Serum Testosterone Level in Type 2 Diabetes Mellitus

Aim: To determine the serum testosterone levels in the type 2 diabetes mellitus and its correlation with biochemical parameters of glycemic and lipid metabolism. Methodology: A cross sectional study was conducted at Department of Biochemistry and Department of Medicine, A cross sectional study was conducted at the Department of Biochemistry, Diabetic outpatient department and Department of Medicine, Liaquat University Hospital. Male type 2 diabetics (n=100) and age matched male (n=100) were included as cases and control for study purpose. Diagnosed cases of type 2 DM, male gender and 40 – 60 years of age were included in the study protocol. Physical examination of male type 2 diabetics was performed by a consultant physician. Sera were separated from blood and stored in refrigerators at Original Research Article – 20 0 C. Blood glucose, A1C, and blood lipids (cholesterol, triglycerides, LDLc and HDLc) were detected by standard laboratory methods. Serum testosterone was measured by ELISA (competitive immuno- assay) assay commercial kit. Data was entered in Statistical software SPSS 21.0 version (IBM, Incorp, USA) for statistical analysis (P≤0.05). Results: Mean±SD age of male type 2 diabetics was 53.2 ± 11.1 years compared to 54.5±10.4 years in control (P=0.056). Serum Testosterone in cases was 10.85±4.7 mmol/L compared to 13.39±3.8mmol/L in control (P=0.0001). Low testosterone level was noted in 46% male.Serum Testosterone shows inverse correlation with RBG (r= -0.31, P=0.003), A1C (r= -0.23, P=0.014), Cholesterol (r= -0.24, P=0.014), TAGs (r= --0.78, P=0.0001) and HDLc(r= -0.70, P=0.0001). Serum testosterone proved positively correlated with LDLc(r= 0.670, P=0.0001). Conclusion: The present study finds low serum testosterone in male type 2 diabetes mellitus patients


INTRODUCTION
Diabetes mellitus (DM) is primarily a disorder of glucose metabolism characterized by chronic hyperglycemia [1,2]. It is caused by relative or absolute deficiency of hormone secreted by the β-cells of Islets of Langerhan's in Pancreas called the insulin. DM is of two types called; the type 1 and type 2 DM. DM is a multifactorial disorder regarding its etiology. Glucose metabolism is characterized primarily by hyperglycemia and secondarily by the hyperlipidemia and dyslipidemia. Type 2 DM (T2DM) affects different organ systems of body in particular the insulin independent cells of body causing end organ damage in nerve cell, kidneys, etc. 1,2 Several studies [3][4][5][6] have reported low testosterone levels in type 2 diabetics resulting in hypogonadism. It has been debated that the testosterone has relationship with onset of DM as risk factor that has not been proved. It has been shown type 2 diabetics have low circulating testosterone levels that might affect the glycemic status of diabetics. Low testosterone levels in type 2 diabetics have been said to predict the onset of diabetes. Testosterone is a cholesterol derived hormone that might be affected by the dyslipidemia and hyperlipidemia in diabetics. It has been reported that the Leydig cells of testes are adversely affected by the diabetic hyperglycemia and dyslipidemia resulting in their dysfunction and reduced secretion of testosterone in male population. As the testosterone plays vital role in different biological functions in the male life hence its low level has been predicted affecting the sexual life of male adversely. Numerous studies [7-9] have narrated negative association of serum testosterone, hyperglycemia and insulin resistance in the male diabetic's. As the DM is an endocrinopathies it may produce other endocrinopathy such as those of low testosterone resulting in disturbed biological functions in the male population. There seems to be highly significant association between these two endocrinopathies influencing each other mutually. Low serum testosterone occurs because of a number of mechanisms, such as hyperglycemia, dyslipidemia, increased oxidants load, Leydig cell injury, enzyme defects, androgen enzymopathy, androgen receptor polymorphism, etc. Visceral obesity of diabetics may contribute to hypogonadism. Sex hormone binding globulins (SHBGs), disturbed gonadotropin synthesis and secretion by Leydig cells have been implicated mechanisms. Oxidative load increases the cytokine secretion of TNF-α (tumor necrosis factor -α), Interleukins (IL-1β, IL-6), disturbed aromatase activity results in estrogen excess and male sex dysfunction [7-9]. Various studies have narrated strong association of low testosterone with metabolic syndrome in Caucasian and Asian men [7-9]. Currently, research has focused on Leydig cell dysfunction in type 2 diabetics, [9] it was concluded that testosterone deficiency and the DM burden is increasing in Pakistan [1,2] hence there is need for further research studies to be conducted at national level to make data available for highlighting the problem of male hypogonadism in type 2 diabetics. The present study was conducted to estimate serum testosterone levels and its correlation with biochemical parameters of glycemic and lipid control in the type 2 male diabetics.

MATERIALS AND METHODS
The present cross sectional study was conducted at the Department of Biochemistry, Diabetic outpatient department and Department of Medicine, Liaquat University Hospital. Study covered duration of January 2018 to February 2019. Sample size was calculated by using Rao software. Diagnosed cases of type 2 DM of male gender were selected according to inclusion and exclusion criteria. One hundred male type 2 diabetics and one hundred age matched male were included in study purpose. Male type 2 diabetics were inducted through non-probability purposive technique. Records of Diabetes mellitus were checked to fulfill the inclusion criteria (dianogsed DMT2, both sexes mal and female). Diagnosed cases of type 2 DM, male gender and 40 -60 years of age were included in the study protocol. Normal healthy male of similar age were included as control. Participants were asked of volunteer inclusion in the study protocol. They were informed interviewed the purpose of study, advantages and disadvantages. They were informed that the study will to improve the male sex function in the future and will not cause any physical or economic loss. Volunteers who gave full voluntary willingness qualified for inclusion in the study protocol. They were informed that the expenses of Laboratory investigations will be paid by the researcher. Physical examination of male type 2 diabetics was performed by a consultant physician. Findings were noted in a prestructured proforma. Participants handling was as per the "Helsinki`s Declaration" of human research. Patient data was kept confidential in lockers. Only principal researcher had access to the personal data. Volunteers were informed of giving consent for blood sampling. Venous blood sample was done by a disposable syringe preferably from the antecubital fossa after applying tourniquet. Five ml venous blood was taken in disposable syringe and divided into two parts. 2 ml was put in the sodium fluoride tubes and 3 ml was centrifuged at x3000 rpm for 15 minutes. Sera were separated from blood and stored in refrigerators at -20 0 C. Blood glucose, A1C, and blood lipids (cholesterol, triglycerides, LDLc and HDLc) were detected by standard laboratory methods. Serum testosterone was measured by ELISA (competitive immunoassay) assay commercial kit. Cobas chemistry analyzer was used for biochemical analysis at the laboratory. Lower limit of serum testosterone was taken as <3 ng/ml [9]. Values of cholesterol (≥200 mg/dl), TAGs (≥150 mg/dl), LDLc (≥100 mg/dl) and HDLc (≤50 mg/dl) were taken as hyperlipidemia/dyslipidemia [10]. Data was entered in Statistical software SPSS 21.0 version (IBM, Incorp, USA) for statistical analysis. Numerical data was analyzed by Student`s t-test and results were presented as mean±SD. Linear Pearson`s correlation was run to check association/ correlation of serum testosterone with the glycemic and lipid parameters. Level of significance was at 95% confidence interval (P≤0.05).

DISCUSSION
The present study is first cross sectional study analyzing the serum testosterone levels in the type 2 diabetes mellitus and its correlation with biochemical parameters of glycemic and lipid metabolism. Major objective of present research was to estimate the serum testosterone levels in type 2 diabetic male cases comparing with the normal healthy age matched control. Serum testosterone is most important gonadal androgen hormone that plays role in mediating the various physiological functions in the human body. Interestingly, the present study found low serum testosterone levels in type 2 male diabetics compared to control (p-value < 0.0001). In present study, the Mean±SD age of male type 2 diabetics was 53.2 ± 11.1 years compared to 54.5±10.4 years in control (P=0.056). Age finding of present study is in agreement with previous studies [6,7,9]. In present study, the Serum Testosterone in cases was 10.85±4.7 mmol/L compared to 13.39±3.8 mmol/L in control (P=0.0001) ( . We found high glycemic and lipid parameters in present study (Table 1) Table 2). The findings are in agreement with previous studies [9][10][11][12][13]. Previous studies [9-12] found inverse association of serum testosterone with fasting and random blood glucose. Kim et al. [12] reported the negative association of serum total testosterone with blood glucose (r=-0.142, P=0.002) and A1C values (r=-0.097, P=0.040) in male diabetics. Findings are in line keeping with the present study. Similar finding of negative linear correlation of fasting blood glucose and A1C with the testosterone is reported by another previous study [10]. They showed a significant negative correlation of serum testosterone level with Fasting glucose (r = −0.252, p = 0.001) and A1C (r = −0.697, p = 0.001) [10]. The findings of present study are worth to report the low serum testosterone occurs in male type 2 diabetics showing association with glycemic and lipid parameters. Limitations of present study are; first-small sample size, and secondcross sectional study design. The study sample belonged to peculiar ethnic group of male diabetic population hence findings cannot be generalized to other settings.

CONCLUSION
The present study finds low serum testosterone in male type 2 diabetes mellitus patients. Negative correlation of serum testosterone is noted with blood glucose, glycemic control, cholesterol, triglycerides and high density lipoprotein. Serum testosterone proved positively correlated with low density lipoprotein cholesterol in present study. Further research with large sample size in indigenous male diabetics is warranted.

DISCLAIMER
The products used for this research are commonly and predominantly use products in our area of research and country. There is absolutely no conflict of interest between the authors and producers of the products because we do not intend to use these products as an avenue for any litigation but for the advancement of knowledge. Also, the research was not funded by the producing company rather it was funded by personal efforts of the authors.

CONSENT
All authors declare that 'written informed consent was obtained from the patient.

ETHICAL APPROVAL
Study was conducted after the approval of Ethics committee.