[A new mutation c.422C>G (p.S141C) in homo- and heterozygous forms of the human leptin gene].

Mutation g.15409C>G, c.422C>G (p.S141C) in homo- and heterozygous forms of the human LEP gene was identified among some patients of the high mountain village of Karaul, Ashkhabad oblast, Turkmenistan, some of which suffer from adiposity. It causes the substitution S120C in the secreted leptin. The mature leptin molecule (146 aa) has only two Cys residues (C96 and C146) forming an S-S bridge, which is important for the hormone function. A third mutation, C120, in the molecule might disturb the correct formation of the S-S bond and could alter the leptin activity.

[The interconnections of molecular mechanisms of hormone actions and their role in pathogenesis of obesity, insulin resistance, and diabetes mellitus].

The various hormones, proteins and other compounds related to developing obesity, insulin resistance and type 2 diabetes are analyzed in the paper. 1) Leptin, ciliary neurutrophic factor, adiponectin, glucagon-like peptide 1, peptide YY, neuromedin S, as well as the protein receptors of these hormones decrease the food consumption, increase the energy turnover, and prevent obesity, insulin resistance, and type 2 diabetes development. The mediators of these hormone and receptor actions are melanocyte stimulating hormone (MSH), corticotropin-releasing hormone (CRH), and the others. 2) Ghrelin, endogenose cannabinoides, galanin-like peptide and the mediators of their actions: neuropeptide Y (NPY) and Agouti gene related protein (AGRP) increase the appetite and food consumption. Peroxisome proliferation-activated receptor (PPAR) performs the similar action on food intake. The activation of the first group compound functioning decreases the obesity, increases the energy turnover, facilitates the insulin action and prevents the insulin resistance and type 2 diabetes. Increasing the activities of the second group, as well as, decreasing the actions of the first one of substances induce the opposite effects and facilitate obesity, insulin resistance, and type 2 diabetes developments. The interconnections of the molecular mechanisms of so many hormone actions make the very complicated tusk to study the various endocrine disorders including diabetes mellitus as well.

[Molecular and genetic study of the role of hormones, receptors, and enzymes in regulation of reproduction, lipid metabolism, and other human physiological functions].

Prevalence of uterine progesterone receptors over estrogen ones, high uterine cAMP level, and low uterine prostaglandin level are necessary conditions of normal pregnancy. In cases of spontaneous and antiprogestin RU486-induced abortions, estrogen receptors prevail over progesterone ones, cAMP level decreases, and prostaglandin concentration in decidual tissue increases. Porcine and bovine beta-lipotropines were the first proteins, whose correct amino acid sequence was first determined in Russia. Several research centers carried out collaborative studies of the nucleotide sequences of human and animal proopiomelanocortin (lipotropin precursor) and prolactin cDNA. Researchers constructed genetic engineering producers of human pre-proinsulin and somatostatin, identified structural genes expressed in pancreatic beta-cells, studied antigenic properties of glutamic acid decarboxylase (GAD), which determine insulin-dependent diabetes, and identified the cholesterase determinant. They revealed mutations in the genes of proopiomelanocortin and melanocortin receptors (MC4-P), which inhibit leptin regulation of appetite and are associated with human obesity.

[Screening of mutations in genes of pro-opiomelanocortin in patients with constitutional exogenous obesity]. / Skrining mutatsii v gene proopiomelanokortina u bol'nykh konstutsional'no-ékzogennym ozhireniem.

Proopiomelanocortin (POMC) is a precursor of ACTH, beta- and gamma-liportopins, alpha-, beta- and gamma-MSH, beta-endorphin. alpha-, beta- and gamma-MSH are synthesized by hypothalamus neurons, and leptin stimulates their synthesis. These hormones regulate food consumption and energy metabolism by via melanocortin receptors (MC3-R and MC4-R) in hypothalamus. Screening mutations in the coding region of human POMC has been carried out with PCR, SSCP and DNA sequencing and the association study of these mutations and human obesity has been performed. Group of patients with the exogenous obesity (BMI 37.8 +/- 6.8 kg/m2) consisted of 228 persons (173 women and 55 men). 145 blood donors (67 women and 78 men) without obesity (BMI J25 kg/m2, 23.1 +/- 2.2 kg/m2) and 170 women without apparent obesity at the beginning of the study were included in the control group. 8 polymorph sites: insertions; missense and silent mutations have been identified in the coding region of POMC. Among them 1) two heterozygous mutations: the insertion of 6 b.p. (GGGCCC) in codon 176 inducing the insertion of two amino acid residues (Arg-Ala) in POMC and nonsense mutation (G-7316-T) in codon 180 of gamma-LTH coding region of the same DNA chain were identified in 4 women (5.8%) out of 69 patients with morbid obesity (BMI 40-53 kg/m2). These mutations were not found in control (n = 315). 2) The new heterozygous mutation T-7130-C (Phe118Leu) in active site of alpha-MSH has been identified in POMC gene of a woman suffering with obesity since the early childhood. 3) Mutation A-7341-G (Glu188Gly) seemed to have a protective effect because it was revealed more frequently in control (3.9%) than in obese patients (0.66%). The results of genetic study of two pedigrees suggested the dominant influence of the first two mutations (1 and 2) on woman obesity.

[Study of the association between constitutional exogenous obesity and polymorphism of the apolipoprotein B gene]. / Issledovanie sviazi konstitutsional'no-ékzogennogo ozhireniia s polimorfizmom v lokuse gena apolipoproteina B.

An attempt was made to associate the insertion-deletion (Ins/Del) polymorphism of the apolipoprotein B gene (apoB) with obesity and to identify alleles and genotypes predisposing to this disorder. The apoB Ins/Del allele frequencies observed in the Russian population were similar to those in West European populations and significantly differed from frequencies reported for Asian populations. Patients with obesity did not differ from healthy individuals in allele and genotype frequencies regardless of whether total or sex-stratified samples were compared. Estimation of relative risk for individuals with genotype Ins/Ins did not reveal a significant association between obesity and this genotype. Thus, constitutional exogenous obesity did not prove to be associated with the Ins/Del polymorphism of the apoB gene in the Russian population.

[Sex hormone receptors, prostaglandins and cAMP, in functional activity of the uterus]. / Rol retseptorov polovykh gormonov, prostaglandinov i TsAMF v funktsionalnoi aktivnosti matki.

The levels of progesterone and estrogen receptors, cAMP and prostaglandins E2 and F2 alpha were studied in the decidua of pregnant women, which was obtained during surgical, spontaneous or prostaglandin-induced abortion or by pregnancy termination with vacuum aspiration after RU-486 treatment (before the onset of abortion). Our data have allowed the authors to suggest that (i) for the maintenance of normal activity of decidua during pregnancy, the following points are considerably important: the constant ratio of progesterone and estrogen receptors in favor of progesterone receptors, low PG levels and high cAMP levels; (ii) both spontaneous and induced abortion are characterized by decreases in the levels of progesterone receptor (which in turn changes the ratio of progesterone and estrogen receptors in favor of the latter), by increased PG levels and decreasing cAMP concentrations.

[Genetic engineering in the bacterial synthesis of somatostatin]. / Genno-inzhenernye podkhody k bakterialnomu syntezu somatostatina.

Series of recombinant plasmids for expression of the synthetic gene somatostatin-14 (SST) as a fusion protein were obtained. The somatostatin gene was fused to chloramphenicol acetyltransferase (cat) or its deleted variant genes. Both parts of the resultant fusion protein were joined through a Met residue. The hybrid gene was expressed under the control of the cat gene promoter (Pcat), the tryptophan operon promoter (Ptrp) or the promoter of bacteriophage T5 (PT5). These fusions gave insoluble polypeptide products amounting from 5-10% of the total cellular protein under constitutive biosynthetic conditions (Pcat) to 5-30% upon induction (Ptrp, PT5). A correlation between the efficiency of expression and the length of cat, the power of the promoter used and the absence or presence of transcription terminators, was studied. The scheme for SST isolation from bacterial cells was developed. SST was liberated from the fused polypeptide by treatment with cyanogen bromide and purified to homogenity by a combination of chromatographic steps: gel filtration, ion-exchange and rpHPLC. The renaturated recombinant SST showed specific biological and immunological activities and had 98% purity. The yield was 1 mg of the purified cyclic SST/1 culture of E.coli.

[Synthesis of somatostatin in Escherichia coli cells: isolation and characteristics]. / Sintez somatostatina v kletkakh Escherichia coli. Vydelenie i kharakteristika.

A synthetic gene coding for somatostatin-14 (SST) was cloned in plasmid expression vectors in frame with the chloramphenicol acetyl transferase (CAT) gene, both genes being divided by a Met residue. The hybrid gene was expressed under the control of the CAT gene promoter (Pcat) or the tryptophan operon promoter (Ptrp). Them fused genes gave insoluble polypeptide products amounting from 5% of the total cellular protein under constitutive biosynthesis conditions (Pcat) to 30% upon induction (Ptrp). SST was liberated from the fused polypeptide by treatment with cyanogen bromide, purified to homogeneity by gel-filtration and reverse phase HPLC, and finally refolded by dilution and air oxidation. The renaturated recombinant SST showed the specific biological and immunological activities of the native peptide.

[Endometrial steroid receptor levels in habitual abortion in advanced pregnancy]. / Soderzhanie steroidnykh retseptorov v éndometrii pri privychnom nevynashivanii beremennosti pozdnikh srokov.

Estrogen and progesterone reception was examined in the endometrial cytoplasmic and nuclear fraction from apparently healthy subjects and patients with habitual late incompetent pregnancy in the early proliferative and late secretory periods of a menstrual cycle. The reception of the two reproductive hormones in the nuclei and the binding of estrogens in the endometrial cytosol were demonstrated to be significantly higher in the female patients than in healthy subjects. The cytosol estrogen to progesterone reception ratio was increased in the patients in the secretory phase of a menstrual cycle than in normal females. There was a complete normalization of endometrial reproductive hormone receptor levels in the patients undergoing hyperbaric oxygenation.

[State of the sex hormone receptor in the endometrium of women with late habitual abortion]. / Sostoianie retseptornogo apparata polovykh gormonov v éndometrii zhenshchin pri privychnom nevynashivanii beremennosti pozdnikh srokov.

A study was made of estrogen and progesterone reception of the endometrial cytoplasmatic and nuclear fractions of healthy women and women suffering from late habitual abortion in the early proliferative and late secretory phases of the menstrual cycle. Reception of both sex hormones in the nuclei and estrogen binding in the endometrial cytosol of women with late habitual abortion was significantly higher than that in healthy women. The estrogen receptors/progesterone receptors ratio in the cytosol in the pathological endometrium at the secretory phase of the menstrual cycle was higher than in the normal one. Hyperbaric therapy given to such patients resulted in complete normalization of endometrial sex hormones reception.

[Plasma levels of sex hormones and their receptors in the endometrium of women with normal menstrual cycle]. / Soderzhanie polovykh gormonov v plazme i ikh retseptorov v éndometrii zhenshchin v normal'nom menstrual'nom tsikle.

Plasma estrogen, progesterone, cortisol and prolactin concentrations were determined by RIA in 20 healthy women on the 7th-10th and 24th-26th days of the menstrual cycle. The content of estrogen and progesterone receptors in the cytosol and nuclei of the endometrium was studied by a method of adsorption on dextran-covered carbon; pathomorphological investigation of the endometrium was performed. Sex hormone concentration was shown to be significantly lower in the middle (proliferative) phase as compared to the late one (secretory). Cortisol and prolactin concentrations in both periods did not change. The content of both sex hormone receptors was significantly higher in the middle (proliferative) phase. On the basis of the content and ratio of plasma sex hormones and receptor content in the endometrium a conclusion was made that the main factor in the regulation of receptor levels by sex hormones was the ratio of their concentrations in the plasma.

[Estrogen and progesterone receptors in the decidual tissue of women administered prostaglandins and experiencing spontaneous abortion]. / Retseptory ésterogenov i progesterona v detsidual'noi tkani zhenshchin pri vvedenii prostaglandinov i samoproizvol'nom preryvanii beremennosti.

A study was made of the content of estrogen and progesterone receptors (ER and PR, respectively) in the cytosol and nuclei of the decidual tissue of women in induced and spontaneous abortion at 7-12 week pregnancy. Abortion was done surgically (control group) and in preliminary cervical dilatation with the help of intravaginal suppositories with a synthetic prostaglandin analog of E1-16.16-dimethyltrans-delta 2-PGE1 (ONO-802) group or laminaria. The level of ER in the cytosol and nuclei of the decidual tissue of the control group was twice as low as that of PR, and the ER/PR ratio in cell fractions as well as in general cell receptors was 0.58 +/- 0.09, 0.42 +/- 0.07 and 0.56 +/- 0.07, respectively. The suppositories with ONO-802 caused a statistically significant decrease in the PR content in each of the cell fractions and cytoplasmic ER resulting in an increase in the ER/PR ratio to 0.87 +/- 0.06 (P less than 0.05) in the cytosol, 0.09 +/- 0.24 (P less than 0.05) in the nuclei, and 0.86 +/- 0.8 (P less than 0.05) in the total cell. A decrease in the PR level in both cell fractions and the ER level in the fraction of the nuclei with an acute increase in the content of cytoplasmic ER was marked in spontaneous abortion. The ER/PR ratio also rose to ER and was 1.98 +/- +/- 0.2 (P less than 0.001) in the cytosol and 1.02 +/- 0.16 (P less than 0.05) in the total cell. It was concluded that an absolute and/or relative decrease in the PR level plays an important role in mechanism of abortion.

[Hormonal regulation of estrogen and progesterone receptor biosynthesis in human endometrial tissue culture]. / Gormonal'naia reguliatsiia biosinteza retseptorov éstrogenov i progesterona v kul'ture tkani éndometriia cheloveka.

Estradiol and progesterone effects on the biosynthesis of estrogen and progesterone cytoplasmic receptors have been studied in the endometrium culture tissue of pregnant women. It was established that in a 48-hour culture estradiol stimulated the biosynthesis of its own and progesterone receptors. Under the same conditions progesterone stimulated the biosynthesis of estrogen receptors but did not affect the biosynthesis of its own receptors.

[Prostaglandin regulation of the sex hormone receptor content in human decidual tissue]. / Regulirovanie prostaglandinami soderzhaniia retseptorov polovykh gormonov v detsidual'noi tkani cheloveka.

Prostaglandin F2 alpha (PGF2 alpha) and its synthetic analog, 15-methyl-PGF2 alpha effects on the receptor content of estrogens and progesterone in the human decidual tissue were studied in vitro and in vivo experiments. The receptor content in vivo experiments was determined by Sketchard's technique and in vitro by the saturation method. Three hours after intravaginal administration of 15-methyl-PGF2 alpha suppositories to pregnant women the level of estrogen and progesterone receptors in decidual tissue cytosol significantly increased. Affinity of receptors with hormones remained unchanged. 10, 20, 50 and 100 mcg/ml concentrations of PGF2 alpha produced a significant and dose-dependent increase in the receptor content of estrogens following 15, 30, 60 and 180 minutes incubation of decidual tissue sections. The receptor level of progesterone remained unchanged. In experiments with inhibitors of protein and RNA biosynthesis, it was shown that an augmentation of the estrogen receptor content under PGF2 alpha action is not associated with biosynthesis of receptors de novo.

[Effect of various 1,3,5(10)-estratrien derivatives on the estrogen receptor transport into the nucleus in the guinea pig uterus]. / Vliianie razlichnykh proizvodnykh 1,3,5(10)-éstratriena na transport retseptora éstrogenov v iadro v matke morskoi svinki.

The capacity of five derivates of 1, 3, 5 (10)-estratrien to be transported together with estradiol receptor from the cytoplasm into the nucleus in the uterus of sexually immature guinea pig was investigated. The extent of this transport was assessed by the amount of estrogen receptor in the nucleus following incubation of the uterine slices in the presence of the compound tested (2.5.10(-7) M). The content of estrogen receptor in the nucleus of the intact guinea pig uterus served as control. Incubation of the uterine slices with compounds containing 17 beta-hydroxyl group (2-brom-1, 3, 5 (10)-estratriendiol-3, 17 beta, 4-brom-1,3,5 (10)-estratriendiol, 3, 17 beta, 1, 3, 5 (10)-estratrienol-17 beta), led to a 1.9--4.2-fold increase in the content of nuclear estrogen receptor, this pointing to the transport of the estrogen receptor complex with these compounds into the nucleus. Two compounds (1, 3, 5 (10) estratrienol-3 beta and 3-methoxy-1, 3, 5 (10)-estratrien) contained no 17 beta-hydroxyl group and failed to increase the contents of the estrogen receptor in the nucleus in comparison with control. 17 beta-hydroxyl group is apparently necessary for the formation of conformative changes in the receptor molecule transforming the steroid-receptor complex into the active one, capable of transporting into the nucleus.

[Sex hormone regulation of progesterone and estradiol receptors in the cytosol of human endometrium in normal and pathologic pregnancy]. / Regulirovanie polovymi gormonami soderzhaniia retseptorov progesterona i estradiola v tsitozole endometriia cheloveka pri normal'noi i patologicheskoi beremennosti.

Hormonal control of progesterone and estradiol receptors was studied in the cytozol of short-lived culture of human endometrium in normal and undeveloping pregnancy. Endometrium was cultivated in the presence of estradiol or progesterone for 16 hours. In cultivation of normal endometrium with estradiol the content of estradiol receptors increased 4-fold in comparison with unstimulated tissue, and of progesteron receptors--3-fold. In cultivation of normal endometrium with progesterone the content of estradiol receptors rose 4--5-fold, and of progesterone receptors--3-fold. In cultivation of pathological endometrium with estradiol or progesterone the number of estradiol receptors was only doubled, and of progesterone receptors--increased only 1 1/2 times, this pointing to a diminished sensitivity of pathological endometrium to the regulating action of sex hormones.

PIP: Hormonal control of progesterone and estradiol receptors was studied in the cytozol of a short-lived culture of human endometrium in both normal and undeveloping pregnancy. Endometrium was cultivated in the presence of estradiol or progesterone for 16 hours. In the cultivation of the normal endometrium with estradiol, the content of estradiol receptors increased 4-fold in comparison with the normal tissue, and the progesterone receptors increased 3-fold. In the culturation of the normal endometrium with progesterone, the content of the estradiol receptors rose 4-5-fold while the progesterone receptors increased 3-fold. In the cultivation of the pathological endometrium with either estradiol or progesterone, the number of estradiol receptors only doubled and the progesterone receptors increased only 1-1/2 times. This points to a diminished sensitivity of a pathological endometrium to the regulating action of sex hormones. (Authors' modified)