Stimulation of the endogenous hydrogen sulfide synthesis suppresses oxidative-nitrosative stress and restores endothelial-dependent vasorelaxation in old rats.

Hydrogen sulfide (H2S) is an endogenous gas transmitter with profound effects on the cardiovascular system. We hypothesized that stimulation of H2S synthesis might alleviate age-associated changes in vascular reactivity. Pyridoxal-5-phosphate (PLP), the coenzyme of H2S-synthesizing enzymes, was administrated to old male Wistar rats per os at a dose of 0.7 mg/kg body mass once a day for 2 weeks. H2S content in the aortic tissue, markers of oxidative stress, inducible nitric oxide synthase (iNOS) and constitutive nitric oxide synthase (cNOS), arginase activities, and endothelium-dependent vasorelaxation of the aortic rings were studied. Our results showed that PLP restored endogenous H2S and low molecular weight S-nitrosothiol levels in old rat aorta to the levels detected in adults. PLP significantly reduced diene conjugate content, hydrogen peroxide and peroxynitrite generation rates, and iNOS and arginase activity in the aortic tissue of old rats. PLP also greatly improved acetylcholine-induced relaxation of old rat aorta (47.7% ± 4.8% versus 18.4% ± 4.1% in old rats, P < 0.05) that was abolished by NO inhibition with N-nitro-l-arginine methyl ester hydrochloride (L-NAME) or H2S inhibition with O-carboxymethylhydroxylamine (O-CMH). Thus, PLP might be used for stimulation of endogenous H2S synthesis and correction of oxidative and nitrosative stress and vessel tone dysfunction in aging and age-associated diseases.

[PYRIDOXAL-5-PHOSPHATE RESTORES HYDROGEN SULFIDE SYNTHES AND REDOX STATE OF HEART AND BLOOD VESSELS TISSUE IN OLD ANIMALS].

It was shown the alterations in hydrogen sulfide (H(2)S) metabolism and the development of oxidative and nitrozative stress in cardiovascular system by aging. The administration of pyridoxal-5-phosphate as cofactor of H(2)S synthesizing enzymes restored endogenous H(2)S level and redox state in the heart and aorta tissues. Under these conditions, the following indicators of oxidative stress were significantly decreased in heart and aorta tissues: superoxide generation rate (·0(2)(-)) and hydroxyl (·OH) anion radicals, compared with significantly elevated levels of these parameters in old animals. We also found the reduction of non-enzymatic (diene conjugates and malonic dialdehyde) and enzymatic (uric acid, LTC(4) and TxB(2)) lipid oxidation products levels in old rats under H(2)S synthesis stimulation that confirms the restriction of oxidative stress. An important consequence of endogenous synthesis stimulation of hydrogen sulfide during aging is a decrease of nitrozative stress, such as iNOS activity and nitrate reductase, as well as recovery of constitutive NO synthase activity, indicating the importance of this gas transmitter in cardiovascular system. Thus, stimulation of hydrogen sulfide endogenous synthesis contributed to reduced production of reactive oxygen species (oxidative stress) and nitrogen (nitrozative stress) in heart and aorta tissues with aging. The presence of a pronounced antioxidant effect and modulating influence of pyridoxal-5- phosphate in the redox state of heart tissue and blood vessels during aging suggests cardioprotective properties of the substance and prospects for future research.

MITOCHONDRIAL DYSFUNCTION IN THE AGING HEART IS ACCOMPANIED BY CONSTITUTIVE NO-SYNTHASES UNCOUPLING ON THE BACKGROUND OF OXIDATIVE AND NITROSATIVE STRESS.

We investigated the sensitivity of mitochondrial permeability transition pore (MPTP) opening to its inductor calcium, as well as indicators of oxidative and nitrosative stress in adult and old rat heart mitochondria and heart tissues. The coupling index of constitutive Ca2+/calmodulin-dependent NO-synthase (сNOS) was calculated based on experimentally found parameters. The aging is characterized by oxidative and nitrosative stress, which accompanied by a decrease of the cNOS coupling index and an increased sensitivity of MPTP to calcium. We found that in the heart mitochondria of old rats such oxidative stress indicators as the rate of generation of superoxide (• O2 - ) and hydroxyl (• OH) anion radicals were significantly increased (in 4 and 2,7 times respectively). Also, increased levels of urea and products of early lipid peroxidation ­ conjugated dienes display the intensification of oxidative stress. Such indicators of nitrosative stress as nitrate reductase and iNOS activity were also enhanced. At the same time under aging the NO2 - pools, generated when the heart oxygenation is normal, nitrosothiols pools and the activity of cNOS were decreased. It is due to the enhanced level of cNOS uncoupling, resulting in increased oxidative stress. It was also shown the lower level of hydrogen sulfide (H2S) in old rat heart mitochondria. Thus, we observed the increased sensitivity of MPTP to calcium, due to decreased concentration of its inhibitors nitric oxide and hydrogen sulfide and increased levels of its inducers ROS and RNS in old rat heart mitochondria.

[New fluorine-containing openers of ATP-sensitive potassium channels flokalin and tioflokalin inhibit calcium-induced mitochondrial pore opening in rat hearts].

In experiments in vitro on the mitochondria isolated from the rat's heart we studied the effects of the openers of ATP-sensitive potassium channels (K(ATP)-channels), flocalin and tioflocalin, on the calcium-induced mitochondrial pore (MPTP) opening. Flocalin and tioflocalin caused moderate Ca(2+)-independent mitochondria swelling, which was prevented by a specific inhibitor of 5-hydroxydecanoate. This allowed to identify these compounds as mitochondrial K(ATP)-channels openers. We found that concentration-dependent inhibitory effects (10(-7) to 10(-4) M) of flocalin (with IC50 = 50 microM) and tioflocalin (with IC50 = 2,7 microM) on Ca(2+)-induced mitochondrial swelling (MPTP opening) in the heart characterized more powerful cardioprotective action of the latter. It was shown that the administration of these compounds in experiments in vivo decreased the sensitivity of the MPTP opening to Ca2+. Thus, under physiological conditions the activators K(ATP)-channels probably provide the membrane-stabilizing effects, thereby effectively increasing the organelles resistance to Ca2+, an inductor of MPTP. The results obtained allowed to characterize the role of the compound studied as cardioprotectors and regulators of the MPTP formation in the heart, indicated their anti-ischemic and anti-apoptotic effects that can be used in order to correct the mitochondrial dysfunction under pathological conditions of the cardiovascular system.