Atrial natriuretic peptide-dependent modulation of hypoxia-induced pulmonary vascular remodeling.

UNLABELLED: Hypoxic stress upsets the balance in the normal relationships between mitogenic and growth inhibiting pathways in lung, resulting in pulmonary vascular remodeling characterized by hyperplasia of pulmonary arterial smooth muscle cells (PASMCs) and fibroblasts and enhanced deposition of extracellular matrix. Atrial natriuretic peptide (ANP) reduces pulmonary vascular resistance and attenuates hypoxia-induced pulmonary hypertension in vivo and PASMC proliferation and collagen synthesis in vitro. The current study utilized an ANP null mouse model (Nppa-/-) to test the hypothesis that ANP modulates the pulmonary vascular and alveolar remodeling response to normobaric hypoxic stress. Nine-10 wk old male ANP null (Nppa-/-) and wild type nontransgenic (NTG) mice were exposed to chronic hypoxia (10% O(2), 1 atm) or air for 6 wks. MEASUREMENT: pulmonary hypertension, right ventricular hypertrophy, and pulmonary arterial and alveolar remodeling were assessed. Hypoxia-induced pulmonary arterial hypertrophy and muscularization were significantly increased in Nppa-/- mice compared to NTG controls. Furthermore, the stimulatory effects of hypoxia on alveolar myofibroblast transformation (8.2 and 5.4 fold increases in Nppa-/- and NTG mice, respectively) and expression of extracellular matrix molecule (including osteopontin [OPN] and periostin [PN]) mRNA in whole lung were exaggerated in Nppa-/- mice compared to NTG controls. Combined with our previous finding that ANP signaling attenuates transforming growth factor (TGF)-beta-induced expression of OPN and PN in isolated PASMCs, the current study supports the hypothesis that endogenous ANP plays an important anti-fibrogenic role in the pulmonary vascular adaptation to chronic hypoxia.

Dominant negative mutation of the TGF-beta receptor blocks hypoxia-induced pulmonary vascular remodeling.

The present study utilized a novel transgenic mouse model that expresses an inducible dominant negative mutation of the transforming growth factor (TGF)-beta type II receptor (DnTGFbetaRII mouse) to test the hypothesis that TGF-beta signaling plays an important role in the pathogenesis of chronic hypoxia-induced increases in pulmonary arterial pressure and vascular and alveolar remodeling. Nine- to 10-wk-old male DnTGFbetaRII and control nontransgenic (NTG) mice were exposed to normobaric hypoxia (10% O2) or air for 6 wk. Expression of DnTGFbetaRII was induced by drinking 25 mM ZnSO4 water beginning 1 wk before hypoxic exposure. Hypoxia-induced increases in right ventricular pressure, right ventricular mass, pulmonary arterial remodeling, and muscularization were greatly attenuated in DnTGFbetaRII mice compared with NTG controls. Furthermore, the stimulatory effects of hypoxic exposure on pulmonary arterial and alveolar collagen content, appearance of alpha-smooth muscle actin-positive cells in alveolar parenchyma, and expression of extracellular matrix molecule (including collagen I and III, periostin, and osteopontin) mRNA in whole lung were abrogated in DnTGFbetaRII mice compared with NTG controls. Hypoxic exposure had no effect on systemic arterial pressure or heart rate in either strain. These data support the hypothesis that endogenous TGF-beta plays an important role in pulmonary vascular adaptation to chronic hypoxia and that disruption of TGF-beta signaling attenuates hypoxia-induced pulmonary hypertension, right ventricular hypertrophy, pulmonary arterial hypertrophy and muscularization, alveolar remodeling, and expression of extracellular matrix mRNA in whole lung.

Induction of estrus in suckled female yaks (Bos grunniens) and synchronization of ovulation in the non-sucklers for timed artificial insemination using progesterone treatments and Co-Synch regimens.

The objectives of the present study were to evaluate the induction of estrus and fertility in yak cows treated with Co-Synch regimens or progesterone (P(4)). In Experiment 1, postpartum suckled yaks were assigned to three treatments: (1) A (n=28), insertion of an intravaginal device containing P(4) (CIDR) on Day 0, PGF(2alpha) (i.m.) on Day 6 and PMSG (i.m.) at the time of CIDR removal on Day 7 (P(4)-PGF(2alpha)-PMSG); (2) B (n=21), PGF(2alpha) (i.m.) on Day 6 and PMSG on Day 7; (3) C (n=26), control group. Seven yak bulls were grazed with the cows for natural breeding. Rate of estrus within 96h of the end of treatment was greater (P<0.05) in A (100.0%) than in B (28.6%) or C (0.0%). First service conception rate (CR) determined by serum P(4) on Day 21 after breeding was greater (P<0.05) in A (78.6%) than in B (22.2%). Also, pregnancy rate (PR) during the breeding season was greater (P<0.05) in A (82.1%) than in B (19.0%) and C (7.7%). In Experiment 2, non-suckled yaks that calved in previous years but not in the current year were assigned to three treatments: (1) A (n=31), GnRH (i.m.) on Day 0, followed by PGF(2alpha) on Day 7 and timed artificial insemination (TAI) concurrently with GnRH treatment on Day 9 (Co-Synch regimen); (2) B (n=50), a CIDR device for 7 days plus PGF(2alpha) and PMSG at the time of CIDR withdrawal on Day 7 and TAI on Day 9 (P(4)-PGF(2alpha)-PMSG); (3) C (n=50), yak cows were artificially inseminated at spontaneous estrus. Frozen semen of Holstein and Jersey were used for insemination in Experiment 2. The CR assessed by rectal palpation 35 days after TAI was not different in A (22.6%), B (30.0%) and C (33.3%), but PR was greater in A and B than in C, when based on those cows presented for estrous synchronization programs. It is concluded that P(4)-PGF(2alpha)-PMSG protocol could efficiently induce estrus and result in an acceptable pregnancy rate in postpartum suckled yak cows. This technique and Co-Synch regimen can be applied successfully for TAI of non-suckled yak cows.

Atrial natriuretic peptide dose-dependently inhibits pressure overload-induced cardiac remodeling.

We hypothesized that a single copy of the proatrial natriuretic peptide gene (Nppa+/-) would not be adequate to protect heterozygous mice against exaggerated cardiac hypertrophy and remodeling after pressure-overload stress. Nppa+/+, Nppa+/-, and Nppa-/- mice were subjected to sham surgery or transverse aortic constriction and fed a basal salt diet. Heart weight varied inversely with Nppa gene load by 1 week after either surgery. Fractional shortening did not differ among genotypes at baseline and fell in Nppa-/- mice only after transverse aortic constriction. There was a graded response in collagen deposition related to atrial natriuretic peptide (ANP) expression after either surgery. A robust interstitial and perivascular fibrosis was noted in Nppa-/- and Nppa+/- but not in Nppa+/+ mice after transverse aortic constriction. Our findings are consistent with a growing body of evidence that ANP is an important modulator of cardiac hypertrophy and remodeling in response to hemodynamic stress. The observation that partial ANP deficiency results in exaggerated hypertrophy and remodeling after pressure overload suggests that genetic or environmental variation in ANP levels may play a role in the development of cardiac hypertrophy, remodeling, and failure in humans.

Volume overload results in exaggerated cardiac hypertrophy in the atrial natriuretic peptide knockout mouse.

OBJECTIVE: Previous studies suggest that atrial natriuretic peptide (ANP) may act as an autocrine/paracrine factor to modulate cardiac hypertrophy in response to various stimuli. The effect of ANP deficiency on the response to volume overload has not previously been studied. We hypothesised that ANP deficient mice would develop excess cardiac hypertrophy in response to volume overload stress. METHODS: Male homozygous ANP deficient (Nppa(-/-)) and wildtype (Nppa(+/+)) male mice maintained on either a normal salt (0.55% NaCl) or low salt (0.05% NaCl) diet from weaning were studied after 2 weeks of volume overload from an aorto-caval fistula (ACF). Unoperated littermates served as controls. Left ventricular (LV) structure and function was evaluated by echocardiography. Heart, LV, and lung weights were determined at sacrifice. Myocyte diameter was measured by morphometric analysis of fixed sections of the left ventricle. RESULTS: BP, heart weight, and LV weight were increased in Nppa(-/-) vs. Nppa(+/+) unoperated mice. Nppa(-/-) mice developed exaggerated heart and LV weight compared to Nppa(+/+) mice following ACF. Increased myocyte diameter paralleled increased echo LV wall thickness following ACF in Nppa(+/+) but not Nppa(-/-) mice fed with 0.55% NaCl, indicating that an alternate mechanism contributed to increased wall thickness in Nppa(-/-) mice. Mid-wall shortening was mildly depressed in the Nppa(-/-) vs. Nppa(+/+) genotype following ACF with fed 0.55% NaCl. A 0.05% NaCl diet from weaning normalized BP, but did not prevent exaggerated cardiac enlargement and LV hypertrophy following ACF in Nppa(-/-) mice. CONCLUSIONS: ANP-deficient mice exhibited an exaggerated increase in heart and LV weight in response to volume overload, which was not prevented by normalization of blood pressure. The findings suggest that ANP is an important physiologic modulator of the cardiac hypertrophy induced by volume overload.

Pressure-independent enhancement of cardiac hypertrophy in atrial natriuretic peptide-deficient mice.

1. Homozygous deletion of the pro-atrial natriuretic peptide (Nppa) gene (ANP-/-) has been associated with both cardiac hypertrophy and salt-sensitive hypertension in mice, suggesting that cardiac hypertrophy in ANP-/- mice may be related, at least in part, to increased afterload. 2. To test the hypothesis that cardiac hypertrophy in ANP-/- mice is independent of blood pressure, male ANP-/- and wild-type ANP+/+ mice were fed a low (0.05%) or basal (0.55%) NaCl diet. Five weeks later, mean arterial pressure (MAP) was measured in conscious mice; the whole heart, atria, left and right ventricles (LV and RV, respectively), brain, lung, kidney, liver and spleen were weighed and fixed for histological analysis. Separate groups of mice were subjected to echocardiographic examination under tribromoethanol anaesthesia. 3. Mean arterial pressure and atrial, LV and RV mass were greater in ANP-/- mice than in ANP+/+ mice fed the basal salt diet. Salt depletion equalized MAP in the two genotypes, but did not alter the relative cardiac hypertrophy in ANP-/- mice. The ANP-/- mice had significant LV cardiomyocyte hypertrophy when fed either basal or low-salt diets. 4. Left ventricle chamber dimensions did not differ between genotypes, but were significantly reduced in mice fed the low-salt diet; LV posterior wall and septal thickness were greater in ANP-/- than ANP+/+ mice and were not altered by diet, indicating a concentric pattern of LV hypertrophy in ANP-/- mice. Left ventricle function (cardiac output, stroke volume, ejection fraction, circumferential wall stress and velocity of circumferential wall shortening) did not differ between strains on either diet; circumferential wall stress was reduced in the low-salt groups; other functional parameters were not altered by diet. 5. These findings indicate that ANP deletion results in cardiomyocyte hypertrophy and biventricular hypertrophy independent of blood pressure, supporting the concept that ANP has direct antihypertrophic effects in the heart.

Effects of pressure overload on extracellular matrix expression in the heart of the atrial natriuretic peptide-null mouse.

This study tested the hypothesis that atrial natriuretic peptide has direct antihypertrophic actions on the heart by modulating expression of genes involved in cardiac hypertrophy and extracellular matrix production. Hearts of male, atrial natriuretic peptide-null and control wild-type mice that had been subjected to pressure overload after transverse aortic constriction and control unoperated hearts were weighed and subjected to microarray, Northern blot, and immunohistochemical analyses. Microarray and Northern blot analyses were used to identify genes that are regulated differentially in response to stress in the presence and absence of atrial natriuretic peptide. Immunohistochemical analysis was used to identify and localize expression of the protein products of these genes. Atrial natriuretic peptide-null mice demonstrated cardiac hypertrophy at baseline and an exaggerated hypertrophic response to transverse aortic constriction associated with increased expression of the extracellular matrix molecules periostin, osteopontin, collagen I and III, and thrombospondin, as well as the extracellular matrix regulatory proteins, matrix metalloproteinase-2 and tissue inhibitor of metalloproteinase-3, and the novel growth factor pleiotrophin compared with wild-type controls. These results support the hypothesis that atrial natriuretic peptide protects against pressure overload-induced cardiac hypertrophy and remodeling by negative modulation of genes involved in extracellular matrix deposition.