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Peripheral artery disease (PAD) affects more than 230 million people worldwide. PAD patients suffer from reduced quality of life and are at increased risk of vascular complications and all-cause mortality. Despite its prevalence, impact on quality of life and poor long-term clinical outcomes, PAD remains underdiagnosed and undertreated compared to myocardial infarction and stroke. PAD is due to a combination of macrovascular atherosclerosis and calcification, combined with microvascular rarefaction, leading to chronic peripheral ischemia. Novel therapies are needed to address the increasing incidence of PAD and its difficult long-term pharmacological and surgical management. The cysteine-derived gasotransmitter hydrogen sulfide (H 2 S) has interesting vasorelaxant, cytoprotective, antioxidant and anti-inflammatory properties. In this review, we describe the current understanding of PAD pathophysiology and the remarkable benefits of H 2 S against atherosclerosis, inflammation, vascular calcification, and other vasculo-protective effects.

Objective: Hydrogen sulfide is a proangiogenic gas produced primarily by the transsulfuration enzyme cystathionine-γ-lyase (CGL). CGL-dependent hydrogen sulfide production is required for neovascularization in models of peripheral arterial disease. However, the benefits of increasing endogenous CGL and its mechanism of action have not yet been elucidated., Methods: Male whole body CGL-overexpressing transgenic (CGL Tg ) mice and wild-type (WT) littermates (C57BL/6J) were subjected to the hindlimb ischemia model (age, 10-12 weeks). Functional recovery was assessed via the treadmill exercise endurance test. Leg perfusion was measured by laser Doppler imaging and vascular endothelial-cadherin immunostaining. To examine the angiogenic potential, aortic ring sprouting assay and postnatal mouse retinal vasculature development studies were performed. Finally, comparative metabolomics analysis, oxidized/reduced nicotinamide adenine dinucleotide (NAD + /NADH) analysis, and quantitative real-time polymerase chain reaction were performed on CGL WT and CGL Tg gastrocnemius muscle., Results: The restoration of blood flow occurred more rapidly in CGL Tg mice. Compared with the CGL WT mice, the median ± standard deviation running distance and time were increased for the CGL Tg mice after femoral artery ligation (159 ± 53 m vs 291 ± 74 m [ P  < .005] and 17 ± 4 minutes vs 27 ± 5 minutes [ P  < .05], respectively). Consistently, in the CGL Tg ischemic gastrocnemius muscle, the capillary density was increased fourfold (0.05 ± 0.02 vs 0.20 ± 0.12; P  < .005). Ex vivo, the endothelial cell (EC) sprouting length was increased in aorta isolated from CGL Tg mice, especially when cultured in VEGFA (vascular endothelial growth factor A)-only media (63 ± 2 pixels vs 146 ± 52 pixels; P  < .05). Metabolomics analysis demonstrated a higher level of niacinamide, a precursor of NAD + /NADH in the muscle of CGL Tg mice (61.4 × 10 6  ± 5.9 × 10 6 vs 72.4 ± 7.7 × 10 6 area under the curve; P  < .05). Similarly, the NAD + salvage pathway gene expression was increased in CGL Tg gastrocnemius muscle. Finally, CGL overexpression or supplementation with the NAD + precursor nicotinamide mononucleotide improved EC migration in vitro (wound closure: control, 35% ± 9%; CGL, 55% ± 11%; nicotinamide mononucleotide, 42% ± 13%; P  < .05)., Conclusions: Our results have demonstrated that CGL overexpression improves the neovascularization of skeletal muscle on hindlimb ischemia. These effects correlated with changes in the NAD pathway, which improved EC migration., (Copyright © 2023 by the Society for Vascular Surgery. Published by Elsevier Inc.)

Therapies to accelerate vascular repair are currently lacking. Pre-clinical studies suggest that hydrogen sulfide (H 2 S), an endogenous gasotransmitter, promotes angiogenesis. Here, we hypothesized that sodium thiosulfate (STS), a clinically relevant source of H 2 S, would stimulate angiogenesis and vascular repair. STS stimulated neovascularization in WT and LDLR receptor knockout mice following hindlimb ischemia as evidenced by increased leg perfusion assessed by laser Doppler imaging, and capillary density in the gastrocnemius muscle. STS also promoted VEGF-dependent angiogenesis in matrigel plugs in vivo and in the chorioallantoic membrane of chick embryos. In vitro , STS and NaHS stimulated human umbilical vein endothelial cell (HUVEC) migration and proliferation. Seahorse experiments further revealed that STS inhibited mitochondrial respiration and promoted glycolysis in HUVEC. The effect of STS on migration and proliferation was glycolysis-dependent. STS probably acts through metabolic reprogramming of endothelial cells toward a more proliferative glycolytic state. These findings may hold broad clinical implications for patients suffering from vascular occlusive diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Macabrey, Joniová, Gasser, Bechelli, Longchamp, Urfer, Lambelet, Fu, Schwarz, Wagnières, Déglise and Allagnat.)

Arterial occlusive disease is the narrowing of the arteries via atherosclerotic plaque buildup. The major risk factors for arterial occlusive disease are age, high levels of cholesterol and triglycerides, diabetes, high blood pressure, and smoking. Arterial occlusive disease is the leading cause of death in Western countries. Patients who suffer from arterial occlusive disease develop peripheral arterial disease (PAD) when the narrowing affects limbs, stroke when the narrowing affects carotid arteries, and heart disease when the narrowing affects coronary arteries. When lifestyle interventions (exercise, diet…) fail, the only solution remains surgical endovascular and open revascularization. Unfortunately, these surgeries still suffer from high failure rates due to re-occlusive vascular wall adaptations, which is largely due to intimal hyperplasia (IH). IH develops in response to vessel injury, leading to inflammation, vascular smooth muscle cells dedifferentiation, migration, proliferation and secretion of extra-cellular matrix into the vessel's innermost layer or intima. Re-occlusive IH lesions result in costly and complex recurrent end-organ ischemia, and often lead to loss of limb, brain function, or life. Despite decades of IH research, limited therapies are currently available. Hydrogen sulfide (H 2 S) is an endogenous gasotransmitter derived from cysteine metabolism. Although environmental exposure to exogenous high H 2 S is toxic, endogenous H 2 S has important vasorelaxant, cytoprotective and anti-inflammatory properties. Its vasculo-protective properties have attracted a remarkable amount of attention, especially its ability to inhibit IH. This review summarizes IH pathophysiology and treatment, and provides an overview of the potential clinical role of H 2 S to prevent IH and restenosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Macabrey, Longchamp, Déglise and Allagnat.)

Background: Intimal hyperplasia (IH) remains a major limitation in the long-term success of any type of revascularisation. IH is due to vascular smooth muscle cell (VSMC) dedifferentiation, proliferation and migration. The gasotransmitter Hydrogen Sulfide (H 2 S), mainly produced in blood vessels by the enzyme cystathionine- γ-lyase (CSE), inhibits IH in pre-clinical models. However, there is currently no H 2 S donor available to treat patients. Here we used sodium thiosulfate (STS), a clinically-approved source of sulfur, to limit IH., Methods: Low density lipoprotein receptor deleted (LDLR -/- ), WT or Cse-deleted (Cse -/- ) male mice randomly treated with 4 g/L STS in the water bottle were submitted to focal carotid artery stenosis to induce IH. Human vein segments were maintained in culture for 7 days to induce IH. Further in vitro studies were conducted in primary human vascular smooth muscle cells (VSMCs)., Findings: STS inhibited IH in WT mice, as well as in LDLR -/- and Cse -/- mice, and in human vein segments. STS inhibited cell proliferation in the carotid artery wall and in human vein segments. STS increased polysulfides in vivo and protein persulfidation in vitro, which correlated with microtubule depolymerisation, cell cycle arrest and reduced VSMC migration and proliferation., Interpretation: STS, a drug used for the treatment of cyanide poisoning and calciphylaxis, protects against IH in a mouse model of arterial restenosis and in human vein segments. STS acts as an H 2 S donor to limit VSMC migration and proliferation via microtubule depolymerisation., Funding: This work was supported by the Swiss National Science Foundation (grant FN-310030&#95;176158 to FA and SD and PZ00P3-185927 to AL); the Novartis Foundation to FA; and the Union des Sociétés Suisses des Maladies Vasculaires to SD, and the Fondation pour la recherche en chirurgie vasculaire et thoracique., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Objective: Hypertension is a major risk factor for intimal hyperplasia (IH) and re-stenosis following vascular and endovascular interventions. Preclinical studies suggest that hydrogen sulphide (H 2 S), an endogenous gasotransmitter, limits re-stenosis. While there is no clinically available pure H 2 S releasing compound, the sulfhydryl containing angiotensin converting enzyme inhibitor zofenopril is a source of H 2 S. Here, it was hypothesised that zofenopril, due to H 2 S release, would be superior to other non-sulfhydryl containing angiotensin converting enzyme inhibitors (ACEi) in reducing intimal hyperplasia., Methods: Spontaneously hypertensive male Cx40 deleted mice (Cx40 -/- ) or wild type (WT) littermates were randomly treated with enalapril 20 mg or zofenopril 30 mg. Discarded human vein segments and primary human smooth muscle cells (SMCs) were treated with the active compound enalaprilat or zofenoprilat. IH was evaluated in mice 28 days after focal carotid artery stenosis surgery and in human vein segments cultured for seven days ex vivo. Human primary smooth muscle cell (SMC) proliferation and migration were studied in vitro., Results: Compared with control animals (intima/media thickness 2.3 ± 0.33 μm), enalapril reduced IH in Cx40 -/- hypertensive mice by 30% (1.7 ± 0.35 μm; p = .037), while zofenopril abrogated IH (0.4 ± 0.16 μm; p < .002 vs. control and p > .99 vs. sham operated Cx40 -/- mice). In WT normotensive mice, enalapril had no effect (0.9665 ± 0.2 μm in control vs. 1.140 ± 0.27 μm; p > .99), while zofenopril also abrogated IH (0.1623 ± 0.07 μm; p < .008 vs. control and p > .99 vs. sham operated WT mice). Zofenoprilat, but not enalaprilat, also prevented IH in human vein segments ex vivo. The effect of zofenopril on carotid and SMCs correlated with reduced SMC proliferation and migration. Zofenoprilat inhibited the mitogen activated protein kinase and mammalian target of rapamycin pathways in SMCs and human vein segments., Conclusion: Zofenopril provides extra beneficial effects compared with non-sulfhydryl ACEi in reducing SMC proliferation and re-stenosis, even in normotensive animals. These findings may hold broad clinical implications for patients suffering from vascular occlusive diseases and hypertension., Competing Interests: Conflict of interest None., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Currently available interventions for vascular occlusive diseases suffer from high failure rates due to re-occlusive vascular wall adaptations, a process called intimal hyperplasia (IH). Naturally occurring hydrogen sulfide (H 2 S) works as a vasculoprotective gasotransmitter in vivo. However, given its reactive and hazardous nature, H 2 S is difficult to administer systemically. Here, we developed a hydrogel capable of localized slow release of precise amounts of H 2 S and tested its benefits on IH. The H 2 S-releasing hydrogel was prepared from a short peptide attached to an S-aroylthiooxime H 2 S donor. Upon dissolution in aqueous buffer, the peptide self-assembled into nanofibers, which formed a gel in the presence of calcium. This new hydrogel delivered H 2 S over the course of several hours, in contrast with fast-releasing NaHS. The H 2 S-releasing peptide/gel inhibited proliferation and migration of primary human vascular smooth muscle cells (VSMCs), while promoting proliferation and migration of human umbilical endothelial cells (ECs). Both NaHS and the H 2 S-releasing gel limited IH in human great saphenous vein segments obtained from vascular patients undergoing bypass surgery, with the H 2 S-releasing gel showing efficacy at a 5x lower dose than NaHS. These results suggest local perivascular H 2 S release as a new strategy to limit VSMC proliferation and IH while promoting EC proliferation, hence re-endothelialization. STATEMENT OF SIGNIFICANCE: Arterial occlusive disease is the leading cause of death in Western countries, yet current therapies suffer from high failure rates due to intimal hyperplasia (IH), a thickening of the vascular wall leading to secondary vessel occlusion. Hydrogen sulfide (H 2 S) is a gasotransmitter with vasculoprotective properties. Here we designed and synthesized a peptide-based H 2 S-releasing hydrogel and found that local application of the gel reduced IH in human vein segments obtained from patients undergoing bypass surgery. This work provides the first evidence of H 2 S efficacy against IH in human tissue, and the results show that the gel is more effective than NaHS, a common instantaneous H 2 S donor., (Copyright © 2019 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)