Capillary Rarefaction Research Articles

The role of the microcirculation in muscle function and plasticity

It is widely acknowledged that maintenance of muscle, size, strength and endurance is necessary for quality of life and the role that skeletal muscle microcirculation plays in muscle health is becoming increasingly clear. Here we discuss the role that skeletal muscle microcirculation plays in muscle function and plasticity. Besides the density of the capillary network, also the distribution of capillaries is crucial for adequate muscle oxygenation. While capillaries are important for oxygen delivery, the capillary supply to a fibre is related to fibre size rather than oxidative capacity. This link between fibre size and capillary supply is also reflected by the similar time course of hypertrophy and angiogenesis, and the cross-talk between capillaries and satellite cells. A dense vascular network may in fact be more important for a swift repair of muscle damage than the abundance of satellite cells and a lower capillary density may also attenuate the hypertrophic response. Capillary rarefaction does not only occur during ageing, but also during conditions as chronic heart failure, where endothelial apoptosis has been reported to precede muscle atrophy. It has been suggested that capillary rarefaction precedes sarcopenia. If so, stimulation of angiogenesis by for instance endurance training before a hypertrophic stimulus may enhance the hypertrophic response. The microcirculation may thus well be a little-explored target to improve muscle function and the success of rehabilitation programmes during ageing and chronic diseases.

Addition of cyclic angiotensin-(1-7) to angiotensin-converting enzyme inhibitor therapy has a positive add-on effect in experimental diabetic nephropathy

The Renin-Angiotensin System (RAS) possesses a counter-regulatory axis composed of angiotensin converting enzyme (ACE)2, angiotensin-(1-7) [Ang-(1-7)] and the Mas receptor, which opposes many AT1-receptor-mediated effects of ligand angiotensin II. Ang-(1-7), as a ligand of the Mas receptor, has inhibitory effects on renal inflammation and fibrosis in experimental diabetes. However, Ang-(1-7) has a short half-life in plasma, which may render it unsuitable for use in clinics. Here, we investigated the effects of the lanthionine-stabilized Ang-(1-7), cyclic (c)Ang-(1-7), a lanthipeptide that is more peptidase-resistant than the linear peptide, in BTBR ob/ob mice with type 2 diabetic nephropathy. BTBR ob/ob mice received vehicle, cAng-(1-7), or the ACE inhibitor lisinopril. The treatment started at ten weeks of age, when the animals had already developed albuminuria, and ended at 19-20 weeks of age. cAng-(1-7) limited albuminuria progression, and limited podocyte dysfunction similarly to lisinopril. cAng-(1-7), unlike lisinopril, reduced glomerular fibrosis and inflammation, and counteracted glomerular capillary rarefaction. Furthermore, when cAng-(1-7) was combined with lisinopril, a superior antiproteinuric effect than with lisinopril alone was found, in association with better preservation of podocyte proteins and amelioration of capillary density. Thus, adding cAng-(1-7) to ACE-inhibitor therapy could benefit those diabetic patients who do not respond completely to ACE-inhibitor therapy.

Oxygen therapy in premature low birth weight infants is associated with capillary loss and increases in blood pressure: a pilot study

Low birth weight (LBW) and premature birth are known risk factors for future cardiovascular disease and in particular essential hypertension (EH). Capillary rarefaction (CR) is an established hallmark of EH and is known to occur in individuals with a history of LBW. We previously reported that LBW infants do not have CR at birth but rather increased capillary density (CD). We hypothesized that LBW infants undergo a process of accelerated CR in early life, triggered in part by oxygen therapy. We studied 26 LBW infants, of whom 10 infants received oxygen therapy, and compared them to 14 normal birth weight (NBW) infants. We measured CD at 1, 5 and 10 days after birth and again after 40 weeks adjusted gestational age equivalent to birth at full term. We confirmed that LBW infants had higher CD at birth compared to NBW infants and found that significant structural CR occurred at term age in LBW infants who had received oxygen therapy (mean difference −22 capillaries/field, p = 0.007) and in those who did not receive oxygen therapy (mean difference −29 capillaries/field, p < 0.001) compared to baseline at birth. Both LBW groups showed a significant rise in BP at 40 weeks adjusted term age and the rise in systolic (mean difference 24 mm Hg, p < 0.0001) and diastolic BP (mean difference 14 mm Hg, p < 0.001) was more pronounced in the oxygen treated group compared to the nonoxygen group (mean difference 14 mm Hg, p = 0.043 and mean difference = 9 mm Hg p = 0.056 respectively). In conclusion, oxygen therapy in premature LBW infants may induce significant increases in their BP in early life.

FoxO3 activation in hypoxic tubules prevents chronic kidney disease.

Acute kidney injury (AKI) can lead to chronic kidney disease (CKD) if injury is severe and/or repair is incomplete. However, the pathogenesis of CKD following renal ischemic injury is not fully understood. Capillary rarefaction and tubular hypoxia are common findings during the AKI to CKD transition. We investigated the tubular stress response to hypoxia and demonstrated that a stress responsive transcription factor, FoxO3, was regulated by prolyl hydroxylase. Hypoxia inhibited FoxO3 prolyl hydroxylation and FoxO3 degradation, thus leading to FoxO3 accumulation and activation in tubular cells. Hypoxia-activated Hif-1α contributed to FoxO3 activation and functioned to protect kidneys, as tubular deletion of Hif-1α decreased hypoxia-induced FoxO3 activation, and resulted in more severe tubular injury and interstitial fibrosis following ischemic injury. Strikingly, tubular deletion of FoxO3 during the AKI to CKD transition aggravated renal structural and functional damage leading to a more profound CKD phenotype. We showed that tubular deletion of FoxO3 resulted in decreased autophagic response and increased oxidative injury, which may explain renal protection by FoxO3. Our study indicates that in the hypoxic kidney, stress responsive transcription factors can be activated for adaptions to counteract hypoxic insults, thus attenuating CKD development.

Bone marrow pericyte dysfunction in individuals with type 2 diabetes

Aims/hypothesisPrevious studies have shown that diabetes mellitus destabilises the integrity of the microvasculature in different organs by damaging the interaction between pericytes and endothelial cells. In bone marrow, pericytes exert trophic functions on endothelial cells and haematopoietic cells through paracrine mechanisms. However, whether bone marrow pericytes are a target of diabetes-induced damage remains unknown. Here, we investigated whether type 2 diabetes can affect the abundance and function of bone marrow pericytes.MethodsWe conducted an observational clinical study comparing the abundance and molecular/functional characteristics of CD146+ pericytes isolated from the bone marrow of 25 individuals without diabetes and 14 individuals with uncomplicated type 2 diabetes, referring to our Musculoskeletal Research Unit for hip reconstructive surgery.ResultsImmunohistochemistry revealed that diabetes causes capillary rarefaction and compression of arteriole size in bone marrow, without changing CD146+ pericyte counts. These data were confirmed by flow cytometry on freshly isolated bone marrow cells. We then performed an extensive functional and molecular characterisation of immunosorted CD146+ pericytes. Type 2 diabetes caused a reduction in pericyte proliferation, viability, migration and capacity to support in vitro angiogenesis, while inducing apoptosis. AKT is a key regulator of the above functions and its phosphorylation state is reportedly reduced in the bone marrow endothelium of individuals with diabetes. Surprisingly, we could not find a difference in AKT phosphorylation (at either Ser473 or Thr308) in bone marrow pericytes from individuals with and without diabetes. Nonetheless, the angiocrine signalling reportedly associated with AKT was found to be significantly downregulated, with lower levels of fibroblast growth factor-2 (FGF2) and C-X-C motif chemokine ligand 12 (CXCL12), and activation of the angiogenesis inhibitor angiopoietin 2 (ANGPT2). Transfection with the adenoviral vector carrying the coding sequence for constitutively active myristoylated AKT rescued functional defects and angiocrine signalling in bone marrow pericytes from diabetic individuals. Furthermore, an ANGPT2 blocking antibody restored the capacity of pericytes to promote endothelial networking.Conclusions/interpretationThis is the first demonstration of pericyte dysfunction in bone marrow of people with type 2 diabetes. An altered angiocrine signalling from pericytes may participate in bone marrow microvascular remodelling in individuals with diabetes.

Energy Metabolism in the Failing Right Ventricle: Limitations of Oxygen Delivery and the Creatine Kinase System.

Pulmonary arterial hypertension (PAH) results in hypertrophic remodeling of the right ventricle (RV) to overcome increased pulmonary pressure. This increases the O2 consumption of the myocardium, and without a concomitant increase in energy generation, a mismatch with demand may occur. Eventually, RV function can no longer be sustained, and RV failure occurs. Beta-adrenergic blockers (BB) are thought to improve survival in left heart failure, in part by reducing energy expenditure and hypertrophy, however they are not currently a therapy for PAH. The monocrotaline (MCT) rat model of PAH was used to investigate the consequence of RV failure on myocardial oxygenation and mitochondrial function. A second group of MCT rats was treated daily with the beta-1 blocker metoprolol (MCT + BB). Histology confirmed reduced capillary density and increased capillary supply area without indications of capillary rarefaction in MCT rats. A computer model of O2 flux was applied to the experimentally recorded capillary locations and predicted a reduction in mean tissue PO2 in MCT rats. The fraction of hypoxic tissue (defined as PO2 < 0.5 mmHg) was reduced following beta-1 blocker (BB) treatment. The functionality of the creatine kinase (CK) energy shuttle was measured in permeabilized RV myocytes by sequential ADP titrations in the presence and absence of creatine. Creatine significantly decreased the KmADP in cells from saline-injected control (CON) rats, but not MCT rats. The difference in KmADP with or without creatine was not different in MCT + BB cells compared to CON or MCT cells. Improved myocardial energetics could contribute to improved survival of PAH with chronic BB treatment.

Functional capillary impairment in patients with ventricular assist devices

The implantation of continuous – flow ventricular assist devices (VAD) is suggested to evoke angiodysplasia contributing to adverse events such as gastrointestinal bleeding. We evaluated in vivo capillary density and glycocalyx dimensions to investigate possible systemic microvascular changes in patients with chronic heart failure and VAD support vs. standard medical treatment. Forty-two patients with VAD support were compared to forty-one patients with ischemic and non-ischemic chronic heart failure (CHF) on standard pharmacotherapy and to a group of forty-two healthy subjects in a prospective cross-sectional study. Sublingual microcirculation was visualized using Sidestream Darkfield videomicroscopy and functional and perfused total capillary densities were quantified. Patients with VAD implantation were followed for one year and bleeding events were recorded. Median time after VAD implantation was 18 months. Patients were treated with centrifugal-flow devices (n = 31) or axial-flow devices (n = 11). Median functional capillary density was significantly lower in patients with VAD therapy as compared to CHF patients (196 vs. 255/mm2, p = 0.042, adjusted p-value). Functional and total capillary densities were 44% and 53% lower (both p < 0.001) in patients with VAD therapy when compared to healthy subjects. Cox regression analysis revealed loss of capillary density as a significant predictor of bleeding events during one -year follow-up of VAD patients (HR: 0.987, CI (95%): 0.977–0.998, p = 0.021 for functional and 0.992, CI (95%): 0.985–0.999, p = 0.03 for total capillary density). In conclusion, patients with VAD support exhibit capillary density rarefaction, which was associated with bleeding events. If confirmed independently, capillary impairment may be evaluated as novel marker of bleeding risk.

Mitochondrial dysfunction-mediated decline in angiogenic capacity of endothelial progenitor cells is associated with capillary rarefaction in patients with hypertension via downregulation of CXCR4/JAK2/SIRT5 signaling.

BackgroundHypertensive patients exhibit decline in capillary density and endothelial progenitor cells (EPCs). However, whether capillary rarefaction in hypertension is associated with defect angiogenesis of EPCs remains unknown. We hypothesized that impaired mitochondrial function of late EPCs in hypertension is associated with the structural lack of capillary microcirculation via deficient CXCR4/JAK2/SIRT5 signaling.MethodsWe performed capillary microcirculation detection in hypertensive patients and healthy subjects. Angiogenic capacity and mitochondrial function of circulating EPCs were evaluated. The underlying mechanisms were further investigated by genetic inhibition and overexpression.FindingsCapillary density of nail fold and eye fundus were significantly reduced in hypertensive patients, which was paralleled to decreased in vitro late EPC function and in vivo angiogenic capacity. Meanwhile the decline of EPC function in hypertension was accompanied by impaired mitochondrial ultrastructure, diminished mitochondrial membrane potential, reduced oxygen consumption, increased ROS generation and NADH level. Rotenone induced inhibition of oxygen consumption rate, excessive ROS generation and loss of MMP, which markedly decreased the in vitro functions of EPCs. Furthermore, SIRT5 expression of EPCs in hypertension was markedly reduced, which was correlated to mitochondrial dysfunction. CXCR4 gene transfer enhanced SIRT5 expression, improved mitochondrial functions and augmented angiogenic capacity of EPCs. The beneficial impacts of SIRT5 up-regulation on late EPC-mediated angiogenesis can be abrogated by blockade of CXCR4/JAK2/SIRT5 signaling pathway.InterpretationMitochondrial dysfunction-mediated fall in angiogenic capacity due to deficient CXCR4/JAK2/SIRT5 signaling of late EPCs is probably responsible for the capillary rarefaction in hypertension. Our findings provide insight into the potential of EPC mitochondria as a novel target for the treatment of hypertension-related loss of microvascular density.FundsNational Nature Science Foundation of China, 973Program, the Nature Science Foundation of Guangdong.

Right Ventricular Reverse Remodeling after Pulmonary Endarterectomy in Patients with Chronic Thrombo Embolic Pulmonary Hypertension

Purpose Chronic thrombo-embolic pulmonary hypertension (CTEPH) leads to right ventricular (RV) adverse remodeling if left untreated. We recently compared RV features between CTEPH patients and healthy controls. We found that capillary rarefaction, myocardial fibrosis, cardiomyocytes hypertrophy and increased macrophages infiltration were associated with impaired ventricular-arterial coupling, RV dysfunction and increased glucose uptake. Pulmonary endarterectomy (PE) is the treatment of choice for CTEPH, resulting in a marked decrease in RV afterload. In this study, we sought to investigate the correlations between preoperative RV pathological features and postoperative RV function and metabolism. Methods We followed 9 CTEPH patients undergoing PE during which we performed RV biopsies. RV phenotype was assessed using cardiac magnetic resonance, resting echocardiography and 18-Fluoro-Deoxi-Glucose Positron Emission Tomography before PE, and at 1- and 6-months post operatively. Results RV systolic pressures decreased in all patients at 1 month follow-up. Similarly, indices of RV function, remodeling, glucose uptake and coupling as assessed by the Stroke Volume/End Systolic Volume ratio significantly improved at 6 months follow-up. Improvement of RV ejection fraction and RV end systolic volume index were correlated to pre-operative cardiomyocytes hypertrophy (Figure 1). There was a trend for an association between RV fibrosis and moderate improvement in RV systolic function after PE. Preoperative macrophage infiltration and capillary density did not impact on postoperative RV functional recovery. Conclusion After PE, RV phenotype was associated with marked improvement in RV systolic function 6 months, which correlated with pre-operative cardiomyocytes hypertrophy. RV fibrosis was not associated with impaired outcomes and did not limit RV functional recovery in this study. Chronic thrombo-embolic pulmonary hypertension (CTEPH) leads to right ventricular (RV) adverse remodeling if left untreated. We recently compared RV features between CTEPH patients and healthy controls. We found that capillary rarefaction, myocardial fibrosis, cardiomyocytes hypertrophy and increased macrophages infiltration were associated with impaired ventricular-arterial coupling, RV dysfunction and increased glucose uptake. Pulmonary endarterectomy (PE) is the treatment of choice for CTEPH, resulting in a marked decrease in RV afterload. In this study, we sought to investigate the correlations between preoperative RV pathological features and postoperative RV function and metabolism. We followed 9 CTEPH patients undergoing PE during which we performed RV biopsies. RV phenotype was assessed using cardiac magnetic resonance, resting echocardiography and 18-Fluoro-Deoxi-Glucose Positron Emission Tomography before PE, and at 1- and 6-months post operatively. RV systolic pressures decreased in all patients at 1 month follow-up. Similarly, indices of RV function, remodeling, glucose uptake and coupling as assessed by the Stroke Volume/End Systolic Volume ratio significantly improved at 6 months follow-up. Improvement of RV ejection fraction and RV end systolic volume index were correlated to pre-operative cardiomyocytes hypertrophy (Figure 1). There was a trend for an association between RV fibrosis and moderate improvement in RV systolic function after PE. Preoperative macrophage infiltration and capillary density did not impact on postoperative RV functional recovery. After PE, RV phenotype was associated with marked improvement in RV systolic function 6 months, which correlated with pre-operative cardiomyocytes hypertrophy. RV fibrosis was not associated with impaired outcomes and did not limit RV functional recovery in this study.

Astaxanthin ameliorates renal interstitial fibrosis and peritubular capillary rarefaction in unilateral ureteral obstruction

Loss of peritubular capillaries is a notable feature of progressive renal interstitial fibrosis. Astaxanthin (ASX) is a natural carotenoid with various biological activities. The present study aimed to evaluate the effect of ASX on unilateral ureteral obstruction (UUO)-induced renal fibrosis in mice. For that purpose, mice were randomly divided into five treatment groups: Sham, ASX 100 mg/kg, UUO, UUO + ASX 50 mg/kg and UUO + ASX 100 mg/kg. ASX was administered to the mice for 7 or 14 days following UUO. The results demonstrated that UUO-induced histopathological changes in the kidney tissue were prevented by ASX. Renal function was improved by ASX treatment, as evidenced by decreased blood urea nitrogen and serum creatinine levels. Furthermore, the extent of renal fibrosis and collagen deposition induced by UUO was suppressed by ASX. The levels of collagen I, fibronectin and α-smooth muscle actin were increased by UUO in mice or by transforming growth factor (TGF)-β1 treatment in NRK-52E cells, and were reduced by ASX administration. In addition, ASX inhibited the UUO-induced decrease in peritubular capillary density by upregulating vascular endothelial growth factor and downregulating thrombospondin 1 levels. Inactivation of the TGF-β1/Smad signaling pathway was involved in the anti-fibrotic mechanism of ASX in UUO mice and TGF-β1-treated NRK-52E cells. In conclusion, ASX attenuated renal interstitial fibrosis and peritubular capillary rarefaction via inactivation of the TGF-β1/Smad signaling pathway.

The effect of two exercise modalities on skeletal muscle capillary ultrastructure in individuals with type 2 diabetes.

Type 2 diabetes is associated with microvascular dysfunction, but little is known about how capillary ultrastructure is affected by exercise training. To investigate the effect of two types of exercise training on skeletal muscle capillary ultrastructure and capillarization in individuals with type 2 diabetes, 21 individuals with type 2 diabetes were allocated (randomized controlled trial) to 11weeks of aerobic exercise training consisting of either moderate-intensity endurance training (END; n=10) or low-volume high-intensity interval training (HIIT; n=11). Skeletal muscle biopsies (m vastus lateralis) were obtained before and after the training intervention. At baseline, there was no difference in capillarization, capillary structure, and exercise hyperemia between the two groups. After the training intervention, capillary-to-fiber ratio increased by 8%±3% in the END group (P<0.05) and was unchanged in the HIIT group with no difference between groups. Endothelium thickness increased (P<0.05), basement membrane thickness decreased (P<0.05), and the capillary lumen tended (P=0.07) to increase in the END group, whereas these structural indicators were unchanged after HIIT. In contrast, skeletal muscle endothelial nitric oxide synthase (eNOS) increased after HIIT (P<0.05), but not END, whereas there was no change in vascular endothelial growth factor (VEGF), superoxide dismutase (SOD)-2, or NADPH oxidase after both training protocols. In contrast to END training, HIIT did not alter capillarization or capillary structure in individuals with type 2 diabetes. In conclusion, HIIT appears to be a less effective strategy to treat capillary rarefaction and reduce basement thickening in type 2 diabetes.

Endothelial cell mineralocorticoid receptors oppose VEGF-induced gene expression and angiogenesis.

Aldosterone is a key factor in adverse cardiovascular remodeling by acting on the mineralocorticoid receptor (MR) in different cell types. Endothelial MR activation mediates hypertrophy, inflammation and fibrosis. Cardiovascular remodeling is often accompanied by impaired angiogenesis, which is a risk factor for the development of heart failure. In this study, we evaluated the impact of MR in endothelial cells on angiogenesis. Deoxycorticosterone acetate (DOCA)-induced hypertension was associated with capillary rarefaction in the heart of WT mice but not of mice with cell type-specific MR deletion in endothelial cells. Consistently, endothelial MR deletion prevented the inhibitory effect of aldosterone on the capillarization of subcutaneously implanted silicon tubes and on capillary sprouting from aortic ring segments. We examined MR-dependent gene expression in cultured endothelial cells by RNA-seq and identified a cluster of differentially regulated genes related to angiogenesis. We found opposing effects on gene expression when comparing activation of the mineralocorticoid receptor in ECs to treatment with vascular endothelial growth factor (VEGF), a potent activator of angiogenesis. In conclusion, we demonstrate here that activation of endothelial cell MR impaired angiogenic capacity and lead to capillary rarefaction in a mouse model of MR-driven hypertension. MR activation opposed VEGF-induced gene expression leading to the dysregulation of angiogenesis-related gene networks in endothelial cells. Our findings underscore the pivotal role of endothelial cell MR in the pathophysiology of hypertension and related heart disease.

I1-imidazoline receptor-mediated cardiovascular and metabolic effects in high-fat diet-induced metabolic syndrome in rats.

The objective of this study was to investigate the effects of a new I1-imidazoline receptor-selective pyrroline compound on the hemodynamic, metabolic and microvascular alterations in a high-fat diet (HFD)-induced model of metabolic syndrome in rats. In total, twenty adult male Wistar rats were fed a high-fat diet (HFD, n = 20) for 20 weeks. Thereafter, the rats received a new pyrroline compound selective for I1-imidazoline receptors (LNP599; 10 mg/kg/day) or vehicle (n = 10/group) orally by gavage for 4 weeks. Functional microcirculation was assessed using intravital video microscopy, and structural microcirculation was evaluated using histochemical analysis. LNP599 induced concomitant reductions in the SBP, HR and plasma catecholamine levels. The animals treated with this new antihypertensive compound also presented an improvement in body weight and the metabolic parameters related to metabolic syndrome, such as the glucose and lipid profiles. These effects were accompanied by a reversal of the functional and structural capillary rarefaction in the skeletal muscle. The modulation of the sympathetic nervous system by a selective agonist for I1-imidazoline receptors improves the hemodynamic and metabolic parameters in an experimental model of metabolic syndrome. LNP599 can also contribute to the restoration of microcirculatory parameters.

Autophagy in FOXD1 stroma-derived cells regulates renal fibrosis through TGF-β and NLRP3 inflammasome pathway

Renal fibrosis is the final common pathway of various renal injuries and it leads to chronic kidney disease. Recent studies reported that FOXD1-lineage pericyte plays a critical role in tubulointerstitial fibrosis (TIF). However the regulatory mechanisms remain unclear. Autophagy is a cellular process of degradation of damaged cytoplasmic components that regulates cell death and proliferation. To investigate the role of autophagy in FOXD1-lineage pericytes on renal TIF, we generated the FOXD1-lineage stromal cell-specific Atg7 deletion (Atg7△FOXD1) mice. FOXD1-lineage stromal cell-specific Atg7 deletion enhanced renal TIF through Smad-dependent transforming growth factor (TGF)-β signaling after unilateral ureteral obstruction (UUO). FOXD1-lineage stromal cell-specific Atg7 deletion increased the accumulation of interstitial myofibroblasts and enhanced the differentiation of pericytes into myofibroblasts after UUO. Peritubular capillary rarefaction was accelerated in Atg7△FOXD1 mice after UUO. Atg7△FOXD1 mice increased the accumulation of SQSTM1/p62-positive aggregates in the obstructed kidney and resulted in increased expression of NLRP3 inflammasome, interleukin (IL) 1-β and caspase-1 signaling pathway, which enhanced apoptosis of interstitial cells after UUO. In summary, our data showed that autophagy in FOXD1-lineage stromal cells plays a protective role in renal TIF through regulating the Smad4 dependent TGF-β an NLRP3 inflammasome signaling pathway.

Arterial stiffness of the forearm is associated with nailfold capillary count in systemic sclerosis: A novel marker of early vasculopathy?

BackgroundMicrovascular disease, with rarefaction of nailfold capillaries, is the hallmark of systemic sclerosis (SSc). Obliteration of the ulnar and radial artery is regularly observed, implicating involvement of the forearm arteries. Pulse wave velocity (PWV) may serve as early biomarker of forearm artery involvement, before occurrence of irreversible arterial obliteration.ObjectivesThe aim was to investigate arterial stiffness of the aorta and the upper extremities in SSc patients and to correlate these findings with nail-fold capillary count and extent of disease.MethodsAortic PWV was defined as carotid-femoral (cf). Upper extremity PWV was measured as carotid-brachial (cb) and carotid-radial (cr), the ratio between cbPWV/crPWV was used as an indication of the relative PWV change in the forearm. Capillary count was the mean capillary count per 3 mm of 8 fingers. The number of SSc classification criteria was used as surrogate for extent of disease. [1]ResultsIn total, 19 SSc patients (median age 51 years, 68% female) were included. CbPWV/crPWV ratio correlated strongly with capillary count (r = −0.55, p = 0.022, figure 1) in SSc patients, with a trend in regards to its relation with the extent of disease (r = 0.48, p = 0.053).ConclusionsOur findings demonstrate that arterial stiffness of the forearm has a relationship with nailfold capillary count and tends to be associated with the extent of disease in patients with SSc. These may suggest that vascular damage also occurs in larger arteries of the forearm, which potentially serves as novel tool for assessing early vascular involvement in SSc.

Induced pluripotent stem cell-derived endothelial progenitor cells attenuate ischemic acute kidney injury and cardiac dysfunction

BackgroundRenal ischemia–reperfusion (I/R) injury is a major cause of acute kidney injury (AKI), which is associated with high morbidity and mortality. AKI is a serious and costly medical condition. Effective therapy for AKI is an unmet clinical need, and molecular mechanisms underlying the interactions between an injured kidney and distant organs remain unclear. Therefore, novel therapeutic strategies should be developed.MethodsWe directed the differentiation of human induced pluripotent stem (iPS) cells into endothelial progenitor cells (iEPCs), which were then applied for treating mouse AKI. The mouse model of AKI was induced by I/R injury.ResultsWe discovered that intravenously infused iEPCs were recruited to the injured kidney, expressed the mature endothelial cell marker CD31, and replaced injured endothelial cells. Moreover, infused iEPCs produced abundant proangiogenic proteins, which entered into circulation. In AKI mice, blood urea nitrogen and plasma creatinine levels increased 2 days after I/R injury and reduced after the infusion of iEPCs. Tubular injury, cell apoptosis, and peritubular capillary rarefaction in injured kidneys were attenuated accordingly. In the AKI mice, iEPC therapy also ameliorated apoptosis of cardiomyocytes and cardiac dysfunction, as indicated by echocardiography. The therapy also ameliorated an increase in serum brain natriuretic peptide. Regarding the relevant mechanisms, indoxyl sulfate and interleukin-1β synergistically induced apoptosis of cardiomyocytes. Systemic iEPC therapy downregulated the proapoptotic protein caspase-3 and upregulated the anti-apoptotic protein Bcl-2 in the hearts of the AKI mice, possibly through the reduction of indoxyl sulfate and interleukin-1β.ConclusionsTherapy using human iPS cell-derived iEPCs provided a protective effect against ischemic AKI and remote cardiac dysfunction through the repair of endothelial cells and the attenuation of cardiomyocyte apoptosis.

Canonical BMP signaling in tubular cells mediates recovery after acute kidney injury

Bone morphogenetic protein (BMP) signaling has been shown to modulate the development of renal fibrosis in animal models of kidney injury, but the downstream mediators are incompletely understood. In wild-type mice, canonical BMP signaling mediated by SMAD1/5/8 transcription factors was constitutively active in healthy renal tubules, transiently down-regulated after ischemia reperfusion injury (IRI), and reactivated during successful tubular regeneration. We then induced IRI in mice with a tubular-specific BMP receptor 1A (BMPR1A) deletion. These mice failed to reactivate SMAD1/5/8 signaling in the post-ischemic phase and developed renal fibrosis after injury. Using unbiased genomic analyses, we identified three genes encoding inhibitor of DNA-binding (ID) proteins (Id1, Id2, and Id4) as key targets of BMPR1A-SMAD1/5/8 signaling. BMPR1A-deficient mice failed to re-induce these targets following IRI. Instead, BMPR1A-deficiency resulted in activation of pro-fibrotic signaling proteins that are normally repressed by ID proteins, namely, p38 mitogen-activated protein kinase and cell cycle inhibitor p27. These data indicate that the post-ischemic activation of canonical BMP signaling acts endogenously to repress pro-fibrotic signaling in tubular cells and may help to prevent the progression of acute kidney injury to chronic kidney disease.

Aristolochic acid inhibits Slit2-induced migration and tube formation via inactivation of Robo1/Robo2-NCK1/NCK2 signaling pathway in human umbilical vein endothelial cells

Robo1/Robo2-NCK1/NCK2 signaling pathway controls endothelial cell sprouting and migration induced by Slit2 or VEGF, but whether it is involved in peritubular capillary (PTC) rarefaction of Aristolochic acid nephropathy (AAN) is unclear. In the present study, we evaluated whether AA exerts antiangiogenic effects by targeting this signaling pathways in HUVECs. HUVECs or lentivirus-mediated NCK1-overexpressing HUVECs were stimulated with AA (1, 2 or 3 μg/ml) in the absence or presence of 6 nM Slit2. Our results showed that AAІ (1–3 μg/ml) dose-dependently inhibited the migration and tube formation of HUVECs. This inhibition was in parallel with down-regulated mRNA and protein expression of Slit2/Robo1/Robo2-NCK1/NCK2 signaling pathway. Importantly, overexpression of NCK1 rescued AAІ-impaired angiogenesis, as evidenced by the increase of cell migration and tube formation of HUVECs in response to Slit2. The down-regulation of NCK2 and decreased activation of Rac1 was also restored by overexpression of NCK1. Taken together, our findings show that AA inhibits Slit2-induced migration and tube formation via inactivation of Robo1/Robo2-NCK1/NCK2 signaling pathway in HUVECs, and NCK1 might be a potential agent for vascular remodeling in AAN and diseases associated with impaired angiogenesis.

A5335 Early Vascular Parameters in the Micro- and Macrocirculation in Type 2 Diabetes

Objectives: Diabetes converts from a metabolic disorder into a predominantly vascular disease, once its duration extends over several years or/and when additional cardiovascular risk factors coexist. In a large cross-sectional analysis we analyzed various vascular parameters in the renal, retinal and systemic circulation, with the goal to identify which vascular parameter of early organ damage is the earliest that can be clinically detected. <ST>Methods:</ST> In 111 patients with type 2 diabetes (T2DM) and 54 subjects without T2DM we assessed urinary albumin creatinine ratio (UACR) and estimated glomerular filtration rate (eGFR), retinal capillary flow (RCF), intercapillary distance (ICD) as parameters of capillary rarefaction, wall-to-lumen ratio (WLR) of the retinal arterioles[all assessed by Scanning Laser Doppler Flowmetry], and central systolic pressure (cSBP) and central pulse pressure (cPP)[measured by pulse wave analysis, Syphygmocor] both reflecting vascular stiffness of large arteries. <ST>Results:</ST> Compared to subjects without T2DM, patients with T2DM (duration: mean 63.9 ± 56.4 months) were older (59.8 ± 7.3vs.43.4 ± 12.9 years, p < 0.001), more females (33.3vs20.4 %, p < 0.001) but 24-hour systolic and diastolic blood pressure did not differ between the two groups (129.3 ± 11.4/78.9 ± 8.3vs.130.4 ± 10.8/77.4 ± 5.6 mmHg). The analysis adjusted for age, gender and cardiovascular risk factors showed that ICD, cPP were significantly higher and eGFR was significantly lower in patients with T2DM than in subjects without T2DM. <ST>Conclusion:</ST> These data suggest that at similar blood pressure capillary rarefication in the retinal circulation (ICD), eGFR in the renal circulation and central pulse pressure (cPP) of large arteries are earlier detectable than vascular remodeling of the micro-(WLR, RCF, UACR) and macrocirculation (cSPB) in patients with T2DM.

Mitochondrial Dysfunction-Mediated Decline in Angiogenic Capacity of Circulating Endothelial Progenitor Cells is Associated with Capillary Rarefaction in Patients with Hypertension

Hypertensive patients exhibit decline in capillary density and endothelial progenitor cells (EPCs). However, whether capillary rarefaction in hypertension is associated with defect angiogenesis of EPCs remains unknown. We hypothesized that impaired mitochondrial function of late EPCs in hypertension is associated with the structural lack of capillary microcirculation via deficient CXCR4/JAK2/SIRT5 signaling. Capillary density of nail fold was significantly reduced in hypertensive patients, which was paralleled to decreased in vitro late EPC function and in vivo angiogenic capacity. Meanwhile the decline of EPC function in hypertension was accompanied by impaired mitochondrial ultrastructure, diminished mitochondrial membrane potential, reduced oxygen consumption, increased ROS generation and NADH level. Furthermore, SIRT5 expression of EPCs in hypertension was markedly reduced, which was correlated to mitochondrial dysfunction. CXCR4 gene transfer enhanced SIRT5 expression, improved mitochondrial functions and augmented angiogenic capacity of EPCs. The beneficial impacts of SIRT5 up-regulation on late EPC-mediated angiogenesis can be abrogated by blockade of CXCR4/JAK2/SIRT5 signaling pathway. Mitochondrial dysfunction-mediated fall in angiogenic capacity due to deficient CXCR4/JAK2/SIRT5 signaling of late EPCs is probably responsible for the capillary rarefaction in hypertension. Our findings provide insight into the potential of EPC mitochondria as a novel target for the treatment of hypertension-related loss of microvascular density. Funding: This work was supported by the National Nature Science Foundation (31530023, 81500205, 81671379) of China, 973 Program (2013CB531200), the Nature Science Foundation (2016A030310184, 2016A020215056) of Guangdong and Science and Technology Planning Project (2017A020215085) of Guangdong. Declaration of Interest: None. Ethical Approval: The study protocol was approved by the Ethics Committee of the First Affiliated Hospital of Sun Yat-sen University (Guangzhou, China).