Diet-induced obesity augments ischemic myopathy and functional decline in a murine model of peripheral artery disease.

Peripheral artery disease (PAD) causes an ischemic myopathy contributing to patient disability and mortality. Most preclinical models to date use young, healthy rodents with limited translatability to human disease. Although PAD incidence increases with age, and obesity is a common comorbidity, the pathophysiologic association between these risk factors and PAD myopathy is unknown. Using our murine model of PAD, we sought to elucidate the combined effect of age, diet-induced obesity and chronic hindlimb ischemia (HLI) on (1) mobility, (2) muscle contractility, and markers of muscle (3) mitochondrial content and function, (4) oxidative stress and inflammation, (5) proteolysis, and (6) cytoskeletal damage and fibrosis. Following 16-weeks of high-fat, high-sucrose, or low-fat, low-sucrose feeding, HLI was induced in 18-month-old C57BL/6J mice via the surgical ligation of the left femoral artery at 2 locations. Animals were euthanized 4-weeks post-ligation. Results indicate mice with and without obesity shared certain myopathic changes in response to chronic HLI, including impaired muscle contractility, altered mitochondrial electron transport chain complex content and function, and compromised antioxidant defense mechanisms. However, the extent of mitochondrial dysfunction and oxidative stress was significantly greater in obese ischemic muscle compared to non-obese ischemic muscle. Moreover, functional impediments, such as delayed post-surgical recovery of limb function and reduced 6-minute walking distance, as well as accelerated intramuscular protein breakdown, inflammation, cytoskeletal damage, and fibrosis were only evident in mice with obesity. As these features are consistent with human PAD myopathy, our model could be a valuable tool to test new therapeutics.

Myoglobinemia, Peripheral Arterial Disease, and Patient Mortality.

BACKGROUND: Peripheral arterial disease (PAD) causes leg muscle damage due to inadequate perfusion and increases cardiovascular events and mortality 2- to 3-fold. It is unclear if PAD is a biomarker for high-risk cardiovascular disease or if skeletal muscle injury harms arterial health. The objective of this work is to test if serum myoglobin levels (myoglobinemia) are a marker of PAD, and if so, whether myoglobin impairs vascular health. STUDY DESIGN: Patient blood samples were collected from PAD and control (no PAD) patients and interrogated for myoglobin concentrations and nitric oxide bioavailability. Patient mortality over time was captured from the medical record. Myoglobin activity was tested on endothelial cells and arterial function. RESULTS: Myoglobin is a biomarker for symptomatic PAD and was inversely related to nitric oxide bioavailability; 200 ng/mL myoglobin in vitro increased endothelial cell permeability in vitro and decreased nitrate bioavailability. Ex vivo, 100 ng/mL myoglobin increased vascular tone in naive murine aortas approximately 1.5 times, impairing absolute vessel relaxation. In vivo, we demonstrated that myoglobinemia caused impaired flow-mediated dilation in a porcine model. Patients presenting with myoglobin levels of 100 ng/mL or greater had significantly more deaths than those with myoglobin levels of less than 100 ng/mL. CONCLUSIONS: Using a combination of patient data, in vitro, ex vivo, and in vivo testing, we found that myoglobin is a biomarker for symptomatic PAD and a potent regulator of arterial health that can increase vascular tone, increase vascular permeability, and cause endothelial dysfunction, all of which may contribute to the vulnerability of PAD patients to cardiovascular events and death.

Skeletal muscle MiR-210 expression is associated with mitochondrial function in peripheral artery disease patients.

Previous studies have demonstrated that circulating microRNA (miR)-210 levels are elevated in peripheral artery disease (PAD) patients. MiR-210 is known to be a negative regulator of mitochondrial respiration; however, the relationship between miR-210 and mitochondrial function has yet to be studied in PAD. We aimed to compare skeletal muscle miR-210 expression of PAD patients to non-PAD controls (CON) and to examine the relationship between miR-210 expression and mitochondrial function. Skeletal muscle biopsies from CON (n = 20), intermittent claudication (IC) patients (n = 20), and critical limb ischemia (CLI) patients (n = 20) were analyzed by high-resolution respirometry to measure mitochondrial respiration of permeabilized fibers. Samples were also analyzed for miR-210 expression by real-time PCR. MiR-210 expression was significantly elevated in IC and CLI muscle compared to CON (P = 0.008 and P < 0.001, respectively). Mitochondrial respiration of electron transport chain (ETC) Complexes II (P = 0.001) and IV (P < 0.001) were significantly reduced in IC patients. Further, CLI patients demonstrated significant reductions in respiration during Complexes I (state 2: P = 0.04, state 3: P = 0.003), combined I and II (P < 0.001), II (P < 0.001), and IV (P < 0.001). The expression of the miR-210 targets, cytochrome c oxidase assembly factor heme A: farnesyltransferase (COX10), and iron-sulfur cluster assembly enzyme (ISCU) were down-regulated in PAD muscle. MiR-210 may play a role in the cellular adaptation to hypoxia and may be involved in the metabolic myopathy associated with PAD.

Endothelial cell-derived pro-fibrotic factors increase TGF-ß1 expression by smooth muscle cells in response to cycles of hypoxia-hyperoxia.

BACKGROUND: The vascular pathology of peripheral artery disease (PAD) encompasses abnormal microvascular architecture and fibrosis in response to ischemia-reperfusion (I/R) cycles. We aimed to investigate the mechanisms by which pathological changes in the microvasculature direct fibrosis in the context of I/R. METHODS: Primary human aortic endothelial cells (ECs) were cultured under cycles of normoxia-hypoxia (NH) or normoxia-hypoxia-hyperoxia (NHH) to mimic I/R. Primary human aortic smooth muscle cells (SMCs) were cultured and treated with media from the ECs. FINDINGS: The mRNA and protein expression of the pro-fibrotic factors platelet derived growth factor (PDGF)-BB and connective tissue growth factor (CTGF) were significantly upregulated in ECs undergoing NH or NHH cycles. Treatment of SMCs with media from ECs undergoing NH or NHH cycles led to significant increases in TGF-ß1, TGF-ß pathway signaling intermediates, and collagen expression. Addition of neutralizing antibodies against PDGF-BB and CTGF to the media blunted the increases in TGF-ß1 and collagen expression. Treatment of SMCs with PAD patient-derived serum also led to increased TGF-ß1 levels. INTERPRETATION: In an in-vitro model of I/R, which recapitulates the pathophysiology of PAD, increased secretion of PDGF-BB and CTGF by ECs was shown to be predominantly driving TGF-ß1-mediated expression by SMCs. These cell culture experiments help elucidate the mechanism and interaction between ECs and SMCs in microvascular fibrosis associated with I/R. Thus, targeting these pro-fibrotic factors may be an effective strategy to combat fibrosis in response to cycles of I/R. FUNDING: National Institute on Aging at the National Institutes of Health grant number R01AG064420. RESEARCH IN CONTEXT: Evidence before this study: Previous studies in gastrocnemius biopsies from peripheral artery disease (PAD) patients showed that transforming growth factor beta 1 (TGF-ß1), the most potent inducer of pathological fibrosis, is increased in the vasculature of PAD patients and correlated with collagen deposition. However, the exact cellular source of TGF-ß1 remained unclear. Added value of this study: Exposing cells to cycles of normoxia-hypoxia-hyperoxia (NHH) resulted in pathological changes that are consistent with human PAD. This supports the idea that the use of NHH may be a reliable, novel in vitro model of PAD useful for studying associated pathophysiological mechanisms. Furthermore, pro-fibrotic factors (PDGF-BB and CTGF) released from endothelial cells were shown to induce a fibrotic phenotype in smooth muscle cells. This suggests a potential interaction between these cell types in the microvasculature that drives increased TGF-ß1 expression and collagen deposition. Thus, targeting these pro-fibrotic factors may be an effective strategy to combat fibrosis in response to cycles of ischemia-reperfusion.

Phytochemicals as Therapeutic Interventions in Peripheral Artery Disease.

Peripheral artery disease (PAD) affects over 200 million people worldwide, resulting in significant morbidity and mortality, yet treatment options remain limited. Among the manifestations of PAD is a severe functional disability and decline, which is thought to be the result of different pathophysiological mechanisms including oxidative stress, skeletal muscle pathology, and reduced nitric oxide bioavailability. Thus, compounds that target these mechanisms may have a therapeutic effect on walking performance in PAD patients. Phytochemicals produced by plants have been widely studied for their potential health effects and role in various diseases including cardiovascular disease and cancer. In this review, we focus on PAD and discuss the evidence related to the clinical utility of different phytochemicals. We discuss phytochemical research in preclinical models of PAD, and we highlight the results of the available clinical trials that have assessed the effects of these compounds on PAD patient functional outcomes.

The Nitric Oxide System in Peripheral Artery Disease: Connection with Oxidative Stress and Biopterins.

Peripheral artery disease (PAD) pathophysiology extends beyond hemodynamics to include other operating mechanisms, including endothelial dysfunction. Oxidative stress may be linked to endothelial dysfunction by reducing nitric oxide (NO) bioavailability. We aimed to investigate whether the NO system and its regulators are altered in the setting of PAD and to assess the relationship between NO bioavailability and oxidative stress. Sera from 35 patients with intermittent claudication (IC), 26 patients with critical limb ischemia (CLI), and 35 non-PAD controls were analyzed to determine levels of tetrahydrobiopterin (BH4), dihydrobiopterin (BH2), nitrate/nitrite (nitric oxides, or NOx), arginine, citrulline, asymmetric dimethylarginine (ADMA), symmetric dimethylarginine (SDMA), and the oxidative stress markers 8-Oxo-2'-deoxyguanosine (8-OHdG), 4-hydroxynonenal (4-HNE), advanced glycation end products (AGEs), and protein carbonyls. NOx was significantly lower in IC and CLI patients compared to controls in association with elevated oxidative stress, with the greatest NOx reductions observed in CLI. Compared with controls, IC and CLI patients had reduced BH4, elevated BH2, and a reduced BH4/BH2 ratio. SDMA, the arginine/SDMA ratio, and the arginine/ADMA ratio were significantly higher in CLI patients. The NO system and its regulators are significantly compromised in PAD. This dysregulation appears to be driven by increased oxidative stress and worsens as the disease progresses from claudication to CLI.

Altered Metabolomic Profile in Patients with Peripheral Artery Disease.

Peripheral artery disease (PAD) is a common atherosclerotic disease characterized by narrowed or blocked arteries in the lower extremities. Circulating serum biomarkers can provide significant insight regarding the disease progression. Here, we explore the metabolomics signatures associated with different stages of PAD and investigate potential mechanisms of the disease. We compared the serum metabolites of a cohort of 26 PAD patients presenting with claudication and 26 PAD patients presenting with critical limb ischemia (CLI) to those of 26 non-PAD controls. A difference between the metabolite profiles of PAD patients from non-PAD controls was observed for several amino acids, acylcarnitines, ceramides, and cholesteryl esters. Furthermore, our data demonstrate that patients with CLI possess an altered metabolomic signature different from that of both claudicants and non-PAD controls. These findings provide new insight into the pathophysiology of PAD and may help develop future diagnostic procedures and therapies for PAD patients.

Role of Transforming Growth Factor-ß in Skeletal Muscle Fibrosis: A Review.

Transforming growth factor-beta (TGF-ß) isoforms are cytokines involved in a variety of cellular processes, including myofiber repair and regulation of connective tissue formation. Activation of the TGF-ß pathway contributes to pathologic fibrosis in most organs. Here, we have focused on examining the evidence demonstrating the involvement of TGF-ß in the fibrosis of skeletal muscle particularly. The TGF-ß pathway plays a role in different skeletal muscle myopathies, and TGF-ß signaling is highly induced in these diseases. In this review, we discuss different molecular mechanisms of TGF-ß-mediated skeletal muscle fibrosis and highlight different TGF-ß-targeted treatments that target these relevant pathways.

Effects of Limb Revascularization Procedures on Oxidative Stress.

Revascularization procedures to treat patients with peripheral artery disease are among the most common operations performed by vascular surgeons. However, there are major limitations to revascularizations, readmission rates due to procedural complications are high, and greater risks of cardiovascular and limb adverse outcomes have been reported for patients with peripheral artery disease undergoing limb revascularization. Specifically, surgical revascularization may be associated with increased generation of reactive oxygen species based on the ischemia reperfusion injury theory, as restored blood flow and reoxygenation of ischemic areas may be accompanied by increased oxidative stress. In this review, we present the current evidence regarding the effects of revascularization procedures on oxidative stress. We also discuss potential therapeutic interventions to prevent ischemia reperfusion injury-mediated tissue damage.

Oxidative Stress and Arterial Dysfunction in Peripheral Artery Disease.

Peripheral artery disease (PAD) is an atherosclerotic disease characterized by a narrowing of the arteries in the lower extremities. Disease manifestations are the result of more than just reduced blood flow, and include endothelial dysfunction, arterial stiffness, and inflammation. Growing evidence suggests that these factors lead to functional impairment and decline in PAD patients. Oxidative stress also plays an important role in the disease, and a growing amount of data suggest a link between arterial dysfunction and oxidative stress. In this review, we present the current evidence for the involvement of endothelial dysfunction, arterial stiffness, and inflammation in the pathophysiology of PAD. We also discuss the links between these factors and oxidative stress, with a focus on nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2)-derived reactive oxygen species (ROS) and decreased nitric oxide (NO) bioavailability. Finally, the potential therapeutic role of NOX2 antioxidants for improving arterial function and functional status in PAD patients is explored.

Oxidative stress and antioxidant treatment in patients with peripheral artery disease.

Peripheral artery disease is an atherosclerotic disease of arterial vessels that mostly affects arteries of lower extremities. Effort induced cycles of ischemia and reperfusion lead to increased reactive oxygen species production by mitochondria. Therefore, the pathophysiology of peripheral artery disease is a consequence of metabolic myopathy, and oxidative stress is the putative major operating mechanism behind the structural and metabolic changes that occur in muscle. In this review, we discuss the evidence for oxidative damage in peripheral artery disease and discuss management strategies related to antioxidant supplementation. We also highlight the major pathways governing oxidative stress in the disease and discuss their implications in disease progression. Potential therapeutic targets and diagnostic methods related to these mechanisms are explored, with an emphasis on the Nrf2 pathway.

Surgical management of carotid body tumors: a 15-year single institution experience employing an interdisciplinary approach.

Cervical paragangliomas are rare neoplasms that arise from extraadrenal paraganglia in close association with the cranial nerves and extracranial arterial system of the head and neck, and therefore surgical extirpation can be challenging. A retrospective study was conducted of all patients undergoing surgical excision of a cervical paraganglioma between 2000 and 2015. The demographic characteristics, clinical features, surgical approach, and outcomes were reviewed. A total of 20 cervical paragangliomas were excised in 17 patients. There were 14 female and 3 male patients with a mean age of 56.6 ± 17.0 at the time of operation. Twelve patients had unilateral tumors and 5 patients had bilateral tumors. Familial involvement was confirmed by history or direct genetic analysis in 8 (47%) of the 17 patients. There were no malignant paragangliomas, and only 3 patients had tumors that were determined to be functional. Tumor size ranged from 1.3 to 6.0 cm. Two patients required combined arterial resection as part of complete excision of the tumor. There were no permanent operative cranial nerve injuries, no recurrences, minimal morbidity, and no mortality. In conclusion, optimal management of cervical paragangliomas should include a thorough preoperative evaluation, accurate definition of the surgical anatomy, and exclusion of synchronous paragangliomas. A combined therapeutic approach by a multidisciplinary team including surgeons and interventional radiologists provides safe and effective management of cervical paragangliomas with very low morbidity and excellent outcomes.

The management of chronic venous insufficiency with ulceration: the role of minimally invasive perforator interruption.

BACKGROUND: The purpose of this study was to analyze the long-term outcomes associated with interruption of incompetent perforator veins (IPV) using minimally invasive techniques as adjunctive therapies in the management of patients with chronic venous insufficiency (CVI). METHODS: This is a retrospective review of a prospectively maintained venous database collected over 6 years (2005-2011). The study cohort included 64 patients with CVI stage C5 or C6 who underwent minimally invasive perforator interruption with subfascial endoscopic perforator surgery (SEPS) or radiofrequency ablation of IPV (RFA-IPV) as part of the management of their CVI. All patients were referred for evaluation after having failed conservative treatment with compression dressings. Relevant patient characteristics and comorbidities were recorded along with symptom resolution, venous ulcer healing, recurrence, and surgical complications. In addition to clinic follow-up examination by a surgical provider, chart notes from other subspecialties were also reviewed. We also conducted telephone assessments in patients who had been lost to clinic follow-up in order to provide complete outcome data. RESULTS: In this subset (n = 64) of patients with CVI who had adjunctive IPV treatment, 41 (64%) underwent SEPS and 23 (36%) patients underwent RFA-IPV along with ablation of the greater saphenous vein for C5 or C6 disease. The mean patient follow-up was 37 months. There were no differences in patient demographics or risk factors. Twenty-three (88%) SEPS and 12 (100%) RFA-IPV patients (P = NS) with C6 disease went on to completely heal their venous ulcers after the procedure with an average healing time of 5.2 (SEPS) and 4.4 (RFA-IPV) months (P = NS). Overall, 7 (17%) SEPS and 6 (23%) RFA-IPV patients (P = NS) developed a recurrent ulcer after surgical treatment. Procedural complications were seen in 14 (34%) SEPS and 2 (9%) RFA-IPV patients (P = NS), mostly minor. Major complications only occurred in the SEPS group consisting of 2 major amputations caused by pain from nonhealing ulcers and 1 deep venous thrombosis. CONCLUSIONS: This study supports the premise that in patients with advanced venous disease, there may be a demonstrable benefit directly attributable to perforator interruption. Our recurrent ulceration rates are acceptable, with low complication rates in patients undergoing RFA-IPV, thereby making this procedure more attractive in patients with multiple comorbidities. We support an aggressive approach to patients with C5/C6 disease that includes perforator elimination when appropriate.

Endovascular repair of a pancreatic allograft mycotic aneurysm: two-year follow-up.

PURPOSE: To describe midterm outcome of endovascular stent-graft repair of a mycotic aneurysm associated with a peripancreatic allograft abscess after transplantation. CASE REPORT: A 46-year-old woman underwent combined kidney and pancreas allograft transplantation under heavy immunosuppression. She developed a peripancreatic allograft abscess with associated mycotic aneurysm of the pancreatic allograft donor iliac artery 2 months after transplantation. Endovascular stent-grafts were used to exclude the aneurysm, retaining normal pancreatic allograft function over the next 2 years. CONCLUSION: Allograft arterial mycotic aneurysm development after transplantation is a rare but potentially life-threatening problem. This case suggests that control of a mycotic aneurysm may be obtained with stent-grafting in the presence of active infection under intense immunosuppression, avoiding allograft pancreatectomy.

The impact of recent European trials on abdominal aortic aneurysm repair: is a paradigm shift warranted?

Recent reports, following upon the reported outcomes of European randomized prospective trials of endovascular abdominal aortic aneurysm repair (EVAR), have brought into question the appropriateness of some of the trials' main conclusions, particularly in patients deemed at high-risk for surgical intervention. Based on the data of these trials, specifically EVAR 2, it has been suggested that EVAR should not be performed in high-risk individuals due to the likelihood of poor outcomes and the lack of improved survival, both associated with higher costs. In addition, certain aspects of the trials involving those deemed fit for open repair (the EVAR 1 and Dutch Randomized Endovascular Aneurysm Management trials) deserve qualified reservations. Although prospective, randomized United States trial data on such patients are not currently available, some large retrospective studies and registry reviews provide a basis for comparison of these trials with US EVAR experiences. In this review, the European EVAR trials are analyzed along with these other US studies and the rationale for modifying some of the conclusions drawn from the trials is presented and general guidelines for the selective management of abdominal aortic aneurysm patients presenting with potential indications for intervention are proposed.