Skeletal Muscle Blood Vessels Research Articles

Vasculogenic Mesodermal Cells Derived From Induced Pluripotent Stem Cells Improve Limb Perfusion and Decrease Necrosis in Murine Models of Hindlimb Ischemia

Critical limb-threatening ischemia (CLTI) is a national health care priority as there has been a 50% increase in amputations from 2010 to 2018. Unlike other cell products that simply secrete proangiogenic growth factors, induced pluripotent stem cell (iPSC)-derived mesodermal cells have the capacity to differentiate into endothelial cells and smooth muscle cells and form functioning blood vessels in vivo. Here we discuss the effects of intramuscular injection of human iPSC mesodermal cells (VSC100) on limb perfusion and limb necrosis. iPSCs were produced using methods including Yamanaka factors (Broxmeyer et al [21393480]). The iPSC cells were differentiated using sequential Activin A, BMP4, vascular endothelial growth factor, and fibroblast growth factor 2 to promote mesodermal and endothelial cell differentiation. Murine CLTI models were created via excision of the common femoral artery in male BALB-C Nude mice, aged 6 to 8 weeks. After creation of the CLTI model mice were randomized to 1 × 106 VSC100 cells labeled with td-tomato (red fluorescence) or vehicle control injected into the gastrocnemius and gracilis muscles (n = 10/group) at 7 to 14 days postinduction of ischemia. Limb perfusion was measured with laser speckle contrast imaging up to day 64, qualitative analysis of VSC100 differentiation into capillaries was assessed with immunohistochemical staining for isolectin B4 and human CD31 and confocal microscopy to detect td-tomato. Degree of tissue loss of the index limb was quantified with established necrosis scores. VSC100-treated BALB-C nude mice showed a significant increase in limb perfusion as compared to vehicle controls with average percent blood flow/contralateral limb ratios of 45% in treated groups compared to 25% in control (P < .001) at day 64 (Fig 1). Limb necrosis scores showed similar results, in that necrosis was statistically significantly reduced in iPSC-treated vs vehicle control treated groups (Fig 2). Immunohistochemical analysis showed qualitative significant deposition of cells of interest in our CLTI models. iPSC-derived VSC100 is a novel approach to treating CTLI as the cells are capable of forming functioning blood vessels in ischemia skeletal muscle, improve tissue perfusion, and mitigate tissue necrosis. These early results and data support further translational work to support development of an investigational new drug for future clinical trial studies.Fig 2Clinical scoring ratio of chronic limb-threatening ischemia (CLTI) in control vs treated groups. Mouse population same as in Fig 1 (n = 10/group).View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Abstract 547: Vasculogenic Mesodermal Cells Derived From Induced Pluripotent Stem Cells Improve Limb Perfusion And Decrease Necrosis in Murine Models of Hindlimb Ischemia

Objectives: Critical Limb Threatening Ischemia (CLTI) is a health care priority with a 50% increase in amputations from 2010 to 2018. Unlike other cell products that simply secrete pro-angiogenic growth factors Induced pluripotent stem cells (iPSC) derived mesodermal cells can differentiate into endothelial and smooth muscle cells and form functioning blood vessels in vivo . We discuss the effects of intramuscular injection of human iPSC mesodermal cells (VSC100) on limb perfusion and limb necrosis. Methods: IPSCs were produced using methods including Yamanaka factors (Broxmeyer H. et al [21393480}). iPSC cells were differentiated using sequential Activin A, BMP4, VEGF, and FGF2 to promote mesodermal and endothelial cell (EC) differentiation. Murine CLTI models were created via excision of the common femoral artery in male BALB-C Nude mice, aged 6-8 weeks. After creation of the CLTI model mice were randomized to 1x10 6 VSC100 cells labeled with td-tomato (red fluorescence) or vehicle control injected into the gastrocnemius and gracilis muscles (N=10/group) at 7-14 days postinduction of ischemia. Limb perfusion was measured with Laser Speckle Contrast Imaging (LSCI) up to Day 64, qualitative analysis of VSC100 differentiation into capillaries was assessed with immunohistochemical staining for Isolectin B4 and human CD31 and confocal microscopy to detect td-tomato. Degree of tissue loss of the index limb was quantified with established necrosis scores. Results: VSC100 treated mice showed a significant increase in limb perfusion compared to controls, average percent blood flow/contralateral limb ratios of 45% in treated groups compared to 25% in control (p &lt; 0.001) at day 64 (Figure 1). Limb necrosis scores showed necrosis was statistically significantly reduced in iPSC treated versus vehicle control treated groups (Figure 2). Immunohistochemical analysis showed qualitative significant deposition of cells of interest in our CLTI models. Conclusions: iPSC derived VSC100 is a novel approach to treating CTLI as the cells can form functioning blood vessels in ischemic skeletal muscle, improve tissue perfusion, and mitigate tissue necrosis. These early results are being used for FDA applications for clinical trials utilizing iPSCs in CLTI patients.

Effects of Exercise Training on PPARβ/δ Expression in Skeletal Muscle of Rats with Spontaneous Hypertension.

ABSTRACTPurposeThis study aimed to identify the relationship and mechanism between skeletal muscle peroxisome proliferator–activated receptor β/δ (PPARβ/δ) and spontaneous hypertension.MethodsRats were divided into four groups (n = 10): spontaneous hypertensive rats exercise group (SHR-E), spontaneous hypertensive rats sedentary group (SHR-S), Wistar-Kyoto control rats exercise group (WKY-E), and Wistar-Kyoto control rats sedentary group (WKY-S). Although the sedentary groups were placed on the treadmill without moving during the training sessions, the exercise groups were forced to run on a treadmill for 8 wk, 1 h·d−1, 5 d·wk−1. After training, the density and area of gastrocnemius microvessels were observed. PPARβ/δ, vascular endothelial growth factor A (VEGFA), superoxide dismutase 2 (SOD-2), and nitric oxide synthase in gastrocnemius were measured by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot.ResultsExcept the sixth week of age, the systolic blood pressure of SHR-S was significantly higher than that of WKY-S at all time periods. Exercise significantly reduced systolic blood pressure in SHR rats. Compared with the SHR-S group, the WKY-S group had significantly higher PPARβ/δ protein level and density of skeletal muscle microvessels. Eight weeks of exercise increased the PPARβ/δ, SOD-2, VEGFA, and microvessel density and area in the skeletal muscle of SHR.ConclusionsExercise training promoted PPARβ/δ mRNA and protein-level expression of PPARβ/δ, SOD-2 and VEGFA in skeletal muscle, thus increasing the density and area of skeletal muscle blood vessels. These regulations contribute to the reduction of peripheral vascular resistance. This may be a potential mechanism of exercise to reduce blood pressure.

EP.125SCN4A mutation causes episodic myotonia and persistent paralysis with internalized blood vessels in skeletal muscle

EP.125SCN4A mutation causes episodic myotonia and persistent paralysis with internalized blood vessels in skeletal muscle

Sex‐specific Conditions Affecting Vascular Tone, Cerebral Blood Flow and Cognition

There are sex and sex‐hormone specific differences in autonomic nervous system control of blood pressure. The sympathetic nerve fibers of the autonomic nervous system innervate skeletal muscle blood vessels, initiating vasoconstriction, and thus regulate blood flow and vascular tone. This regulation of vascular tone is different between men and women, such that estrogen increases the activity of the b2‐adrenergic receptor activity (causing vasodilation) which offset sympathetically‐mediated vasoconstriction and results in lower blood pressure values in young women. Higher sympathetic nerve activity is associated with greater augmented aortic blood pressure in young men, but lower augmented aortic blood pressure in young women. Such differences in blood pressure regulation will likely have effects that extend beyond blood pressure, affecting blood flow, and ultimately tissue structure and function. For example, higher aortic blood pressure and greater arterial stiffness of the central arteries are associated with white matter hyperintensity fraction in normotensive postmenopausal women and older adults. In addition, there are conditions unique to women, menopause and pregnancy, that further affect blood pressure and blood flow regulation. Pregnancy constitutes a stress that challenges the regulation of blood pressure and blood flow in women. Hypertensive conditions of pregnancy, such as preeclampsia, carry life‐long risk for developing cardiovascular and cerebrovascular diseases. We evaluated the influence of pregnancy history in postmenopausal women on vascular function, cerebral blood flow, and cognition 35 years after pregnancy. Aortic blood pressure was associated with lower cognitive function scores in women with a history of preeclampsia. Additionally, women with a history of preeclampsia had lower cerebral blood flow responses to a vasodilatory stimulus and this was associated with greater intravascular cellular activation. Furthermore, women with a history of preeclampsia and with late‐life hypertension had atrophy in specific regions of the brain, suggesting that pregnancy history may later impact brain structure. Therefore, conditions that a woman may experience during pregnancy may be informative for future risk of disease. In summary, understanding sex‐specific physiological mechanisms, and how this may change over the lifespan is crucial for identification and treatment of individuals at accelerated risk of cardiovascular and cerebrovascular disease.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Application of recombinant peroxisome proliferator-activated receptor-γ coactivator-1α mediates neovascularization in the retina.

Peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) is able to induce the expression of vascular endothelial growth factor (VEGF), promoting the formation of new blood vessels in skeletal muscle. The aim of the current study was to determine whether PGC-1α is able to regulate angiogenesis in human retinal vascular endothelial cells (hRVECs) in vitro and in retinas in vivo. hRVECs treated with recombinant PGC-1α were incubated for 24 h and then placed into a normoxic (20% O2) or hypoxic (1% O2) environment for a further 16 h. Following this, VEGF mRNA and protein levels were significantly increased. Cellular proliferation was enhanced by treatment with recombinant PGC-1α in normoxic and hypoxic conditions. At 24 h following recombinant PGC-1α treatment, hRVECs were plated into Matrigel-coated plates and cultured under normoxic (20% O2) or hypoxic (1% O2) conditions for a further 24 h. Recombinant PGC-1α-treated cells were observed to form significantly greater numbers of tubes. In a C57BL/6J mouse model of ischemic retinopathy, mice received an intravitreal injection of recombinant PGC-1α, resulting in a significant increase in VEGF mRNA and protein levels in the retina. Retinal neovascular tufts and neovascular nuclei were investigated by angiographic and cross-sectional analysis and were observed to be significantly increased in the PGC-1α group compared with the control group. These results indicate that PGC-1α is able to induce angiogenesis in hRVECs and retinas, and suggests that PGC-1α is a potential anti-angiogenic target in retinal neovascularization.

Hindlimb unweighting does not alter vasoconstrictor responsiveness and nitric oxide-mediated inhibition of sympathetic vasoconstriction.

Physical inactivity increases the risk of cardiovascular disease and may alter sympathetic nervous system control of vascular resistance. Hindlimb unweighting (HU), a rodent model of physical inactivity, has been shown to diminish sympathetic vasoconstrictor responsiveness and reduce NO synthase expression in isolated skeletal muscle blood vessels. Our understanding of the effects of HU on sympathetic vascular regulation in vivo is very limited. The present findings demonstrate that HU did not alter sympathetic vasoconstrictor responsiveness and NO-mediated inhibition of sympathetic vasoconstriction in resting and contracting skeletal muscle. This study suggests that short-term physical inactivity does not alter in vivo sympathetic vascular control in the skeletal muscle vascular bed at rest and during contraction. We tested the hypothesis that physical inactivity would increase sympathetic vasoconstrictor responsiveness and diminish NO-mediated inhibition of sympathetic vasoconstriction in resting and contracting skeletal muscle. Sprague-Dawley rats (n=33) were randomly assigned to sedentary time control (S) or hindlimb unweighted (HU) groups for 21days. Following the intervention, rats were anaesthetized and instrumented for measurement of arterial blood pressure and femoral artery blood flow and stimulation of the lumbar sympathetic chain. The percentage change of femoral vascular conductance (%FVC) in response to sympathetic chain stimulation delivered at 2 and 5Hz was determined at rest and during triceps surae muscle contraction before (control) and after NO synthase blockade with l-NAME (5mgkgi.v.). Sympathetic vasoconstrictor responsiveness was not different (P>0.05) in S and HU rats at rest (S, 2Hz, -26±8% and 5Hz, -46±12%; and HU, 2Hz, -29±9% and 5Hz, -51±10%) and during contraction (S, 2Hz, -10±7% and 5Hz, -23±11%; and HU, 2Hz, -9±5% and 5Hz, -22±7%). Nitric oxide synthase blockade caused a similar increase (P>0.05) in sympathetic vasoconstrictor responsiveness in HU and S rats at rest (S, 2Hz, -41±7% and 5Hz, -58±8%; and HU, 2Hz, -43±6% and 5Hz, -63±8%) and during muscle contraction (S, 2Hz, -15±6% and 5Hz, -31±11%; and HU, 2Hz, -12±5% and 5Hz, -29±8%). Skeletal muscle NO synthase expression and ACh-mediated vasodilatation were also not different between HU and S rats. These data suggest that HU does not alter sympathetic vasoconstrictor responsiveness and NO-mediated inhibition of sympathetic vasoconstriction in resting and contracting skeletal muscle.

Cardiac arrest in athletes and resuscitation.

Participation in competitive sports is not completely free from risk. The death rate for marathon runners in London is approximately 1 in 80,000 finishersand among high school athletes playing competitive sport in the USA, it is approximately 1 in 200,000. Although rare, because they are so unexpected, these deaths hugely affect not only the family but the wider community as well. Kim et al. reported a survival rate of just 29% following cardiac arrest in long distance runners and this dropped to only 7% in those younger than 40 years of age. One likely reason for this is the higher incidence of undiagnosed hypertrophic cardiomyopathy (HCMP) in younger athletes. However, cardiac arrest during competitive sports is also complicated by several physiological factors, which we believe should be addressed during resuscitation. Compared to the oxygen consumption of about 250mlmin 1 at rest, the oxygen consumption in a male marathon runner can reach 5100mlmin . During heavy exercise, within a few minutes athletes develop oxygen debt, which has to be ‘repaid’ once the exercise is over. Hence, during the recovery period for about 30–40min, the oxygen uptake remains considerably higher than normal. The temperature of athletes also rises and may reach up to 40 C or even higher. A cardiac arrest in such a case would worsen the metabolic acidosis much faster than in a resting person. Therefore, establishing adequate ventilation and oxygenation promptly could be a critical factor in athletes. Presently, Advanced Life Support (ALS) guidelines do not have any specific recommendations for doing cardiopulmonary resuscitation (CPR) in athletes. In a study on out of hospital cardiac arrests, Wang et al. reported that endotracheal intubation (ETT) was associated with improved outcomes compared to supraglottic airways. It is conceivable that early use of ETT may be even more important in athletes. During extreme exercise, athletes can lose up to 3–5 kg of their weight in an hour largely because of sweating. Their skeletal muscle blood vessels are also maximally dilated. This degree of dehydration and vasodilatation may create a relative deficit of circulatory volume and contribute to failure to resuscitate during CPR. In a situation such as this, a rapid intravenous infusion of 1–2 l of fluids such as Hartmann’s solution can improve circulatory volume, help in cooling and may improve the chances of a successful resuscitation. Third, since HCMP could be a cause of cardiac arrest in young athletes, use of magnesium may prove to be beneficial in these cases. In HCMP, sarcoplasmic mutations alter calcium balance by increasing the affinity of troponin C for calcium in sarcoplasm, resulting in increased calcium levels during diastole. Sarcomeric mutations also increase the energy requirements of myosin ATPase and reduce the activity of other ATP consuming processes such as ion pumps. During cardiac arrest, presence of sustained hypoxia causes a failure to extrude calcium from cytosol which is a highly energy-dependent process. It is easy to see how this process is likely to be amplified in presence of HCMP. Magnesium is a physiological antagonist of calcium and drives calcium into the sarcoplasmic reticulum, reduces mitochondrial overload and competes with calcium for binding to troponin C. It also increases the threshold stimulus required to provoke ventricular arrhythmias. Although magnesium has not been shown to improve outcome in the general population having cardiac arrest, perhaps in athletes, where the underlying cause may be undetected HCMP, early intravenous administration of 4–5 g magnesium could be beneficial and worth an investigation. There is a need for further research to improve outcome in this group. Present ALS guidelines do not have specific recommendations for athletes suffering cardiac arrest on the field. We believe that cardiac arrest in athletes should be given specific consideration in the ALS guidelines.

Sixty Years since Vasopressin Synthetic Preparation Followed by a Nobel Prize. Is it Time to Augment its Use in Cardiac Surgery?

hormone”. This hormone has been shown to have effects within minutes when administrated intravenously. AVP rapidly distributes from plasma into the extracellular fluid volume. It is metabolized by the liver and the kidneys, and a small proportion is eliminated with the urine. The plasma half-life is very short (4-20 min), resulting in the necessity of continuous infusion in order to maintain its effects. Vasopressin causes vasoconstriction in skin, skeletal muscle, and mesenteric blood vessels mediated via V1 receptors. Nonetheless, some studies have also suggested additional vasodilatory effects at low concentrations in selected vascular beds, including coronary, pulmonary, and cerebral arteries. Circulating AVP, acting via specific V1 receptors in the area postrema, modulates central cardiovascular regulation by augmenting the baroreflex inhibition of efferent sympathetic nerve activity, and therefore counterbalances the caused increase in peripheral resistance. Moreover, there is growing evidence for the existence of AVP receptors located on the presynaptic terminals of central sympathetic efferent nerves in the spinal cord, the stimulation of which may decrease sympathetic excitability. For more than 25 years vasopressin indications were restricted to treating polyuria in patients with diabetes insipidus, reducing blood loss in patients with gastrointestinal bleeding and increasing the mobility of the gastrointestinal system. It was only in the 1990s that it was first used for its potent vasopressor effects in patients with cardiac arrest [2], septic shock [3], drug intoxication [4] and for surgery of carcinoid tumors [5]. Since then there are several studies that have demonstrated the augmented vasoconstrictor action of vasopressin, which can persist for up to 2 h, in patients with hypotension not responding to high-dose norepinephrine, dopamine and fluid resuscitation [5,6]. Another important advantage of vasopressin is the less prominent vasoconstriction in the coronary and cerebral circulation compared to the rest of the circulation. In addition to the double beneficial action of vasopressin in the myocardium and brain, there is a protective action to the kidneys as well. The increased urine output is a remarkable result of infused vasopressin that, according to several studies, is caused by the increased MAP which, consequently, improves the glomerular filtration rate [7].

Defective Homocysteine Metabolism: Potential Implications for Skeletal Muscle Malfunction

Hyperhomocysteinemia (HHcy) is a systemic medical condition and has been attributed to multi-organ pathologies. Genetic, nutritional, hormonal, age and gender differences are involved in abnormal homocysteine (Hcy) metabolism that produces HHcy. Homocysteine is an intermediate for many key processes such as cellular methylation and cellular antioxidant potential and imbalances in Hcy production and/or catabolism impacts gene expression and cell signaling including GPCR signaling. Furthermore, HHcy might damage the vagus nerve and superior cervical ganglion and affects various GPCR functions; therefore it can impair both the parasympathetic and sympathetic regulation in the blood vessels of skeletal muscle and affect long-term muscle function. Understanding cellular targets of Hcy during HHcy in different contexts and its role either as a primary risk factor or as an aggravator of certain disease conditions would provide better interventions. In this review we have provided recent Hcy mediated mechanistic insights into different diseases and presented potential implications in the context of reduced muscle function and integrity. Overall, the impact of HHcy in various skeletal muscle malfunctions is underappreciated; future studies in this area will provide deeper insights and improve our understanding of the association between HHcy and diminished physical function.

The significance of an isoprenaline-like metabolite in the interpretation of the responses of blood vessels in skeletal muscle to adrenaline.

Abstract A depressor substance, which resembled isoprenaline in Rf value, was demonstrated after chromatography of plasma collected from chloralose-anaesthetised cats during the blood pressure responses to intravenous injections of adrenaline, but the results obtained did not substantiate the hypothesis of a functional metabolite. The amount of vasodilatation that the “metabolite” would have caused in the skeletal muscles of the hind limb could not, for example, be correlated with the increases in blood flow produced by the original intravenous doses of adrenaline. The fact that adrenaline was largely vasoconstrictor in acutely denervated hind limbs was taken to indicate that the original vasodilator effects were of reflex nervous origin. It is suggested that the afferent source of this and related reflexes involves the stimulation of chemoreceptors as opposed to mechanoreceptors and that increases in flow through denervated muscles in response to adrenaline are caused by a direct action on the walls of the blood vessels.

Hypoxia and reoxygenation-induced oxidant production increase in microvascular endothelial cells depends on connexin40

Connexins (Cx) are recognized as structural constituents of gap-junctional intercellular communication (GJIC). However, their function may extend beyond facilitating the exchange of metabolites and electrical signals between cells. In this study we asked if increased production of reactive oxygen species (ROS) in microvascular endothelial cells challenged by hypoxia/reoxygenation (H/R) requires Cx40, independent of GJIC. Because we showed that this ROS increase depends on NADPH oxidase, we also asked if Cx40 function (i.e., Cx40-dependent reduction in interendothelial electrical coupling after H/R) requires NADPH oxidase. ROS increase was assessed in confluent monolayers of cultured endothelial cells derived from skeletal muscle blood vessels of wild-type (WT) and Cx40 −/− mice and in monolayers of GJIC-deficient SKHep1 cells overexpressing GFP-tagged Cx40. Electrical coupling was assessed in WT cells and in cells lacking the NADPH oxidase subunit gp91phox or p47phox. H/R elicited a 70–80% ROS increase in WT but not in Cx40 −/− cells. The increase was not affected by the gap junction blocker 18α-glycyrrhetinic acid or by preventing the cells from establishing cell-to-cell contact. H/R increased ROS in SKHep1 cells expressing Cx40–GFP, but not in cells expressing the control vector. Finally, H/R reduced electrical coupling in WT and gp91phox −/− but not in p47phox −/− cells. Our data indicate that (i) the H/R-induced ROS increase in microvascular endothelial cells requires Cx40, independent of its role in GJIC, and (ii) p47phox rather than NADPH oxidase-derived ROS affects modulation of intercellular coupling. Together, the results raise an intriguing possibility that H/R-induced signaling in endothelial cells involves a cross-talk between Cx40 and NADPH oxidase.

Not so free associations

According to PubMed, there have been 62 872 genetic association studies published, about 2 for every gene in the human genome. That assessment is of course incorrect. Both the notorious insertion/deletion polymorphism in the angiotensin-converting enzyme (ACE) gene and polymorphisms in the angiotensinogen gene have representative papers numbering in the thousands. However, the adrenergic receptors have not been sleeping in this regard; their polymorphism paper number currently stands at 1334 reports. In this issue of Nephrol Dial Transplant appears the next such paper. Fung et al. [1] reported that adrenergic beta1 receptor (ADB1R) genetic variation predicts the rate of estimated glomerular filtration rate (eGFR) decline in participants of the African American Study of Kidney Disease and Hypertension (AASK) study cohort. The authors obtained a similar result in a second cohort from San Diego, CA, USA, termed the ‘replication’ cohort. Do we now have the genetic basis for the decline in eGFR over time in patients with hypertension-induced kidney damage? AASK was a (US National Institutes of Health sponsored) drug study in black Americans [2]. The study was heroic in that participants underwent a renal biopsy to verify that they indeed had hypertension-associated nephrosclerosis. The study compared two levels of blood pressure reduction and three antihypertensive drug classes and tested the effect on eGFR. Briefly, AASK detected no additional benefit of slowing progression of hypertensive nephrosclerosis with the lower blood pressure goal. The authors stated that ACE inhibitors appeared to be more effective than beta-blockers or dihydropyridine calcium channel blockers in slowing GFR decline. However, they also admitted that none of the drug group comparisons showed consistent significant differences in the eGFR slope. Disappointing surely for the participants was the fact that in many AASK patients, eGFR declined relentlessly although they controlled their blood pressure to contemporary guideline goals. eGFR declined relentlessly even though they did everything their doctors asked. With such a result, we could conclude either that blood pressure control and drug classes are not as important as we all believe or that something else (genes for instance) is at fault. Large drug trials, irrespective of the sponsors, are favourite for polymorphism studies and most, if not all, feature one or more genetic ‘spin-off’ studies. For instance, the Antihypertensive and Lipid-Lowering Treatment to Prevent Heart Attack Trial (ALLHAT) featured an entire polymorphism consortium disingenuously entitled GENHAT. This group published the presence of an absence of an interaction between the ACE gene insertion–deletion polymorphism and pravastatin in cardiovascular disease in ALLHAT patients [3]. The AASK study has already been milked for polymorphisms. A paper on G-protein-coupled receptor kinase 4 polymorphisms and response to metoprolol in the AASK cohort has recently been published. Several authors on the current report also appear on that paper [4]. Why did the authors pick ADB1R? The hypothesis is not directly stated. However, the senior author is specifically interested in adrenergic mechanisms in hypertension and perhaps that is the reason. Did the authors make a good choice in selecting polymorphisms? Yes, they did. First, adrenergic tone is decidedly increased with diminished renal function. The issue has been studied with direct measurement of muscle sympathetic nerve activity. Converse et al. recorded the rate of postganglionic sympathetic-nerve discharge to the blood vessels in skeletal muscle by means of microelectrodes inserted into the peroneal nerve in 18 patients with native kidneys who were undergoing long-term treatment with haemodialysis, 5 patients receiving haemodialysis who had undergone bilateral nephrectomy and 11 normal subjects [5]. They showed unequivocally that chronic renal failure is accompanied by reversible sympathetic activation, which appears to be mediated by an afferent signal arising in the failing kidneys. The literature on the issue is extensive. A recent review outlines all the findings in detail [6]. Too bad that the sympatholytic drugs are not renoprotective. Metoprolol, a predominantly ADB1R blocker, did not seem to help the AASK participants very much. The authors got their best mileage out of the Ser49Gly polymorphism in ADB1R, with minimal decline in persons homozygous for Gly49Gly. This finding is a bit surprising, because the most highly touted polymorphism, at least for a risk of heart failure in ADB1R has been Arg389Gly [7], which was only in partial linkage disequilibrium with Ser49Gly according to the authors [1]. The authors’ polymorphism is located towards the aminoterminus of the receptor, at the other end compared to the Arg389Gly

Immediate Effect of Slow PaceBhastrika Pranayamaon Blood Pressure and Heart Rate

The objective of this study was to evaluate the immediate effect of slow pace bhastrika pranayama (respiratory rate 6/min) for 5 minutes on heart rate and blood pressure and the effect of the same breathing exercise for the same duration of time (5 minutes) following oral intake of hyoscine-N-butylbromide (Buscopan), a parasympathetic blocker drug. Heart rate and blood pressure of volunteers (n = 39, age = 25-40 years) was recorded following standard procedure. First, subjects had to sit comfortably in an easy and steady posture (sukhasana) on a fairly soft seat placed on the floor keeping head, neck, and trunk erect, eyes closed, and the other muscles reasonably loose. The subject is directed to inhale through both nostrils slowly up to the maximum for about 4 seconds and then exhale slowly up to the maximum through both nostrils for about 6 seconds. The breathing must not be abdominal. These steps complete one cycle of slow pace bhastrika pranayama (respiratory rate 6/min). During the practice the subject is asked not to think much about the inhalation and exhalation time, but rather was requested to imagine the open blue sky. The pranayama was conducted in a cool, well-ventilated room (18-20 degrees C). After 5 minutes of this breathing practice, the blood pressure and heart rate again were recorded in the aforesaid manner using the same instrument. The other group (n = 10) took part in another study where their blood pressure and heart rate were recorded following half an hour of oral intake of hyoscine-N-butylbromide 20 mg. Then they practiced the breathing exercise as stated above, and the abovementioned parameters were recorded again to study the effect of parasympathetic blockade on the same pranayama. It was noted that after slow bhastrika pranayamic breathing (respiratory rate 6/min) for 5 minutes, both the systolic and diastolic blood pressure decreased significantly with a slight fall in heart rate. No significant alteration in both blood pressure and heart rate was observed in volunteers who performed the same breathing exercise for the same duration following oral intake of hyoscine-N-butylbromide. Pranayama increases frequency and duration of inhibitory neural impulses by activating pulmonary stretch receptors during above tidal volume inhalation as in Hering Bruer reflex, which bring about withdrawal of sympathetic tone in the skeletal muscle blood vessels, leading to widespread vasodilatation, thus causing decrease in peripheral resistance and thus decreasing the diastolic blood pressure. After hyoscine-N-butylbromide, the parasympathetic blocker, it was observed that blood pressure was not decreased significantly as a result of pranayama, as it was observed when no drug was administered. Vagal cardiac and pulmonary mechanisms are linked, and improvement in one vagal limb might spill over into the other. Baroreceptor sensitivity can be enhanced significantly by slow breathing (supported by a small reduction in the heart rate observed during slow breathing and by reduction in both systolic and diastolic pressure). Slow pace bhastrika pranayama (respiratory rate 6/min) exercise thus shows a strong tendency to improving the autonomic nervous system through enhanced activation of the parasympathetic system.

Cerebral sympathetic nerve activity has a major regulatory role in the cerebral circulation in REM sleep

Sympathetic nerve activity (SNA) in neurons projecting to skeletal muscle blood vessels increases during rapid-eye-movement (REM) sleep, substantially exceeding SNA of non-REM (NREM) sleep and quiet wakefulness (QW). Similar SNA increases to cerebral blood vessels may regulate the cerebral circulation in REM sleep, but this is unknown. We hypothesized that cerebral SNA increases during phasic REM sleep, constricting cerebral vessels as a protective mechanism against cerebral hyperperfusion during the large arterial pressure surges that characterize this sleep state. We tested this hypothesis using a newly developed model to continuously record SNA in the superior cervical ganglion (SCG) before, during, and after arterial pressure surges occurring during REM in spontaneously sleeping lambs. Arterial pressure (AP), intracranial pressure (ICP), cerebral blood flow (CBF), cerebral vascular resistance [CVR = (AP - ICP)/CBF], and SNA from the SCG were recorded in lambs (n = 5) undergoing spontaneous sleep-wake cycles. In REM sleep, CBF was greatest (REM > QW = NREM, P < 0.05) and CVR was least (REM < QW = NREM, P < 0.05). SNA in the SCG did not change from QW to NREM sleep but increased during tonic REM sleep, with a further increase during phasic REM sleep (phasic REM > tonic REM > QW = NREM, P < 0.05). Coherent averaging revealed that SNA increases preceded AP surges in phasic REM sleep by 12 s (P < 0.05). We report the first recordings of cerebral SNA during natural sleep-wake cycles. SNA increases markedly during tonic REM sleep, and further in phasic REM sleep. As SNA increases precede AP surges, they may serve to protect the brain against potentially damaging intravascular pressure changes or hyperperfusion in REM sleep.

Resident Endothelial Precursors in Muscle, Adipose, and Dermis Contribute to Postnatal Vasculogenesis

A novel population of tissue-resident endothelial precursors (TEPs) was isolated from small blood vessels in dermal, adipose, and skeletal muscle of mouse based on their ability to be grown as spheres. Cellular and molecular analyses of these cells revealed that they were highly related regardless of the tissue of origin and distinct from embryonic neural stem cells. Notably, TEPs did not express hematopoietic markers, but they expressed numerous characteristics of angiogenic precursors and their differentiated progeny, such as CD34, Flk-1, Tie-1, CD31, and vascular endothelial cadherin (VE-cadherin). TEPs readily differentiated into endothelial cells in newly formed vascular networks following transplantation into regenerating skeletal muscle. Taken together, these experiments suggest that TEPs represent a novel class of endothelial precursors that are closely associated with small blood vessels in muscle, adipose, and dermal tissue. This finding is of particular interest since it could bring new insight in cancer angiogenesis and collateral blood vessels developed following ischemia. Disclosure of potential conflicts of interest is found at the end of this article.

Mural cells paint a new picture of muscle stem cells

Despite being largely a post-mitotic tissue, adult skeletal muscle exhibits a remarkable capacity for regeneration. A new study has shown that cells derived from mural cells (pericytes) from blood vessels in postnatal human skeletal muscle contribute to robust skeletal muscle regeneration after intra-arterial delivery into a dystrophic mouse model.

Pericytes of human skeletal muscle are myogenic precursors distinct from satellite cells

Cells derived from blood vessels of human skeletal muscle can regenerate skeletal muscle, similarly to embryonic mesoangioblasts. However, adult cells do not express endothelial markers, but instead express markers of pericytes, such as NG2 proteoglycan and alkaline phosphatase (ALP), and can be prospectively isolated from freshly dissociated ALP(+) cells. Unlike canonical myogenic precursors (satellite cells), pericyte-derived cells express myogenic markers only in differentiated myotubes, which they form spontaneously with high efficiency. When transplanted into severe combined immune deficient-X-linked, mouse muscular dystrophy (scid-mdx) mice, pericyte-derived cells colonize host muscle and generate numerous fibres expressing human dystrophin. Similar cells isolated from Duchenne patients, and engineered to express human mini-dystrophin, also give rise to many dystrophin-positive fibres in vivo. These data show that myogenic precursors, distinct from satellite cells, are associated with microvascular walls in the human skeletal muscle, may represent a correlate of embryonic 'mesoangioblasts' present after birth and may be a promising candidate for future cell-therapy protocols in patients.

VEGF gradients, receptor activation, and sprout guidance in resting and exercising skeletal muscle

Extensive experimental studies have identified vascular endothelial growth factor (VEGF) concentrations and concentration gradients as major factors in angiogenesis; however, localized in vivo measurements of these parameters have not been possible. We developed a three-dimensional computational model of skeletal muscle fibers, blood vessels, and interstitial space. Here it is applied to rat extensor digitorum longus. VEGF isoforms are secreted by myocytes, diffuse through extracellular matrix and basement membranes, and bind endothelial cell surface receptors on blood vessels. In addition, one isoform, VEGF164, binds to proteoglycans in the interstitial space. VEGF secretion rate is determined from the predicted tissue oxygen level through its effect on the hypoxia inducible factor-1alpha transcription factor. We estimate VEGF secretion and its concentrations and gradients in resting muscle and for different levels of exercise. The effects of low levels of inspired oxygen are also studied. We predict that the high spatial heterogeneity of muscle fiber VEGF secretion in hypoxic tissue leads to significant gradients of VEGF concentration and VEGF receptor activation. VEGF concentration gradients are predicted to be significant in both resting and exercising muscle (4% and 6-8% change in VEGF over 10 microm, respectively), sufficient for chemotactic guidance of 50-microm-long sprout tip cells. VEGF gradients also result in heterogeneity in VEGF receptor activation--a possible explanation for the stochasticity of sprout location. In the absence of interstitial flow, gradients are 10-fold steeper in the transverse direction (i.e., perpendicular to the muscle fibers) than in the longitudinal direction. This may explain observed perpendicular anastomoses in skeletal muscle.

α‐Adrenergic Control of Skeletal Muscle Circulation at Rest and During Exercise in Aging Humans

Aging is associated with many changes in autonomic nervous system function that often lead to impairments in the normal ability to respond to physiological stressors commonly encountered in daily life. In addition, many of these chronic age-related changes in autonomic-circulatory function can potentially predispose the older adult to elevated risk for acute and chronic cardiovascular complications. One of the most pronounced and repeatable findings with respect to changes in the autonomic nervous system with human aging is the progressive increase in basal muscle sympathetic nerve activity (MSNA) directed to skeletal muscle vascular beds. Although the mechanism(s) underlying this sustained age-associated increase in MSNA are not completely understood, several changes in sympathetic a-adrenergic function occur with age. In this review, the authors discuss how aging affects (1) a-adrenergic control of skeletal muscle vascular tone under resting conditions and the differences that exist in this control of the upper and lower limbs (forearm vs leg circulation); (2) vasoconstrictor responsiveness to endogenous norepinephrine release, as well as the specific responsiveness of postjunctional a(1)- and a(2)-adrenergic receptors; and (3) sympathetic a-adrenergic control of muscle blood flow and vascular tone during exercise in humans. Further, they discuss how these changes in sympathetic a-adrenergic control of skeletal muscle blood vessels have important physiological and clinical implications for the aging human.