Animals ; Diethylhexyl Phthalate ; toxicity ; Female ; Ovary ; drug effects ; metabolism ; ultrastructure ; Rats, Sprague-Dawley ; Receptors, LH ; metabolism ; Receptors, LHRH ; metabolism ; Toxicity Tests

At present, there is no reliable in vitro assembled prepubertal testis-like biomimetic organ culture system designed to assess the functional effects of human gonadotropins on Sertoli and Leydig cells. Spermatogenesis is regulated by endocrine, paracrine, and juxtacrine factors (testicular cross-talk), mainly orchestrated by gonadotropins such as luteinizing hormone (LH) and follicle-stimulating hormone (FSH) that play a pivotal role by stimulating Leydig and Sertoli cells, respectively. The aim of our study was to set up an in vitro prepubertal porcine bioengineered construct as a new model for experimental studies on reassembled Sertoli and Leydig cells. We have evaluated Sertoli and Leydig cells obtained from 15- to 20-day-old neonatal pig testes in terms of purity and function. Subsequently, purified Sertoli and enriched Leydig cells were subjected to coincubation to obtain an in vitro prepubertal porcine testis-like culture system. We performed enzyme-linked immunosorbent assay (ELISA) for anti-Müllerian hormone (AMH), inhibin B, and testosterone secretion in the medium, and Real-Time PCR analysis of AMH, inhibin B, FSH-r, aromatase, LHr, and 3β-HSD mRNA expression levels. This in vitro testis-like system was highly responsive to the effects of human gonadotropins and testosterone. AMH mRNA expression and secretion declined, and inhibin-B increased, while FSH-receptor expression was downregulated upon FSH/LH exposure/treatment. Finally, the production of testosterone was increased selectively upon LH treatment. In summary, our proposed model could help to better determine the action of human gonadotropins on Sertoli and Leydig cells. The potential usefulness of the system for shedding light into male infertility-related issues is evident.
3-Hydroxysteroid Dehydrogenases/metabolism* ; Animals ; Animals, Newborn ; Anti-Mullerian Hormone/metabolism* ; Aromatase/metabolism* ; Cell Culture Techniques ; Enzyme-Linked Immunosorbent Assay ; Follicle Stimulating Hormone/pharmacology* ; Hormones/pharmacology* ; In Vitro Techniques ; Inhibins/metabolism* ; Leydig Cells/metabolism* ; Luteinizing Hormone/pharmacology* ; Male ; Models, Biological ; Real-Time Polymerase Chain Reaction ; Receptors, FSH/metabolism* ; Receptors, LH/metabolism* ; Sertoli Cells/metabolism* ; Swine ; Testis/metabolism* ; Testosterone/metabolism*

The aim of the study was to provide a descriptive analysis of familial male-limited precocious puberty (FMPP), which is a rare inherited disease caused by heterozygous constitutively activating mutations of the luteinizing hormone/choriogonadotropin receptor gene (LHCGR). The patient was a ten-month-old boy, presenting with penile enlargement, pubic hair formation, and spontaneous erections. Based on the clinical manifestations and laboratory data, including sexual characteristics, serum testosterone levels, GnRH stimulation test, and bone age, this boy was diagnosed with peripheral precocious puberty. Subsequently the precocious puberty-related genes were analyzed by direct DNA sequencing of amplified PCR products from the patient and his parents. Genetic analysis revealed a novel heterozygous missense mutation c.1732G>C (Asp578His) of the LHCGR gene exon11 in the patient, which had never been reported. His parents had no mutations. After combined treatment with aromatase inhibitor letrozole and anti-androgen spironolactone for six months, the patient's symptoms were controlled. The findings in this study expand the mutation spectrum of the LHCGR gene, and provide molecular evidence for the etiologic diagnosis as well as for the genetic counseling and prenatal diagnosis in the family.
Heterozygote ; Humans ; Infant ; Male ; Mutation ; Puberty, Precocious ; drug therapy ; genetics ; Receptors, LH ; chemistry ; genetics

OBJECTIVETo assess the association of rs13405728 polymorphism of luteinizing hormone receptor (LHR) gene with slow ovarian response during assisted reproductive technology (ART).

METHODSTwo hundred and thirty-six women were enrolled and grouped according to their genotypes. The rs13405728 polymorphism was genotyped by DNA sequencing.

RESULTSNo signifiicant difference was found in antral follicle count and anti-Mullerian hormone between the three genotypes (P>0.05). The incidence of slow response in genotype GG was lower than in the other two genotypes (P<0.05). There was no significant difference in the amount of follicle stimulating hormone required, the number of follicles ≥14 mm on human chorionic gonadotrophin day, oocytes, mature oocytes, available embryos, and the clinical pregnancy rate among the three genotypes (P>0.05). There was an independent correlation between slow ovarian response with the genotypes of rs13405728, the initial dose of gonadotropin, and the dose of luteinizing hormone required (P<0.05).

CONCLUSIONRs13405728 of the LHR gene may be associated with slow ovarian response in ART. Various mechanisms may be involved in the poor response and slow response.

Adult ; Female ; Fertilization in Vitro ; methods ; Gene Frequency ; Genotype ; Humans ; Logistic Models ; Ovarian Reserve ; genetics ; Ovulation Induction ; methods ; Polymorphism, Single Nucleotide ; Receptors, LH ; genetics ; Reproductive Techniques, Assisted ; Sequence Analysis, DNA ; methods

The effect of high concentrations of testosterone on ovarian follicle development was investigated. Primary follicles and granulosa cells were cultured in vitro in media supplemented with a testosterone concentration gradient. The combined effects of testosterone and follicle-stimulating hormone (FSH) on follicular growth and granulosa cell gonadotropin receptor mRNA expression were also investigated. Follicle growth in the presence of high testosterone concentrations was promoted at early stages (days 1-7), but inhibited at later stage (days 7-14) of in vitro culture. Interestingly, testosterone-induced follicle development arrest was rescued by treatment with high concentrations of FSH (400 mIU/mL). In addition, in cultured granulosa cells, high testosterone concentrations induced cell proliferation, and increased the mRNA expression level of FSH receptor (FSHR), and luteinized hormone/choriogonadotropin receptor. It was concluded that high concentrations of testosterone inhibited follicle development, most likely through regulation of the FSH signaling pathway, although independently from FSHR downregulation. These findings are an important step in further understanding the pathogenesis of polycystic ovary syndrome.
Androgens ; pharmacology ; Animals ; Cell Proliferation ; drug effects ; Female ; Follicle Stimulating Hormone ; genetics ; metabolism ; pharmacology ; Gene Expression Regulation, Developmental ; Granulosa Cells ; cytology ; drug effects ; metabolism ; Mice ; Ovarian Follicle ; cytology ; drug effects ; growth & development ; metabolism ; Primary Cell Culture ; RNA, Messenger ; genetics ; metabolism ; Receptors, FSH ; genetics ; metabolism ; Receptors, Gonadotropin ; genetics ; metabolism ; Receptors, LH ; genetics ; metabolism ; Signal Transduction ; drug effects ; genetics ; Testosterone ; antagonists & inhibitors ; pharmacology

OBJECTIVETo investigate the effects of Morina Officinalis How (MOH) on the abnormal levels of serum luteotrophic hormone (LH) and LH receptor (LHR) in the testis tissue induced by cellphone radiation (CPR) in rats.

METHODSFifty adult male SD rats were randomly divided into five groups of equal number: sham CPR, untreated CPR, negative double distilled water (DDW) control, aqueous MOH extract, and alcohol MOH extract. All the animals were exposed to mobile phone radiation except those of the sham CPR group. Then, the rats of the latter two groups were treated intragastrically with MOH at 20 g per kg of the body weight per day in water and alcohol, respectively. After 2. weeks of treatment, all the rats were sacrificed for measurement of the levels of serum LH and LHR in the testis tissue.

RESULTSThe levels of serum LH and LHR were 30.15 ± 8.71 and 33.28 ± 6.61 in the aqueous MOH group and 0.96 ± 0.06 and 0.94 ± 0.08 in the alcohol MOH group, both significantly decreased as compared with the negative DDW controls (P < 0.05), but with no remarkable difference between the two MOH groups (P > 0.05).

CONCLUSIONMOH can improve CPR-induced abnormality of LH and LHR in adult male rats.

Animals ; Cell Phone ; Electromagnetic Radiation ; Luteinizing Hormone ; blood ; drug effects ; radiation effects ; Male ; Morinda ; chemistry ; Radiation Injuries, Experimental ; blood ; drug therapy ; Random Allocation ; Rats ; Receptors, LH ; blood ; drug effects ; radiation effects ; Testis ; radiation effects

OBJECTIVEFamilial male-limited precocious puberty (FMPP) is due to constitutive activation of a mutant luteinizing hormone/choriogonadotropin receptor (LH/CGR) leading to elevated testosterone synthesis in testicular Leydig cells. In the present study, we have analyzed the LHCGR gene for members of a Chinese FMPP family.

METHODSPhysical examinations have included assessment of penile length, testicular volume and pubic hair. Bone age assessment, levels of testosterone and gonadotropin-releasing hormone (GnRH) stimulations tests were measured. DNA was extracted from blood samples of the proband and his parents using an QIAGEN Blood DNA Mini Kit. The 11 exons of LHCGR gene were amplified using an AmpliTaq PCR system, and the PCR products were sequenced using an ABI3130xl Genetic Analyzer.

RESULTSThe affected boy was 3 year and 1 month old and showed typical clinical manifestation of peripheral precocious puberty. His height was 116.8cm (+5.1s) and Tanner stages were PH 2. Testicular volume was 8 mL bilaterally, penile was 8.5 cm × 2.5 cm. Basal testosterone was 2310 ng/L and bone age was 9 years. GnRH stimulation test revealed a prepubertal response to gonadotropin. The peak of LH was 2.66 IU/L, and the peak of FSH was 1.03 IU/L. Upon sequencing exon 11 of the LHCGR, a heterozygous point mutation of nucleotide 1703 from C to T was detected, which resulted in an amino acid transition from Ala (GCC) to Val (GTC) at position 568. Thus the mutation of LHCGR gene was confirmed to be constitutively active. After treating with aromatase inhibitors for half a year, the patient showed an increase in bone age and height by half a year and 4 cm, respectively. The same point mutation was detected in the patient's father, but did not have any influence on his puberty development.

CONCLUSIONA novel point mutation of the LHCGR gene has been identified in a family affected with FMPP. The c.1703C>T mutant LHCGR was confirmed to be constitutively active, which has led to maturation and proliferation of Leydig cells. The variable phenotype within the family suggested variable expressivity of the disease.

Adult ; Amino Acid Substitution ; Base Sequence ; Child, Preschool ; Codon ; Exons ; Humans ; Male ; Models, Molecular ; Mutation ; Protein Conformation ; Puberty, Precocious ; diagnosis ; genetics ; Receptors, LH ; chemistry ; genetics

Endometritis is one of the primary reasons for reproductive failure. In order to investigate endometritis-associated marker proteins, proteomic analysis was performed on bovine endometrium with endometritis. In bovine endometritis, desmin, alpha-actin-2, heat-shock protein (HSP) 27, peroxiredoxin-6, luteinizing hormone receptor isoform 1, collectin-43 precursor, deoxyribonuclease-I (DNase-I), and MHC class I heavy chain (MHC-Ih) were up-regulated. In contrast, transferrin, interleukin-2 precursor, hemoglobin beta subunit, and potassium channel tetramerisation domain-containing 11 (KCTD11) were down-regulated in comparison to normal endometrium. The proteomic results were validated by semiquantitative-PCR and immunoblot analysis. The mRNA levels of desmin, transferrin, alpha-actin-2, HSP27, KCTD11, and MHC-Ih were up-regulated by over 1.5-fold, and showed a pattern similar to their proteomic profiles. Desmin and alpha-actin-2 protein showed positive correlations between proteomic analysis and immunoblot analysis. These results suggest that desmin and alpha-actin-2 may play important roles in endometritis-related function, and could be useful markers for the diagnosis of bovine endometritis.
Actins ; Collectins ; Desmin ; Endometritis ; Endometrium ; Female ; Heat-Shock Proteins ; Hemoglobins ; Interleukin-2 ; Potassium Channels ; Proteins ; Proteomics ; Receptors, LH ; RNA, Messenger ; Transferrin

Introduction: In Vietnam, the andrology field has been developed in recent years. Some studies have indicated the reasons for male infertility. Amongst them, the most common reason is abnormal semen. Asthenozoospermia rate was found at a higher ratio in abnormal semen analysis. \r\n', u'Objectives: To identify the characteristics of sperm and concentration of FSH, LH and Testosterone in serum; and the association between the sperm morphology and sperm motility as well as the concentration of FSH, LH and Testosterone in serum of asthenozoospermia men. \r\n', u'Subjects and methods: 90 asthenozoospermia male partners of infertile couples. The semen was analyzed at the Laboratory for Tissue and Embryo Preservation, Hanoi Medical University. The hormone concentration was measured by the Automated Chemiluminescence System ACS 180. \r\n', u'Results: The mean semen volume: 3.0 \xb1 1.1 (ml); sperm density: 104.7 \xb1 86.7 million/ml; rapid motility rate: 5.9 \xb1 6.4 (%); sperm vitality rate: 67.4 \xb1 20.1 (%); normal morphology rate: 14.4 \xb1 6.9 (%). In serum, the concentration of FSH was 5.3 \xb1 2.5 lUlL; LH: 4.1 \xb1 1.8 IU/L. Testosterone: 18.3 \xb1 9.4 nmol/L. Conclusions: Among the studied asthenozoospermia men, normal semen volume was found at 91.11 %; normal sperm vitality parameter: 84.4%; normal morphology parameter: 45.56%; normal level of FSH concentration: 47.7%; normal level of LH concentration: 26.66%; normal level of Testosterone concentration: 78.89%. There was a strong association between the sperm morphology and sperm motility as well as serum Testosterone concentration. \r\n', u'
Semen analysis ; Asthenozoospermia ; FSH ; LH ; Testosteron

PURPOSE: Ethane 1,2-Dimethane sulfonate (EDS), an alkylating agent, has been widely used to create the testosterone withdrawal rat model. The present study was carried out to test the effect of EDS administration on the expression of steroidogenesis-related genes in the rat testis and on epididymal sperm counts. MATERIALS AND METHODS: Adult male Sprague-Dawley rats (300~350 g B.W.) were injected with a single dose of EDS (75 mg/kg, i.p.) and sacrificed on days 0, 7, 14, 21, 28, 35, 42, and 49. Tissue weights (testis, epididymis and seminal vesicle) were measured, and serum LH levels were determined by specific radioimmunoassay. The transcriptional activities of LH receptor (LH-R), 3beta-hydroxysteroid dehydrogenase (3beta-HSD), and steroidogenic acute regulatory protein (StAR) were evaluated by semi-quantitative RT-PCR. RESULTS: Weights of the reproductive and accessory organs declined progressively after the EDS treatment (weeks 1~3). After this, the decrease stopped, with a gradual return towards normal. Full recovery was observed in testis and seminal vesicle evaluations on weeks 5 and 6, respectively. Only 70% recovery was found in epididymis during weeks 5~7. A more dramatic drop was observed in caput epididymal sperm count, and the maximum recovery was 40% on week 7. Serum LH level increased significantly on week 2 after EDS treatment, then gradually decreased during weeks 3~5. The transcripts for the steroidogenesis-related genes in testis declined sharply during weeks 1~2, then returned to normal on week 4. CONCLUSIONS: Our results demonstrate that EDS might directly induce severe damage, such as tissue destruction and decreased sperm counts, in epididymis compared to those in testis and seminal vesicles. Changes in the activities of testicular steroidogenesis-related genes caused by abrupt death and repopulation of Leydig cells in EDS-treated rats were in good correlation with other parameters shown in this and previouslypublished data. Taken together, the EDS injection model might be useful to understand not only the mechanism of differentiation of testicular somatic and germ cells but also the function of the epididymis in the aging process.
Adult* ; Aging ; Animals ; Epididymis ; Ethane* ; Germ Cells ; Humans ; Leydig Cells ; Male ; Models, Animal ; Oxidoreductases ; Radioimmunoassay ; Rats* ; Rats, Sprague-Dawley ; Receptors, LH ; Seminal Vesicles ; Sperm Count* ; Spermatozoa* ; Testis ; Testosterone ; Weights and Measures