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Pericytes are perivascular cells imbedded within the basement membrane of microvascular endothelial cells. Importantly, pericytes are critical for microvascular integrity especially endothelial barrier integrity and immune quiescence and may participate in organ disease notably vascular dementia or heart failure. However, little is known about pericyte homeostasis in adult tissues. Our aim is to investigate the behavior of pericytes after they have been reduced by 50% in the brain and heart of adult mice. To deplete pericytes in mice, we used diphtheria toxin (DTA)-mediated conditional cell ablation. More specifically, we used Pdgfrb-Cre/ERT2; Rosa-DTA mice, in which DTA expression is induced specifically in pericytes by Pdgfrb promoter-driven expression of Cre/ERT2 and may be temporally controlled by tamoxifen injections. To induce a 50% pericyte depletion, adult mice were injected with 1 mg tamoxifen for 2 consecutive days and sacrificed 4, 7, 14, 21 and 28 days after the first injection to follow their fate over time. We found pericytes are renewed both in the brain and in the heart. Their renewal follows the same kinetic in both organs, i.e. their number starts to increase between 7 and 14 days after the first tamoxifen injection and pericytes are fully renewed 21 days after they were depleted. The origin of the new pericytes is still under investigation, however, what we found is that pericytes being renewed, transiently express smooth muscle actin alpha 2 (ACTA2), at least in the heart. This study indicates for the first time that pericytes can undergo extensive remodeling after tissue injury in adults. [ABSTRACT FROM AUTHOR]

Heart failure with preserved ejection fraction (HFpEF) is proposed to be caused by endothelial dysfunction in cardiac small vessels. We previously identified Hhipl1, as a gene upregulated in the coronary vasculature of Leptin receptor deficient mice (Leprdb/db) a well-established mouse model of HFpEF. Importantly, Hhipl1 encodes for a decoy receptor of Desert Hedgehog (DHH) which is known to be critical for endothelial integrity. Our objective is to investigate the functional consequences of impaired Hedgehog (HH) signaling in the adult heart in order to identify novel mechanisms underlying the development of diastolic dysfunction. To do so, Cdh5-Cre/ERT2, DhhFlox/Flox (DhhECKO) mice and their control littermates were administered with tamoxifen at 8 weeks of age to induce Dhh KO. Their cardiac function, exercise tolerance, and the phenotype of their coronary vasculature were assessed one month later. DhhECKO mice presented significantly reduced exercise tolerance, increased end-diastolic pressure (EDP) and Tau, with no change in their ejection fraction consistent with diastolic dysfunction. At molecular and cellular level, impaired cardiac relaxation in DhhECKO mice was associated with a significantly decreased phospholamban phosphorylation on Thr17 and an alteration of sarcomeric shortening in ex-vivo. Besides, as expected, DhhECKO mice exhibited phenotypic changes in their coronary vasculature including a prominent pro-thrombotic phenotype (63 ± 6.2 vs. 25 ± 5.2 thrombi/mm2; P < 0.001) leading to an impaired capillary perfusion and local hypoxia. Notably, antiaggregant therapies (aspirin and clopidogrel) prevented the occurrence of both diastolic dysfunction and exercise intolerance in DhhECKO mice demonstrating for the first time that thrombosis may promote diastolic dysfunction. Importantly, we confirmed the critical role of thrombosis in Leprdb/db mice which also displayed increased cardiac small vessel thrombosis in comparison to control mice. Alike DhhECKO mice, we found that antiaggregants decreased EDP (6.3 ± 0.4 mmHg in aspirin-treated vs. 11.3 ± 0.79 in control mice; P = 0.001) and improved exercise tolerance in Leprdb/db mice (34 ± 2.52 min in aspirin-treated vs. 24 ± 3.55 in control mice; P = 0.004). Altogether, these results demonstrate that small vessel thrombosis may participate in the pathophysiology of heart failure with preserved ejection fraction. [ABSTRACT FROM AUTHOR]

Chronic limb threatening ischemia (CLTI), which is one of the major complications of diabetes, is associated with poor limb and cardiovascular prognoses, along with a dramatic decrease in life expectancy. However, the pathophysiology of this disease is poorly understood and therapeutic targets are missing. Our objective is to explore the contribution of endothelial dysfunction in the development of CLTI in diabetic mice. Hind limb ischemia (HLI) was induced by ligation and resection of the femoral artery in C57BL6/J male mice, in which insulin resistance was induced by a high fat diet, and hyperglycaemia was induced one month later by low dose streptozotocin administrations. According to our previous investigations, ischemic foot re-perfusion, assessed via laser doppler perfusion imaging, was significantly reduced in diabetic animal 28 days after HLI surgery was performed (ratio blood flow in the ischemic leg vs non ischemic leg = 0.27 ± 0.09 vs 0.44 ± 0.21 in control mice; P = 0.03), even though angiogenesis was identical in both groups. On the contrary, we found that impaired ischemic foot re-perfusion was associated with an increased endothelial cell activation attested by an increased ICAM1 expression (P < 0.0001). We then hypothesized that ICAM1, by interacting with white blood cells (WBC), may compromise the perfusion of capillaries with a diameter smaller than a WBC. Accordingly, we found that WBC circulation velocity in the microvasculature was significantly diminished in diabetic mice (533 ± 169 μm/s vs 960 ± 188 in control mice; P = 0.004) and associated with a decreased percentage of capillaries perfusion by BS-1 lectin (78.5 ± 10.5% vs 96.1 ± 5.7 in control mice; P = 0.03). With the aim to test whether ICAM1 overexpression may be responsible for impaired ischemic foot re-perfusion, diabetic mice were administered with anti-ICAM1 antibodies or isotype control for 14 days, starting 14 days after HLI surgery was performed. We found that anti-ICAM1 therapy significantly increased WBC circulation velocity within the microvasculature (779 ± 113 μm/s vs 447 ± 107 in mice treated with isotype control; P < 0.0001) and the percentage of perfused capillary (P = 0.009). Altogether, our results demonstrate that ICAM1 overexpression may compromise hind limb ischemia recovery in diabetic mice by decreasing WBC circulation velocity and impairing capillary perfusion. [ABSTRACT FROM AUTHOR]

Purpose: To assess the feasibility and safety of concomitant intra-articular (IA) knee injection of mesenchymal stem cells (MSCs) and platelet-rich plasma (PRP) under fluoroscopic guidance to treat patellofemoral osteoarthritis (OA).Materials and Methods: This prospective study included 19 consecutive patients referred for fluoroscopically guided IA MSC and PRP injection for symptomatic patellofemoral chondropathy in which conservative treatment had failed. Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) score and magnetic resonance (MR) data, including T2 mapping sequence, were prospectively collected before and 6 months after treatment. Clinical data without MR imaging were collected until 12 months after the procedure.Results: WOMAC scores were significantly lower after IA injection of MSCs and PRP at 6 months and during 12-months follow-up compared with baseline (mean score decreased from 34.3 to 14.2; P < .0018). Patients reported no complications. Concerning MR imaging follow-up, there were no significant differences in grade, surface, or T2 value of the chondral lesions (P > .375).Conclusions: IA injection of MSCs and PRP in early patellofemoral OA appears to allow functional improvement. [ABSTRACT FROM AUTHOR]

Coronary microvascular disease has been proposed to be responsible for heart failure with preserved ejection fraction (HFpEF) about 10 years ago. However, to date the role and phenotype of the coronary microvasculature have still been poorly considered and investigated in animal models of HFpEF. To characterize the phenotype of the coronary microvasculature in male and female mice with diastolic dysfunction. We assessed cardiac function and characterized the coronary microvasculature in two mouse models of HFpEF: mice fed with a high fat diet (HFD) + L-NAME regimen and Leptin receptor deficient (Leprdb/db) mice. Notably, our study was done in males, females and ovariectomized (OVX) female mice in order to search for possible sexual dysmorphism. Upon a HFD + L-NAME regimen, both males and OVX females but not non OVX females develop diastolic dysfunction attested by an increased end diastolic pressure. In Leprdb/db mice, both male and non OVX female mice develop diastolic dysfunction. Notably, female mice have reduced estradiol and progesterone level mice mimicking ovariectomy. We found that both OVX and non OVX females but not males display increased endothelial activation attested by increased ICAM1 expression, endothelium leakage attested by increased Fibrinogen and IgG extravasation and decreased arteriole diameter suggesting vasoconstriction. The same results were found in Leprdb/db mice. Diastolic dysfunction is not always associated with cardiac small vessel disease since Leprdb/db males and C57BL/6 J males fed with a high fat diet (HFD) + L-NAME regimen develop diastolic dysfunction in the absence of endothelial dysfunction. Also endothelial dysfunction may not be sufficient to induce diastolic dysfunction since non OVX female mice fed with a high fat diet (HFD) + L-NAME regimen display endothelial dysfunction while they do not develop diastolic dysfunction. [ABSTRACT FROM AUTHOR]

Cardiovascular diseases which are the leading cause of death worldwide are promoted by well-known risk factors that include high blood pressure, age, sex, diabetes and/or obesity. Pericytes are mural cells wrapped around microvessels in many organs. They are mostly described as essential for microvascular integrity in the brain and retina. While the detrimental effects of cardiovascular risk factors on endothelial cells have been extensively studied, little is known about their impact on the biology of pericytes, especially in the heart. To determine how cardiac pericytes are modified by some cardiovascular risk factors. To do so, we used mice that were exposed to a high-fat diet (HFD) only to recapitulate obesity, to a HFD together with low dose-streptozotocin injections to mimic type 2 diabetes or to a HFD and L-NAME to recapitulate obesity with a mild hypertension. Mice were sacrificed 3 or 6 months after establishing these risk factors for histological and gene expression analyses. In mice exposed to the HFD + L-NAME regimen for 3 months, the percentage of capillaries covered by pericytes was not modified. However their phenotype appeared to be modified. Indeed, we isolated pericytes (CD31–, CD146+ cells) in the heart of both HFD L-NAME fed mice and control mice using magnetic beads and performed a bulk RNA-sequencing analysis on these cells which identified 3 genes of which the expression was significantly different in the 2 groups. In particular, Angiopoietin-like 4 mRNA expression was increased in the pericytes of HFD + L-NAME fed mice. This protein plays an important role in the regulation of lipid metabolism which makes it attractive to explore. In mice exposed to either HFD alone of HFD + streptozotocin for 6 months, the percentage of capillaries covered by pericytes was significantly diminished suggesting pericyte death. Cardiovascular risk factors, such as obesity and diabetes, may change the phenotype and/or the number of pericytes in the heart suggesting that these cells, known to be critical for microvascular integrity, could contribute to the onset of cardiac diseases. [ABSTRACT FROM AUTHOR]

Lower-limb peripheral artery disease (PAD) is a prevalent and challenging complication of diabetes, requiring innovating therapies. Praliciguat is an orally available stimulator of soluble guanylate cyclase (sGC) reported to have favorable effects on metabolic and hemodynamic endpoints in preclinical and clinical studies, suggesting the potential for benefit in PAD. We evaluated the effect of praliciguat on hindlimb ischemia recovery in a mouse model of diabetes and investigated the molecular mechanism of sGC stimulation. Hindlimb ischemia was induced in db/db mice by ligation and excision of the left femoral artery. Praliciguat 10 mg/kg/day (n = 10 mice) or vehicle (n = 10) were administered in the diet for 31 days starting 3 days before surgery. Foot perfusion was assessed with a Laser Doppler Imager and reported as a ratio in the ischemic versus non-ischemic limb. Ischemic leg function was assessed with a 4-point scale: 0, plantar/toe flexion in response to tail traction; 1, plantar but not toe flexion; 2, no flexion; 3, foot dragging. Ischemic foot perfusion and function were better 28 days after surgery in praliciguat than in vehicle treated mice (mean ± SD: perfusion 1.05 ± 0.23 vs. 0.38 ± 0.16; P < 0.0001; function 1.14 ± 0.36 vs. 1.73 ± 0.80; P = 0.03). Praliciguat did not impact angiogenesis. Arteriolar diameter was higher (9.88 ± 0.96 vs. 8.57 ± 0.64 μm; P = 0.004) and ICAM-1 expression lower (186 ± 41 vs. 403 ± 177% of healthy muscle; P = 0.005) in the ischemic limb of mice treated with praliciguat than with vehicle. Praliciguat attenuated accumulation of oxidative pro-angiogenic and pro-inflammatory muscle fibers. In cultured myoblasts praliciguat significantly downregulated Myh2 and Cxcl12 mRNA expression. Conditioned medium from praliciguat-treated myoblast decreased ICAM-1 mRNA expression in HUVECs. These results suggest that praliciguat restored perfusion and function in the ischemic muscle of db/db mice by increasing arteriolar diameter and decreasing ICAM-1 expression in endothelial cells via downregulation of Cxcl12 in myocytes. [ABSTRACT FROM AUTHOR]

Heart failure with preserved ejection fraction is proposed to be caused by endothelial dysfunction in cardiac small vessels. Our goal is to demonstrate that endothelial dysfunction is sufficient to induce diastolic dysfunction using mice with genetically induced endothelial dysfunction. We used Desert Hedgehog (Dhh) endothelial deficient mice (DhhECKO) as such a model. Indeed, we recently demonstrated that Dhh is critical for endothelial integrity. Practically, DhhECKO mice were administered with tamoxifen at 8 weeks of age to induce Dhh KO and endothelial dysfunction. Mice were sacrificed one month later. As expected, DhhECKO mice exhibited phenotypic changes in their cardiac small vessels characterized by a pro-inflammatory phenotype, a pro-thrombotic phenotype and abnormal endothelium permeability. Notably, these changes are sufficient to induce diastolic dysfunction since DhhECKO mice presented significantly increased end-diastolic pressures, while their left ventricular ejection fraction was comparable to control mice. Moreover, DhhECKO have a reduced exercise tolerance, suggesting a heart failure. Importantly, Aspirin therapy prevented the occurrence of both diastolic dysfunction and exercise intolerance in these mice. Altogether, these results demonstrate that small vessel thrombosis may participate in the pathophysiology of diastolic dysfunction. [ABSTRACT FROM AUTHOR]

Schiattarella et al. introduced a new mouse model mimicking heart failure with preserved ejection fraction (HFpEF). In this model, diastolic dysfunction is induced by a high fat diet (HFD) regimen with L-NAME treatment. Small vessel disease is currently proposed to be responsible for HFpEF. Moreover, blocking Cdon, a negative co-receptor of Hedgehog signalling pathway, in endothelial cells in mice promote endothelium integrity in inflammatory conditions. Consequently, CdonECKO mice represent a good model of endothelial protection. To determine whether prevent endothelial dysfunction prevent diastolic dysfunction. Ovariectomized CdonECKO female mice were fed with HFD from 8 weeks of age. L-NAME treatment was initiated 2 weeks later. Cardiac function was assessed via echocardiography and left ventricle (LV) catheterization at 20 weeks of age. Immediately after, mice were sacrificed for histological and gene expression analyses. Consistently with our previous results, CdonECKO mice treated with HFD and L-NAME display a decreased permeability, a decreased expression of ICAM1 and a decreased infiltration of CD45+ leukocytes compared to their controls. Capillary density and mural cell coverage were not modified. Despite their endothelial improvement, mice displayed increased end diastolic pressure with a preserved LV ejection fraction. Diastolic dysfunction was associated with cardiac hypertrophy and fibrosis as in their controls. Prevention of endothelial dysfunction observed in HFpEF mice models does not prevent diastolic dysfunction in HFD + L-NAME treated mice supporting that small vessel disease is not the only key factor in the pathophysiology of HFpEF. [ABSTRACT FROM AUTHOR]

The article presents a study that determined the role of the inflammatory chemokine osteopontin (OPN) in myocarditis, cardiomyopathy, and heart failure. Results of the study revealed that OPN in the heart instigates in vivo T-cell recruitment and activation causing chronic myocarditis leading into dilated cardiomyopathy. It is concluded that OPN can therefore be a possible therapeutic target as a means to treat heart failure.

Context Angioplasty with stent placement, used to treat symptomatic atherosclerosis plaques, is frequently complicated by restenosis. This pathology is characteried by fibroproliferative and immuno-inflammatory mechanisms of the arterial wall, called intimal hyperplasia (IH). Aims Our previous results demonstrated that phosphoinositide 3-kinase (??(PI3K(??) was a key mediator of inflammatory processes of the arterial wall leading to atherosclerosis. We now proposed this enzyme as a potential clinical target to prevent vascular damages occuring after arterial lesion leading to IH. As cellular and molecular aspects of IH are still incompletely described, we intended to offer a better understanding ofimmune responses involved in this pathology. Results Using a mouse model of arterial mechanical injury, we showed that PI3K(?-deficient mice and mice expressing a catalytically-inactive PI3K(?(PI3K(?KD) showed reduced arterial occlusion and accumulation of monocytes and T cells around site of vascular lesion. The transfer of PI3K(?KD CD4 + T cells into Rag2-deficient mice greatly reduced vascular occlusion compared to WT cells, clearly demonstrating the involvement of PI3K(?in CD4 + T cells during IH formation. In addition, we found that IH is associated with increased levels of Th1 and Th17 cytokines. A specific decrease in the Th1 response was observed in the absence of PI3K(?activity, leading to decreased chemokine production by smooth muscle cells. Finally, we show that a short period of treatment with a PI3K(?inhibitor decreased IH development in arterial mechanical injury in mice but also in a rat carotid artery balloon injury model demonstrating the therapeutic potential of this inhibitor. Conclusion Our work pintpoint PI3K(?as a good target for IH prevention and brings new insights in the immune biology of the disease. [ABSTRACT FROM AUTHOR]

Background In elderly and diabetic patients, the increased risk of developing ischemic disease is associated with impaired regenerative properties of most tissues including skeletal muscle, which make those patients a challenging population to treat. We recently demonstrated that in skeletal muscle, Hedgehog (Hh) signaling promotes ischemia-induced angiogenesis by maintaining peripheral nerve and by promoting myogenesis. The objective of this study is to investigate the functionality of Hh dependent regulation of peripheral nerve survival and myogenesis in aged mice. Methods and Results In the present study, we used 12 week old (young mice) and 20 to 24 month old C57BL/6 mice (old mice) to investigate the activity of Hh signaling in the setting of ischemic skeletal muscle regeneration. In this model, delayed ischemic muscle repair observed in old mice was associated with an impaired upregulation of Gli1. Sonic Hedgehog expression was not different in old mice compared to young mice while Desert Hedgehog (Dhh) expression was downregulated in skeletal muscle of old mice both in healthy and ischemic conditions. Rescue of Dhh expression by gene therapy in old mice increased nerve density and promoted ischemia-induced angiogenesis, nevertheless it failed to promote myogenesis. After further investigation, we found that, in addition to Dhh knockdown, Smoothened (Smo) expression was significantly downregulated in old mice. We used mice in which Hh signaling is specifically disrupted in myocytes (HSA Cre:Smo flox/flox ) and demonstrated that Smo knockdown is sufficient to impair ischemiainduced myogenesis. Conclusion The present study demonstrates that Hh signaling is impaired in aged mice which leads to impaired peripheral nerve survival and myogenesis in acute ischemic stress condition. Moreover, this study brings the new concept that it is necessary to restore both peripheral nerve integrity and myogenesis in elderly to promote revascularisation of ischemic tissues. [ABSTRACT FROM AUTHOR]