Application of near-infrared spectroscopy to assess the effect of the cupping size on the spatial hemodynamic response from the area inside and outside the cup of the biceps.

Cupping therapy is a popular intervention for improving muscle recovery after exercise although clinical evidence is weak. Previous studies demonstrated that cupping therapy may improve microcirculation of the soft tissue to accelerate tissue healing. However, it is unclear whether the cupping size could affect the spatial hemodynamic response of the treated muscle. The objective of this study was to use 8-channel near-infrared spectroscopy to assess this clinical question by assessing the effect of 3 cupping sizes (35, 40, and 45 mm in inner diameter of the circular cup) under -300 mmHg for 5 min on the muscle hemodynamic response from the area inside and outside the cup, including oxyhemoglobin and deoxy-hemoglobin in 18 healthy adults. Two-way factorial design was used to assess the interaction between the cupping size (35, 40, and 45 mm) and the location (inside and outside the cup) and the main effects of the cupping size and the location. The two-way repeated measures ANOVA demonstrated an interaction between the cupping size and the location in deoxy-hemoglobin (P = 0.039) but no interaction in oxyhemoglobin (P = 0.100), and a main effect of the cup size (P = 0.001) and location (P = 0.023) factors in oxyhemoglobin. For the cupping size factor, the 45-mm cup resulted in a significant increase in oxyhemoglobin (5.738±0.760 µM) compared to the 40-mm (2.095±0.312 µM, P<0.001) and 35-mm (3.134±0.515 µM, P<0.01) cup. Our findings demonstrate that the cupping size and location factors affect the muscle hemodynamic response, and the use of multi-channel near-infrared spectroscopy may help understand benefits of cupping therapy on managing musculoskeletal impairment.

Alginate-Encapsulated Mesenchymal Stromal Cells Improve Hind Limb Ischemia in a Translational Swine Model.

BACKGROUND: Cellular therapies have been investigated to improve blood flow and prevent amputation in peripheral artery disease with limited efficacy in clinical trials. Alginate-encapsulated mesenchymal stromal cells (eMSCs) demonstrated improved retention and survival and promoted vascular generation in murine hind limb ischemia through their secretome, but large animal evaluation is necessary for human applicability. We sought to determine the efficacy of eMSCs for peripheral artery disease-induced limb ischemia through assessment in our durable swine hind limb ischemia model. METHODS AND RESULTS: Autologous bone marrow eMSCs or empty alginate capsules were intramuscularly injected 2 weeks post-hind limb ischemia establishment (N=4/group). Improvements were quantified for 4 weeks through walkway gait analysis, contrast angiography, blood pressures, fluorescent microsphere perfusion, and muscle morphology and histology. Capsules remained intact with mesenchymal stromal cells retained for 4 weeks. Adenosine-induced perfusion deficits and muscle atrophy in ischemic limbs were significantly improved by eMSCs versus empty capsules (mean±SD, 1.07±0.19 versus 0.41±0.16, P=0.002 for perfusion ratios and 2.79±0.12 versus 1.90±0.62 g/kg, P=0.029 for ischemic muscle mass). Force- and temporal-associated walkway parameters normalized (ratio, 0.63±0.35 at week 3 versus 1.02±0.19 preligation; P=0.17), and compensatory footfall patterning was diminished in eMSC-administered swine (12.58±8.46% versus 34.85±15.26%; P=0.043). Delivery of eMSCs was associated with trending benefits in collateralization, local neovascularization, and muscle fibrosis. Hypoxia-cultured porcine mesenchymal stromal cells secreted vascular endothelial growth factor and tissue inhibitor of metalloproteinase 2. CONCLUSIONS: This study demonstrates the promise of the mesenchymal stromal cell secretome at improving peripheral artery disease outcomes and the potential for this novel swine model to serve as a component of the preclinical pipeline for advanced therapies.

Impact of capillary and sarcolemmal proximity on mitochondrial structure and energetic function in skeletal muscle.

Mitochondria within skeletal muscle cells are located either between the muscle contractile apparatus (interfibrillar mitochondria, IFM) or beneath the cell membrane (subsarcolemmal mitochondria, SSM), with several structural and functional differences reported between IFM and SSM. However, recent 3D imaging studies demonstrate that mitochondria are particularly concentrated in the proximity of capillaries embedded in sarcolemmal grooves rather than in proximity to the sarcolemma itself (paravascular mitochondria, PVM). To evaluate the impact of capillary vs. sarcolemmal proximity, we compared the structure and function of skeletal muscle mitochondria located either lateral to embedded capillaries (PVM), adjacent to the sarcolemma but not in PVM pools (SSM) or interspersed between sarcomeres (IFM). Mitochondrial morphology and interactions were assessed by 3D electron microscopy coupled with machine learning segmentation, whereas mitochondrial energy conversion was assessed by two-photon microscopy of mitochondrial membrane potential, content, calcium, NADH redox and flux in live, intact cells. Structurally, although PVM and SSM were similarly larger than IFM, PVM were larger, rounder and had more physical connections to neighbouring mitochondria compared to both IFM and SSM. Functionally, PVM had similar or greater basal NADH flux compared to SSM and IFM, respectively, despite a more oxidized NADH pool and a greater membrane potential, signifying a greater activation of the electron transport chain in PVM. Together, these data indicate that proximity to capillaries has a greater impact on resting mitochondrial energy conversion and distribution in skeletal muscle than the sarcolemma alone. KEY POINTS: Capillaries have a greater impact on mitochondrial energy conversion in skeletal muscle than the sarcolemma. Paravascular mitochondria are larger, and the outer mitochondrial membrane is more connected with neighbouring mitochondria. Interfibrillar mitochondria are longer and have greater contact sites with other organelles (i.e. sarcoplasmic reticulum and lipid droplets). Paravascular mitochondria have greater activation of oxidative phosphorylation than interfibrillar mitochondria at rest, although this is not regulated by calcium.

Age and sex differences in microvascular responses during reactive hyperaemia.

Microvascular impairments are typical of several cardiovascular diseases. Near-infrared spectroscopy (NIRS) combined with a vascular occlusion test provides non-invasive insights into microvascular responses by monitoring skeletal muscle oxygenation changes during reactive hyperaemia. Despite increasing interest in the effects of sex and ageing on microvascular responses, evidence remains inconsistent. Therefore, the present study aimed to investigate the effects of sex and age on microvascular responsiveness. Twenty-seven participants (seven young men and seven young women; seven older men and six older women; aged 26 ± 1, 26 ± 4, 67 ± 3 and 69 ± 4 years, respectively) completed a vascular occlusion test consisting of 5 min of arterial occlusion followed by 5 min reperfusion. Oxygenation changes in the vastus lateralis were monitored by near-infrared spectroscopy. The findings revealed that both women (referring to young and older women) and older participants (referring to both men and women) exhibited lower microvascular responsiveness. Notably, both women and older participants demonstrated reduced desaturation (-38% and -59%, respectively) and reperfusion rates (-24% and -40%, respectively) along with a narrower range of tissue oxygenation (-39% and -39%, respectively) and higher minimal tissue oxygenation levels (+34% and +21%, respectively). Women additionally displayed higher values in resting (+12%) and time-to-peak (+15%) tissue oxygenation levels. In conclusion, this study confirmed decreased microvascular responses in women and older individuals. These results emphasize the importance of considering sex and age when studying microvascular responses. Further research is needed to uncover the underlying mechanisms and clinical relevance of these findings, enabling the development of tailored strategies for preserving vascular health in diverse populations.

Differential changes in blood flow and oxygen utilization in active muscles between voluntary exercise and electrical muscle stimulation in young adults.

The physiological effects on blood flow and oxygen utilization in active muscles during and after involuntary contraction triggered by electrical muscle stimulation (EMS) remain unclear, particularly compared with those elicited by voluntary (VOL) contractions. Therefore, we used diffuse correlation and near-infrared spectroscopy (DCS-NIRS) to compare changes in local muscle blood flow and oxygen consumption during and after these two types of muscle contractions in humans. Overall, 24 healthy young adults participated in the study, and data were successfully obtained from 17 of them. Intermittent (2-s contraction, 2-s relaxation) isometric ankle dorsiflexion with a target tension of 20% of maximal VOL contraction was performed by EMS or VOL for 2 min, followed by a 6-min recovery period. DCS-NIRS probes were placed on the tibialis anterior muscle, and relative changes in local tissue blood flow index (rBFI), oxygen extraction fraction (rOEF), and metabolic rate of oxygen (rMRO2) were continuously derived. EMS induced more significant increases in rOEF and rMRO2 than VOL exercise but a comparable increase in rBFI. After EMS, rBFI and rMRO2 decreased more slowly than after VOL and remained significantly higher until the end of the recovery period. We concluded that EMS augments oxygen consumption in contracting muscles by enhancing oxygen extraction while increasing oxygen delivery at a rate similar to the VOL exercise. Under the conditions examined in this study, EMS demonstrated a more pronounced and/or prolonged enhancement in local muscle perfusion and aerobic metabolism compared with VOL exercise in healthy participants.NEW & NOTEWORTHY This is the first study to visualize continuous changes in blood flow and oxygen utilization within contracted muscles during and after electrical muscle stimulation (EMS) using combined diffuse correlation and near-infrared spectroscopy. We found that initiating EMS increases blood flow at a rate comparable to that during voluntary (VOL) exercise but enhances oxygen extraction, resulting in higher oxygen consumption. Furthermore, EMS increased postexercise muscle perfusion and oxygen consumption compared with that after VOL exercise.

Endothelial-Specific Reduction in Arf6 Impairs Insulin-Stimulated Vasodilation and Skeletal Muscle Blood Flow Resulting in Systemic Insulin Resistance in Mice.

BACKGROUND: Much of what we know about insulin resistance is based on studies from metabolically active tissues such as the liver, adipose tissue, and skeletal muscle. Emerging evidence suggests that the vascular endothelium plays a crucial role in systemic insulin resistance; however, the underlying mechanisms remain incompletely understood. Arf6 (ADP ribosylation factor 6) is a small GTPase that plays a critical role in endothelial cell function. Here, we tested the hypothesis that the deletion of endothelial Arf6 will result in systemic insulin resistance. METHODS: We used mouse models of constitutive endothelial cell-specific Arf6 deletion (Arf6f/- Tie2Cre+) and tamoxifen-inducible Arf6 knockout (Arf6f/f Cdh5CreER+). Endothelium-dependent vasodilation was assessed using pressure myography. Metabolic function was assessed using a battery of metabolic assessments including glucose and insulin tolerance tests and hyperinsulinemic-euglycemic clamps. We used a fluorescence microsphere-based technique to measure tissue blood flow. Skeletal muscle capillary density was assessed using intravital microscopy. RESULTS: Endothelial Arf6 deletion impaired insulin-stimulated vasodilation in white adipose tissue and skeletal muscle feed arteries. The impairment in vasodilation was primarily due to attenuated insulin-stimulated nitric oxide bioavailability but independent of altered acetylcholine-mediated or sodium nitroprusside-mediated vasodilation. Endothelial cell-specific deletion of Arf6 also resulted in systematic insulin resistance in normal chow-fed mice and glucose intolerance in high-fat diet-fed obese mice. The underlying mechanisms of glucose intolerance were reductions in insulin-stimulated blood flow and glucose uptake in the skeletal muscle and were independent of changes in capillary density or vascular permeability. CONCLUSIONS: Results from this study support the conclusion that endothelial Arf6 signaling is essential for maintaining insulin sensitivity. Reduced expression of endothelial Arf6 impairs insulin-mediated vasodilation and results in systemic insulin resistance. These results have therapeutic implications for diseases that are associated with endothelial cell dysfunction and insulin resistance such as diabetes.

Blood flow restriction augments the cross-education effect of isometric handgrip training.

INTRODUCTION: The application of blood flow restriction (BFR) to low-intensity exercise may be able to increase strength not only in the trained limb but also in the homologous untrained limb. Whether this effect is repeatable and how that change compares to that observed with higher intensity exercise is unknown. PURPOSE: Examine whether low-intensity training with BFR enhances the cross-education of strength compared to exercise without BFR and maximal efforts. METHODS: A total of 179 participants completed the 6-week study, with 135 individuals performing isometric handgrip training over 18 sessions. Participants were randomly assigned to one of four groups: 1) low-intensity (4 × 2 min of 30% MVC; LI, n = 47), 2) low-intensity with blood flow restriction (LI + 50% arterial occlusion pressure; LI-BFR, n = 41), 3) maximal effort (4 × 5 s of 100% MVC; MAX, n = 47), and 4) non-exercise control (CON, n = 44). RESULTS: LI-BFR was the only group that observed a cross-education in strength (CON: 0.64 SD 2.9 kg, LI: 0.95 SD 3.6 kg, BFR-LI: 2.7 SD 3.3 kg, MAX: 0.80 SD 3.1 kg). In the trained hand, MAX observed the greatest change in strength (4.8 SD 3.3 kg) followed by LI-BFR (2.8 SD 4.0 kg). LI was not different from CON. Muscle thickness did not change in the untrained arm, but ulna muscle thickness was increased within the trained arm of the LI-BFR group (0.06 SD 0.11 cm). CONCLUSION: Incorporating BFR into low-intensity isometric training led to a cross-education effect on strength that was greater than all other groups (including high-intensity training).

Sex-specific microvascular and hemodynamic responses to passive limb heating in young adults.

OBJECTIVE: We examined sex-specific microvascular reactivity and hemodynamic responses under conditions of augmented resting blood flow induced by passive heating compared to normal blood flow. METHODS: Thirty-eight adults (19 females) completed a vascular occlusion test (VOT) on two occasions preceded by rest with or without passive heating in a randomized, counterbalanced order. Skeletal muscle tissue oxygenation (StO2, %) was assessed with near-infrared spectroscopy (NIRS), and the rate of desaturation and resaturation as well as maximal StO2 (StO2max) and prolonged hypersaturation (area under the curve, StO2AUC) were quantified. Before the VOT, brachial artery blood flow (BABF), vascular conductance, and relative BABF (BABF normalized to forearm lean mass) were determined. Sex × condition ANOVAs were used. A p-value ≤.05 was considered statistically significant. RESULTS: Twenty minutes of heating increased BABF compared to the control (102.9 ± 28.3 vs. 36.0 ± 20.9 mL min-1; p < .01). Males demonstrated greater BABF than females (91.9 ± 34.0 vs. 47.0 ± 19.1 mL min-1; p < .01). There was no sex difference in normalized BABF. There were no significant interactions for NIRS-VOT outcomes, but heat did increase the rate of desaturation (-0.140 ± 0.02 vs. -0.119 ± 0.03% s-1; p < .01), whereas regardless of condition, males exhibited greater rates of resaturation and StO2max than females. CONCLUSIONS: These results suggest that blood flow is not the primary factor causing sex differences in NIRS-VOT outcomes.

A machine learning-based approach to identify peripheral artery disease using texture features from contrast-enhanced magnetic resonance imaging.

Diagnosing and assessing the risk of peripheral artery disease (PAD) has long been a focal point for medical practitioners. The impaired blood circulation in PAD patients results in altered microvascular perfusion patterns in the calf muscles which is the primary location of intermittent claudication pain. Consequently, we hypothesized that changes in perfusion and increase in connective tissue could lead to alterations in the appearance or texture patterns of the skeletal calf muscles, as visualized with non-invasive imaging techniques. We designed an automatic pipeline for textural feature extraction from contrast-enhanced magnetic resonance imaging (CE-MRI) scans and used the texture features to train machine learning models to detect the heterogeneity in the muscle pattern among PAD patients and matched controls. CE-MRIs from 36 PAD patients and 20 matched controls were used for preparing training and testing data at a 7:3 ratio with cross-validation (CV) techniques. We employed feature arrangement and selection methods to optimize the number of features. The proposed method achieved a peak accuracy of 94.11% and a mean testing accuracy of 84.85% in a 2-class classification approach (controls vs. PAD). A three-class classification approach was performed to identify a high-risk PAD sub-group which yielded an average test accuracy of 83.23% (matched controls vs. PAD without diabetes vs. PAD with diabetes). Similarly, we obtained 78.60% average accuracy among matched controls, PAD treadmill exercise completers, and PAD exercise treadmill non-completers. Machine learning and imaging-based texture features may be of interest in the study of lower extremity ischemia.

An exercise stress test for contrast-enhanced duplex ultrasound assessment of lower limb muscle perfusion in patients with peripheral arterial disease.

OBJECTIVE: The aim of the present study was to develop a standardized contrast-enhanced duplex ultrasound (CE-DUS) protocol to assess lower-extremity muscle perfusion before and after exercise and determine relationships of perfusion with clinical and functional measures. METHODS: CE-DUS (EPIQ 5G, Philips) was used before and immediately after a 10-minute, standardized bout of treadmill walking to compare microvascular perfusion of the gastrocnemius muscle in older (55-82 years) patients with peripheral arterial disease (PAD) (n = 15, mean ankle-brachial index, 0.78 ± 0.04) and controls (n = 13). Microvascular blood volume (MBV) and microvascular flow velocity (MFV) were measured at rest and immediately following treadmill exercise, and the Modified Physical Performance Test (MPPT) was used to assess mobility function. RESULTS: In the resting state (pre-exercise), MBV in patients with PAD was not significantly different than normal controls (5.17 ± 0.71 vs 6.20 ± 0.83 arbitrary units (AU) respectively; P = .36); however, after exercise, MBV was ∼40% lower in patients with PAD compared with normal controls (5.85 ± 1.13 vs 9.53 ± 1.31 AU, respectively; P = .04). Conversely, MFV was ∼60% higher in patients with PAD compared with normal controls after exercise (0.180 ± 0.016 vs 0.113 ± 0.018 AU, respectively; P = .01). There was a significant between-group difference in the exercise-induced changes in both MBV and MFV (P ≤ .05). Both basal and exercise MBV directly correlated with MPPT score in the patients with PAD (r = 0.56-0.62; P < .05). CONCLUSIONS: This standardized protocol for exercise stress testing of the lower extremities quantifies calf muscle perfusion and elicits perfusion deficits in patients with PAD. This technique objectively quantifies microvascular perfusion deficits that are related to reduced mobility function and could be used to assess therapeutic efficacy in patients with PAD.

Physiological and perceptual responses to acute arm cranking with blood flow restriction.

INTRODUCTION: Lower-body aerobic exercise with blood flow restriction (BFR) offers a unique approach for stimulating improvements in muscular function and aerobic capacity. While there are more than 40 reports documenting acute and chronic responses to lower-body aerobic exercise with BFR, responses to upper-body aerobic exercise with BFR are not clearly established. PURPOSE: We evaluated acute physiological and perceptual responses to arm cranking with and without BFR. METHODS: Participants (N = 10) completed 4 arm cranking (6 × 2 min exercise, 1 min recovery) conditions: low-intensity at 40%VO2peak (LI), low-intensity at 40%VO2peak with BFR at 50% of arterial occlusion pressure (BFR50), low-intensity at 40%VO2peak with BFR at 70% of arterial occlusion pressure (BFR70), and high-intensity at 80%VO2peak (HI) while tissue oxygenation, cardiorespiratory, and perceptual responses were assessed. RESULTS: During exercise, tissue saturation for BFR50 (54 ± 6%), BFR70 (55 ± 6%), and HI (54 ± 8%) decreased compared to LI (61 ± 5%, all P < 0.01) and changes in deoxyhemoglobin for BFR50 (11 ± 4), BFR70 (15 ± 6), and HI (16 ± 10) increased compared to LI (4 ± 2, all P < 0.01). During recovery intervals, tissue saturation for BFR50 and BFR70 decreased further and deoxyhemoglobin for BFR50 and BFR70 increased further (all P < 0.04). Heart rate for BFR70 and HI increased by 9 ± 9 and 50 ± 15b/min, respectively, compared to LI (both P < 0.02). BFR50 (8 ± 2, 1.0 ± 1.0) and BFR70 (10 ± 2, 2.1 ± 1.4) elicited greater arm-specific perceived exertion (6-20 scale) and pain (0-10 scale) compared to LI (7 ± 1, 0.2 ± 0.5, all P < 0.05) and pain for BFR70 did not differ from HI (1.7 ± 1.9). CONCLUSION: Arm cranking with BFR decreased tissue saturation and increased deoxyhemoglobin without causing excessive cardiorespiratory strain and pain.

Pedicled Peroneus Brevis Muscle Flaps as an Alternative to Fasciocutaneous Rotational Flaps for Lower-Extremity Soft Tissue Defects.

OBJECTIVES: To report our experience using a peroneus brevis flap (PBF) for soft tissue defects of the distal third of the tibia, ankle, and hindfoot in resource-challenged environments. DESIGN: Retrospective review. SETTING: Rural outpatient surgical facility in Honduras. PATIENT SELECTION CRITERIA: Patients who sustained tibia, ankle, or hindfoot fractures or traumatic degloving, with critical-sized soft tissue defects treated with either a proximally based or distally based pedicled PBF to achieve coverage of the middle and distal third of the leg, ankle, and/or hindfoot. OUTCOME MEASURES AND COMPARISONS: Flap healing, complications, and reoperations. RESULTS: Twenty-three patients, 4 with proximally based and 19 with distally based PBF flaps were included. The mean patient age was 37.3 (SD = 18.3; range 18-75 years). Duration of follow-up averaged 14.7 months (SD = 11.4; range 4-46). The PBF successfully covered the defect without the need for additional unplanned surgical flap coverage in all but 2 patients. Thirty percent of the PBFs received a split thickness skin graft, while the remainder granulated successfully without skin graft. Four flaps were partially debrided without additional flap mobilization, while 1 flap was lost completely. Ten patients had successful re-elevation of their flaps for secondary procedures such as implant removal, spacer exchange, deep debridements, and bone grafting. All donor site incisions healed without complication. CONCLUSIONS: The pedicled PBF allows coverage of distal leg, ankle, and hindfoot wounds using muscle in patients who may otherwise require free tissue flaps or transfer to another institution for coverage. PBFs can be learned and implemented without the use of microvascular techniques. LEVEL OF EVIDENCE: Therapeutic Level IV. See Instructions for Authors for a complete description of levels of evidence.

Co-Delivery of Bioengineered Exosomes and Oxygen for Treating Critical Limb Ischemia in Diabetic Mice.

Diabetic patients with critical limb ischemia face a high rate of limb amputation. Regeneration of the vasculature and skeletal muscles can salvage diseased limbs. Therapy using stem cell-derived exosomes that contain multiple proangiogenic and promyogenic factors represents a promising strategy. Yet the therapeutic efficacy is not optimal because exosomes alone cannot efficiently rescue and recruit endothelial and skeletal muscle cells and restore their functions under hyperglycemic and ischemic conditions. To address these limitations, we fabricated ischemic-limb-targeting stem cell-derived exosomes and oxygen-releasing nanoparticles and codelivered them in order to recruit endothelial and skeletal muscle cells, improve cell survival under ischemia before vasculature is established, and restore cell morphogenic function under high glucose and ischemic conditions. The exosomes and oxygen-releasing nanoparticles, delivered by intravenous injection, specifically accumulated in the ischemic limbs. Following 4 weeks of delivery, the exosomes and released oxygen synergistically stimulated angiogenesis and muscle regeneration without inducing substantial inflammation and reactive oxygen species overproduction. Our work demonstrates that codelivery of exosomes and oxygen is a promising treatment solution for saving diabetic ischemic limbs.

Exercise and Ischemia-Activated Pathways in Limb Muscle Angiogenesis and Vascular Regeneration.

Exercise has a profound effect on cardiovascular disease, particularly through vascular remodeling and regeneration. Peripheral artery disease (PAD) is one such cardiovascular condition that benefits from regular exercise or rehabilitative physical therapy in terms of slowing the progression of disease and delaying amputations. Various rodent pre-clinical studies using models of PAD and exercise have shed light on molecular pathways of vascular regeneration. Here, I review key exercise-activated signaling pathways (nuclear receptors, kinases, and hypoxia inducible factors) in the skeletal muscle that drive paracrine regenerative angiogenesis. The rationale for highlighting the skeletal muscle is that it is the largest organ recruited during exercise. During exercise, skeletal muscle releases several myokines, including angiogenic factors and cytokines that drive tissue vascular regeneration via activation of endothelial cells, as well as by recruiting immune and endothelial progenitor cells. Some of these core exercise-activated pathways can be extrapolated to vascular regeneration in other organs. I also highlight future areas of exercise research (including metabolomics, single cell transcriptomics, and extracellular vesicle biology) to advance our understanding of how exercise induces vascular regeneration at the molecular level, and propose the idea of "exercise-mimicking" therapeutics for vascular recovery.

Regulation of Skeletal Muscle Resistance Arteriolar Tone: Integration of Multiple Mechanisms.

INTRODUCTION: Physiological system complexity represents an imposing challenge to gaining insight into how arteriolar behavior emerges. Further, mechanistic complexity in arteriolar tone regulation requires that a systematic determination of how these processes interact to alter vascular diameter be undertaken. METHODS: The present study evaluated the reactivity of ex vivo proximal and in situ distal resistance arterioles in skeletal muscle with challenges across the full range of multiple physiologically relevant stimuli and determined the stability of responses over progressive alterations to each other parameter. The five parameters chosen for examination were (1) metabolism (adenosine concentration), (2) adrenergic activation (norepinephrine concentration), (3) myogenic activation (intravascular pressure), (4) oxygen (superfusate PO2), and (5) wall shear rate (altered intraluminal flow). Vasomotor tone of both arteriole groups following challenge with individual parameters was determined; subsequently, responses were determined following all two- and three-parameter combinations to gain deeper insight into how stimuli integrate to change arteriolar tone. A hierarchical ranking of stimulus significance for establishing arteriolar tone was performed using mathematical and statistical analyses in conjunction with machine learning methods. RESULTS: Results were consistent across methods and indicated that metabolic and adrenergic influences were most robust and stable across all conditions. While the other parameters individually impact arteriolar tone, their impact can be readily overridden by the two dominant contributors. CONCLUSION: These data suggest that mechanisms regulating arteriolar tone are strongly affected by acute changes to the local environment and that ongoing investigation into how microvessels integrate stimuli regulating tone will provide a more thorough understanding of arteriolar behavior emergence across physiological and pathological states.

Venous congestion affects neuromuscular changes in pigs in terms of muscle electrical activity and muscle stiffness.

Early detection of venous congestion (VC)-related diseases such as deep vein thrombosis (DVT) is important to prevent irreversible or serious pathological conditions. However, the current way of diagnosing DVT is only possible after recognizing advanced DVT symptoms such as swelling, pain, and tightness in affected extremities, which may be due to the lack of information on neuromechanical changes following VC. Thus, the goal of this study was to investigate acute neuromechanical changes in muscle electrical activity and muscle stiffness when VC was induced. The eight pigs were selected and the change of muscle stiffness from the acceleration and muscle activity in terms of integral electromyography (IEMG) was investigated in three VC stages. Consequently, we discovered a significant increase in the change in muscle stiffness and IEMG from the baseline to the VC stages (p < 0.05). Our results and approach can enable early detection of pathological conditions associated with VC, which can be a basis for further developing early diagnostic tools for detecting VC-related diseases.

Evidence of impaired functional sympatholysis in patients with heart failure with preserved ejection fraction.

Exercising muscle blood flow is reduced in patients with heart failure with a preserved ejection fraction (HFpEF), which may be related to disease-related changes in the ability to overcome sympathetic nervous system (SNS)-mediated vasoconstriction during exercise, (i.e., "functional sympatholysis"). Thus, in 12 patients with HFpEF (69 ± 7 yr) and 11 healthy controls (Con, 69 ± 4 yr), we examined forearm blood flow (FBF), mean arterial pressure (MAP), and forearm vascular conductance (FVC) during rhythmic handgrip exercise (HG) at 30% of maximum voluntary contraction with or without lower-body negative pressure (LBNP, -20 mmHg) to increase SNS activity and elicit peripheral vasoconstriction. SNS-mediated vasoconstrictor responses were determined as LBNP-induced changes (%Δ) in FVC, and the "magnitude of sympatholysis" was calculated as the difference between responses at rest and during exercise. At rest, the LBNP-induced change in FVC was significantly lesser in HFpEF compared with Con (HFpEF: -9.5 ± 5.5 vs. Con: -21.0 ± 8.0%; P < 0.01). During exercise, LBNP-induced %ΔFVC was significantly attenuated in Con compared with rest (HG: -5.8 ± 6.0%; P < 0.05) but not in HFpEF (HG: -9.9 ± 2.5%; P = 0.88). Thus, the magnitude of sympatholysis was lesser in HFpEF compared with Con (HFpEF: 0.4 ± 4.7 vs. Con: -15.2 ± 11.8%; P < 0.01). These data demonstrate a diminished ability to attenuate SNS-mediated vasoconstriction in HFpEF and provide new evidence suggesting impaired functional sympatholysis in this patient group.NEW & NOTEWORTHY Data from the current study suggest that functional sympatholysis, or the ability to adequately attenuate sympathetic nervous system (SNS)-mediated vasoconstriction during exercise, is impaired in patients with heart failure with preserved ejection fraction (HFpEF). These observations extend the current understanding of HFpEF pathophysiology by implicating inadequate functional sympatholysis as an important contributor to reduced exercising muscle blood flow in this patient group.

Estrogen-Related Receptor Gamma Gene Therapy Promotes Therapeutic Angiogenesis and Muscle Recovery in Preclinical Model of PAD.

Background Peripheral arterial disease and critical limb ischemia are cardiovascular complications associated with vascular insufficiency, oxidative metabolic dysfunction, and myopathy in the limbs. Estrogen-related receptor gamma (ERRγ) has emerged as a dual regulator of paracrine angiogenesis and oxidative metabolism through transgenic mouse studies. Here our objective was to investigate whether postischemic intramuscular targeting of ERRγ via gene therapy promotes ischemic recovery in a preclinical model of peripheral arterial disease/critical limb ischemia. Methods and Results Adeno-associated virus 9 (AAV9) Esrrg gene delivery vector was developed and first tested via intramuscular injection in murine skeletal muscle. AAV9-Esrrg robustly increased ERRγ protein expression, induced angiogenic and oxidative genes, and boosted capillary density and succinate dehydrogenase oxidative metabolic activity in skeletal muscles of C57Bl/6J mice. Next, hindlimb ischemia was induced via unilateral femoral vessel ligation in mice, followed by intramuscular AAV9-Esrrg (or AAV9-green fluorescent protein) gene delivery 24 hours after injury. ERRγ overexpression increased ischemic neoangiogenesis and markers of endothelial activation, and significantly improved ischemic revascularization measured using laser Doppler flowmetry. Moreover, ERRγ overexpression restored succinate dehydrogenase oxidative metabolic capacity in ischemic muscle, which correlated with increased mitochondrial respiratory complex protein expression. Most importantly, myofiber size to number quantification revealed that AAV9-Esrrg restores myofibrillar size and mitigates ischemia-induced myopathy. Conclusions These results demonstrate that intramuscular AAV9-Esrrg delivery rescues ischemic pathology after hindlimb ischemia, underscoring that Esrrg gene therapy or pharmacological activation could be a promising strategy for the management of peripheral arterial disease/critical limb ischemia.