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We have previously established that human adipose cells and the human adipose cell line LS14 express the calcium sensing receptor (CaSR) and that its expression is elevated upon exposure to inflammatory cytokines that are typically elevated in obese humans. Research in recent years has established that an important part of the adverse metabolic and cardiovascular consequences of obesity derive from a dysfunction of the tissue, one of the mechanisms being a disordered secretion pattern leading to an excess of proinflammatory cytokines and chemokines. Given the reported association of the CaSR to inflammatory processes in other tissues, we sought to evaluate its role elevating the adipose expression of inflammatory factors. We exposed adipose tissue and in-vitro cultured LS14 preadipocytes and differentiated adipocytes to the calcimimetic cinacalcet and evaluated the expression or production of the proinflammatory cytokines IL6, IL1β and TNFα as well as the chemoattractant factor CCL2. CaSR activation elicited an elevation in the expression of the inflammatory factors, which was in part reverted by SN50, an inhibitor of the inflammatory mediator NFκB. Our observations suggest that CaSR activation elevates cytokine and chemokine production through a signaling pathway involving activation of NFκB nuclear translocation. These findings confirm the relevance of the CaSR in the pathophysiology of obesity-induced adipose tissue dysfunction, with an interesting potential for pharmacological manipulation in the fight against obesity- associated diseases.

The study of the mechanisms that trigger inflammation in adipose tissue is key to understanding and preventing the cardiometabolic consequences of obesity. We have proposed a model where activation of the G protein-coupled calcium sensing receptor (CaSR) leads to inflammation and dysfunction in adipose cells. Upon activation, CaSR can mediate the expression and secretion of proinflammatory factors in human preadipocytes, adipocytes, and adipose tissue explants. One possible pathway involved in CaSR-induced inflammation is the activation of the NLR family, pyrin domain-containing 3 (NLRP3) inflammasome, that promotes maturation and secretion of interleukin (IL)-1β. The present work aimed to study whether CaSR mediates the activation of NLRP3 inflammasome in the human adipose cell model LS14. We assessed NLRP3 inflammasome priming and assembly after cinacalcet-induced CaSR activation and evaluated if this activation is mediated by downstream ERK1/2 signaling in LS14 preadipocytes. Exposure to 2 μM cinacalcet elevated mRNA expression of NLRP3, CASP-1, and IL-1β, as well as an increase in pro-IL-1β protein. In addition, CaSR activation triggered NLRP3 inflammasome assembly, as evidenced by a 25% increase in caspase-1 activity and 63% IL-1β secretion. CaSR silencing (siRNA) abolished the effect. Upstream ERK pathway inhibition decreased cinacalcet-dependent activation of NLRP3 inflammasome. We propose CaSR-dependent NLRP3 inflammasome activation in preadipocytes through ERK signaling as a novel mechanism for the development of adipose dysfunction, that may favor the cardiovascular and metabolic consequences of obesity. To the best of our knowledge, this is the first report linking the inflammatory effect of CaSR to NLRP3 inflammasome induction in adipose cells.

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Obesity is a major current public health problem worldwide due to the severe co-morbid conditions that this disease entails. The development of obesity-related cardiometabolic disorders is in direct association with adipose tissue inflammation that leads to its functional impairment. Activation of the Calcium-Sensing Receptor (CaSR) in adipose tissue contributes to inflammation and adipose dysfunction. Autophagy, a process of cell component degradation, is closely related to inflammation in many diseases, however, whether autophagy is associated with CaSR-induced inflammation remains unknown. Using LS14 and SW872 preadipose cell lines as well as primary human preadipocytes, we show that CaSR activation with the allosteric activator cinacalcet induces autophagosome formation. Cinacalcet-induced LC3II content elevation was precluded by knockdown of the CaSR and enhanced by CaSR overexpression, indicating a specific effect. Autophagy inhibition using 3-methyladenine prevented CaSR-induced TNFα production, indicating that autophagy contributes to CaSR-induced inflammation in human preadipocytes. Our results suggest that modulation of CaSR-induced autophagy is an attractive target in obese inflamed adipose tissue, to prevent the development of diseases triggered by adipose dysfunction. We describe a novel mechanism and possible new target to modulate and prevent adipose inflammation and hence the resulting disease-generating adipose tissue dysfunction.

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SUMMARY The distribution of 16 antigens of the HLA-A and 15 antigens of the HLA-B series of HLA system, the blood groups ABO, and Rh antigens were studied in 40 alcoholics with cirrhosis, 18 alcoholics without cirrhosis, and in normal control subjects. The group of alcoholics with cirrhosis showed a significantly high frequency of HLA-B13 (corrected P < 0-01) when compared with normal subjects, while the frequency of HLA-B13 was similar to normal in alcoholics without cirrhosis. On the basis of these findings, its seems that the carriers of HLA-B1 3 are more susceptible to liver damage caused by alcohol. Both groups of alcoholics and the normal controls had a similar distribution of ABO blood groups and Rh antigens.Many alcoholics do not develop hepatic disease no matter how long and how much alcohol they have ingested (Insunza et al., 1969). This seems to indicate a different individual resistance and/or susceptibility to hepatic damage related to diet, genetic background, or other possible factors. An important role has been assigned to the human leucocyte antigens (HLA) system in the susceptibility of the host to certain diseases, so that this system could well be a genetic marker in this respect (NIH, 1975;Sasazuki et al., 1977).On the other hand, an association between the blood groups of the ABO system and different diseases has also been reported (Doll et al., 1960;Kingsbury, 1971).In the present study, we examined the relationship between antigens of the HLA system and blood groups of the ABO system and Rh antigens and the heavy drinker's susceptibility to chronic liver damage. MethodsHLA, blood groups, and Rh antigens were assayed in 58 subjects who were drinking or had been drinking more than the equivalent of 100 g ethanol daily for over 10 years. These 58 subjects were divided

We established that human adipose cells and the human adipose cell line LS14 express the calcium-sensing receptor (CaSR) and that its activation induces inflammatory cytokine production. Also, its expression is enhanced upon exposure to obesity-associated proinflammatory cytokines. We have thus proposed that CaSR activation may be associated with adipose dysfunction. Here, we evaluated a possible effect on adipogenesis. We induced adipose differentiation of primary and LS14 human preadipocytes with or without the simultaneous activation of CaSR, by the exposure to the calcimimetic cinacalcet. Activation of the receptor for 24 h decreased by 40 % the early differentiation marker CCAAT/enhancer-binding protein β. However, upon longer-term (10 day) exposure to the adipogenic cocktail, cinacalcet exerted the opposite effect, causing a dose-response increase in the expression of the mature adipose markers adipocyte protein 2, adiponectin, peroxisome proliferator-activated receptor γ, fatty acid synthase, and glycerol-3-phosphate dehydrogenase. To assess whether there was a time-sensitive effect of CaSR activation on adipogenesis, we evaluated the 10 day effect of cinacalcet exposure for the first 6, 24, 48 h, 6, and 10 days. Our observations suggest that regardless of the period of exposure, 10 day adipogenesis is elevated by cinacalcet. CaSR activation may interfere with the initial stages of adipocyte differentiation; however, these events do not seem to preclude adipogenesis from continuing. Even though adipogenesis (particularly in subcutaneous depots) is associated with insulin sensitivity and adequate adipose function, the implications of our findings in visceral adipocytes, especially in the context of inflamed AT and overnutrition, remain to be established.