Intracellular and Tissue Levels of Vitamin B12 in Hepatocytes Are Modulated by CD320 Receptor and TCN2 Transporter.

The liver mass constitutes hepatocytes expressing receptors for vitamin B12 (B12)-bound transporters in circulation. However, intrahepatic and circulating B12 interrelationship levels remain unclear. We assessed the intracellular B12 levels at various circulating B12 concentrations in human HepG2 cell-line and liver tissue levels of B12 in the C57BL/6 mouse model. In HepG2 cells treated with a range of B12 concentrations, the intracellular and circulatory B12 levels, transcript and protein levels of B12 receptor (CD320) and transporter (TCN2) were determined using immunoassays, qRT-PCR and Western blot, respectively. Similar assessments were done in plasma and liver tissue of C57BL/6 mice, previously fed a diet of either a high or low B12 (30.82 µg B12/kg and 7.49 µg B12/kg, respectively) for 8-10 weeks. The physiological B12 status (0.15-1 nM) resulted in increased levels of intracellular B12 in HepG2 cells compared to supraphysiological levels of B12 (>1 nM). Gene and protein expression of CD320 and TCN2 were also higher at physiological levels of B12. Progressively increasing extracellular B12 to supraphysiological levels led to relative decreased levels of intracellular B12, lower expression of gene and protein levels of CD320 and TCN2. Similar results were observed in liver tissue from mice fed on a low B12 diet verses high B12 diet. These findings suggest that unlike supraphysiological B12, physiological levels of B12 in the extracellular media or circulation accelerates active transport of B12, and expression of CD320 and TCN2, resulting in higher relative uptake of B12 in hepatocytes.

Low Vitamin B12 and Lipid Metabolism: Evidence from Pre-Clinical and Clinical Studies.

Obesity is a worldwide epidemic responsible for 5% of global mortality. The risks of developing other key metabolic disorders like diabetes, hypertension and cardiovascular diseases (CVDs) are increased by obesity, causing a great public health concern. A series of epidemiological studies and animal models have demonstrated a relationship between the importance of vitamin B12 (B12) and various components of metabolic syndrome. High prevalence of low B12 levels has been shown in European (27%) and South Indian (32%) patients with type 2 diabetes (T2D). A longitudinal prospective study in pregnant women has shown that low B12 status could independently predict the development of T2D five years after delivery. Likewise, children born to mothers with low B12 levels may have excess fat accumulation which in turn can result in higher insulin resistance and risk of T2D and/or CVD in adulthood. However, the independent role of B12 on lipid metabolism, a key risk factor for cardiometabolic disorders, has not been explored to a larger extent. In this review, we provide evidence from pre-clinical and clinical studies on the role of low B12 status on lipid metabolism and insights on the possible epigenetic mechanisms including DNA methylation, micro-RNA and histone modifications. Although, there are only a few association studies of B12 on epigenetic mechanisms, novel approaches to understand the functional changes caused by these epigenetic markers are warranted.

Seminal but not Serum Levels of Holotranscobalamin are Altered in Morbid Obesity and Correlate with Semen Quality: A Pilot Single Centre Study.

Vitamin B12 (cobalamin) is an essential cofactor in the one-carbon metabolism. One-carbon metabolism is a set of complex biochemical reactions, through which methyl groups are utilised or generated, and thus plays a vital role to many cellular functions in humans. Low levels of cobalamin have been associated to metabolic/reproductive pathologies. However, cobalamin status has never been investigated in morbid obesity in relation with the reduced semen quality. We analysed the cross-sectional data of 47-morbidly-obese and 21 lean men at Careggi University Hospital and evaluated total cobalamin (CBL) and holotranscobalamin (the active form of B12; holoTC) levels in serum and semen. Both seminal and serum concentrations of holoTC and CBL were lower in morbidly obese compared to lean men, although the difference did not reach any statistical significance for serum holoTC. Seminal CBL and holoTC were significantly higher than serum levels in both groups. Significant positive correlations were observed between seminal holoTC and total sperm motility (r = 0.394, p = 0.012), sperm concentration (r = 0.401, p = 0.009), total sperm number (r = 0.343, p = 0.028), and negative correlation with semen pH (r = -0.535, p = 0.0001). ROC analysis supported seminal holoTC as the best predictor of sperm number (AUC = 0.769 ± 0.08, p = 0.006). Our findings suggest that seminal rather than serum levels of holoTC may represent a good marker of semen quality in morbidly obese subjects.

Massive Weight Loss Obtained by Bariatric Surgery Affects Semen Quality in Morbid Male Obesity: a Preliminary Prospective Double-Armed Study.

OBJECTIVES: The aim of this study is to evaluate the effect of massive weight loss on the seminal parameters at 6 months from bariatric surgery. DESIGN: Two-armed prospective study performed in 31 morbidly obese men, undergoing laparoscopic roux-en-Y-gastric bypass (n = 23) or non-operated (n = 8), assessing sex hormones, conventional (sperm motility, morphology, number, semen volume), and non-conventional (DNA fragmentation and seminal interleukin-8), semen parameters, at baseline and after 6 months from surgery or patients' recruitment. RESULTS: In operated patients only, a statistically significant improvement in the sex hormones was confirmed. Similarly, a positive trend in the progressive/total sperm motility and number was observed, though only the increase in semen volume and viability was statistically significant (Δ = 0.6 ml and 10%, P < 0.05, respectively). A decrease in the seminal interleukin-8 levels and in the sperm DNA fragmentation was also present after bariatric surgery, whereas these parameters even increased in non-operated subjects. Age-adjusted multivariate analysis showed that the BMI variations significantly correlated with the changes in the sperm morphology (ß = -0.675, P = 0.025), sperm number (ß = 0.891, P = 0.000), and semen volume (r = 0.618, P = 0.015). CONCLUSION: The massive weight loss obtained with bariatric surgery was associated with an improvement in some semen parameters. The correlations found between weight loss and semen parameter variations after surgery suggest that these might occur early downstream of the testis and more slowly than the changes in the sex hormones.

Effect of liraglutide on proliferation and differentiation of human adipose stem cells.

Glucagon-Like Peptide-1 (GLP-1) receptor agonists, used as glucose-lowering drugs, also induce weight loss by inhibiting food intake. The present study was aimed at the assessment of the in vitro effects of the GLP-1 receptor agonist liraglutide on proliferation and differentiation of human adipose stem cells (ASC) obtained from subcutaneous adipose tissue of morbidly obese subjects undergoing bariatric surgery. Liraglutide (10-100 nM) significantly inhibited ASC proliferation and viability, with a maximum effect at 6 days of culture (45% and 50%, for liraglutide 10 and 100 nM, respectively); the effect was reverted by exendin 9-39. Glucose uptake was significantly reduced by liraglutide in a dose dependent manner. Treatment with liraglutide reduced intracellular lipid accumulation in differentiating ASC, together with FABP-4 mRNA expression (-18%, -23%, -46%, for 1 nM, 10 nM and 100 nM, respectively), whereas it stimulated adiponectin (APN) expression (1.86-, 2.64-, 2.28-fold increase, for 1 nM, 10 nM and 100 nM, respectively). Liraglutide exerts effects on human adipose cell precursors, inhibiting proliferation and differentiation, while stimulating the expression of the insulin-sensitizing adipokine APN. These effects could contribute to the actions of GLP-1 receptor agonists on body weight and insulin sensitivity.

Hypogonadism as an additional indication for bariatric surgery in male morbid obesity?

OBJECTIVE: Male obesity is often associated with reduced levels of circulating total (TT) and calculated free testosterone (cFT), with normal/reduced gonadotropins. Bariatric surgery often improves sex steroid and sex hormone-binding globulin (SHBG) levels. The aim of this study was to assess the effects of bariatric surgery on waist circumference (WC) and BMI, and on TT levels, in morbidly obese men, stratified, according to the gonadal state, in eugonadal and hypogonadal (TT<8 nmol/l) subjects. DESIGN: A cohort of morbidly obese patients (29 with hypogonadism (HG) and 26 without) undergoing bariatric surgery (37, 10, 6, and 2, with Roux-en-Y gastric bypass, laparoscopic adjustable gastric banding, biliopancreatic diversion and gastric sleeve, respectively) was studied at 6 and 12 months from the operation. METHODS: Anthropometric parameters (weight, BMI, WC) and sex hormones (gonadotropins, TT, cFT, estradiol (E2), SHBG) were assessed. RESULTS: WC was the only parameter significantly correlated with androgens, but not with E2, SHBG, and gonadotropins, at baseline. After surgery, a significant increase in TT, cFT, and SHBG, accompanied by a decrease in E2, was evident in the two groups. However, both TT and cFT, but not E2, SHBG, and gonadotropin variations, were significantly higher in the hypogonadal group at follow-up, with an overall 93% complete recovery from HG. Reduction in WC, but not BMI, was significantly greater in hypogonadal men (ΔWC=-29.4±21.6 vs -14.4±17.4 at 12 months, P=0.047). CONCLUSIONS: Recovery from obesity-associated HG is one of the beneficial effects of bariatric surgery in morbidly obese men. The present findings suggest that the gonadal state is a predictor of WC decrease after bariatric surgery.

Acrosome reaction is impaired in spermatozoa of obese men: a preliminary study.

OBJECTIVE: To compare spontaneous (Sp-AR) and P-induced acrosome reaction (AR) in spermatozoa of obese and lean subjects. SETTING: Bariatric unit at a university hospital. DESIGN: Prospective, observational study. PATIENT(S): Twenty-three obese (mean±SD body mass index [BMI], 44.3±5.9 kg/m2) and 25 age-matched lean (BMI, 24.2±3.0 kg/m2) subjects. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Spontaneous and P-induced AR in spermatozoa of obese and lean subjects. RESULT(S): A statistically significant difference was found between obese and lean cohorts in total T and calculated free T, E2, glycated hemoglobin, and high-density lipoproteins, whereas among the routine semen parameters analyzed, only immotile sperm percentage and ejaculate volume differed significantly. Spermatozoa of obese (n=13) vs. lean men (n=19) showed a higher Sp-AR (17.9%±7.2% vs. 8.3%±4.2%), which resulted in a reduced ability to respond to P evaluated as the AR-after-P-challenge parameter (3.5%±3.2% vs. 17.6%±9.2%). Multivariate analysis adjusted for age revealed a significant correlation between BMI, waist, E2, and glycated hemoglobin with both Sp-AR (age-adjusted r=0.654, r=0.711, r=0.369, and r=0.644, respectively) and AR-after-P-challenge (age-adjusted r=-0.570, r=-0.635, r=-0.507, and r=-0.563, respectively). A significant difference in sperm cholesterol content was reported between obese and lean men (29.8±19.5 vs. 19.1±14.6 ng/µg of proteins). CONCLUSION(S): Sperm AR is impaired in obese men, showing reduced response to P and elevated Sp-AR, associated with altered circulating levels of E2 and sperm cholesterol content.

Age as a predictive factor of testosterone improvement in male patients after bariatric surgery: preliminary results of a monocentric prospective study.

BACKGROUND: Male obesity can be associated with symptomatic alterations in sex hormones resulting in hypogonadism and impaired fertility. Surgical-induced weight loss can improve the sex hormone profile in men. The aim of the present study is to evaluate the levels of sex hormones in obese males before and after 6 months from bariatric surgery. Possible mechanisms and clinical implications are also discussed. METHODS: We evaluated levels of serum total testosterone (TT), sex hormone-binding globulin (SHBG), calculated free testosterone (cFT), follicular-stimulating hormone (FSH), luteinizing hormone (LH), and total estradiol (E2) in 20 male patients at the baseline and 6 months after bariatric surgery. RESULTS: Median [interquartile range] age at the time of surgery was 40.5 [27.2-46.7] years with a median [interquartile range] BMI of 43.6 [40.9-48.7] kg/m(2). The median baseline levels of TT, SHBG, cFT, LH, and FSH were reduced; levels of E2 were elevated. At 6 months from surgery, the median BMI dropped to 34.8 [31.7-40.5] kg/m(2), TT, SHBG, cFT, LH, and FSH increased, while levels of E2 decreased. The improvement in the sex hormone profile was more evident in younger patients, with a statistically significant difference in cFT following surgery and in the raise of TT and cFT between the groups of patients below and above 35 years. At multivariate analysis, the age was the best predictive factor of the postoperative variations of TT. CONCLUSIONS: These preliminary results confirm the general improvement in sex hormone profile in obese men after bariatric surgery and introduce the age as a possible contributing factor to this improvement.

Determinants of testosterone recovery after bariatric surgery: is it only a matter of reduction of body mass index?

OBJECTIVE: To explore the correlation models between body mass index (BMI) and sex hormones constructed from a male cross-sectional survey and evaluate the effects of surgery-induced weight loss on sex hormones in morbidly obese subjects that are not predicted by the constructed BMI correlation models. DESIGN: Cross-sectional population and longitudinal studies. SETTING: Bariatric surgery center in a university hospital. PATIENT(S): A cross-sectional survey of a male general population of 161 patients (BMI median [interquartile range] = 29.2 [24.8-41.9] kg/m(2)) in addition to 24 morbidly obese subjects (BMI = 43.9 [40.8-53.8] kg/m(2)) who were undergoing bariatric surgery were prospectively studied for 6 and 12 months. INTERVENTION(S): Bariatric surgery on 24 morbidly obese men. MAIN OUTCOME MEASURE(S): Cross-sectional population: construction of the best-fitting models describing the relationship between baseline BMI with total (TT) and calculated free (cFT) testosterone, E2, sex hormone-binding globulin (SHBG), FSH, and LH levels. Longitudinal study deviation between the observed sex hormone levels at 6- and 12-month follow-up and those expected on BMI bases. RESULT(S): The correlation of BMI with sex hormones was not univocally linear (E2), but the best-fitting model was exponential for TT, cFT, FSH, LH, and TT/E2 and power for SHBG. In addition to the significant improvement of all parameters observed after surgery in the longitudinal cohort, the increase in TT and SHBG, but not in cFT, was significantly higher than expected from the corresponding weight loss at 6 months from surgery (14.80 [12.30-19.00] nM vs. 12.77 [10.92-13.64] nM and 40.0 [28.9-54.5] nM vs. 24.7 [22.5-25.8] nM for TT and SHBG, respectively), remaining rather stable at 12 months. CONCLUSION(S): The increase in TT and SHBG, but not the increase in cFT, after bariatric surgery is greater than expected based on weight loss.