Carnitine deficiency is associated with late-onset hypogonadism and depression in uremic men with hemodialysis

Abstract

Late-onset hypogonadism (LOH) and depression contribute to cardiovascular disease (CVD) in male hemodialysis (HD) patients. Carnitine deficiency is frequently observed in HD patients, playing a role in CVD. We examined whether carnitine deficiency was independently associated with LOH and depression in these patients. Twenty-six male HD patients underwent determinations of serum levels of free carnitine and testosterone. Status of LOH and depression were evaluated by questionnaires using aging male symptoms’ (AMS) scale and self-rating depression scale (SDS), respectively. Free carnitine and testosterone levels in male HD patients were significantly lower than those in age-matched healthy male subjects. Linear regression analysis showed that AMS scale was positively associated with SDS. Univariate regression analysis revealed that total carnitine (inversely), free carnitine (inversely) and HD duration were correlated with AMS scale. Multiple stepwise regression analysis revealed that free carnitine was an independent determinant of AMS scale. Furthermore, free carnitine was also independently correlated with SDS in male HD patients. This study demonstrated that decreased free carnitine levels were independently associated with AMS scale and SDS in male HD patients. The observations suggest that decreased free carnitine levels could be a marker and therapeutic target of LOH and depression in uremic men with HD.

• AMS scale
• carnitine deficiency
• depression
• hemodialysis
• late-onset hypogonadism
• SDS

Introduction

Late-onset hypogonadism (LOH) has been linked to poor quality of life and increased mortality in patients with end-stage renal disease [1]. About 52% of male hemodialysis (HD) patients were diagnosed as hypogonadism on the basis of low concentration of serum testosterone levels [2]. Furthermore, low testosterone levels have also been associated with the risk of cardiovascular disease (CVD) and all-cause mortality in male patients on HD [2,3]. On the other hand, depression, a mental disorder with a high personal, societal and economic impact, affects at least 20–30% of patients receiving HD therapy [4]. Depression could decrease the adherence to medication and perceived quality of life in patients undergoing HD [5,6]. In addition, it is not only associated with suicide risk but also correlated with the increased risk for CVD in patients on HD as well [7,8]. Although several factors such as malnutrition, inflammation, erythropoietin resistance, environmental stress and atherosclerosis have been proposed to contribute to the progression of LOH and depression in uremic men [9–12], the precise underlying mechanism is not fully understood. Moreover, therapeutic options for LOH and depression in male HD subjects are far from satisfactory [5,13,14]. Therefore, to identify a novel therapeutic target that could ameliorate LOH and depression is urgently needed for improving quality of life and preventing the progression of CVD in male HD patients.

Carnitine is a natural substance, which is supplied by the intake of protein-rich foods and also synthesized by the liver, kidney, skeletal, cardiac muscles, brain, epididymis and testis in humans [15,16]. Carnitine is involved in fatty acid β-oxidation and energy production by transporting long-chain fatty acids from the cytoplasm to mitochondria [16]. We, along with others, have reported that serum carnitine levels are significantly decreased in HD patients [17,18]. Since carnitine supplementation has been shown to improve aging-related sexual dysfunction assessed by nocturnal penile tumescence and International Index of Erectile Function Score and ameliorate cancer-related depression [19,20], carnitine deficiency may be an important causative factor for the progression of LOH and depression in male HD patients. However, it remains unclear which anthropometric, metabolic, clinical and biochemical variables, including serum carnitine, are independently correlated with LOH and depression in male patients with HD. Therefore, in this study, we examined (1) the correlation between LOH and depression status assessed by aging male symptoms’ (AMS) scale and self-rating depression scale (SDS), respectively, and (2) which anthropometric valuables were independently associated with AMS scale and SDS in male HD subjects.

Methods

Patients

Twenty-six male patients receiving chronic HD (mean age: 63.4 ± 10.0 years; mean duration of HD: 51.3 (10–185) months) and age-matched 15 healthy male subjects (mean age: 57.6 ± 7.1 years) underwent a complete history, physical examination and determinations of blood chemistries, including serum total, free and acylcarnitine and free testosterone. Patients were dialyzed for 4–5 h with high-flux dialyzers three times a week. Ten patients had diabetes mellitus, 23 patients received inhibitors of renin–angiotensin system (RAS) and five statins for the treatment of dyslipidemia.

Informed consent was obtained from all patients, and the study protocol was approved by the Institutional Ethics Committees of Kurume University School of Medicine, Japan.

Data collection

The medical history was ascertained by a questionnaire. Blood pressure was measured in the sitting position using an upright standard sphygmomanometer just before starting HD. Vigorous physical activity and smoking were avoided for at least 30 min before blood pressure measurement.

General symptoms for LOH were judged according to the AMS scale in all male patients [21]. Nineteen patients were further examined depression status by SDS [22]. The AMS scale is designed as self-administrated scale to assess symptoms of aging which consists of general well-being, joint pain and muscular ache, excessive sweating, sleep problems, increased need for sleep and often feeling tired, irritability, nervousness, anxiety, physical exhaustion, decrease in muscular strength, depressive mood, feeling burnt out, decrease in beard growth, decrease in ability/frequency to perform sexually, morning erections and sexual desire/libido [21]. Each question is scored on a scale of one through five. The SDS is designed to assess the level of depression for patients diagnosed with depressive disorder [22]. The SDS is a short self-administered survey to quantify the depressed status of a patient. There are 20 items on the scale that rate affective, psychological and somatic symptoms associated with depression. There are 10 positively worded and 10 negatively worded questions. Each question is scored on a scale of one through four.

Blood was drawn from arteriovenous shunt just before starting HD sessions for the determinations of hemoglobin, total protein, lipids (low-density lipoprotein-cholesterol and triglycerides), blood urea nitrogen, creatinine, uric acid, calcium and phosphate. Intact parathyroid hormone (PTH) was evaluated by an immunoradiometric assay (Allegro I-PTH; Nichols Institute, San Juan Capistrano, CA). β₂-microglobulin was measured by a latex immunoagglutination assay (Eiken Chemical Co., Ltd. Tokyo, Japan). Serum carnitine levels were determined as described previously [23]. Free testosterone levels were measured by electro-chemiluminescence immunoassay and radioimmunoassay, respectively (SRL Inc., Tokyo, Japan). Other blood chemistries were measured at a commercially available laboratory (Wako Pure Chemical Industries, Ltd, Osaka, Japan) as described previously [24]. Efficacy of HD was also evaluated by a single-pool fractional clearance of body water for urea (Kt/V) as described previously [25].

Statistical analysis

Data are presented as mean ± standard deviation. Use of RAS inhibitors and statins and the presence or absence of diabetes mellitus were coded as dummy variables. Since HD duration, triglycerides and intact PTH levels were not normally distributed, log-transformed values were used for statistical analysis. To compare clinical values between healthy controls and HD patients, unpaired t-test was performed. To evaluate the correlation between free carnitine and AMS scale and SDS or AMS scale and SDS, univariate regression analysis was performed. To determine the independent correlates of AMS scale and SDS, multiple stepwise regression analysis was performed. Statistical significance was defined as p < 0.05. All statistical analyses were performed with SPSS version 20 (Chicago, IL) system.

Results

Demographic data

Demographic data are shown in Table 1. Total and free carnitine and free testosterone levels in male HD patients were significantly lower, whereas acylcarnitine levels and acyl-to-free carnitine ratio were significantly higher than those in age-matched healthy subjects (total carnitine: 43.1 ± 10.4 versus 59.0 ± 6.0 µmol/l, p < 0.001, free carnitine: 25.2 ± 5.7 versus 48.1 ± 5.7  µmol/l, p < 0.001, free testosterone: 6.06 ± 2.48 versus 10.13 ± 3.54 pg/ml, p < 0.001, acylcarnitine: 17.9 ± 5.7 versus 15.4 ± 5.8 µmol/l, p < 0.001, acyl/free carnitine ratio: 0.72 ± 0.18 versus 0.23 ± 0.05, p < 0.001). AMS scale and SDS in HD patients were higher than normal reference ranges (36.8 ± 11.8, 43.9 ± 10.2, respectively, normal reference ranges: AMS scale <27, SDS: 20–44).

Table 1. Clinical characteristics of the patients.

	Control subjects	HD patients	p Value
No. of patients	15	26
Age (years old)	57.6 ± 7.1	63.4 ± 10.0	0.056
HD duration (months)  (range)^a		51.3 (10–185)
Body mass index (kg/m^2)	22.3 ± 2.4	21.3 ± 1.9	0.156
Systolic BP (mmHg)	131 ± 21	151 ± 15	0.001
Diastolic BP (mmHg)	81 ± 10	84 ± 11	0.450
Hemoglobin (g/dl)	15.6 ± 0.8	11.5 ± 0.9	<0.001
Total protein (g/dl)	7.37 ± 0.36	6.85 ± 0.49	0.001
LDL-cholesterol (mg/dl)	132.6 ± 38.3	83.2 ± 21.3	<0.001
Triglycerides^a (mg/dl)  (range)	106.9 (38–200)	93.4 (31–442)	0.953
BUN (mg/dl)	14.6 ± 3.5	66.1 ± 15.8	<0.001
Serum Cr (mg/dl)	0.8 ± 0.1	11.9 ± 2.3	<0.001
Uric acid (mg/dl)	5.38 ± 1.40	8.39 ± 1.47	<0.001
Corrected Ca (mg/dl)	8.65 ± 0.24	9.32 ± 0.50	<0.001
Phosphate (mg/dl)	3.22 ± 0.42	4.60 ± 1.14	<0.001
Intact-PTH (pg/ml) (range)^a	50.0 (31–94)	93.5 (25–250)	<0.001
β2-MG (mg/l)	1.6 ± 0.4	28.1 ± 6.2	<0.001
Total carntine (µmol/l)	59.0 ± 6.0	43.1 ± 10.4	<0.001
Free carnitine (µmol/l)	48.1 ± 5.7	25.2 ± 5.7	<0.001
Acyl carnitine (µmol/l)	11.0 ± 2.2	17.9 ± 5.7	<0.001
Acyl/free carnitine ratio	0.23 ± 0.05	0.72 ± 0.18	<0.001
Free testosterone (pg/ml)	10.13 ± 3.54	6.06 ± 2.48	<0.001
Kt/V		1.57 ± 0.10
AMS scale		36.8 ± 11.8
SDS		43.9 ± 10.2
DM (No.) (±)	15/0	16/10
Medication
RAS inhibitors (No.) (±)	15/0	3/23
Statins (No.) (±)	15/0	21/5

Values are shown as mean ± SD or range. No., number; HD, hemodialysis; BP, blood pressure; LDL, low-density lipoprotein; BUN, blood urea nitrogen; Cr, creatinine; Ca, calcium; PTH, parathyroid hormone; β₂-MG, β₂-microglobulin; AMS, aging male symptom; SDS, self-rating depression scale; DM, diabetes mellitus; and RAS, renin–angiotensin system.

^aThese variables are shown in the original scale after using log-transformed values.

Correlates of AMS scale

Univariate regression analysis showed that AMS scale was positively associated with SDS in HD patients (r² = 0.366, p = 0.006; Figure 1). Furthermore, HD duration (p = 0.048), total carnitine (inversely, p = 0.013) and free carnitine (inversely, p = 0.003, Figure 2A) were significantly associated with AMS scale (Table 2). Because these significant parameters could be closely correlated with each other, multiple regression analysis was performed. Multiple stepwise regression analysis showed that free carnitine (β = −0.559, p = 0.003) was a sole independent correlate of AMS scale (r² = 0.313; Table 2).

Figure 1. Correlation between AMS scale and SDS in male HD patients (n = 19). AMS, aging male symptom; SDS, self-rating depression scale; and HD, hemodialysis.

Figure 2. (A) Correlation between AMS scale and free carnitine levels in male HD patients (n = 26). (B) Correlation between SDS and free carnitine levels in male HD patients (n = 19). AMS, aging male symptom; SDS, self-rating depression scale; and HD, hemodialysis.

Table 2. Univariate and multiple stepwise regression analysis for the correlates of AMS scale.

	Univariate			Multiple stepwise regression
Variables	β	SE	p Value	β	SE	p Value
Age	0.023	0.240	0.910
HD duration^a	0.391	2.899	0.048
Body mass index	−0.083	1.245	0.685
Systolic BP	0.067	0.161	0.743
Diastolic BP	0.049	0.219	0.810
Hemoglobin	−0.026	2.754	0.899
Total protein	0.087	4.929	0.672
LDL-cholesterol	0.251	0.110	0.217
Triglyceride^a	−0.060	3.652	0.770
BUN	−0.160	0.151	0.435
Serum Cr	−0.089	1.069	0.667
Uric acid	0.234	1.597	0.249
Corrected Ca	0.139	4.783	0.499
Phosphate	0.067	2.120	0.746
Intact-PTH^a	0.046	4.940	0.825
β2-MG	0.114	0.388	0.579
Total carnitine	−0.483	0.204	0.013
Free carnitine	−0.559	0.350	0.003	−0.559	0.313	0.003
Acyl carnitine	−0.312	0.400	0.121
Acyl/free ratio	0.148	13.009	0.472
Free testosterone	−0.078	0.973	0.706
Kt/V	−0.062	23.385	0.765
DM	0.269	4.692	0.184
RAS inhibitors	−0.119	7.366	0.563
Statins	−0.061	6.003	0.768

β, standardized regression coefficients; SE, standard error; AMS, aging male symptom; HD, hemodialysis; BP, blood pressure; LDL, low-density lipoprotein; BUN, blood urea nitrogen; Cr, creatinine; Ca, calcium; PTH, parathyroid hormone; β₂-MG, β₂-microglobulin; DM, diabetes mellitus; RAS, renin–angiotensin system.

^aThese variables are shown in the original scale after using log-transformed values. R² = 0.313.

Correlates of SDS

We next examined which clinical variables are independent determinants of SDS in our subjects. Univariate regression analysis revealed that intact PTH (p = 0.028) and free carnitine (inversely, p = 0.022; Figure 2B) were correlated with SDS in HD patients (Table 3). In the multiple stepwise regression analysis, free carnitine was found to be independently associated with SDS (r² = 0.270; Table 3).

Table 3. Univariate and multiple stepwise regression analysis for the correlates of SDS.

	Univariate			Multiple stepwise regression
Variables	β	SE	p Value	β	SE	p Value
Age	−0.183	0.248	0.454
HD duration^a	0.110	3.024	0.653
Body mass index	0.081	1.278	0.742
Systolic BP	−0.011	0.198	0.964
Diastolic BP	0.178	0.229	0.466
Hemoglobin	−0.190	2.512	0.435
Total protein	0.058	4.901	0.812
LDL-cholesterol	0.306	0.124	0.202
Triglyceride^a	−0.315	3.341	0.188
BUN	0.063	0.149	0.797
Serum Cr	−0.105	1.064	0.699
Uric acid	0.360	1.534	0.130
Corrected Ca	−0.018	4.707	0.941
Phosphate	0.215	1.905	0.376
Intact-PTH^a	0.503	4.096	0.028
β2-MG	−0.110	0.368	0.655
Total carnitine	−0.343	0.215	0.150
Free carnitine	−0.520	0.340	0.022	−0.520	0.340	0.022
Acyl carnitine	−0.082	0.426	0.739
Acyl/free ratio	0.400	11.501	0.089
Free testosterone	−0.148	1.111	0.546
Kt/V	−0.248	20.130	0.305
DM	0.253	4.696	0.297
RAS inhibitors	−0.309	6.250	0.198
Statins	−0.033	5.439	0.893

β, standardized regression coefficients; SE, standard error; SDS, self-rating depression scale; HD, hemodialysis; BP, blood pressure; LDL, low-density lipoprotein; BUN, blood urea nitrogen; Cr, creatinine; Ca, calcium; PTH, parathyroid hormone; β₂-MG, β₂-microglobulin; DM, diabetes mellitus; RAS, renin–angiotensin system.

^aThese variables are shown in the original scale after using log-transformed values. R² = 0.270.

Discussion

We demonstrated In this study that (1) serum free testosterone and free carnitine levels were significantly decreased and that AMS scale and SDS were higher in men with HD; (2) there was a positive correlation between AMS scale and SDS in HD patients; and (3) free carnitine was a sole independent determinant of AMS scale and SDS in male subjects on HD.

There are a couple of papers to suggest the correlation between LOH and depression status in aging male [26,27]. Patients with LOH had significantly higher scores for Beck Depression Inventory and Beck Anxiety Inventory [26]. Testosterone replacement therapy has been reported to not only improve the low testosterone-related symptoms but also ameliorate depression subscale in LOH patients in Chinese population [27]. Furthermore, depression score was found an independent risk factor for the development of erectile dysfunction in HD patients [28]. In this study, since free carnitine, but not testosterone levels, were an independent determinant of both AMS scale and SDS, carnitine deficiency rather than decreased free testosterone values may be a common soil for the progression of LOH and depression in male HD patients. These observations suggest that L-carnitine administration may be a novel therapeutic target for preventing the progression of LOH and depression mode in male uremic patients with HD.

In this study, total and free carnitine levels were negatively associated with AMS scale in HD patients. Carnitine is expressed in the epididymis and testis in humans and its levels in the seminal plasma were associated with sperm motility [15]. Furthermore, carnitine supplementation has been shown to ameliorate aging-related sexual dysfunction in aged men [19] and augment the efficacy of sildenafil, an inhibitor of phosphodiesterase-5, which could restore the sexual potency after bilateral nerve-sparing radical retropubic prostatectomy [29]. In addition, supplementation of carnitine has been reported to inhibit the development and progression of CVD in animal models [30], free fatty acid-induced endothelial dysfunction [31] and decline in mental performances [32] in humans. Therefore, carnitine supplementation might also prevent the progression of LOH and subsequently reduce the risk of CVD in male HD patients.

We did not know in this study the exact mechanisms for the correlation between low free carnitine levels and depression in male HD subjects. However, L-carnitine supplementation significantly improved depression mode and fatigue in carnitine-deficient patients with cancer [20]. Moreover, intensive L-acetylcarnitine treatment has been reported to ameliorate main mental parameters of the senile brain in humans [33]. Since depression status could also be closely correlated with LOH, carnitine supplementation may act on central nerve system by partly improving the gonadal function, which could lead to the amelioration of depression mode in male HD patients.

Limitations

There are several limitations in this study. First, sample size of this study was small. Second, it was a cross-sectional study, and therefore, could not assess the question whether deficiency of serum carnitine levels was a cause or consequence of LOH syndrome or depression. In uremic patients with HD, more than 80% of free carnitine was eliminated from the blood via HD [18]. So, our observations suggest that free carnitine levels could be a marker of LOH and depression status in male HD patients. However, it remains unclear whether carnitine could also be a marker and therapeutic target in female HD patients. Further longitudinal and/or interventional studies are needed to clarify whether carnitine supplementation could improve LOH and depression mode and subsequently reduce the risk of future CVD in male patients with HD.

Declaration of interest

The authors have no conflicts of interest to declare.

This work was supported, in part, by a Grant-in-Aid for Welfare and Scientific Research (C) (no. 25461239) from the Ministry of Education, Culture, Sports, Science and Technology of Japan (K.F) and by Grants of MEXT-Supported Program for the Strategic Research Foundation at Private Universities, the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan (S. Y.).