Background-Patients with pulmonary arterial hypertension (PAH) have reduced expression of apolipoprotein E (apoE)and peroxisome proliferator-activated receptor-␥ in lung tissues, and deficiency of both has been linked to insulin resistance. ApoE deficiency leads to enhanced platelet-derived growth factor signaling, which is important in the pathobiology of PAH. We therefore hypothesized that insulin-resistant apoE-deficient (apoE Ϫ/Ϫ ) mice would develop PAH that could be reversed by a peroxisome proliferator-activated receptor-␥ agonist (eg, rosiglitazone). Methods and Results-We report that apoEϪ/Ϫ mice on a high-fat diet develop PAH as judged by elevated right ventricular systolic pressure. Compared with females, male apoE Ϫ/Ϫ were insulin resistant, had lower plasma adiponectin, and had higher right ventricular systolic pressure associated with right ventricular hypertrophy and increased peripheral pulmonary artery muscularization. Because male apoE Ϫ/Ϫ mice were insulin resistant and had more severe PAH than female apoE Ϫ/Ϫ mice, we treated them with rosiglitazone for 4 and 10 weeks. This treatment resulted in markedly higher plasma adiponectin, improved insulin sensitivity, and complete regression of PAH, right ventricular hypertrophy, and abnormal pulmonary artery muscularization in male apoE Ϫ/Ϫ mice. We further show that recombinant apoE and adiponectin suppress platelet-derived growth factor-BB-mediated proliferation of pulmonary artery smooth muscle cells harvested from apoE Ϫ/Ϫ or C57Bl/6 control mice. Conclusions-We have shown that insulin resistance, low plasma adiponectin levels, and deficiency of apoE may be risk factors for PAH and that peroxisome proliferator-activated receptor-␥ activation can reverse PAH in an animal model. Key Words: apolipoproteins Ⅲ glucose Ⅲ hypercholesterolemia Ⅲ hypertension, pulmonary Ⅲ insulin Ⅲ metabolism Ⅲ PPAR gamma A lthough insulin resistance is associated with systemic cardiovascular disease, 1-3 it has not been implicated as a predisposing factor in pulmonary arterial hypertension (PAH). Several findings, however, support such an association. Patients with idiopathic PAH have reduced pulmonary mRNA expression of peroxisome proliferator-activated receptor gamma (PPAR␥), 4 a ligand-activated nuclear receptor and transcription factor that regulates adipogenesis and glucose metabolism. [5][6][7] They also have reduced pulmonary mRNA expression of apolipoprotein E (apoE), 8 a protective factor known to reduce circulating oxidized low-density lipoprotein and atherogenesis in the vessel wall. 9 Deficiency of both PPAR␥ and apoE has been linked to insulin resistance and the metabolic syndrome. 7,9 Elevated levels of several circulating factors that are normally repressed by PPAR␥ are associated with insulin resistance 3 and implicated in the pathobiology of PAH. These include interleukin-6, 10,11 fractalkine, 12,13 monocyte chemoattractant protein-1, 14 endothelin-1 (ET-1), [15][16][17] and the endogenous nitric oxide synthase inhibitor asymmetric dimethylarginine (ADM...

Apelin and its cognate G protein-coupled receptor APJ constitute a signaling pathway with a positive inotropic effect on cardiac function and a vasodepressor function in the systemic circulation. The apelin-APJ pathway appears to have opposing physiological roles to the renin-angiotensin system. Here we investigated whether the apelin-APJ pathway can directly antagonize vascular disease-related Ang II actions. In ApoE-KO mice, exogenous Ang II induced atherosclerosis and abdominal aortic aneurysm formation; we found that coinfusion of apelin abrogated these effects. Similarly, apelin treatment rescued Ang II-mediated increases in neointimal formation and vascular remodeling in a vein graft model. NO has previously been implicated in the vasodepressor function of apelin; we found that apelin treatment increased NO bioavailability in ApoE-KO mice. Furthermore, infusion of an NO synthase inhibitor blocked the apelin-mediated decrease in atherosclerosis and aneurysm formation. In rat primary aortic smooth muscle cells, apelin inhibited Ang II-mediated transcriptional regulation of multiple targets as measured by reporter assays. In addition, we demonstrated by coimmunoprecipitation and fluorescence resonance energy transfer analysis that the Ang II and apelin receptors interacted physically. Taken together, these findings indicate that apelin signaling can block Ang II actions in vascular disease by increasing NO production and inhibiting Ang II cellular signaling.

Given their self-renewing and pluripotent capabilities, human embryonic stem cells (hESCs) are well poised as a cellular source for tissue regeneration therapy. However, the host immune response against transplanted hESCs is not well characterized. In fact, controversy remains as to whether hESCs have immune-privileged properties. To address this issue, we used in vivo bioluminescent imaging to track the fate of transplanted hESCs stably transduced with a double-fusion reporter gene consisting of firefly luciferase and enhanced GFP. We show that survival after transplant is significantly limited in immunocompetent as opposed to immunodeficient mice. Repeated transplantation of hESCs into immunocompetent hosts results in accelerated hESC death, suggesting an adaptive donor-specific immune response. Our data demonstrate that transplanted hESCs trigger robust cellular and humoral immune responses, resulting in intragraft infiltration of inflammatory cells and subsequent hESC rejection. Moreover, we have found CD4 ؉ T cells to be an important modulator of hESC immunemediated rejection. Finally, we show that immunosuppressive drug regimens can mitigate the anti-hESC immune response and that a regimen of combined tacrolimus and sirolimus therapies significantly prolongs survival of hESCs for up to 28 days. Taken together, these data suggest that hESCs are immunogenic, trigger both cellular and humoral-mediated pathways, and, as a result, are rapidly rejected in xenogeneic hosts. This process can be mitigated by a combined immunosuppressive regimen as assessed by molecular imaging approaches. molecular imaging ͉ immunological response ͉ immunosuppression

Background-Embryonic

Hypothesis: Hyperbaric oxygen (HBO) therapy increases vascular endothelial growth factor (VEGF) levels in wounds.Design: Wounds were monitored for oxygen delivery during HBO treatment, and wound fluids were analyzed for VEGF and lactate on days 2, 5, and 10 following wounding.Setting: Experimental animal model.Interventions: Rats were randomized to HBO therapy and control groups. The HBO therapy was administered for 90 minutes, twice daily with 100% oxygen at 2.1 atmospheres absolute. Treatment was administered for 7 days following wounding.Main Outcome Measures: Vascular endothelial growth factor, PO 2 , and lactate levels in wound fluid were measured on days 2, 5, and 10.

Studies have shown significant cardiovascular effects of exogenous apelin administration, including the potent activation of cardiac contraction. However, the role of the endogenous apelin-APJ pathway is less clear. To study the loss of endogenous apelin-APJ signaling, we generated mice lacking either the ligand (apelin) or the receptor (APJ). Apelin-deficient mice were viable, fertile, and showed normal development. In contrast, APJ-deficient mice were not born in the expected Mendelian ratio, and many showed cardiovascular developmental defects. Under basal conditions, both apelin and APJ null mice that survived to adulthood manifested modest decrements in contractile function. However, with exercise stress both mutant lines demonstrated consistent and striking decreases in exercise capacity. To explain these findings, we explored the role of autocrine signaling in vitro using field stimulation of isolated left ventricular cardiomyocytes lacking either apelin or APJ. Both groups manifested less sarcomeric shortening and impaired velocity of contraction and relaxation with no difference in calcium transient. Taken together, these results demonstrate that endogenous apelin-APJ signaling plays a modest role in maintaining basal cardiac function in adult mice with a more substantive role during conditions of stress. In addition, an autocrine pathway seems to exist in myocardial cells, the ablation of which reduces cellular contraction without change in calcium transient. Finally, differences in the developmental phenotype between apelin and APJ null mice suggest the possibility of undiscovered APJ ligands or ligand-independent effects of APJ.

Introduction A comparative analysis of the efficacy of different cell candidates for the treatment of heart disease remains to be described. This study is designed to evaluate the therapeutic efficacy of 4 cell types in a murine model of myocardial infarction. Methods Bone marrow mononuclear cells (MN), mesenchymal stem cells (MSC), skeletal myoblasts (SkMb) and fibroblasts (Fibro) were isolated from male L2G transgenic mice (FVB background) that constitutively express firefly luciferase (Fluc) and green fluorescence protein (GFP). Cells were characterized by flow cytometry, bioluminescence imaging (BLI), and luminometry. Female FVB mice (n=60) underwent LAD ligation and were randomized into 5 groups to intramyocardially receive one cell type (5 × 105) or PBS as control. Cell survival was measured in vivo by BLI and ex vivo by TaqMan PCR at week 6. Cardiac function was assessed by echocardiography and invasive hemodynamic measurements were made at week 6. Results Fluc expression correlated with the cell number in all groups (r2 >0.93). In vivo BLI revealed acute donor cell death of MSC, SkMb, and Fibro within 3 weeks after transplantation. By contrast, cardiac signals were still present after 6 weeks in the MN group, as confirmed by TaqMan PCR (P<0.01). Echocardiography showed significant preservation of fractional shortening in the MN group compared to controls (P<0.05). Measurements of left ventricular end-systolic/diastolic volumes revealed that the least amount of ventricular dilatation occurred in the MN group (P<0.05). Histology confirmed the presence of MN, although there was no evidence of transdifferentiation by donor MN into cardiomyocytes. Conclusion This is the first study to directly compare a variety of cell candidates for myocardial therapy. Compared to MSC, SkMB, and Fibro, our results suggest that MN cells exhibit a more favorable survival pattern, which translates into a more robust preservation of cardiac function.

Background— Cardiac cell transplantation is limited by poor graft viability. We aimed to enhance the survival of transplanted cardiomyoblasts using growth factor-supplemented collagen matrices. Methods and Results— H9c2 cardiomyoblasts were lentivirally transduced to express firefly luciferase and green fluorescent protein (GFP). Lewis rats underwent ligation of the left anterior descending artery (LAD) ligation to induce an anterior wall myocardial infarction. Hearts (n=9/group) were harvested and restored ex vivo with 1×10 6 genetically labeled H9c2 cells either in (1) saline-suspension, or seeded onto (2) collagen-matrix (Gelfoam [GF];), (3) GF/Matrigel (GF/MG), (4) GF/MG/VEGF (10 μg/mL), or (5) GF/MG/FGF (10 μg/mL). Hearts were then abdominally transplanted into syngeneic recipients (working heart model). Controls (n=6/group) underwent infarction followed by GF implantation or saline injection. Cell survival was evaluated using optical bioluminescence on days 1, 5, 8, 14, and 28 postoperatively. At 4 weeks, fractional shortening and ejection fraction were determined using echocardiography and magnetic resonance imaging, respectively. Graft characteristics were assessed by immunohistology. Bioluminescence signals on days 5, 8, and 14 were higher for GF-based grafts compared with plain H9c2 injections ( P <0.03). Signals were higher for GF/MG grafts compared with GF alone ( P <0.02). GFP-positive, spindle-shaped H9c2 cells were found integrated in the infarct border zones at day 28. Left ventricular (LV) function of hearts implanted with collagen-based grafts was better compared with controls ( P <0.05). Vascular endothelial growth factor or fibroblast growth factor did not further improve graft survival or heart function. Conclusions— Collagen matrices enhance early survival of H9c2 cardiomyoblasts after transplantation into ischemic hearts and lead to improved LV function. Further optimization of the graft design should make restoration of large myocardial infarctions by tissue engineering approaches effective.