Regulation of Vascular Tone, Angiogenesis and Cellular Bioenergetics by the 3-Mercaptopyruvate Sulfurtransferase/H2S Pathway: Functional Impairment by Hyperglycemia and Restoration by dl-α-Lipoic Acid

Abstract

Hydrogen sulfide (H 2 S), as a reducing agent and an antioxidant molecule, exerts protective effects against hyperglycemic stress in the vascular endothelium. The mitochondrial enzyme 3-mercaptopyruvate sulfurtransferase (3-MST) is an important biological source of H 2 S. We have recently demonstrated that 3-MST activity is inhibited by oxidative stress in vitro and speculated that this may have an adverse effect on cellular homeostasis. In the current study, given the importance of H 2 S as a vasorelaxant, angiogenesis stimulator and cellular bioenergetic mediator, we first determined whether the 3-MST/H 2 S system plays a physiological regulatory role in endothelial cells. Next, we tested whether a dysfunction of this pathway develops during the development of hyperglycemia and diabetes-associated vascular complications. Intraperitoneal (IP) 3-MP (1 mg/kg) raised plasma H 2 S levels in rats. 3-MP (10 μmol/L to 1 mmol/L) promoted angiogenesis in vitro in bEnd3 microvascular endothelial cells and in vivo in a Matrigel assay in mice (0.3-1 mg/kg). In vitro studies with bEnd3 cell homogenates demonstrated that the 3-MP-induced increases in H 2 S production depended on enzymatic activity, although at higher concentrations (1-3 mmol/L) there was also evidence for an additional nonenzymatic H 2 S production by 3-MP. In vivo, 3-MP facilitated wound healing in rats, induced the relaxation of dermal microvessels and increased mitochondrial bioenergetic function. In vitro hyperglycemia or in vivo streptozotocin diabetes impaired angiogenesis, attenuated mitochondrial function and delayed wound healing; all of these responses were associated with an impairment of the proangiogenic and bioenergetic effects of 3-MP. The antioxidants DL-α-lipoic acid (LA) in vivo, or dihydrolipoic acid (DHLA) in vitro restored the ability of 3-MP to stimulate angiogenesis, cellular bioenergetics and wound healing in hyperglycemia and diabetes. We conclude that diabetes leads to an impairment of the 3-MST/H 2 S pathway, and speculate that this may contribute to the pathogenesis of hyperglycemic endothelial cell dysfunction. We also suggest that therapy with H 2 S donors, or treatment with the combination of 3-MP and lipoic acid may be beneficial in improving angiogenesis and bioenergetics in hyperglycemia.

Regulation and role of endogenously produced hydrogen sulfide in angiogenesis

“…As has been described for other agents that enhance endothelial cell proliferation, H 2 S was reported to inhibit smooth muscle cell growth [73]. Similarly to what was seen with L-cysteine, endothelial cells grown in the presence of 3-MP, the 3MST substrate, exhibit higher growth rates [74]. On the other hand, silencing of 3-MST via short hairpin RNA (shRNA) lentiviral particles, resulted in a significant inhibition of ECs proliferation rate [74].…”

“…Similarly to what was seen with L-cysteine, endothelial cells grown in the presence of 3-MP, the 3MST substrate, exhibit higher growth rates [74]. On the other hand, silencing of 3-MST via short hairpin RNA (shRNA) lentiviral particles, resulted in a significant inhibition of ECs proliferation rate [74]. The above findings taken together provide strong evidence that H 2 S is an endogenous endothelial cell mitogen and indicate that the growth-promoting properties of H 2 S are independent of the enzymatic source of the gasotransmitter.…”

Delayed Treatment with Sodium Hydrosulfide Improves Regional Blood Flow and Alleviates Cecal Ligation and Puncture (CLP)-Induced Septic Shock

“…The concentration of H 2 S in plasma was quantified with the H 2 S-specific fluorescent probe 7-azido-4-methylcoumarine (AzMC) as described (13). Whole blood was collected 30 min later in K 2 EDTA blood collection tubes and centrifuged at 4 °C for 15 min at 2000g.…”

The role of H2S bioavailability in endothelial dysfunction

“…However, CSE mRNA is unaltered in the thoracic aorta of diabetic rats [54]. There are also no changes in the expression of CSE, CBS or MST in endothelial cells exposed to elevated extracellular glucose, or in the thoracic aorta of streptozotocin-diabetic rats [50, 55]. In fact, CSE expression in cerebral microvessels was increased in type I diabetes [56].…”

“…Recent data indicate that the endothelial MST was inhibited or inactivated during hyperglycemia, leading to impaired endothelial cell H 2 S production, suppressed angiogenesis and attenuation of mitochondrial function [55]. Moreover, cellular levels of H 2 S are determined by the balance of its production and consumption/elimination [57].…”