiac fibrosis, apoptosis, induction of hypertrophy and contractile dysfunction. This study investigates whether TGF-beta1 plays a role in laminin receptor 37/67 (37/67 LR)-dependent regulation of cardiac performance. Therefore, isolated adult cardiomyocytes were stimulated with TGF-beta1, the expression of the 37/67 LR was determined and cell shortening was investigated on cells attached to a non-specific, serum-based attachment substrate or to specific, laminin-coated dishes. The role of the MAP kinases in TGF-beta1-dependent induction of the 37/67 LR was examined by addition of PD98059, SB202190 and SP600125. Finally, the expression of receptor mRNA was investigated in transgenic mice constitutively over-expressing TGF-beta1 and the relationship to distress score and lung wet weight-to-body weight was analysed. TGF-beta1 induced a significant increase of the 37/67 LR mRNA and protein expression. The cytokine induced p38 MAP kinase and JNK, but not ERK. Inhibition of either p38 MAP kinase or JNK attenuated the TGF-beta1-dependent increase in 37/67 LR expression. TGF-beta1 induced a loss of cell shortening in cells attached to a non-specific substrate, but not in cells on a pre-coated laminin matrix. Inhibition of JNK attenuated the protective effect of laminin receptor up-regulation on cardiac performance. Inhibition of p38 MAP kinase attenuated the depressive effect of TGF-beta1 on basal cell shortening. In transgenic mice over-expressing TGF-beta1 a strong induction of laminin receptor expression attenuated the severeness of the mice' symptoms. This study shows a new and protective role of TGF-beta1-dependent up-regulation of the 37/67 LR in cardiomyocytes in cardiac remodelling with increased laminin expression.