| Erectile dysfunctiom(ED)is a common clinical manifestation.The prevalence of ED is 26.1% in China.Penile erection is a complex process affected by many factors.In recent years,most studies have focused on the effects of hypothyroidism in women,especially in pregnant women.The effects on male gonadal function are rarely studied.Objective:The aim of this study was to investigate sexual functioning and gonad function in men with hypothyroidism and evaluate the benefit from L-thyroxine treatment.Methods:1.Grouping: Sixty patients with primary hypothyroidism admitted in the second Affiliated Hospital,Anhui Medical University from January 2013 to January 2020 were enrolled into this study,including 30 patients with clinical hypothyroidism group and 30 patients with subclinical hypothyroidism group.Thirty patients with euthyroid from physical examination center.All included patients completed a questionnaires evaluating male sexual function(International Index of Erectile Function-5: IIEF-5).The determination of results was as follows: ≥22 points,normal erectile function;12–21points,mild erectile dysfunction;8–11 points,moderate erectile dysfunction;and 5–7points,severe erectile dysfunction.The exclusion criteria were as follows: diabetes,multiple sclerosis,prostatitis,psychiatric problems,cardiovascular disease,impaired renal or hepatic function,vasculogenic or neurogenic disorders known to impair male sexual function,developmental or acquired anomalies of the male reproductive system.2.Treatment: Patients with both clinical and subclinical hypothyroidism were treated with L-thyroxine.Adjust the dose(25-50 ug once)according to thyroid function.Patients with euthyroid after treatment at 12 weeks were enrolled and other patients without euthyroid were dropped out.The treatment was maintained untill 24 weeks.3.Laboratory tests:Venous blood samples were drawn from the antecubital vein between 7:00 and 9:00 a.m..Serum levels of thyrotropin,thyroxine,triiodothyronine,luteinizing hormone,follicle stimulating hormone,prolactin,testosterone,triglyceride,total cholesterol and low density lipoprotein were measured.Patients with both clinical and subclinical hypothyroidism completed questionnaires evaluating male sexual function(International Index of Erectile Function-5: IIEF-5)before and after 24 weeks of levothyroxine treatment.In the clinical hypothyroidism group,pituitary MRI was performed at baseline,and in patients with pituitary lesions,pituitary MRI was reexamined 24 weeks after treatment.4.Statistical analysis:SPSS19.0 software was used for statistical analysis.The measurement data are presented as the mean ± standard deviation(x± S).The comparison of measurement data that was normally distributed among groups was performed by one-way ANOVA followed by LSD post-hoc test.The non-parametric test was used for statistical analysis of non-normally distributed data or non-uniform variance among groups(Kruskal Wallis H-rank sum test).Comparison of normally distributed measurement data before and after treatment was performed by the paired sample t-test.The measurement data of the two groups were compared by independent sample t-test.Correlation analysis of measurement data was performed by Pearson’s correlation analysis.The chi-square test was used to compare the count data among groups.P < 0.05 was considered statistically significant.Results:1.Comparison of thyroid hormone,sex hormone levels,serum lipid among the three groups before treatment(1)Serum lipid:Total cholesterol and low density lipoprotein was significantly higher in men with clinical hypothyroidism than these in control group(P < 0.05).Total cholesterol was significantly higher in men with clinical hypothyroidism than these in subclinical hypothyroidism group(P<0.05).(2)Thyroid hormone:Patients with clinical hypothyroidism exhibited increaseing levels of thyrotropin and lower levels of thyroid hormones and lower total testosterone than men with subclinical hypothyroidism and healthy controls(P<0.05).(3)Sex hormone levels:Patients with clinical hypothyroidism exhibited higher serum levels of prolactin than patients in control group(P<0.05).Moreover,circulating levels of thyrotropin and prolactin were higher,while circulating levels of triiodothyronine and total testosterone were lower in patients with subclinical hypothyroidism than these in control group(P<0.05).There were no significant differents of LH and FSH between three groups(P>0.05).2.Comparison of thyroid hormone,sex hormone levels,serum lipid among the three groups after treatment:There were no significant differences between serum levels of lipid,thyrotropin,thyroid hormone and gonadal hormone among three groups(P>0.05).3.Comparison of serum lipid,thyroid function and sex hormone levels before and after treatment: A significant decrease in levels of the serum total cholesterol,low density lipoprotein,thyrotropin and prolactin were observed after treatment.Thyroid hormones and total testosterone was significantly increased after levothyroxine treatment(P< 0.05).4.Rate of erectile dysfunctiom: The rate of ED was higher in clinical hypothyroidism group than that in subclinical hypothyroidism group and control group,as well as that was higher in subclinical hypothyroidism group than that in control group(P<0.05).A significant decreases in rate of ED were observed in clinical hypothyroidism group and subclinical hypothyroidism group after treatment(P<0.05).5.IIEF-5(International Index of Erectile Function-5: IIEF-5):The scores of IIEF-5was lower in clinical hypothyroidism group than that in subclinical hypothyroidism group and control group,as well as that was lower in subclinical hypothyroidism group than that in control group(P<0.05).After levothyroxine treatment,the scores of IIEF-5 were higher in clinical hypothyroidism and subclinical hypothyroidism groups than control group.After levothyroxine treatment,the IIEF scores were significantly lower in clinical hypothyroidism groups and in subclinical hypothyroidism groups than in control group(P < 0.05).Scores were lower in clinical hypothyroidism than in subclinical hypothyroidism(P < 0.05).Change in IIEF-5 after 24 weeks increased in clinical hypothyroidism than in subclinical hypothyroidism.6.Comparison of erectile dysfunction grade among three groups:Clinical hypothyroidism group had six cases without erectile dysfunction,siteen cases with mild erectile dysfunction and eight cases with moderate erectile dysfunction before treatment.The number of patients without erectile dysfunction were fifteen and the number with mild and moderate erectile dysfunction respectively ten and five in subclinical hypothyroidism group.The number of patients with non-erectile dysfunction and mild erectile dysfunction respectively twenty-eight and two in control group before treatment(P<0.05).There was no patient with severe erectile dysfunction in any of the three groups.After treatment,there were twenty-one cases without erectile dysfunction,nine cases with mild erectile dysfunction in clinical hypothyroidism group.In subclinical hypothyroidism group,there were twenty-three cases of erectile dysfunction,seven cases of mild erectile dysfunction.There were no patient with mild or severe erectile dysfunction in any of the three groups after treatment(P<0.05).7.Correlation analysis: IIEF-5 scores were positively correlated with thyroid hormones and total testosterone levers(r=0.302,P=0.006;r=0.53,P<0.001;r=0.660,P<0.001)and negatively with hormothyrin,prolactin,total cholesterol,low density lipoprotein levers at baseline(r=-0.647,P < 0.001;r=-0.292,P=0.008;r=-0.357,P=0.001;r=-0.314,P=0.004).Total testosterone levels positively correlated with thyroid hormones and luteotropin levels(r=0.233,P=0.035;r=504,P<0.001;r=0.220,P=0.047)and negatively with hormothyrin,prolactin,total cholesterol,low density lipoprotein levels at baseline(r=-0.531,P < 0.001;r=-0.289,P=0.009;r=-0.430,P <0.001;r=-0.226,P=0.041;r=-0.415,P < 0.001).Total cholesterol levels were positively correlated with hormothyrin(r=0.439,P<0.001),negatively with thyroid hormones at baseline(r=-0.282,P=0.010;r=0.386,P < 0.001).Low density lipoprotein levels were positively correlated with hormothyrin(r=0.315,P < 0.004),negatively with tetraiodothyronine(r=-0.254,P=0.021)at baseline.Change in IIEF-5 after 24 weeks were negatively correlated with T3,T4 and TT(r=-0.240,P=0.03;r=-0.448,P<0.001;r=-0.577,P<0.001)and positively correlated with TSH and TC at baseline(r=0.612,P<0.001;r=0.350,P=0.001).8.The clinical and serum hormone characteristics of patients with pituitary lesions :Compared with those without pituitary lesions,age,serum testosterone level and prolactin level were decreased in patients with pituitary lesions(P < 0.05);Pituitary lesions were recovered in all patients after treatment.Conclusions:Primary hypothyroidism in men is characterized by sexual disturbances and pituitary hyperplasia.Hypothyroid patients with sexual dysfunction are benefit from Lthyroxine treatment.Screening for thyroid dysfunction in men presenting with ED is recommended.Primary hypothyroidism effect on hypothalamic-pituitary-gonadal axis in the central nervous systemIn addition to regulating the normal development and metabolic balance of the body,thyroid hormone and its receptors also play an important biological role in animal reproduction activities.In recent years,a large number of clinical data and experimental studies have proved that an appropriate amount of thyroid hormone can help maintain the stability of the pituitary-gonad axis.Thyroid dysfunction can affect the stability of sex hormone secretion levels,which in turn affects the stability of the male and female reproductive systems.The kisspeptin encoded by the KISS1 gene and its receptor G-protein-coupled receptor 54(GPR54)have been shown to play a key role in regulating the release of gonadotropin-releasing hormone(Gn RH)and luteinizing Hormone(LH).The study found that after the injection of kisspeptin in rats,the levels of LH and folliclestimulating hormone(FSH)in the pituitary portal blood surged,and it was found that Gn RH antagonists can block this group of secretions of kisspeptin.The direct effect of thyroid hormone on Gn RH neurons and the co-expression of thyroid hormone receptors in Gn RH neurons have been confirmed.Studies have shown that thyroid hormones are necessary for the pulsatility of Gn RH and LH.Although Gn RH is closely related to the pulsatile secretion of kisspeptin,the potential role of thyroid hormones in the regulation of the kisspeptin system has not been studied.Gonadotropin inhibitory hormone(Gn IH)is a newly discovered hypothalamic neuropeptide that can effectively inhibit the synthesis and release of gonadotropin.After the discovery of Gn IH,it has been proved that Gn IH is conserved in various vertebrates,such as mammals and primates including humans.In mammals and primates,Gn IH is also called RF-amide related petide(RFRP).Current research shows that RFPR-3 can inhibit Gn RH in the hypothalamus and thus affect the synthesis and release of LH.The researchers found that thyroid dysfunction altered the expression of Gn IH in the hypothalamus by changing the chromatin modification of the Gn IH promoter region in female mice.This is a new function of Gn IH as an intermediary between the HPT axis and the HPG axis.Importantly,female mice with hypothyroidism showed a delay in puberty as Gn IH expression increased.Objective:To study the effect of hypothyroidism on the expression of gonadotropin-releasing hormone and its upstream regulatory genes in rats.Meterials:1.Construct a hypothyroid rat animal model: 60 male SD rats were randomly divided into 2 groups,30 rats were the normal drinking water group;30 rats were fed drinking water containing 0.05% propylthiouracil(PTU)Water,induced adult hypothyroidism rat model for 6 weeks.Five rats were randomly selected from both groups to measure the level of thyroid hormone to determine whether the model of hypothyroidism was successfully established or not.2.Specimen collection: After 12-hour fasting,rats were made intraperitoneal anesthesia,blood was taken from the abdominal aorta,and samples of hypothalamus and pituitary tissues were harvested.One of part of the samples was immediately extracted for protein,and the other parts were placed in cryopreservation tubes and stored in liquid nitrogen for RNA extraction and immunohistochemistry.3.Thyroid hormone determination: Enzyme linked immunosorbent assay(ELISA)method was used to determine the serum levels of Triiodothyronine(T3),Thyroxine(T4)and Thyroid Stimulating Hormone(TSH).4.Determination of rat sex hormone related indicators:ELISA method was used todetermine the serum levels of LH,FSH,Prolactin(PRL),Estradiol(E2),Progesterone(P)and Testosterone(TT).5.Determination of the expression of rat gonadotropin-releasing hormone and its upstream regulatory genes:Quantitative Real-time(q RT-PCR)was used to detect the expression levels of Gn RH,RFRP-3,kisspeptin m RNA in hypothalamus and gonadotropin releasing hormone receptor(Gn RHR)in pituitary;the ELISA method was used to detect the expression of Gn RH,RFRP-3 and kisspeptin proteins Levels in hypothalamus and pituitary;immunohistochemical methods were used to observe the expression levels of Gn RH,RFRP-3,kisspeptin protain in hypothalamus and Gn RHR protain in pituitary.6.Statistical method: SPSS19.0 software was used for statistical analysis.The measurement data are presented as the mean ± standard deviation(x± S).Independent sample T test was performed.After anova of experimental data,Tukey test was used to compare the statistical differences between the two.Image-pro Plus 6.0 software was used for optical density analysis of images,and the relationship between P value and0.05 was compared to judge the statistical significance.Results:1.Changes in thyroid function: Compared with the control group,the serum T3 and T4 levels of rats in the PTU drinking water group decreased(P<0.05),and the TSH level increased(P<0.05).2.Sex hormone changes: ELISA results showed that compared with the control group,the serum FSH and LH levels of the rats in the 0.05% PTU group decreased,the TT level decreased(P<0.05),and the PRL and E2 levels increased(P<0.05).3.The expression levels of Gn RH,RFRP-3 and kisspeptin mRNA in the hypothalamus: q RT-PCR results showed that compared with the control group,in the hypothalamus of the PTU drinking water group,the kisspeptin expression decreased(P<0.05)and RFRP-3 level increased(P<0.05).However,the difference was notstatistically significant in the comparison of Gn RH m RNA level(P>0.05).4.The expression levels of Gn RH,RFRP-3 and kisspeptin proteins in the hypothalamus:(1)ELISA results show that compared with the control group,the expression of Gn RH in the hypothalamus of rats in the PTU drinking water group was not statistically significant(P>0.05),the expression of kisspeptin decreased(P<0.05),and the expression of RFRP-3 increased(P<0.05).(2)The results of immunohistochemistry showed that the number of RFRP-3 positive cells in the hypothalamus of rats in the PTU drinking water group increased(43.13±2.14 vs 40.08±1.13,P<0.05),the number of kisspeptin positive cells decreased(54.685±1.89 vs65.71±1.44,P<0.05),and the number of Gn RH positive cells was not statistically significant(69.28±0.95 vs 69.51±0.75,P>0.05).5.The expression levels of Gn RH,RFRP-3 and kisspeptin proteins in the pituitary gland: ELISA results showed that in the pituitary gland of the PTU drinking water group,the expression of Gn RH was not statistically significant(P>0.05),the expression of kisspeptin was decreased(P<0.05),and the expression of RFRP-3 was increased(P<0.05).6.The expression of Gn RHR in the pituitary gland:(1)The results of qRT-PCR showed that compared with normal drinking water,Gn RHR in the PTU drinking water group was not statistically significant(P ﹥ 0.05).(2)The results of immunohistochemistry showed that compared with normal drinking water,Gn RHR in the PTU drinking water group was not statistically significant(28.68±5.95 vs 31.92± 5.85,P﹥0.05).Conclusions:Hypothyroidism in rats changes the expression of RFRP-3 and Kisspeptin in the hypothalamic-pituitary axis,which may be involved in the regulation of Gn RH and LH expression. |
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