Prostaglandins and calprotectin are genetically and functionally linked to the Inflammatory Bowel Diseases.

BACKGROUND: Genome wide association studies (GWAS) have identified and validated more than 200 genomic loci associated with the inflammatory bowel disease (IBD), although for most the causal gene remains unknown. Given the importance of myeloid cells in IBD pathogenesis, the current study aimed to uncover the role of genes within IBD genetic loci that are endogenously expressed in this cell lineage. METHODS: The open reading frames (ORF) of 42 genes from IBD-associated loci were expressed via lentiviral transfer in the THP-1 model of human monocytes and the impact of each of these on the cell's transcriptome was analyzed using a RNA sequencing-based approach. We used a combination of genetic and pharmacologic approaches to validate our findings in the THP-1 line with further validation in human induced pluripotent stem cell (hiPSC)-derived-monocytes. RESULTS: This functional genomics screen provided evidence that genes in four IBD GWAS loci (PTGIR, ZBTB40, SLC39A11 and NFKB1) are involved in controlling S100A8 and S100A9 gene expression, which encode the two subunits of calprotectin (CP). We demonstrated that increasing PTGIR expression and/or stimulating PTGIR signaling resulted in increased CP expression in THP-1. This was further validated in hiPSC-derived monocytes. Conversely, knocking-down PTGIR endogenous expression and/or inhibiting PTGIR signaling led to decreased CP expression. These analyses were extended to the known IBD gene PTGER4, whereby its specific agonist also led to increased CP expression. Furthermore, we demonstrated that the PTGIR and PTGER4 mediated control of CP expression was dependent on signaling via adenylate cyclase and STAT3. Finally, we demonstrated that LPS-mediated increases in CP expression could be potentiated by agonists of PTGIR and PTGER4, and diminished by their antagonists. CONCLUSION: Our results support a causal role for the PTGIR, PTGER4, ZBTB40, SLC39A11 and NFKB1 genes in IBD, with all five genes regulating the expression of CP in myeloid cells, as well as potential roles for the prostacyclin/prostaglandin biogenesis and signaling pathways in IBD susceptibility and pathogenesis.

Cardiac resident nestin(+) cells participate in reparative vascularisation.

The rodent heart contains a population of nestin((+)) cells derived from the embryonic neural crest and migrate to the scar after myocardial infarction (MI). The present study tested the hypothesis that intron 2 of the nestin gene drives expression and a subpopulation of nestin((+)) cells participate in reparative vascularisation. The directed expression of the green fluorescent protein (GFP) by the second intron of the nestin gene identified GFP/nestin((+)) cells intercalated among ventricular myocytes in the heart of normal transgenic mice. Ischemic injury led to the migration of GFP((+)) cells to the scar and a subpopulation was detected in CD31/nestin((+)) endothelial cells of newly formed blood vessels. The direct contribution to reparative vascularisation provided the impetus to test the hypothesis that increasing the population of nestin((+)) cells in the infarcted heart will improve scar healing. Skin-derived cells isolated from E18 Sprague-Dawley rats grew as spheres, expressed nestin, sox2, neural crest-related transcriptional genes and a panel of peptide growth factors. Skin-derived cells transplanted in the non-infarcted left ventricle of 3-day post-MI rats migrated to the peri-infarct/infarct region and remained engrafted for 21 days. A significantly smaller infarct, increased number of small calibre blood vessels and improved ventricular function were observed in engrafted infarcted rat hearts. Thus, the second intron of the nestin gene drives expression in the mouse heart and a subpopulation of GFP/nestin((+)) cells directly participate in reparative vascularisation. Increasing the population of nestin((+)) cells via the transplantation of skin-derived cells represents a potential approach to limit ischemic damage to the heart.

Nestin expression is lost in ventricular fibroblasts during postnatal development of the rat heart and re-expressed in scar myofibroblasts.

Studies have reported that the intermediate filament protein nestin was expressed in various non-stem/progenitor cells during development, downregulated during postnatal growth and re-expressed following injury. The present study tested the hypothesis that an analogous paradigm was prevalent for ventricular fibroblasts. In the neonatal rat heart, nestin protein levels were significantly higher than the adult heart and the isolation of cardiac cells revealed a selective expression in ventricular fibroblasts. In adult ventricular fibroblasts, nestin protein expression was markedly lower compared to neonatal ventricular fibroblasts. Following ischemic damage to the rat heart, nestin staining was detected in a subpopulation of scar myofibroblasts (37%) and the percentage of immunoreactive cells was greater than adult ventricular fibroblasts (7%) but significantly lower than neonatal ventricular fibroblasts (86%). Moreover, dissimilar rates of (3)H-thymidine uptake were observed among the fibroblast populations and may be related in part to the disparate percentage of nestin(+) cells. To assess the role of nestin in DNA synthesis, neonatal ventricular fibroblasts were infected with a lentivirus containing a shRNAmir directed against the intermediate filament protein. The partial depletion of nestin expression in neonatal ventricular fibroblasts significantly reduced basal DNA synthesis, in the absence of an apoptotic response. Thus, postnatal development of the rat heart was associated with a selective loss of nestin expression in ventricular fibroblasts and subsequent induction in a subpopulation of myofibroblasts following ischemic injury. The re-expression of nestin in scar myofibroblasts may represent an adaptive response to enhance their proliferative rate and accelerate the healing process.

The plating of rat scar myofibroblasts on matrigel unmasks a novel phenotype; the self assembly of lumen-like structures.

During tissue healing, the primary role of myofibroblasts involves the synthesis and deposition of collagen. However, it has also been reported that selective populations of myofibroblasts can acquire the phenotype and/or differentiate to other cells types. The present study tested the hypothesis that myofibroblasts isolated from the scar of the ischemically damaged rat heart can recapitulate an endothelial cell-like response when plated in a permissive in vitro environment. Scar myofibroblasts, neonatal and adult ventricular fibroblasts express smooth muscle α-actin, collagen α(1) type 1 and a panel of pro-fibrotic and pro-angiogenic peptide growth factor mRNAs. Myofibroblasts plated alone on matrigel led to the self assembly of lumen-like structures whereas neonatal and adult rat ventricular fibroblasts were unresponsive. Myofibroblasts labeled with the fluorescent cell tracker CM-DiI were injected in the viable myocardium of 3-day post-myocardial infarcted Sprague-Dawley rats and sacrificed 7 days later. Injected CM-DiI-labeled myofibroblasts were detected predominantly in the peri-infarct/infarct region, highlighting their migration to the damaged region. However, engrafted myofibroblasts in the peri-infarct/infarct region were unable to adopt an endothelial cell-like phenotype or lead to the de novo formation of CM-DiI-labeled blood vessels. The non-permissive nature of the infarct region may be attributed at least in part to the presence of growth-promoting stimuli as TGF-ß and the ß-adrenergic agonist isoproterenol inhibited the self assembly of lumen-like structures by myofibroblasts. Thus, when plated in a permissive in vitro environment, scar myofibroblasts can self assemble and form lumen-like structures providing an additional novel phenotype distinguishing this population from normal ventricular fibroblasts.

Nestin (+) stem cells independently contribute to neural remodelling of the ischemic heart.

Recent studies have revealed the existence of multipotent nestin-immunoreactive cells in the adult mammalian heart. These cells were recruited to infarct site following ischemic injury and differentiated to a vascular lineage leading to de novo blood vessel formation. Here, we show that a sub-population of cardiac resident nestin((+)) cells can further differentiate to a neuronal-like fate in vivo following myocardial infarction. In the ischemically damaged rat heart, neurofilament-M((+)) fibres were detected innervating the peri-infarct/infarct region and the preponderance of these fibres were physically associated with processes emanating from nestin((+)) cells. One week after isogenic heterotopic cardiac transplantation, the beating transplanted rat heart was devoid of neurofilament-M((+)) fibre staining. The superimposition of an ischemic insult to the transplanted heart led to the de novo synthesis of neurofilament-M((+)) fibres by cardiac resident nestin((+)) cells. Nerve growth factor infusion and the exposure of normal rats to intermittent hypoxia significantly increased the density of neurofilament-M((+)) fibres in the heart. However, these newly formed neurofilament-M((+)) fibres were not physically associated with nestin((+)) processes. These data highlight a novel paradigm of reparative fibrosis as a subpopulation of cardiac resident nestin((+)) cells directly contributed to neural remodelling of the peri-infarct/infarct region of the ischemically damaged rat heart via the de novo synthesis of neurofilament-M fibres.

Functional screen of inflammatory bowel disease genes reveals key epithelial functions.

BACKGROUND: Genetic studies have been tremendously successful in identifying genomic regions associated with a wide variety of phenotypes, although the success of these studies in identifying causal genes, their variants, and their functional impacts has been more limited. METHODS: We identified 145 genes from IBD-associated genomic loci having endogenous expression within the intestinal epithelial cell compartment. We evaluated the impact of lentiviral transfer of the open reading frame (ORF) of these IBD genes into the HT-29 intestinal epithelial cell line via transcriptomic analyses. By comparing the genes in which expression was modulated by each ORF, as well as the functions enriched within these gene lists, we identified ORFs with shared impacts and their putative disease-relevant biological functions. RESULTS: Analysis of the transcriptomic data for cell lines expressing the ORFs for known causal genes such as HNF4a, IFIH1, and SMAD3 identified functions consistent with what is already known for these genes. These analyses also identified two major clusters of genes: Cluster 1 contained the known IBD causal genes IFIH1, SBNO2, NFKB1, and NOD2, as well as genes from other IBD loci (ZFP36L1, IRF1, GIGYF1, OTUD3, AIRE and PITX1), whereas Cluster 2 contained the known causal gene KSR1 and implicated DUSP16 from another IBD locus. Our analyses highlight how multiple IBD gene candidates can impact on epithelial structure and function, including the protection of the mucosa from intestinal microbiota, and demonstrate that DUSP16 acts a regulator of MAPK activity and contributes to mucosal defense, in part via its regulation of the polymeric immunoglobulin receptor, involved in the protection of the intestinal mucosa from enteric microbiota. CONCLUSIONS: This functional screen, based on expressing IBD genes within an appropriate cellular context, in this instance intestinal epithelial cells, resulted in changes to the cell's transcriptome that are relevant to their endogenous biological function(s). This not only helped in identifying likely causal genes within genetic loci but also provided insight into their biological functions. Furthermore, this work has highlighted the central role of intestinal epithelial cells in IBD pathophysiology, providing a scientific rationale for a drug development strategy that targets epithelial functions in addition to the current therapies targeting immune functions.

The cardiac neural stem cell phenotype is compromised in streptozotocin-induced diabetic cardiomyopathy.

Neural stem cells were identified in the rat heart and during scar formation and healing participated in sympathetic fiber sprouting and angiogenesis. In the setting of diabetes, impaired wound healing represents a typical pathological feature. These findings provided the impetus to test the hypothesis that experimental diabetes adversely influenced the phenotype of cardiac neural stem cells. Streptozotocin (STZ)-induced diabetic rats were associated with elevated plasma glucose levels, significant loss of body weight and left ventricular contractile dysfunction. In the heart of STZ-diabetic rats, the density of nestin immunoreactive processes emanating from cardiac neural stem cells were reduced. The latter finding was reaffirmed as nestin protein expression was significantly decreased in the heart of STZ-diabetic rats and associated with a concomitant reduction of nestin mRNA. Employing the TUNEL assay, the loss of nestin expression in STZ-diabetic rats was not attributed to widespread cardiac neural stem cell apoptosis. Insulin administration to STZ-diabetic rats with established hyperglycaemia led to a modest recovery of nestin protein expression in cardiac neural stem cells. By contrast, the administration of insulin immediately after STZ injection improved plasma glucose levels and significantly attenuated the loss of nestin protein expression. These data highlight the novel observation that nestin protein expression in cardiac neural stem cells was significantly reduced in STZ-induced type I diabetic rats. The aberrant cardiac neural stem cell phenotype may compromise their biological role and predispose the diabetic heart to maladaptive healing following ischemic injury.

Infarct size is increased in female post-MI rats treated with rapamycin.

Rapamycin represents a recognized drug-based therapeutic approach to treat cardiovascular disease. However, at least in the female heart, rapamycin may suppress the recruitment of putative signalling events conferring cardioprotection. The present study tested the hypothesis that rapamycin-sensitive signalling events contributed to the cardioprotective phenotype of the female rat heart after an ischemic insult. Rapamycin (1.5 mg/kg) was administered to adult female Sprague-Dawley rats 24 h after complete coronary artery ligation and continued for 6 days. Rapamycin abrogated p70S6K phosphorylation in the left ventricle of sham rats and the noninfarcted left ventricle (NILV) of 1-week postmyocardial-infarcted (MI) rats. Scar weight (MI 0.028 +/- 0.006, MI+rapamycin 0.064 +/- 0.004 g) and surface area (MI 0.37 +/- 0.08, MI+rapamycin 0.74 +/- 0.03 cm2) were significantly larger in rapamycin-treated post-MI rats. In the NILV of post-MI female rats, rapamycin inhibited the upregulation of eNOS. Furthermore, the increased expression of collagen and TGF-beta3 mRNAs in the NILV were attenuated in rapamycin-treated post-MI rats, whereas scar healing was unaffected. The present study has demonstrated that rapamycin-sensitive signalling events were implicated in scar formation and reactive fibrosis. Rapamycin-mediated suppression of eNOS and TGF-beta3 mRNA in post-MI female rats may have directly contributed to the larger infarct and attenuation of the reactive fibrotic response, respectively.

The rat heart contains a neural stem cell population; role in sympathetic sprouting and angiogenesis.

Nestin-expressing cells were identified in the normal rat heart characterized by a small cell body and numerous processes and following an ischemic insult migrated to the infarct region. The present study was undertaken to identify the phenotype, origin and biological role of nestin-expressing cells during reparative fibrosis. A neural stem cell phenotype was identified based on musashi-1 expression, growth as a neurosphere, and differentiation to a neuronal cell. Using the Wnt1-cre; Z/EG transgenic mouse model, which expresses EGFP in embryologically-derived neural crest cells, the reporter signal was detected in nestin-expressing cells residing in the heart. In infarcted human hearts, nestin-expressing cells were detected in the viable myocardium and the scar and morphologically analogous to the population identified in the rat heart. Following either an ischemic insult or the acute administration of 6-hydroxydopamine, sympathetic sprouting was dependent on the physical association of neurofilament-M immunoreactive fibres with nestin-positive processes emanating from neural stem cells. To specifically study the biological role of the subpopulation in the infarct region, neural stem cells were isolated from the scar, fluorescently labelled and transplanted in the heart of 3-day post-MI rats. Injected scar-derived neural stem cells migrated to the infarct region and were used as a substrate for de novo blood vessel formation. These data have demonstrated that the heart contains a resident population of neural stem cells derived from the neural crest and participate in reparative fibrosis. Their manipulation could provide an alternative approach to ameliorate the healing process following ischemic injury.

Female rats fed a high-fat diet were associated with vascular dysfunction and cardiac fibrosis in the absence of overt obesity and hyperlipidemia: therapeutic potential of resveratrol.

It remains presently unknown whether vascular reactivity is impaired and whether maladaptive cardiac remodeling occurs before the onset of overt obesity and in the absence of hyperlipidemia. Normal female rats were fed a high-fat diet for 8 weeks and were associated with a modest nonsignificant increase of body weight (standard diet, 300 +/- 10, versus high-fat diet, 329 +/- 14 g) and a normal plasma lipid profile. In rats fed a high-fat diet, systolic (171 +/- 7 mm Hg) and diastolic blood pressures (109 +/- 3) were increased compared to a standard diet (systolic blood pressure, 134 +/- 8; diastolic blood pressure, 96 +/- 5 mm Hg), and acetylcholine-dependent relaxation of isolated aortic rings (high-fat diet, 22 +/- 5%, versus standard diet, 53 +/- 8%) was significantly reduced. Furthermore, perivascular fibrosis was detected in the heart of rats fed a high-fat diet. The exogenous addition of resveratrol (trans-3,5,4'-trihydroxystilbene) (0.1 microM) to aortic rings isolated from rats fed a high-fat diet restored acetylcholine-mediated relaxation (47 +/- 9%). The administration of resveratrol (20 mg/kg/day for 8 weeks) to rats fed a high-fat diet prevented the increase in blood pressure and preserved acetylcholine-dependent relaxation of isolated aortic rings. However, resveratrol therapy failed to attenuate the perivascular fibrotic response. These data have demonstrated that a high-fat diet fed to normal female rats can elicit a hypertensive response and induce perivascular fibrosis before the development of overt obesity and in the absence of hyperlipidemia. Resveratrol therapy can prevent the hypertensive response in female rats fed a high-fat diet but is without effect on the progression of perivascular fibrosis.

Dexamethasone treatment of post-MI rats attenuates sympathetic innervation of the infarct region.

Sympathetic fiber innervation of the damaged region following injury represents a conserved event of wound healing. The present study tested the hypothesis that impaired scar healing in post-myocardial infarction (post-MI) rats was associated with a reduction of sympathetic fibers innervating the infarct region. In 1-wk post-MI rats, neurofilament-M-immunoreactive fibers (1,116 +/- 250 microm(2)/mm(2)) were detected innervating the infarct region and observed in close proximity to a modest number of endothelial nitric oxide synthase-immunoreactive scar-residing vessels. Dexamethasone (Dex) treatment (6 days) of post-MI rats led to a significant reduction of scar weight (Dex + MI 38 +/- 4 mg vs. MI 63 +/- 2 mg) and a disproportionate nonsignificant decrease of scar surface area (Dex + MI 0.54 +/- 0.06 cm(2) vs. MI 0.68 +/- 0.06 cm(2)). In Dex-treated post-MI rats, the density of neurofilament-M-immunoreactive fibers (125 +/- 47 microm(2)/mm(2)) innervating the infarct region was significantly reduced and associated with a decreased expression of nerve growth factor (NGF) mRNA (Dex + MI 0.80 +/- 0.07 vs. MI 1.11 +/- 0.08; P < 0.05 vs. MI). Previous studies have demonstrated that scar myofibroblasts synthesize NGF and may represent a cellular target of Dex. The exposure of 1st passage scar myofibroblasts to Dex led to a dose-dependent suppression of [(3)H]thymidine uptake and a concomitant attenuation of NGF mRNA expression (untreated 3.47 +/- 0.35 vs. Dex treated 2.28 +/- 0.40; P < 0.05 vs. untreated). Thus the present study has demonstrated that impaired scar healing in Dex-treated post-MI rats was associated with a reduction of neurofilament-M-immunoreactive fibers innervating the infarct region. The attenuation of scar myofibroblast proliferation and NGF mRNA expression may represent underlying mechanisms contributing to the diminished neural response in the infarct region of Dex-treated post-MI rats.

Effects of resveratrol (trans-3,5,4'-trihydroxystilbene) treatment on cardiac remodeling following myocardial infarction.

Resveratrol (RES; trans-3,5,4'-trihydroxystilbene) has been shown to improve health and slow the progression of disease in various models. Several cardioprotective mechanisms have been identified including antioxidant, anti-inflammatory, and antifibrotic actions. Each of these actions is thought to have the ability to attenuate the pathophysiology underlying the deleterious cardiac structural remodeling that results from acute myocardial infarction (MI). Therefore, we evaluated the effect of resveratrol treatment on the progression of cardiac remodeling after MI. Four groups of rats (sham, n = 6; sham + RES, n = 21; MI, n = 26; MI + RES, n = 24) were treated for 13 weeks, starting 7 days before ligation of the left anterior descending coronary artery. Serial transthoracic echocardiography revealed that resveratrol had no effect on MI-induced left-ventricular and left-atrial dilatation or reduction in left-ventricular fractional shortening. Consistent with these findings, resveratrol did not improve the deterioration of hemodynamic function or reduce infarct size at 12 weeks post-MI. Resveratrol-treated animals did, however, show preserved cardiac contractile reserve in response to dobutamine administration. Radioligand binding revealed that MI reduced beta-adrenergic receptor density. Resveratrol administration increased beta-adrenoceptor density, so that resveratrol-treated MI rats had beta-adrenoceptor densities similar to normal rats. Real-time reverse transcription-polymerase chain reaction revealed that MI-induced changes in sarcoplasmic reticulum Ca2+-ATPase 2 and transforming growth factor beta-1 expression were unaltered by resveratrol, whereas MI-induced increases in atrial natriuretic factor (ANF) and connective tissue growth factor (CTGF) expression were attenuated. Resveratrol treatment does not improve cardiac remodeling and global hemodynamic function post-MI but does preserve contractile reserve and attenuate ANF and CTGF up-regulation.

Disparate regulation of signaling proteins after exercise and myocardial infarction.

INTRODUCTION: The signaling proteins extracellular signal-regulated kinase (ERK1/2) and protein kinase B (PKB) were implicated in the development of pathological cardiac hypertrophy. The present study examined whether the progression of physiological eccentric cardiac hypertrophy was associated with ERK1/2 and PKB recruitment. METHODS AND RESULTS: Following 1 and 3 wk of voluntary exercise, female Sprague-Dawley rats ran a total distance of 55 +/- 10 and 195 +/- 19 km, respectively. Left ventricular hypertrophy was detected in 3-wk-exercised rats, albeit prepro-ANP protein expression was unchanged. ERK1/2 was not recruited in the left ventricle (LV) of either 1-wk-exercised rats or the hypertrophied LV of 3-wk-exercised rats. In 1-wk-exercised rats, PKB Thr308 and Ser473 phosphorylation were significantly reduced, whereas a selective increase of PKB Ser473 phosphorylation was observed in the hypertrophied LV of 3-wk-exercised rats. In both 1- and 3-wk-exercised rats, an upward electrophoretic mobility band shift of p70 ribosomal S6 kinase (p70 S6K) was detected. In 1-wk post-myocardial-infarcted (MI) female Sprague-Dawley rats, scar formation was associated with increased left ventricular end-diastolic pressure. In the hypertrophied noninfarcted left ventricle (NILV), ERK1/2, p70 S6K, PKB Ser473, and Thr308 phosphorylation were increased. CONCLUSIONS: These data support the premise that ERK1/2 and PKB were differentially regulated during the development of eccentric physiological and pathological cardiac hypertrophy. It remains to be determined whether the chronic activation of either ERK1/2 and/or PKB in the NILV of post-MI rats may contribute in part to maladaptive cardiac remodelling.

Comparison of the effects of an angiotensin-converting enzyme inhibitor and a vasopeptidase inhibitor after myocardial infarction in the rat.

OBJECTIVES: The goal of this study was to compare the effects of the vasopeptidase inhibitor omapatrilat and the angiotensin-converting enzyme inhibitor (ACEI) captopril in the postmyocardial infarction (MI) rat model. BACKGROUND; The cardioprotective effects of ACEIs after MI are thought to be partially due to an increase in bradykinin (BK). Vasopeptidase inhibitors inhibit both ACE and neutral endopeptidase (NEP), further reduce BK metabolism and increase natriuretic peptides, which may result in better cardioprotective effects than with ACEIs after MI. METHODS: Myocardial infarction was induced in 514 Wistar male rats by ligation of the anterior coronary artery. Rats surviving 4 h after MI (n = 282) were assigned to omapatrilat (40 or 80 mg/kg/day), captopril (160 mg/kg/day) or no treatment. After 56 days, neurohumoral, hemodynamic, ventricular remodeling, morphometry, immunohistochemistry and cardiac cytokine expression were measured. RESULTS: Omapatrilat and captopril resulted in similarly improved survival, cardiac hemodynamics and reduced cardiac fibrosis and hypertrophy after MI. The pattern of left ventricular (LV) remodeling differed, omapatrilat causing less attenuation of the rightward shift of the LV pressure-volume relation at lower filling pressures than captopril. Both interventions reduced messenger ribonucleic acid expression of the profibrotic cytokine transforming growth factor-beta(1); neither effected the anti-inflammatory cytokine interleukin-10, and only captopril reduced the proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha). Expression of TNF-alpha was in cardiomyocytes. Both medications reduced circulating endothelin-1, angiotensin II and catecholamines, but only omapatrilat increased atrial natriuretic peptides. CONCLUSIONS: This study indicates that both omapatrilat and captopril markedly improve post-MI survival, cardiac function and cardiac remodeling in the rat. It would appear that the addition of NEP inhibition to those of ACEIs does not result in significant further benefit after MI.

AT1 receptor antagonist therapy preferentially ameliorated right ventricular function and phenotype during the early phase of remodeling post-MI.

1. The influence of AII on contractile dysfunction, regulation of the tyrosine kinase-dependent signaling molecule extracellular signal-regulated kinase (ERK), and natriuretic peptide gene expression were examined in the noninfarcted left ventricle (NILV) and right ventricle (RV) during the early phase of remodeling post-myocardial infarct (MI) in the rat. The selective AT(1) receptor antagonist irbesartan was administered <10 h following coronary artery ligation, and rats were killed either at 4-day or 2-week post-MI. 2. At 4 days post-MI, left ventricular systolic pressure (LVSP: sham=125+/-12, MI=91+/-4 mmHg) was decreased, whereas left ventricular end-diastolic pressure (LVEDP: sham=9+/-2, MI=17+/-2 mm Hg), right ventricular systolic (RVSP: sham=26+/-1, MI=34+/-2 mm Hg), and end-diastolic pressures (RVEDP: sham=3+/-0.5, MI=7+/-1 mm Hg) were increased. ERK phosphorylation was significantly elevated in the NILV and RV. 3. Irbesartan (40 mg x kg(-1)/day(-1)) administration did not improve left ventricular function, or suppress increased ERK phosphorylation in the 4-day post-MI rat. By contrast, irbesartan therapy normalized RVSP (MI+irbesartan=25+/-1 mm Hg), RVEDP (MI+irbesartan=3+/-0.3 mm Hg), and reduced ERK1 (MI=3.0+/-0.6, MI+irbesartan=2.0+/-0.3-fold increase), and ERK2 (MI=3.8+/-0.8, MI+irbesartan=2.2+/-0.5-fold increase) phosphorylation. 4. In 2-week post-MI rats, biventricular dysfunction was associated with increased prepro-ANP, and prepro-BNP mRNA expression. Irbesartan therapy normalized RVSP, attenuated RVEDP, and abrogated natriuretic peptide mRNA expression (prepro-ANP; MI=9+/-2, MI+irbesartan=2+/-1-fold increase, prepro-BNP; MI=6+/-2, MI+irbesartan=1+/-1-fold increase), whereas both transcripts remained elevated in the NILV despite the partial attenuation of LVEDP. 5. These data suggest that the therapeutic benefit of irbesartan treatment during the early phase of remodeling post-MI was associated with the preferential amelioration of RV contractile function and phenotype.

Elevated mean arterial pressure in the ovariectomized rat was normalized by ET(A) receptor antagonist therapy: absence of cardiac hypertrophy and fibrosis.

1. The influence of menopause on ventricular function and remodelling remains undefined. The following study examined the effect of ovariectomy on ventricular contractility, cardiac hypertrophy and extracellular matrix protein expression. 2. Elevated circulating levels of the vasoconstrictor endothelin-1 have been reported in post-menopausal women. Moreover, endothelin-1 has been shown to influence blood pressure, ventricular function and cardiac remodelling. In this regard, the potential pathophysiological role of endothelin-1 in the ovariectomized rat was assessed via the administration of the selective endothelin(A) receptor (ET(A)) antagonist BMS-182874. 3. In 3 and 6 week ovariectomized female Sprague - Dawley rats, uterus atrophy was associated with a significant increase in mean arterial pressure, and left ventricular systolic pressure, as compared to sham. By contrast, right ventricular contractile indices were normal in the ovariectomized rat. Despite increased systolic load, left ventricular hypertrophy was not evident, prepro-atrial natriuretic peptide (prepro-ANP) mRNA levels and collagen protein content were similar to sham. 4. The treatment of ovariectomized rats with BMS-182874 (60 mg kg(-1) per day) did not reverse uterus atrophy. However, BMS-182874 normalized mean arterial pressure, and left ventricular systolic pressure in the ovariectomized rat. 5. Thus, despite elevated blood pressure, ovariectomized rats were not associated with either cardiac hypertrophy or fibrosis. Lastly, endothelin-1, acting via the stimulation of the ET(A) receptor represents an integral mechanism implicated in the increase of mean arterial pressure following ovariectomy.

A high-fat diet increases risk of ventricular arrhythmia in female rats: enhanced arrhythmic risk in the absence of obesity or hyperlipidemia.

Obesity increases the incidence of cardiac arrhythmias and impairs wound healing. However, it is presently unknown whether a high-fat diet affects arrhythmic risk or wound healing before the onset of overt obesity or hyperlipidemia. After 8 wk of feeding a high-fat diet to adult female rats, a nonsignificant increase in body weight was observed and associated with a normal plasma lipid profile. Following ischemia/reperfusion injury, scar length (standard diet 0.29 +/- 0.09 vs. high-fat 0.32 +/- 0.13 cm), thickness (standard diet 0.047 +/- 0.02 vs. high-fat 0.059 +/- 0.01 cm), and collagen alpha(1) type 1 content (standard diet 0.21 +/- 0.04 vs. high-fat 0.20 +/- 0.04 arbitrary units/mm(2)) of infarcted hearts were not altered by the high-fat diet. However, the mortality rate was greatly increased 24 h postinfarction (from 5% to 46%, P < 0.01 for ischemia/reperfusion rats; from 20% to 89%, P < 0.0001, in complete-occlusion rats) in high-fat fed rats, in association with a higher prevalence of ventricular arrhythmias. Ventricular arrhythmia inducibility was also significantly increased in noninfarcted rats fed a high-fat diet. In the hearts of rats fed a high-fat diet, connexin-40 expression was absent, connexin-43 was hypophosphorylated and lateralized, and neurofilament-M immunoreactive fiber density (standard diet 2,020 +/- 260 vs. high-fat diet 2,830 +/- 250 microm(2)/mm(2)) and tyrosine hydroxylase protein expression were increased (P < 0.05). Thus, in the absence of overt obesity and hyperlipidemia, sympathetic hyperinnervation and an aberrant pattern of gap junctional protein expression and regulation in the heart of female rats fed a high-fat diet may have contributed in part to the higher incidence of inducible cardiac arrhythmias.

The phenotype and potential origin of nestin+ cardiac myocyte-like cells following infarction.

Nestin+ cardiac myocyte-like cells were detected in the peri-infarct/infarct region of the ischemically damaged heart. The present study was undertaken to elucidate the phenotype and potential origin of nestin+ cardiac myocyte-like cells and identify stimuli implicated in their appearance. In the infarcted human and rat heart, nestin+ cardiac myocyte-like cells were morphologically and structurally immature, exhibited a desmin-immunoreactive striated phenotype, expressed the beta(1)-adrenergic receptor, and associated with an aberrant pattern of connexin-43 expression and/or organization. Nestin+ cardiac myocyte-like cells were detected 24 h postischemic injury and persisted in the infarcted rat heart for 9 mo. In the normal rat heart, cardiac progenitor transcriptional factors Nkx2.5/GATA4 were detected in a subpopulation of nestin+ neural stem cells. Following an ischemic insult, nestin+/Nkx2.5+ neural stem cells migrated to the peri-infarct/infarct region and appeared to be in a primordial state of differentiation to a nestin+ cardiac myocyte-like cell. The exposure of adult male rats to normobaric hypoxia (12% O2) for 10 days failed to promote the appearance of nestin+ cardiac myocyte-like cells. Following osmotic pump delivery of isoproterenol to normal adult rats, nestin+ cardiac myocyte-like cells were detected, albeit the response was modest and secondary to tissue loss. Thus ischemia-induced appearance of nestin+ cardiac myocyte-like cells apparently represents an adaptive response to heal the infarcted heart. Nkx2.5/GATA4 expression in a subpopulation of resident neural stem cells provides the appropriate phenotype for their potential differentiation to a nestin+ cardiac myocyte-like cell.

Tamoxifen treatment of myocardial infarcted female rats exacerbates scar formation.

Hormonal replacement therapy in postmenopausal women was associated with an increased incidence of nonfatal myocardial infarction. Selective estrogen receptor modulators were considered an alternative pharmacological approach. However, selective estrogen receptor modulators acting via estrogen receptor-dependent and receptor-independent mechanisms may negatively influence cardiac remodeling. The present study tested the hypothesis that tamoxifen (TAM) treatment after coronary artery ligation compromised scar formation. TAM administration (10 mg kg(-1) day(-1) for 3 weeks) to postmyocardial infarcted (MI) female adult rats significantly increased scar surface area (TAM+MI = 0.67 +/- 0.08 vs MI = 0.45 +/- 0.06 cm(2)) and weight (TAM+MI = 0.071 +/- 0.007 vs MI = 0.050 +/- 0.006 grams). In the infarct region, a significant decrease (p < 0.05) of small calibre vessels (lumen diameter <50 microm) was observed in TAM treated post-MI rats (4.5 +/- 0.8 vessels/mm(2)), as compared to untreated MI rats (7 +/- 0.7 vessels/mm(2)). Consistent with the latter finding, 4-OH TAM caused a dose-dependent suppression of vascular endothelial growth factor (VEGF)-stimulated (10(-9) mol/l) capillarity-like tubule formation by rat aortic endothelial cells in vitro via an estrogen receptor-independent mechanism. These data have demonstrated that TAM treatment of post-MI female rats exacerbated scar formation and may have occurred at least in part via the attenuation of new vessel formation in the infarct region.

Antagonism of stromal cell-derived factor-1alpha reduces infarct size and improves ventricular function after myocardial infarction.

To examine the biological impact of locally expressed stromal cell-derived factor-1alpha (SDF-1alpha) during the acute phase of remodeling after myocardial infarction (MI), rats were treated with the selective CXCR4 receptor antagonist AMD3100 (1 mg/kg; given 24 h post-MI and continued for 6 days). In 1-week post-MI rats, intense SDF-1 immunoreactivity was detected in scar-residing vessels, and SDF-1alpha messenger ribonucleic acid (mRNA) levels were significantly greater in the infarct region compared to the noninfarcted left ventricle (NILV). AMD3100 treatment of post-MI rats reduced infarct size, improved systolic function, and partially suppressed the increased expression of atrial natriuretic peptide mRNA in the NILV. The latter finding indirectly suggests that SDF-1alpha may have contributed to the hypertrophic response of the NILV. SDF-1alpha treatment of neonatal rat ventricular myocytes (NNVMs) failed to promote protein synthesis. However, in hypertrophied NNVMs, SDF-1alpha treatment further augmented (3)H-leucine uptake, and AMD3100 selectively inhibited the increase in protein synthesis. Collectively, these data support the existence of an SDF-1alpha gradient in the damaged rat myocardium increasing toward the infarct region and highlight the novel observation that AMD3100 antagonism of the SDF-1alpha/CXCR4 axis reduced scar expansion and improved contractility. In vitro data further suggest that SDF-1alpha may have contributed to the hypertrophic response of the NILV.