[SIGNALING PATHWAYS INVOLVED IN THE REGULATION OF PROTEIN METABOLISM IN SKELETAL MUSCLE].

The ability of muscle cells to respond to certain signaling molecules depends on the presence of specific proteins-receptors, ion channels and transporters, which create a system of metabolic inputs. All cells in the human body have a transmembrane signaling system that allows to get information from extracellular stimuli such as hormones, neurotransmitters, proteins, peptides, derivatives of amino acids and fat acids, sensor molecules. The results of researches published in the last two years, which expand knowledge of action of various factors on protein metabolism in skeletal muscles are presented in the review. The main attention is turned to the discovery and expansion of signaling pathways involved in the regulation of protein metabolism in skeletal muscle in different functional states of the person.

[THE ROLE OF microRNAs IN THE REGULATION OF MUSCLE METABOLTSM].

MicroRNAs, class of the small RNA containing approximately 22 nucleotides take part in regulation of metabolism of skeletal muscles as enzymes, hormones and transcription factors. They are able to regulate the intensity of the translation process through complementary. interaction with mRNA. The review presents the results of studies published in the last two years, which increase the knowledge of the effects of microRNAs on the key stages of protein synthesis in skeletal muscle. The main attention is paid to achievements which open and specify a microRNA role in regulation of the signaling pathways that coordinate intracellular metabolism of skeletal muscles at different finctionni onneiilinnc nf then

[Molecular mechanisms of skeletal muscle hypertrophy].

Enzymes Akt, AMPK, mTOR, S6K and PGC-1a coactivator take part in skeletal muscles in the regulation of synthesis of proteins. The expression of these proteins is regulated by growth factors, hormones, nutrients, mechanical loading and leads to an increase in muscle mass and skeletal muscle hypertrophy. The review presents the results of studies published in the past four years, which expand knowledge on the effects of various factors on protein synthesis in skeletal muscle. The attention is focused on the achievements that reveal and clarify the signaling pathways involved in the regulation of protein synthesis in skeletal muscle. The central place is taken by mTOR enzyme which controls and regulates the main stages of the cascade of reactions of muscle proteins providing synthesis in the conditions of human life. coactivator PGC-1a.

[The role of mTOR in the regulation of skeletal muscle metabolism].

mTOR enzyme belongs to a family of specific serine/threonine protein kinases. mTOR plays an important role in the transmission of extracellular signals by phosphorylation of various substrates in many metabolic reactions in humans. Protein mTOR, having protein kinase activity, is encoded by a gene FRAP1, which is localized on chromosome 1 (1p36.2) in skeletal muscle. It exists in two mTOR protein complexes mTORC1 and mTORC2 with various sensitivity to the inhibitory effect of rapamycin. mTOR regulates metabolism in skeletal muscle by phosphorylation of various of protein metabolizing enzymes as well as transcription and translation factors. mTOR expression occurs in response to metabolic requests of muscles and leads to increased metabolism of proteins. The data of recent studies suggest an important role of mTOR in the regulation of intracellular metabolism and point to the need of studying the molecular mechanisms involved in physiological function of skeletal muscles.

[Involvement of Akt in regulation of skeletal muscle metabolism].

The enzyme Akt 1 is a member of serine/threonine specific protein kinases family. Akt plays an important role in the transmission of extracellular signals and mechanical stimuli via phosphorylation of its numerous substrates involved in anabolic and catabolic processes as well as mechanical responses of skeletal muscle. Expression of the Akt is increased during skeletal muscle hypertrophy and decreased during muscle atrophy. The G205T polymorphism of Akt is associated with transcriptional activity of the gene in muscle cells and, respectively, with skeletal muscle phenotypes.

[Participation AMPK in the regulation of skeletal muscles metabolism].

Enzyme AMPK is a part of the family of serine/threonine specific protein kinases. AMPK plays important role in the transfer extracellular signals through phosphorylation of multiple substrates in different metabolic reactions of skeletal muscles. AMPK is geterotrimetric complex, consisting of the catalytic subunit (AMPKalpha) and two regulatory subunits (AMPKbeta and AMPKgamma), which are encoded by seven different high-homologous genes (alpha1, alpha2, beta1, beta2, gamma1, gamma2, gamma3). AMPK regulates skeletal muscle metabolism through phosphorylation of various enzymes such as carbohydrate, lipid and protein metabolism, as well as factors of transcription and initiation. The AMPK expression occurs in response to a changing metabolic requests muscle cells and it leads to increased energy metabolism. The data of recent studies suggest the important role of AMPK in the regulation of intracellular metabolism and point to the need to study the molecular mechanisms involved in the regulation of gene expression in skeletal muscle.

[Association of the mitochondrial transcription factor (TFAM) gene polymorphism with physical performance of athletes].

The aim of the study was to investigate allelic distribution of the mitochondrial transcription factor gene (TFAM) Ser12Thr polymorphism in athletes (n = 1537) and controls (n = 1113), and to find interrelation between genotypes and aerobic capacity in rowers (n = 90). Genotyping was performed by restriction fragment length polymorphism analysis. Aerobic capacity (maximal oxygen consumption (VO2max) and maximal power production capacity (Wmax)) was determined using an incremental test to exhaustion by rower ergometer. The frequency of TFAM 12Thr allele was significantly higher in endurance-oriented athletes (n = 588) than in controls (14.0% vs. 9.1%; p <0.0001), and increased with the growth of skills. Furthermore, TFAM 12Thr allele was associated with high values of aerobic performance (when Wmax and VO2max were measured). Thus, TFAM gene Ser12Thr polymorphism is associated with physical performance of athletes.

The ACTN3 R577X polymorphism in Russian endurance athletes.

OBJECTIVE: The functional 577R allele of the alpha-actinin-3 (ACTN3) gene has been reported to be associated with elite power athlete status, while the nonfunctional 577XX genotype (predicts an alpha-actinin-3 deficient phenotype) has been hypothesised as providing some sort of advantage for endurance athletes. In the present study, the distribution of ACTN3 genotypes and alleles in Russian endurance-oriented athletes were examined and association between ACTN3 genotypes and the competition results of rowers were sought. METHODS: 456 Russian endurance-oriented athletes of regional or national competitive standard were involved in the study. ACTN3 genotype and allele frequencies were compared with 1211 controls. The data from the Russian Cup Rowing Tournament were used to search for possible association between the ACTN3 genotype and the long-distance (approximately 6 km) rowing results of 54 athletes. DNA was extracted from mouthwash samples. Genotyping for the R577X variant was performed by PCR and restriction enzyme digestion. RESULTS: The frequencies of the ACTN3 577XX genotype (5.7% vs 14.5%; p<0.0001) and 577X allele (33.2% vs 39.0%; p = 0.0025) were significantly lower in endurance-oriented athletes compared with the controls, and none of the highly elite athletes had the 577XX genotype. Furthermore, male rowers with ACTN3 577RR genotype showed better results (1339 (11) s) in long-distance rowing than carriers of 577RX (1386 (12) s) or 577XX (1402 (10) s) genotypes (p = 0.016). CONCLUSION: Our data show that the ACTN3 577X allele is under-represented in Russian endurance athletes and is associated with the rowers' competition results.

[The analysis of PPARGC1B gene polymorphism in athletes].

The aim of the study was to investigate allelic distribution of the peroxisome proliferator-activated receptor gamma coactivator 1beta gene (PPARGC1B; Ala203Pro polymorphism) in athletes (n = = 1535) and controls (n = 1113), and to find interrelation between genotypes and aerobic performance of athletes. Genotyping was performed by restriction of the fragment length polymorphism. Aerobic capacity was determined using incremental test to exhaustion by rower ergometer. The frequency of PPARGC1B 203Pro allele was significantly higher in endurance-oriented athletes (n = 582; 7.2 %, p = 0.007), athletes with acyclic activity (n = 488; 8.1%, p = 0.0005), and power-oriented athletes (n = 372; 7.1%, p = 0.026) than in controls (4.9%), and it increased with the growth of skills (p = 0.046). Furthermore, we found that PPARGCIB 203Pro allele was associated with higher values of aerobic performance (when oxygen pulse and maximal power production capacity were measured). Thus, the PPARGC1B gene Ala203Pro polymorphism is associated with physical performance of athletes.

[The influence of calcineurin gene polymorphism on morphofunctional characteristics of cardiovascular system of athletes].

The aim of the study was to investigate allelic distribution of calcineurin gene (CNB; 5I/5D polymorphism) in athletes of cyclic sports (n = 673) and controls (n = 1073), and to find interrelation between genotypes and risk of left ventricular hypertrophy and physical performance of athletes. Genotyping was performed by restriction fragment length polymorphism. Echocardiography was performed by ultrasound scanner Aloka-3500. Physiological parameters were evaluated by spiroergometry using MetaMax 3B Gas Analyzer. The frequency of CNB D allele was significantly lo- wer in athletes than in controls (5.5 vs. 8.8 %; p = 0.0005), and decreased with the growth of skills. We found that CNB D allele was associated with increased left ventricular mass index and low values of physical performance (when maximal power production capacity and maximal oxygen consumption were measured). Thus, CNB gene 5I/5D polymorphism is associated with the risk of left ventricular hypertrophy and physical performance of athletes.

PPARG Gene polymorphism and locomotor activity in humans.

The distribution of PPARG gene allele frequencies (Pro/Ala polymorphism) was studied in sportsmen specialized in speed and force athletics. A relationship between genotypes and human muscle transverse section area was evaluated. The PPARG Ala allele was significantly more incident in athletes than in controls, the incidence increasing with higher athletic qualification. A hypertrophic effect of PPARG Ala allele on muscle fibers was detected. Hence, the PPARG Pro12Ala polymorphism is associated with human motor activity.

Effect of HIF1A gene polymorphism on human muscle performance.

Allele distribution of hypoxia-inducible factor gene (HIF1A; Pro582Ser polymorphism) was studied in power-oriented athletes and controls practicing no athletics; genotype relationships with muscle fiber composition were studied in speed skaters. Genotyping was carried out by PCR. The composition of muscle fibers was evaluated by the immunohistochemical analysis of m. vastus lateralis. The incidence of HIF1A Ser allele was significantly higher in weight-lifters than in controls (17.9 vs. 8.5%; p=0.001) and increased with athletic skill improvement. A relationship between HIF1A Ser allele and predominance of fast-twitch muscle fibers was shown (Pro/Ser 46.2 (13.8)%, Pro/Pro 31.4 (8.2)%; p=0.007). Hence, HIF1A Pro582Ser polymorphism is associated with muscle activity in humans.

[Association of regulatory genes polymorphisms with aerobic and anaerobic performance of athletes].

The aim of study was to investigate an allelic distribution of PPARA (G/C polymorphism), PPARG (Pro/Ala), PPARD (+294T/C) and PGCIA (Gly482Ser) genes in rowers (n=205) and controls (n=659), and to find correlation between genotypes and physiological parameters. Genotyping was performed by restriction fragment length polymorphism analysis. Physiological parameters were evaluated by PM 3 Rower Ergometer and MetaMax 3B Gas Analyzer. The frequencies ofPPARA G (90.1% vs. 83.6%) and PPARG Ala (23.1% vs. 16.2%) alleles in elite athletes, and of PPARD C (19.1% vs. 10.5%) and PGC1A Gly (75.4% vs. 66.5%) alleles in sub-elite athletes were significantly higher than in controls. Moreover, PPARA G (when oxygen pulse was measured) and PGC 1A Gly (when maximal aerobic power and anaerobic threshold (%) of VO2max were measured) alleles were associated with high values of aerobic performance. Thus, PPARA G, PPARG Ala, PPARD C and PGCIA Gly alleles can be considered as genetic markers associated with enhanced physical performance.

[Association of PPARD gene polymorphism with human physical performance].

Peroxisome proliferator-activated receptor delta (PPARdelta) regulates expression of genes involved in lipid and carbohydrate metabolism. To examine the association of a functional +294T/C polymorphism of PPARD gene with human physical performance, we have studied the distribution of PPARD alleles and genotypes in a cohort of athletes (n=1256; stratified by specialty and skill level) and controls (n=610). We found that the frequency of PPARD C allele (with higher transcriptional activity compared to T allele) in a group of endurance-oriented athletes (n=898) is significantly higher than in controls (18.3% vs. 12.1%; p < 0.0001). Moreover, in the group of endurance-oriented athletes with cyclic activity we revealed an increasing frequency of PPARD C allele with the rising of athletes' skill level. Thus, PPARD C allele is associated with predisposition to endurance performance.

[The association of gene polymorphisms with the muscle fiber type composition].

Muscle fiber composition of m. vastus lateralis has significant individual variability mainly depending on genetic factors. Present study shows analysis of association between polymorphisms of three muscle performance-related genes and muscle fiber type composition in 48 young healthy men. DNA was obtained from mouthwash samples by alkaline extraction. Polymorphism determination of PPARalpha, ACE and ACTN3 genes was performed using polymerase chain raction. Muscle fiber typing from m. vastus lateralis was performed using immunohistochemistry method. We found an association of increased frequency of intron 7 G allele of PPARalpha gene (93.9% vs 60.0%) and D allele of ACE gene (68.8% vs 34.4%) in the group with the highest proportion of slow-twitch fibers (56-70%) compared to the group with the lowest proportion (25-43%). Thus, PPARalpha and ACE genes can be considered as potential candidate genes for muscle fiber type determination.

Properties of free and occupied androgen receptor in rat skeletal muscle cytosol: effect of testosterone.

Our aim was to investigate the effect of a single testosterone (T) injection on the androgen receptor (AR) in rat skeletal muscle (SM) cytosol. The properties of AR were studied in order to establish the protocol for differential determination of free and hormone-occupied AR in SM cytosols from non-hormone-deficient animals. Using the developed ligand-exchange protocol, we demonstrated that injection of T (1 mg/kg) caused alternating changes of the total AR binding. The binding minimum (23% of the control) was measured 1 h after the injection. It was followed by pronounced and lasting elevation of the AR binding. In the control cytosols, AR complexes constituted approximately 25% of the total receptor content. Changes of their relative content immediately after T administration were consistent with rapid nuclear translocation of the AR. Inhibition of protein synthesis by cycloheximide (CHI) injection demonstrated that delayed and lasting increase of the AR binding after T injection partially depended on the stimulated protein synthesis. Altogether, the obtained evidence supports the assumption that the AR mediates elevation of its own gene expression in SM upon administration of T.

Neutrophil antiserum response to decrease in proteolytic activity in loaded rat muscle.

The leukocytes that are found in skeletal muscles after intense muscular activity have been shown to infiltrate areas of injury in skeletal muscle tissue. We believe that leukocyte enzymes, which appear in the tissue after the degranulation or destruction of leukocytes, could play a role in tissue enzyme activities. Neutrophil proteinases were investigated. Histological data provided evidence of rat muscular tissue infiltration by leukocytes after intense physical loading. To reduce the influx of leukocytes in rat muscles, a rabbit antiserum against rat peritoneal leukocytes was injected into rats after muscle loading. This resulted in a reduction in muscle cytosol proteolytic activity as compared to control animals (who received saline injections). The levels of proteolytic activities of media conditioned by the soleus muscle isolated from antiserum-treated rats were also reduced. These data provide evidence that the increase in proteolytic activity observed in rat skeletal muscles after physical loading is partially induced by neutrophil proteinases.

The angiotensin converting enzyme I/D polymorphism in Russian athletes.

The deletion (D) allele of the human ACE gene is associated with higher ACE activity than the insertion (I) allele. There is controversy as to whether the ACE genotype may be associated with elite athletic status; recent studies have identified no significant associations amongst those drawn from mixed sporting disciplines. However, such lack of association may reflect the mixed nature of such cohorts, given that an excess frequency of the I allele has been reported amongst elite endurance athletes, and an excess of the D allele amongst those engaged in more power-orientated sports. We examined this hypothesis by determining ACE I/D allele frequency amongst 217 Russian athletes (swimmers, skiers, triathletes and track-and-field participants) prospectively stratified by performance ('outstanding' or 'average'), and the duration of their event (SDA (<1 min), MDA (1 to 20 min), and LDA (>20 min): short, middle and long distance athletes respectively). ACE genotype and allele frequencies were compared to 449 controls. ACE genotype frequency amongst the whole cohort, or the outstanding athletes alone, was no different to that amongst sedentary controls. However, there was an excess of the D allele (frequency 0.72, P=0.001) amongst the outstanding SDA group, and an excess of the I allele (frequency 0.63, P=0.032) amongst the outstanding MDA group. These findings were replicated in the outstanding swimmers, with track and field SDA similarly demonstrating an excess of the D allele (P=0.01). There was no association found between the outstanding LDA and ACE genotype (P=0.27). These data not only confirm an excess of the D allele in elite SDA, and I allele in elite MDA, but also offer an explanation as to why any such association may be hard to detect amongst a heterogeneous cohort of mixed athletic ability and discipline.