Summary

Hypogonadotropic hypogonadism is one of cause in the pathogenesis of erectile dysfunction (ED) in type 2 diabetic men. Oxidative stress has been defined an important factor associated with the complications of diabetes (DM). Therefore, we aim to investigate the correlation of oxidative stress parameters with gonadotropins and testosterone in type 2 diabetic men with ED. Fifty two diabetic men participated in this study and classified according to ED status as Group 1 (with ED), and Group 2 (without ED). We compared Group 1 with Group 2 and control group. ED was found four times greater in DM than in control group. Advance age, longer diabetes duration, worse metabolic control, microvascular complications, greater BMI and treatment with insulin were found significantly high in Group 1 comparing with the men in Group 2. Serum LH, FSH, testosterone and DHEA-S levels in Group1 were significantly lower than those in Group 2 and control group. Significantly higher levels of lipid peroxidation and lower total antioxidant status and erythrocyt SOD activity and negative correlation between MDA with gonadotropins and testosterone was found in Group 1. In conclusion, increase in lipid peroxidation may be one of the factor responsible for the disruption of the normal feedback mechanism in Hipothalamus-Pituitary-Gonadal (HPG) axis. We considered that antioxidant treatments, glycemic control and loosing weight should be reduced oxidative damage on HPG axis and also in the development of microvascular complications which are major organic causes lead to ED in DM.
Keywords: Erectile dysfunction, oxidative stress, type 2 diabetic men, endocrinologic causes, hypogonadotrophic hypogonadism

Summary

Hypogonadotropic hypogonadism is one of cause in the pathogenesis of erectile dysfunction (ED) in type 2 diabetic men. Oxidative stress has been defined an important factor associated with the complications of diabetes (DM). Therefore, we aim to investigate the correlation of oxidative stress parameters with gonadotropins and testosterone in type 2 diabetic men with ED. Fifty two diabetic men participated in this study and classified according to ED status as Group 1 (with ED), and Group 2 (without ED). We compared Group 1 with Group 2 and control group. ED was found four times greater in DM than in control group. Advance age, longer diabetes duration, worse metabolic control, microvascular complications, greater BMI and treatment with insulin were found significantly high in Group 1 comparing with the men in Group 2. Serum LH, FSH, testosterone and DHEA-S levels in Group1 were significantly lower than those in Group 2 and control group. Significantly higher levels of lipid peroxidation and lower total antioxidant status and erythrocyt SOD activity and negative correlation between MDA with gonadotropins and testosterone was found in Group 1. In conclusion, increase in lipid peroxidation may be one of the factor responsible for the disruption of the normal feedback mechanism in Hipothalamus-Pituitary-Gonadal (HPG) axis. We considered that antioxidant treatments, glycemic control and loosing weight should be reduced oxidative damage on HPG axis and also in the development of microvascular complications which are major organic causes lead to ED in DM.
Keywords: Erectile dysfunction, oxidative stress, type 2 diabetic men, endocrinologic causes, hypogonadotrophic hypogonadism

Introduction

Erectile dysfunction (ED) is the most common sexual disorder affects the diabetic men an average of 10-15 years earlier than it does his nondiabetic counterpart. Vasculopathy and neuropathy are the major organic pathophysiologic mechanisms leading to persistent ED in type 2 diabetic men [1]. On the other hand, previous studies demonstrated abnormal endocrinologic factors, primarily hypogonadotropic hypogonadism with diabetic ED. These conclusions were based on the presence of both pituitary and gonadal disorders [2-3]. Endocrinologic causes of ED in diabetics haven’t been clearly elucidated yet. Diabetes has a known pathologic effect on peripheral tissue innervation and vascularisation, both of which are critical for erectile function. Oxidative stress has been reported to play an important role in the development of both vascular and neural complications of diabetes [1,4]. This study was designed to find the correlation of the serum oxidative stress parameters with gonadotropins and testosterone in the type 2 diabetic men with ED.

Methods

Fifty two type 2 diabetic men aged between 35-60 years were randomly selected among consecutive patients followed in the Division of Endocrinology of the University Hospital, between 2003-2005. Diabetic men were classified according to ED status as Group 1 (with ED), and Group 2 (without ED). Group 1 was compared with Group 2 and control group in terms of gonadotropins, testosterone, and oxidative stress parameters.
The inclusion criteria were age below 60 years and married or living a stable relationship at least six months or more. The exclusion criteria were: [1] concurrent endocrine disorders other than diabetes, hypertension and/or dyslipidaemia, or surgical procedures likely to impair sexual function; [2] alcoholism or other substance abuse; [3] intake of diuretics, beta blockers, or other drugs (phosphodiesterase type 5 inhibitors intake and/or intracavernous prostaglandin E1 ) which have been shown to have effects on ED.
Eligible patients were included in the study after obtaining their informed consent. Information on patients’ height, weight, duration of diabetes, and mode of treatment, presence of diabetic microvascular and macrovascular complications, presence of ED, hypertension, and drug therapy was obtained. Body mass index (BMI:weight kg/ height m2) and Waist to Hip Ratio (WHR) was evaluated. Blood pressure was measured on the right arm of the seated patients after a rest of at least 10 minutes. Heavy smokers (≥20 cigarettes/day), patients drinking alcohol (10≥g/day), and have concomitant disease were excluded from the study.

Diabetic complications assessment

Diabetic nephropathy was evaluated in the morning using urinary albumin/creatinine ratio in a spot urine test. The ratio of more than 30 µg of albumin per miligram of creatinine was accepted as microalbuminuria. All patients were examined by a neurologist. In case of positive complaints (e.g. numbness, tingling, or burning) or physical examination findings, neuropathy was evaluated by electromyography. Retinopathy was evaluated by an ophthalmologist. Patients were grouped as being normal, or having background retinopathy (hard exudates, cotton-wool spots, hemorrhages, dilated veins, arteriovenous shunts, microaneurysms, capilary closure and dilation, or increased capillary permeability) or proliferative retinopathy (new vessels, scar, vitreal hemorrhage, or retinal detachment).

Erectile dysfunction

The presence of ED was investigated according to modified International Index for Erectile Function 5 (IIEF-5) criteria (5). Participants were asked to answer these questions including how often he had experienced problems in achieving and maintaining an erection during the past six months, with responses calibrated on a 5-level scale (from never to more than once per week. The IIEF-5 score was stratified into four group; normal (18–25), mild ED (14–17), moderate ED (10–13), and complete ED (5–9)(6).

Biochemical analysis

All blood samples were taken between 8.00 and 9.00 a.m. After the overnight fasting, 10 ml venous blood were drawn into heparin tubes. Immediately centrifuged and each sample was divided in two parts plasma and erythrocytes packet and stored at (-80 ‘C) studied at least within ten days.

Oxidative Stress Parameters:

Total antioxidant status (TAS) test was applied on the Olympus AU 600 analyser with the Randox Kit. Results were expressed mmol/l.
Malondialdehyde (MDA), an end product of unsaturated fatty acid peroxidation, can react with thiobarbituric acid to form a colored complex called thiobarbituric acid-reactive substances (TBARS). It was assayed in RBC and expressed as nmol/L.
SOD activity was measured by determining the reduction of nitroblue tetrazolium (NBT) by super-oxide anion produced with xanthine/xanthine oxi-dase system. One unit of SOD is defined as the amount of protein that inhibits the rate of NBT reduction by 50%. Results were expressed as units per milliliter plasma.

Hormones:

Radioimmunoassay was used to measure testosterone (the intra- and interassay coefficient of variation 7.5% and 7.8% respectively). LH (the intra- and interassay coefficient of variation 4.6% and 1.9% respectively) and FSH (the intra- and interassay coefficient of variation 4.3% and 3.2% respectively) were measured by Microparticle Enzyme Immunoassay.

Others:

HbA1c levels were determined by ion-exchange high performance liquid chromatography (normal range 4,3-6,1%). Blood glucose, cholesterol, and tryglyceride levels were determined by hexokinase method using an Olympus AU 600 autoanalyzer (Olympus Diagnostica, Hamburg, Germany).

Statistical analysis

Analysis was performed with a statistical package (SPSS version 11.0 SPSS, Inc., Chicago, IL). The results were expressed as mean ± standard deviation (SD). Comparisons between two independent samples were analyzed by with the Mann Whitney U-tests (metric variables), or Fisher’s exact t-test (dichotomous variables). The correlation between data were examined using Pearsons’ correlation. p values of less than 0.05 were considered to indicate significance.

Results

Fifty two men with type 2 diabetes mellitus were compared with age matched healthy men as a control group. The clinical characteristics of diabetic patients and control subjects are shown in (Table 1). Age, BMI, and blood pressure were similiar in diabetics and control group.
Table 1: Comparisons of clinical characteristics of the diabetic men and control group
The prevalence of ED among men with diabetes (69%) was nearly four times greater that of men without diabetes (18%). When the diabetic subjects were stratified according to their ED status, off the 36 patients, 19(53%) had mild ED, 13(36%) had moderate ED, and others 4(11%) had complete ED (Table 1).
Advance age, longer diabetes duration, worse metabolic control, microvascular complications (neuropathy, nephropathy and retinopathy), BMI and treatment with insulin were significantly higher in Group 1 compared with men in Group 2 (Table 1).
Serum LH, FSH, testosterone and DHEA-S levels in Group1 were significantly lower than the patients in Group 2 and control group (Table 2). Serum estradiol levels were slightly high in diabetics but not statistically significant for three groups. We observed diabetic patients more exposed to oxidative stress. In addition to, significantly higher levels of lipid peroxidation (p=0.0001) and lower total antioxidant status (p=0,0001) and erythrocyt SOD activity (p=0.0001) was found in Group 1 compared with the data of Group 2 (Table 2).
Table 2: Comparison of plasma hormones and oxidative stress parameters in diabetic men with control group
There was a negative correlation between MDA with LH (r=-0.31; p=0.03), FSH (r=-0.32; p=0.02), TAS (r=-0.46; p=0.001), SOD(r=-0.39; p=0.003) in diabetic men with ED (Table 3).
Table 3: Correlation analysis between MDA with serum hormone levels and oxidative stress parameters in diabetic men with ED.

Discussion

It is well known that diabetes is significantly associated with an elevated risk of ED. It occurs in younger diabetic men more than in the general population and is more frequently reported in the early stages of diabetes mellitus The prevalence of ED in men with diabetes mellitus has been reported to range from 20 to 71%. [1].
We observed the frequency of ED in diabetic men four times greater than those in healthy men. Moreover, ED was found significantly high in diabetic men with advanced age, longer diabetes duration, worse metabolic control, treatment with insulin, elevated BMI and having microvascular complications. Results of our study were in agreement with the previous studies [1,4-5].
Etiology of ED in diabetes is thought to be multifactorial including vascular, neurological, psychological and endocrinologic components. Hypogonadism is one of the factor which may also contribute to erectile dysfunction. The prevalence of hypogonadism was reported 24.5% in diabetic men versus 12.6% in normal fasting glucose subjects. Hypogonadism was defined as circulating total testosterone below (10.4nmol/l) associated with typical hypogonadism-related symptoms, such as reduction in sexual desire, leading to a decreased number of sexual attempts, and with higher depressive symptomatology [2].
The age-related reduction in serum testosterone levels in normal population is associated with increases in average concentrations of gonadotropins. However, we observed that diabetic men with ED had significantly lower levels of serum LH, FSH, testosterone and DHEA-S than the diabetic men without ED and control group. This hormonal profile in diabetic men was suggested it was not only testicular dysfunction also disruption in the normal feedback mechanism [1-3].
In addition to, both clinical and experimental studies had proven a definitive association of diabetic hypogonadism with alterations in the Hipothalamus-Pituitary-Gonadal (HPG) axis and Leydig cellular dysfunctions [2,7-9].
Serum estradiol levels were slightly higher in diabetics than that of the control subjects. Diabetic men and subjects in control group had a similar BMI. However, among all diabetic patients, ED was significantly found with a grater BMI. Moderate obesity results in reduction in SHBG (Sex Hormone Binding Protein) and total testo-sterone with normal free testosterone levels because of the increase in aromatase enzyme activity in adipose tissue [10]. Estradiol may contribute the negative feedback mechanism of gonadotropins.
It is known that the generation of reactive oxygen species and the reduction of antioxidant defense in diabetes playing an important role in the etiology of vascular and neurologic complications. Hyperglycemia catalysis oxidative stress so reduction in the superoxide dismutase (SOD), catalase (CAT), peroxidase (Px) and glutathione peroxidase (GSH-Px) activities was found in red blood cells (RBC) and other tissues of diabetics. Diabetic patients have significant defects in antioxidant protection and more prone to lipid peroxidation [1-4,11].
We observed that diabetic men with ED were more exposed to oxidative stress than the diabetics without ED and control group. Moreover, we found a negative correlation between MDA with LH, FSH, TAS, and SOD (Table 3). Findings of this study was similiar with the other literatures [11,19].
Previous studies reported that oxidative stress induced by chronic hyperglycemia may contribute to nervous tissue damage both central and peri-pheral neurons because of their high polyun-saturated lipid contents [5-10]. The importance of antioxidant treatment in the prevention of neuronal damage has been recently emphasised [12,14-16]. Studies had shown high luteinizing hormone (LH) content and reduced LH secretion in the pituitary of adult diabetic rats [17]. It may suggest that the disruption was on the receptor level.
Moreover, glycosylation is required for the biological activity of FSH and LH. They act via receptors located in the cell membrane. Accumulation of lipid peroxides lead to loss of membrane fludity and integrity, and inactivite receptors [18,19]. The increase in lipid peroxidation associated with the decrease in antioxidative defense system may be one of the factors responsible for the impairment of HPG axis.
In another study, the conflict result indicated that reactive oxygen radicals were not involved in possible testicular complications of diabetes because diabetes-induced activations of mitochondrial enzymes provided protection against oxidative stress [19,20].
In conclusion, the prevalence of complete ED was approximately four times higher than in the general population. Age, duration of diabetes, worse metabolic control, vascular and neural complications, obesity were positively associated with ED. We observed an positive correlation between lipid peroxidation and hypogonadism. It may be one of the factors responsible for the disruption of the normal feedback mechanism. The results of this present study together with previous studies have led us to think that antioxidant treatments, glycemic control and loosing weight should be reduced oxidative damage on HPG axis and also in the development of microvascular complications which are major organic causes leading to ED in type 2 diabetic men.

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