GnRH Gene Research Articles

Development of the gonadotropin-releasing hormone system.

This review summarizes the current understanding of the development of the neuroendocrine gonadotropin‐releasing hormone (GnRH) system, including discussion on open questions regarding (1) transcriptional regulation of the Gnrh1 gene; (2) prenatal development of the GnRH1 system in rodents and humans; and (3) paracrine and synaptic communication during migration of the GnRH cells.

Epigenetic and transcriptional regulation of GnRH gene under altered metabolism and ageing

Epigenetic and transcriptional regulation of GnRH gene under altered metabolism and ageing

Effect of exercise intensity and duration on the levels of stress hormones and hypothalamic-pituitary–gonadal axis in adult male rats: an experimental study

The effect of exercise on stress has been demonstrated in several studies which have shown that exercise intensity and duration have various effects on the reproductive axis. This study evaluated the effect of different intensities and durations of exercise on the hormonal indices of stress, such as corticosterone (CORT), norepinephrine (NEP), and also reproductive performance indices, including gonadotropin-releasing hormone (GnRH), luteinizing hormone (LH), and testosterone (T). In this experimental study, 30 adult Wistar rats were randomly divided into five groups as follows: no-exercise, RME-1 (regular moderate exercise for 1month), RME-6 (regular moderate exercise for 6months), RIE-1 (regular intensive exercise for 1month), and RIE-6 (regular intensive exercise for 6months). At the end of the experiment, the serum levels of the abovementioned hormones and hypothalamic expressionof the Gnrh gene were measured using the enzyme-linked immunosorbent assay and the real-time polymerase chain reaction method, respectively. The levels of stress hormones, including CORT and NEP, increased only in the RIE-1 group compared with the no-exercise group. In addition, an increase was observed in T hormone levels in the RME-1 group compared with those in the no-exercise group, whereas LH and T hormone levels showed a greater decrease in the RIE-6 group than in the no-exercise group. Gnrh expression levels showed an increase and a decrease in the RME-1 and RIE-6 groups compared with the no-exercise group, respectively. These results confirmed the effects of different intensities and durations of exercise on sex hormone levels.

Short-term regular moderate exercise improved male hypothalamic-pituitary-gonadal axis function via the reduction of hypothalamic neurokinin B expression in adult rats

Introduction: In the arcuate nucleus, kisspeptin, neurokinin-B and pro-dynorphin (KNDy) neurons control the function of gonadotropin-releasing hormone (GnRH) neurons. Early investigations indicated that exercise with various intensities affects luteinizing hormone (LH) and testosterone (T) in different ways. Meanwhile the molecular mechanisms underlying its function not yet been fully understood. Accordingly, the present study evaluated the role of alterations in the levels of KNDy mRNA upstream of GnRH neurons in conveying the effects of various short-term exercise intensities on the male hypothermic-pituitary-gonadal (HPG) axis. Methods: Twenty-one adult Wistar rats were randomly divided into 3 groups: control, one-month regular moderate exercise (ME) and one-month regular intensive exercise (IE). In ME (22m/min) and IE (35m/min) groups, the rats were treated 5 days a week for 60min each day. Finally, we assessed serum levels of LH and T using the ELIZA technique and KNDy and Gnrh mRNA expression by the real-time PCR method. Results: The results revealed that in ME group the expression of Nkb was reduced and the expression of Gnrh mRNA and the LH and T serum levels were increased. However, intensive exercise did not change the serum levels of LH and T or the relative expression of kiss1, Nkb, Pdyn and Gnrh genes. Conclusion: The results suggested that monthly moderate exercise improved male reproductive axis function, while intensive exercise did not have an adverse effect on the reproductive axis. These various effects on the male HPG axis may be propagated by the change in hypothalamic Nkb gene expression.

In-Vitro Exposure to Endocrine Disruptors Alters Inflammatory Markers in Whole Hypothalami and Immortalized GnRH Neurons

Bisphenol A (BPA), a monomer of polycarbonate plastics, and Benzophenones (BPs), used as UV-filters, are endocrine disrupting chemicals (EDC) found in everyday products. Previously, we showed that the in-vitro exposure to BPA decreased Kiss-induced GnRH expression in GN11 cells (donated by Dr. Susan Wray, NIH), immature GnRH neurons, and that exposure to all the EDC decreased Kiss-induced GnRH gene expression in GT1-7 cells (donated by Dr. Pamela Mellon, UCSD), mature GnRH neurons. In this study, we analyzed the effect of in-vitro exposure to the same EDC (BPA, BP2 or BP3, Sigma, 1x10-9 M) or medium as control (C) in mature GnRH neurons (GT1-7 cells), and isolated hypothalami from adult Balb/c males on Glial fibrillary acidic protein (GFAP) and cytokine gene expression. Cells were exposed to the compounds for 12 or 24 h, in DMEM (high glucose) with charcoal-stripped FBS, and the hypothalami for 6 h to the EDC, in Krebs-Ringer buffer. After the incubations, RNA was extracted using Tri-Reagent (Molecular Research Center, OH, USA), 1-2 µg RNA was reverse transcribed and Real-Time PCR performed using specific primers. Results were expressed as Mean±SE and analyzed by T-test or ANOVA using Statistica v12 (StatSoft Inc, USA) Twenty-four hour BPA exposure increased il18 in GT1-7 cells (C=1.0±0.04, BPA=1.2±0.1, T-test p<0.05, n=7), whereas neither BP2 nor BP3 had an effect on il18 expression (ANOVA: ns, n=7). When il6 was analyzed, 24-hour BPA decreased its expression relative to C and to 12-hour BPA, whereas BP3 had a dual effect depending on the time-point analyzed, increasing the expression at 12-hour stimulation and decreasing it after 24-hour stimulation (DMSO-12h=0.82±0.08, DMSO-24h= 1.01±0.13, BPA-12h=1.00±0.11, BPA-24h=0.68±0.06, BP2-12h=0.75±0.12, BP2-24h=0.82±0.18, BP3-12h=1.20±0.10, BP3-24h=0.83±0.05; Repeated Measures Two-way ANOVA: BPA-24h different from DMSO-24h and BPA-12h p<0.05, BP3-12h different from DMSO-12h and from BP3-24h p<0.05, n=4). In the hypothalami, 6-hour BPA exposure increased gfap gene expression (C=0.8±0.2, BPA=1.7±0.4; T-test p<0.05, n=9), whereas neither BP2 nor BP3 had any significant effect (C=0.8±0.2, BP2=1.4±0.3, BP3=1.0±0.3, ANOVA ns, n=9). There was no significant change in il18, il6 or il1b gene expression in whole hypothalami with any of the EDC tested (ANOVA ns). Our results show that the EDC herein studied have the potential to alter the inflammatory state of mature GnRH neurons and to activate astrocytes in the hypothalamus. The pattern for cytokine expression in the whole tissue could be different from the one observed in GnRH neurons, as the hypothalamus contain multiple cell types, and effects can be different in the different cells. More experiments are needed to dissect the mechanisms involved in the effects observed. Funding: CONICET, ANPCyT, UBA, International Society for Neurochemistry, Asoc. ORT Arg., Fund. R. Barón, Fund. Williams.

Multiplicity of gonadotropin-releasing hormone in fish

Gonadotropin-releasing hormone (GnRH) is the major neuroendocrine hormone regulating reproduction in vertebrates, including finfish. In recent years, presence of GnRH and their function in reproduction has been confirmed in shellfish, crustaceans and molluscs. Interestingly, shellfish GnRH do not cluster within finfish GnRH, suggesting diversity in gnrh genes and their evolution in animal kingdom. GnRH analogues have been synthesized chemically and successfully used in induced reproduction of finfish. Studies on GnRH potency in induced maturation and spawning of shellfish are limited. The present paper report the recent information of shellfish GnRH cDNAs that have been successfully isolated in number of molluscan species, and would help in the farming of shellfish under controlled conditions. Also, the paper highlights the scintometric mapping of GnRH research in fish, suggesting rapid accumulation of information on fish GnRH and their importance in fish reproductive biology and endocrinology.

Maternal Exposure to Dibutyl Phthalate (DBP) or Diethylstilbestrol (DES) Leads to Long-Term Changes in Hypothalamic Gene Expression and Sexual Behavior.

Xenobiotic exposure during pregnancy and lactation has been linked to perinatal changes in male reproductive outcomes and other endocrine parameters. This pilot study wished to assess whether brief maternal exposure of rats to xenobiotics dibutyl phthalate (DBP) or diethylstilbestrol (DES) might also cause long-term changes in hypothalamic gene expression or in reproductive behavior of the resulting offspring. Time-mated female Sprague Dawley rats were given either DBP (500 mg/kg body weight, every second day from GD14.5 to PND6), DES (125 µg/kg body weight at GD14.5 and GD16.5 only), or vehicle (n = 8–12 per group) and mild endocrine disruption was confirmed by monitoring postnatal anogenital distance. Hypothalamic RNA from male and female offspring at PND10, PND24 and PND90 was analyzed by qRT-PCR for expression of aromatase, oxytocin, vasopressin, ER-alpha, ER-beta, kisspeptin, and GnRH genes. Reproductive behavior was monitored in male and female offspring from PND60 to PND90. Particularly, DES treatment led to significant changes in hypothalamic gene expression, which for the oxytocin gene was still evident at PND90, as well as in sexual behavior. In conclusion, maternal xenobiotic exposure may not only alter endocrine systems in offspring but, by impacting on brain development at a critical time, can have long-term effects on male or female sexual behavior.

Brain-derived neurotrophic factor (BDNF) affects the activity of the gonadotrophic axis in sheep

This study aimed to examine the hypothesis that BDNF modulates the activity of the gonadotrophic axis in sheep. Central infusions of BDNF were administered to sexually mature Polish Merino sheep. The sheep were randomly divided into three groups: the control group received intracerebroventricular (ICV) infusions of the vehicle, the BDNF 10 group received ICV infusions of BDNF at 10 μg/480 μL/day, and the BDNF 60 group was infused with BDNF at 60 μg/480 μL/day. A series of four infusions on three consecutive days was performed. Blood samples were collected on days 0 and 3 of the infusions. Immediately after the experiment, all the sheep were slaughtered, and selected structures of the hypothalamus and pituitaries were collected for Real Time RT-qPCR analysis. The collected plasma samples, as well as parts of pituitaries were stored for radioimmunoassay analysis of LH and FSH.Central treatment with exogenous BDNF stimulated GnRH mRNA expression in the preoptic area, as well as GnRH-R mRNA in the pituitary. Furthermore, substantial changes in the KNDy mRNA expression in the mediobasal hypothalamus were observed after the ICV BDNF administration. Additionally, central BDNF infusion caused a decrease in LH concentration and a simultaneous increase in FSH concentration in peripheral blood. Neither pulse amplitude nor pulse frequency for any gonadotrophin was affected in both groups of sheep that received BDNF infusion.Our results revealed that exogenous BDNF affects GnRH and KNDy gene expression and changes in the LH and FSH pituitary cell secretory activities. These findings suggest that BDNF may participate in the mechanism modulating the activity of the gonadotrophic axis at the central level in female sheep.

Clinical characteristics and molecular genetic analysis of a cohort with idiopathic congenital hypogonadism.

Hypogonadism is defined as inadequate sex hormone production due to defects in the hypothalamic-pituitary-gonadal axis. In recent years, rare single gene defects have been identified in both hypergonadotropic hypogonadism (Hh), and hypogonadotropic hypogonadism (HH) cases with no chromosomal anomalies. The aim of the present study is to investigate the underlying molecular genetic etiology and the genotype-phenotype relationship of a series of patients with Hh and HH. In total, 27 HH and six Hh cases were evaluated. Clinical and laboratory features are extracted from patients' hospital files. Whole exome sequencing (WES) analysis was performed. A total of 27 HH cases (15 female) (mean age: 15.8±2.7 years) and six Hh patients (six females) (mean age: 14.9±1.2 years) were included. In molecular genetic analysis, a pathogenic/likely pathogenic variant was identified in five (two patients from the same family) of 27 HH cases (two novel) and three of the six Hh. In HH group variants (pathogenic, likely pathogenic and variant of uncertain significance) were identified in KISS1R (n=2), PROK2 (n=1), FGFR1 (n=1), HS6ST1 (n=1), GNRH1 (n=1) genes. In the Hh group, splice-site mutations were detected in DCAF17 (n=1) and MCM9 (n=2) genes. HH and Hh cases are genetically heterogeneous diseases due to oligogenic inheritance, incomplete penetrance, and variable expressivity. We found rare variants in CHH related genes in half of our HH cases, whereas they classified as pathogenic/likely pathogenic according to ACMG criteria in only about 15% of HH cases. Using advanced genetic analysis methods such as whole-genome sequencing and long-read sequencing may increase the mutation detection rate, which should always be associated with and expert genetic counseling to interpret the data.

Characterization and evaluation of the tissue distribution of CRH, apelin, and GnRH2 reveal responses to feeding states in Schizothorax davidi.

Schizothorax davidi is a rare fish in Southwest China and is considered a promising species for aquaculture. Compared with other teleosts, little is known about the endocrine regulation of feeding in this species. In this study, we identified the CRH, apelin, and GnRH2 genes in S. davidi and assessed the effects of different energy statuses on CRH, apelin, and GnRH2 expression. Our results showed that the full-length cDNA sequences of CRH, apelin, and GnRH2 of S. davidi were 995, 905, and 669bp long, respectively. Furthermore, CRH was mainly expressed in the hypothalamus, telencephalon, and myelencephalon; apelin was highly expressed in the spleen and heart; and GnRH2 mRNA was widely distributed in all examined tissues, with the highest level in the hypothalamus. Notably, the levels of CRH and GnRH2 increased in the hypothalamus at 1h and 3h post-feeding, while hypothalamic apelin levels decreased. Conversely, CRH and GnRH2 expression in the hypothalamus significantly decreased after fasting for 7 days and returned to the control levels after re-feeding for 3 or 5 days. In contrast, fasting increased apelin levels in the hypothalamus. Overall, this study suggests that CRH, apelin, and GnRH2 play critical roles in appetite regulation in S. davidi. These results provide an essential groundwork to elucidate the appetite regulatory systems in S. davidi as well as in other teleosts.

Phoenixin(smim20), a gene coding for a novel reproductive ligand, is expressed in the brain of adult zebrafish

Gonadotropin-releasing hormone (GnRH) is a highly conserved neuroendocrine decapeptide that is essential for the onset of puberty and the maintenance of the reproductive state. In addition to its role as hypothalamic releasing hormone, GnRH has multiple functions including modulator of neural activity within the nervous system and of resulting behaviors. These multiple functions are reflected by the existence of multiple isoforms. Despite its importance as a critical hypothalamic releasing hormone, the gnrh1 gene has been lost in zebrafish, and its reproductive function is not compensated for by other GnRH isoforms (GnRH2 and GnRH3), suggesting that, surprisingly, zebrafish do not use any of the GnRH peptides to control reproduction and fertility. Previously we proposed that Phoenixin/SMIM20, a novel peptide identified in mammals and the ligand for the orphan GPR173, is a potential candidate to control the initiation of sexual development and fertility in the zebrafish. Here we confirm the sequence of the zebrafish phoenixin/smim20 gene and by RT-PCR show that it is expressed early in development through adulthood. Subsequently we show that phoenixin/smim20 is expressed in the adult brain including the regions of the hypothalamus important in the control of fertility and reproduction.

Multigenerational reproduction and developmental toxicity, and HPG axis gene expression study on environmentally-relevant concentrations of nonylphenol in zebrafish

Nonylphenol (NP) is a toxic xenobiotic compound, which is persistent in the aquatic environment and is extremely toxic to aquatic organisms. Although the exact molecular mechanisms of its toxic effect are well understood, the multigenerational reproduction and multigenerational - gene expression changes caused by NP still remain unclear. The following work investigated the effect of NP on four consecutive generations of zebrafish by examining their growth and several reproductive parameters, the degree of gonad damage, and the expression of related reproduction related genes. The results showed that high concentrations (20 and 200 μg·L−1) of NP could decrease growth and induce gonad damage in zebrafish. In addition, gnrh2 and gnrh3 genes were up-regulated, and fshβ and lhβ genes were downregulated in the hypothalamus in male zebrafish; while in female fish, the fshβ and lhβ were upregulated in P and F1 generations, and then down-regulated in the F2 generation. Meanwhile, the cyp19a1a gene was downregulated in the gonad of male fish, while the genes of fshr, lhr and esr showed a downward trend in females. Compared to P generation, F2 generation was more tolerant to higher NP concentrations (20 and 200 μg·L−1), as was also more sensitive to lower concentrations of NP (2 μg·L−1). Consequently, stress and damage caused by environmentally-relevant concentrations of aquatic pollutants in a vertebrate model were measured and predicted. Prevention and control measures can be actively and effectively proposed, which might be transversal to other exposed organisms, including humans. After several generations, typical transgenerational genetic phenomena might occur, which should be addressed by further studies.

Forkhead transcription factor FOXO1 is involved in hypoxia/reoxygenation-induced gonadotropin-releasing hormone decline.

Previously, it has been demonstrated that aging is associated with nuclear factor-κB (NF-κB)-mediated hypothalamic gonadotropin-releasing hormone (GnRH) decrease. The hypothalamus is one of the brain regions that are vulnerable to ischemia-reperfusion injury. However, it is unclear whether ischemia-reperfusion has an influence on the hypothalamic GnRH release. In the current study, GT1-7 cells, which are a cell line of hypothalamic GnRH neurons, were subjected to hypoxia-reoxygenation to mimic ischemia-reperfusion. The effect of hypoxia-reoxygenation on the hypothalamic GnRH release was investigated. It was found that GnRH secretion from GT1-7 cells was decreased under the hypoxia-reoxygenation condition. Mechanistic studies revealed that hypoxia-reoxygenation activated nuclear factor-κB (NF-κB) via the protein kinase B (Akt)/forkhead box protein O1 (FOXO1) pathway, thereby inhibiting gnrh1 gene. The results of the current study suggested that hypoxia-reoxygenation injury may facilitate the hypothalamic programming of system aging through impairment of hypothalamic GnRH release.

Association Analysis between Polymorphism of GnRH-1 Genes and Carcass Traits of Quail (Coturnix coturnix)

Background: GnRH-1 mainly exists in hypothalamus and it is vital to gonad development and sexual maturity of animals. This study mainly analyzed the association between GnRH-1 polymorphism and carcass traits of quails, Research conclusions can provide references for breeding of new species of quail.Methods: From 2018 to 2019, 31 female samples of Chinese Yellow quail, beijingbai quail and Korean quail were collected in Luolong District, Luoyang City, Henan Province, China.SNP mutation sites of GnRH-1 gene were investigated in China yellow quail, Beijing white quail and Korean quail through PCR amplification and DNA sequencing technologys. Moreover, correlation analysis between polymorphism sites of GnRH-1 gene and carcass traits of quail was performed. Result: According to research results, a total of 14 SNP mutation sites of GnRH-1 were detected in China yellow quail, Beijing white quail and Korean quail, which were C71T, C108T, C168T, C178T, A184G, C206T, A209C, C215T, A252G, A279T, C281T, C293G, C339T and C458T. For A209C and C281T, only 2 genotypes were detected in China yellow quail and Beijing white quail, while for the remaining 12 SNP mutation sites, 3 genotypes were detected in three quail species. Correlation analysis showed that A209C, A279T, C281T and C339T were significantly correlated with liver weight (P less than 0.05), A209C, C281T and C339T were significantly correlated with dressing percentage (P less than 0.05), C215T and A279T were significantly correlated with breast muscle weight(whole) (P less than 0.05), A209C was significantly correlated with body weight, dressed carcass weight and whole net carcass weight (P less than 0.05).

The Effect of Vitex Agnuse Castus Fruit Powder on Hypothalamic GnRH Gene Expression in Laying Hens

The Effect of Vitex Agnuse Castus Fruit Powder on Hypothalamic GnRH Gene Expression in Laying Hens

Association of GnRH1 Gene with Growth Traits in Two Breeds of Sheep

The present study was conducted to identify the associations between the genetic polymorphisms of GnRH1 gene and growth traits in Awassi and Karakul breeds of sheep. Two GnRH1 amplicons of 313 bp and 323 bp were designed to cover exon 1, exon 2, and their flanking regions, respectively. Genotyping experiments were performed using PCR-SSCP and DNA sequencing. No polymorphism was observed in 323 bp amplicons, while two SSCP-banding patterns were detected in 313 bp amplicons with the presence of two genotypes (AA and AB). Two novel mutations in AB genotype were observed, namely 5′-UTR:50 A > C and intron1:264 G > C. In addition to these detected genotypes, results indicated that several growth traits were significantly associated with breed, sex, and dam-age in the first 6 months of birth. However, a tendency of individuals with AA genotype to exert better performance in the majority of measured growth traits than individuals with AB genotype was also observed. With the advent of the month-9 and onward, this tendency developed to be significant (P < 0.05), in which the association of all growth traits with the GnRH1 genotypes was dominated over other factors. Furthermore, more significant associations were observed in the month-12, in which individuals with AA genotype had higher growth traits measurements than individuals with AB genotype (P < 0.01). In conclusion, we detected a tight linkage between the polymorphisms of the GnRH1 gene and growth traits at sexual maturity. Therefore, this study strongly suggests the GnRH1 gene as a new candidate in marker-assisted selection for growth traits in sheep.

Cellular identity and Ca2+ signaling activity of the non-reproductive GnRH system in the Ciona intestinalis type A (Ciona robusta) larva

Tunicate larvae have a non-reproductive gonadotropin-releasing hormone (GnRH) system with multiple ligands and receptor heterodimerization enabling complex regulation. In Ciona intestinalis type A larvae, one of the gnrh genes, gnrh2, is conspicuously expressed in the motor ganglion and nerve cord, which are homologous structures to the hindbrain and spinal cord, respectively, of vertebrates. The gnrh2 gene is also expressed in the proto-placodal sensory neurons, which are the proposed homologue of vertebrate olfactory neurons. Tunicate larvae occupy a non-reproductive dispersal stage, yet the role of their GnRH system remains elusive. In this study, we investigated neuronal types of gnrh2-expressing cells in Ciona larvae and visualized the activity of these cells by fluorescence imaging using a calcium sensor protein. Some cholinergic neurons and dopaminergic cells express gnrh2, suggesting that GnRH plays a role in controlling swimming behavior. However, none of the gnrh2-expressing cells overlap with glycinergic or GABAergic neurons. A role in motor control is also suggested by a relationship between the activity of gnrh2-expressing cells and tail movements. Interestingly, gnrh2-positive ependymal cells in the nerve cord, known as a kind of glia cells, actively produced Ca2+ transients, suggesting that active intercellular signaling occurs in the glia cells of the nerve cord.

Expression profile of kisspeptin2 and gonadotropin-releasing hormone2 mRNA during photo-thermal and melatonin treatments in the female air-breathing catfish Heteropneustes fossilis.

In seasonally breeding vertebrates, extrinsic factors like photoperiod and temperature are major determinants, controlling the annual reproductive cycle. In teleosts, kisspeptin, which occurs in two molecular forms: kisspeptin1 (Kiss1) and kisspetin2 (Kiss2), has been reported to alter gonadotropin (Lh and Fsh) secretion but its effect on gonadotropin-releasing hormone (Gnrh) secretion is not unequivocally proved. In the catfish Heteropneustes fossilis, we isolated and characterized kiss2 and gnrh2 cDNAs and the present work reports effects of altered photo-thermal conditions and melatonin (MT, a pineal hormone) on their expressions in the brain. The exposure of the catfish to long photoperiod (LP, 16h light) at normal temperature (NT) or high temperature (HT, 28°C) at normal photoperiod (NP) for 14 or 28days stimulated both kiss2 and gnrh2 expression in both gonad resting and preparatory phases with the combination of LP + HT eliciting maximal effects. Short photoperiod (SP, 8h light) under NT or HT altered the gene expression according to the reproductive phase and temperature. MT that mediates photo-thermal signals to the brain inhibited brain kiss2 and gnrh2 gene expression in the NP + HT, LP + NT, and SP + NT groups. The altered photo-thermal conditions elicited changes in steroidogenic pathway as evident from changes in plasma E2, progesterone, and testosterone levels. The results show that brain kiss2-gnrh2 signaling is involved in photo-thermal-mediated mechanisms controlling reproduction.

The effects of common variants in MDM2 and GNRH2 genes on the risk and survival of osteosarcoma in Han populations from Northwest China

Accumulating evidence has shown that both MDM2 and GNRH2 might be related to Osteosarcoma (OS) susceptibility. The study aimed to evaluate the effects of common variants in MDM2 and GNRH2 genes on the risk and survival of osteosarcoma in Han populations from Northwest China. In the study, we recruited 2292 subjects including 596 OS patients and 1696 healthy controls and genotyped 16 selected tag SNPs (6 from GNRH2 and 10 from MDM2). Genetic association analyses were performed at the genotypic and allelic levels. Survival curves were made for OS patients with different genotypes. Two SNPs, rs1690916 (MDM2, P = 0.0002) and rs3761243 (GNRH2, P = 0.0004), were identified to be significantly associated with OS risk. Moreover, SNP rs3761243 was strongly associated with pathological fracture (P = 2.61 × 10–14), metastasis (P < 2.2 × 10–16), and Enneking stage (P < 2.2 × 10–16) in the OS group. Furthermore, survival curves based on different genotypes of SNP rs3761243 were found to be significantly different (P = 0.0003), suggesting increased risk with more copies of C alleles. Our results provide supportive evidence for genetic associations of MDM2 and GNRH2 genes with susceptibility to OS, and for the positive correlation of SNP rs3761243 in GNRH2 with the survival status of OS patients in Han populations from Northwest China.

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