Growth Hormone Improve The Outcomes Of Infertile Women In IVF Cycles And Potential Mechanisms

Chapter One:Effects of different growth hormone addition protocols on the outcomes of infertile patients in IVF cycles:A systematic review and meta-analysisObjectives:In an effort to improve outcomes of in-vitro fertilization(IVF)cycles,the use of growth hormone has been considered.Improving the outcomes of IVF is especially important for infertile women who are considered ’poor responders’.To assess the effectiveness of adjuvant growth hormone in IVF protocols using approaches of systematic review and meta-analysis.Methods:We searched the CBM,WanFang,CNKI,VIP Databases,PubMed,EMbase,Web of Science and the Cochrane Library.We searched the databases,using the terms "growth hormone","GH","IVF" and "in vitro fertilization." All randomised controlled trials were included if they addressed the research strategy and provided outcome data for intervention and control participants.The primary outcomes included clinical pregnancy rate and live birth rate,the secondary outcomes included the number of collected oocytes,MII stage oocytes,fertilization rate,implantation rate and adverse pregnancy outcomes(including miscarriage rate).Extracted the data from the corresponding articles,random-effects model or fixed-effects model was used.Assessment of trial risk of bias and extraction of relevant data was performed independently by two reviewers.RevMan 5.3 software was used for meta-analysis.Results:Nine studies(586 infertile women)were included.Results of the meta-analysis demonstrated a statistically significant difference in collected oocytes number[MD=1.50,95%CI(1.19-1.81)],MII stage oocytes number[MD=2.10,95%CI(1.82-2.38)],clinical pregnancy rate[OR=1.53,95%CI(1.04-2.26)]and live birth rate[OR=1.73,95%CI(1.04-2.88)]favouring the use of adjuvant growth hormone in IVF protocols in women who are considered poor responders without increasing adverse events(P<0.05).However,no difference in fertilization rate[OR=1.18,95%CI(0.93-1.50)],implantation rate[OR=1.19,95%CI(0.99-1.41)]and miscarriage rate[OR=1.47,95%CI(0.64-3.38)]was found in the routine use of GH(P>0.05).The recommended pretreatment dose of GH was low/medium dose(4IU-12IU),and the preferred periods were from the 21st day of last menstruation to the trigger day of HCG or from the 1st day of ovarian stimulation to the trigger day of HCG.Conclusion:The use of growth hormone in poor responders and aging women has been found to show a significant improvement in the outcomes of IVF cycles.GRADE system for evaluation of evidence quality recommended strongly for GH treatment combined with ovarian stimulation in IVF.However,we were unable to identify which sub-group of poor responders would benefit most from adjuvant growth hormone because of the small sample size and inconsistent including criteria of the included trials.Therefore,further research with large samples and conducted in multi-centre is necessary to fully define its role.Chapter Two:Evidence that growth hormone can improve mitochondrial function in oocytes from aged miceObjective:Decline in successful conception decreases more rapidly after 38 years of age owing to follicular depletion and decreased oocyte quality.However,limited information is available regarding the underlying mechanism and the useful treatment.This study aimed to evaluate the effects of growth hormone supplementation before ovarian stimulation on oocyte maturation in vivo in aged and young mice and to determine its effect on mitochondrial function.Methods:Influence of three different doses of recombinant human growth hormone(rhGH)for 8 weeks before ovarian stimulation was analyzed.The female C57BL/6J mice were randomized devided into 5 groups respectively:WT,saline group,rhGH low-dose(0.4 mg/kg/d),medium-dose(0.8 mg/kg/d)and high-dose(1.6 mg/kg/d)group.Superovulated oocytes were released from the oviduct of Twelve-week-old(12W)and fourty-week-old(40W)female C57BL/6J mice 14-16 h after administration of human chorionic gonadotropin(HCG).Serum and ovarian tissue were collected 8h after HCG injection.Serum anti-Mullerian hormone(AMH)level,ovarian follicle and morphological analysis based on hematoxylin and eosin(HE)staining,total oocytes number,MII stage oocytes number and oocyte maturation parameters were then evaluated.Growth differentiation factor 9,glutathione peroxidase 4 mRNA levels were detected by qPCR.Mitochondrial distribution,mitochondrial membrane potential,total ROS levels of oocyte and mitochondrial specific oxidative damage were accessed by immunofluorescence staining.At last,the ATP levels,mtDNA copy number and cumulus expansion gene PTX3、TNFAIP6 mRNA expression levels of oocyte were evaluated.Results:The ovarian volume of aged mice significantly increased after rhGH pretreatment.Medium-and high-dose rhGH significantly increases antral follicles in aged mice(P<0.05)but preantral follicles.Anti-Mullerian hormone(AMH)levels of young mice were three times higher than aged mice.AMH levels,GDF-9 and GPx4 gene mRNA expression levels were not significantly different after rhGH treatment compared with the control groups in aged mice(P>0.05).Furthermore,derived oocytes,MII stage oocyte number,MII stage oocyte rate and adenosine triphosphate(ATP)levels in single oocyte were significantly increased in medium-and high-dose rhGH treatment groups in aged mice(P<0.05),moreover,MII stage oocyte rate and ATP levels came close to the levels of young mice.Mitochondrial membrane potential(ΔΨm)and frequencies of homogeneous mitochondrial distribution increased significantly after rhGH treatment in aged mice(P<0.05).In young mice,mitochondrial membrane potential(ΔΨm)of medium-and high-dose rhGH treatement groups significantly increased compared with the control(P<0.05).In contrast,in both aged and young mice,the mitochondrial DNA(mtDNA)copy numbers and the total ROS levels per oocyte were similar before rhGH administration,and upon saline administration,they did not differ significantly(P>0.05).The predominant reactive oxygen species levels in mitochondria per oocyte in aged mice were significantly higher than the control group of young mice(P<0.05).However,the predominant reactive oxygen species levels in mitochondria per oocyte were significantly decreased(P<0.05),and TNFAIP6 gene mRNA expression level were significantly increased after medium-dose rhGH treatment for 8 weeks in aged mice(P<0.05).Conclusion:The medium rhGH dose administration significantly enhances the MII stage oocyte rate,ATP content,and mitochondrial membrane potential in oocytes of aged mice.However,ovarian reserves and mtDNA copy number were not significantly increased in any group.In young mice,most of the evaluated parameters were not significantly altered except for mitochondrial membrane potential.This study therefore demonstrates,for the first time,that medium-dose rhGH supplementation for 8 weeks before standard ovarian stimulation regimens improves oocyte quality in aged females,potentially by enhancing mitochondrial functionality.However,additional large-scale studies on rhGH dosing,length of pretreatment,and safety of clinical outcomes are necessary before its clinical use.