Modern data on biochemical patterns of nitric oxide biosynthesis in mammal cells from l-arginine in normoxic conditions is described. The attention of the authors is given to the results of the recent years on the structure and regulation features isoforms of No-synthase. The emphasis is put on the latest conception of the compartmentalization of certain isoforms of these enzymes in cells and on the possibility of the directed transport of nitric oxide in the vascular wall. The central place in the review is devoted to issues on the endogenous formation of NO in mitochondria and its potential physiological significance. Our own results on the identification of NO in mitochondria of the uterine smooth muscle, biochemical characteristics of this process and NO possible role in Са 2+ transport regulation by organelles are presented and discussed. K e y w o r d s: nitric oxide, mitochondria, No-synthases, cell compartments, calcium, smooth muscle. I n foreign scientific literature and, partly, in national periodicals, there are numerous review papers devoted to various aspects of the formation and biological activity of nitric oxide and its derivatives (nitric active forms). In view of this, in the presented article the authors tried to provide general ideas about the biosynthesis regularities and the role of NO in a cell in the most summarized version, primarily focusing on the problems that have become relevant in the last period of research, such as compartmentalization of certain isoforms of NO-synthase in cells and the possibility of directed transport of nitric oxide in the vascular wall. Particular attention is paid to the less highlighted and investigated problem -mitochondria own synthesis of nitric oxide and its possible functional activity in this compartment. Available material relates only to the synthesis of NO from L-arginine in the presence of O 2 and does not include important issues on reductase activity in hypoxic conditions and the functioning of the nitric oxide cycle in the mammalian organism. Numerous works by prof. Reutov V. P. and, for example, a review article in Ukrainian are devoted to these problems [1].Nitric oxide (NO) is a structurally simple low molecular amphiphilic free radical molecule. In biosystems NO has a relatively short time of existence (up to 5 sec depending on microenvironment) and can migrate at short distances from the generation sites what is determined by the rate of oxidation [2]. This limitation of diffusion implies the specificity of NO effect in individual cell compartments due to the colocalization of target proteins with NO sources in a multiprotein complex of signalosomes.Synthesis of NO in a cell is provided by the family of isoforms of NO-synthase (NOS), which, with the participation of NADPH as an electron source and the presence of O 2 , carry out a five-electron two-stage oxidation of the L-arginine guanidine group with the formation of NO and L-citrulline [2][3][4][5][6][7].NO is an almost universal messenger and regulatory molecule. NO signaling is carrie...
Aim. To demonstrate the possibility of NO synthesis in intact myocytes of uterus. Methods. Confocal scanning microscopy method, NO-sensitive fl uorescent probe DAF-FM, MitoTracker Orange CM-H 2 T-MRos. Results. The basal production of NO in intact myocytes was shown using DAF-FM. Incubation of myocytes with NO donor -sodium nitroprusside (SNP) -led to an increase of the DAF-FM-T fl uorescent signal. On the contrary, the addition of NO-synthase inhibitor -N-nitro-L-arginine (NA) -results in the reduction of fl uorescent intensity. It was demonstrated colocalizition of specifi c probe for mitochondria MitoTracker Orange CM-H 2 TMRos and NO-sensitive dye DAF-FM. Conclusions. For the fi rst time it has been demonstrated the presence of NO in smooth muscle cell mitochondria using laser confocal microscopy, NO-sensitive probe DAF-FM and specifi c marker of the functionally active mitochondria MitoTracker Orange CM-H 2 TMRos. K e y w o r d s: nitric oxide, myometrium, DAF-FM, confocal microscopy.
the present study was undertaken to evaluate [ca 2+ ] i signals that occur in human sperm cells exposed in vitro to three diverse compounds; progesterone, 4-aminopyridine (a highly effective inducer of hyperactivation in human sperm) and tetraethylammonium. the [ca 2+ ] i reached after the extracellular calcium treatment was always higher in normozoospermic samples pretreated with progesterone than in pathozoospermic samples pretreated with progesterone. there were no changes in calcium signal in spermatozoa pretreated with progesterone from patients with oligozoospermia and leucocytospermia compared to control samples (without progesterone). [ca 2+ ] i . was always higher in pathozoospermic samples without 4-aminopyridine and always lower in pathozoospermic samples with 4-aminopyridine compared to these values in normozoospermic men. the 2 mM extracellular calcium administration to spermatozoa pretreated with tetraethylammonium did not result in a detectable increase in [ca 2+ ] i in normo-and pathozoospermic samples. the mechanisms of progesterone-dependent activation of the ca 2+ -entry and the functioning of the voltage gated ca 2+ -channels of plasmalemma are disturbed in pathologies -there was no increase in the ca 2+ level in the conditions of k + -depolarization (in the presence of inhibitors of k + -channels). k e y w o r d s: calcium, spermatozoa, progesterone, inhibitors of k + channels. R eproductive problems occur in 15% of couples. Male infertility is up to 50% of all infertility cases. Oligozoospermia and leukocytospermia are the most common causes of male infertility. Oligozoospermia is low spermatozoa concentration in ejaculate (< 20·10 6 cells/ml). Leukocytospermia (the presence of leucocytes ≥ 1·10 6 cells/ml) is a well-known indicator of infectious process in the genitourinary tract which affects male reproductive function and fertility [1]. Up to now the data regarding ionic aspects of male infertili ty remain poorly understood. Calcium is the major second messenger which plays an important role in sperm physiology. Fluctuations in intracellular Ca 2+ concentration ([Ca 2+ ] i ) mediate transmission of information from receptors on plasmatic mem-brane to nucleus or other internal response systems [2]. The ability of [Ca 2+ ] i to encode and mediate rapid transmission of information depends on the ability of the cell to generate calcium signals of sufficient precision and complexity. In particular, regulation of sperm motility, hyperactivation and chemotaxis depends on intracellular calcium level [3]. Hyperactivation facilitates progressive movement of spermatozoa and enables them to penetrate oocyte. It is also associa ted with capacitation leading to acrosome reac tion. Disturbances in Ca 2+ signalling in sperm are associated with male subfertility [4]. In fact, abnormal motility might be explained by abnormally low cytoplasmic calcium [5]. Sperm cells are not able to perform either of these crucial functions in the absence of extracellular Ca 2+ [6]. However, mo-
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