Calcium In Adipocytes Research Articles

Calcium and obesity: effect size and clinical relevance

The connection between calcium intake and body weight was first described by McCarron et al.,1 who found an inverse relationship between these two variables in the National Health and Nutrition Examination Survey (NHANES) data. This observation was largely ignored for the next 16 years, until Zemel et al.2 described the weight-loss effect of calcium supplementation in a hypertension trial and suggested a cell biologic basis for the action of calcium in adipocytes. Since that time, there have been well over 100 reports of both experimental and observational studies evaluating the role of calcium intake in energy balance and body composition in humans. As summarized in 2009 in this journal,3 most of these studies found that increased calcium intake augmented the weight loss of energy-restricted diets, protected lean body mass, and reduced age-related weight gain, or that it had null effect. A very few reported negative findings (i.e., high calcium intakes were associated with weight gain), but the preponderance of the evidence tilted clearly toward the side of calcium intake favoring weight loss. In the present issue of this journal, Onakpoya et al.4 present yet another meta-analysis of this topic, with the results also favoring calcium. They report a highly significant, positive effect of calcium supplementation on total body weight loss and a somewhat larger effect on fat loss (suggesting some protection of lean tissue mass, which is a frequent casualty … [↵][1] RP Heaney , Creighton University, 2500 California Plaza, Omaha, NE 68178, USA. E-mail: rheaney{at}creighton.edu [1]: #xref-corresp-1-1

Calcium Mobilization Inhibits Lipid Accumulation During the Late Adipogenesis via Suppression of PPARγ and LXRα Signalings

Calcium plays a role as a signaling molecule in various cellular events. It has been reported that calcium suppresses adipocyte differentiation only in the early phase of adipogenesis. Herein, we demonstrate that treatment of A23187, a mobilizer of intracellular calcium, on day 4 post adipocyte differentiation could still reduce lipid accumulation in differentiating 3T3- L1 cells for 48 h. In addition, luciferase reporter gene and RT-Q-PCR assays demonstrate that A23187 can selectively inhibit transcriptional activities and expression of PPARγ and LXRα, suggesting that A23187 may reduce lipid accumulation in the late phase of adipogenesis via downregulation of PPARγ and LXRα expression and transactivation. Moreover, inhibition of HDAC activity by trichostatin A (TSA) partially blocked A23187-mediated downregulation of transcriptional activities of PPARγ and LXRα. Together, our data demonstrate that calcium mobilization inhibits expression and transcriptional activities of PPARγ and LXRα, resulting in reduced lipid accumulation in differentiating adipocytes, and thus, mobilization of intracellular calcium in adipocytes may serve as a new preventive and therapeutic approach for obesity.

Receptors for NPY and PACAP differ in expression and activity during adipogenesis in the murine 3T3‐L1 fibroblast cell line

Neuropeptides are involved in the regulation of food intake in the central nervous system, but they might also act on peripheral fat tissue via neuropeptide receptors. We investigated the receptor expression and activity of pituitary adenylate cyclase-activating polypeptide (PACAP) and of neuropeptide Y at the mRNA and protein levels in the 3T3-L1 fibroblast line during differentiation into adipocytes. Intracellular calcium concentration was measured by calcium imaging. The PACAP receptors PAC(1) and VPAC(2) as well as the neuropeptide Y(1) receptor were expressed at the mRNA level in fibroblasts, pre-adipocytes and adipocytes. The mRNA profile of the PAC(1) receptor isoforms showed the HOP sequence, whereas the HIP-isoform was present in subconfluent 3T3-L1 fibroblasts only. At the protein level, the mature 3T3-L1 adipocytes produced the PAC(1) and Y(1) receptors; only the PAC(1) receptor showed carbohydrate residues. Both neuropeptides induced an increase of intracellular calcium in mature adipocytes, which was absent in the precursor cells. These changes in calcium were mediated by Y(1) and PAC(1) receptors as demonstrated by the effects of specific receptor agonists and antagonists. As the PAC(1)-HOP receptor variant seems to be responsible for PACAP-mediated calcium influx in many cell types, the HOP sequence might also mediate the increase in intracellular calcium in adipocytes. Because a high intracellular calcium level is associated with lipogenesis, peptidergic innervation of adipose tissue might be involved in stress-induced obesity.

Increasing intracellular calcium in adipocytes increases fatty acid oxidation via the activation of AMP‐activated protein kinase

Increased dietary calcium intake is associated with reduced adiposity in humans. We hypothesized that increased intracellular Ca++ would augment fatty oxidation in 3T3‐L1 adipocytes in an AMP‐activated protein kinase (AMPK)‐dependent manner. Adipocytes were grown under euglycemic (EG) or hyperglycemic (HG) conditions and treated with or without calcium ionophores (4x10‐6 M A23187 and 4x10‐6 M Ionomycin, [AI]). After 15–60 min, AI increased intracellular Ca++ concentrations by 250% (P<0.001) and 150% (P<0.05), in EG and HG cells respectively, vs. controls. P‐AMPK and P‐Acetyl CoA carboxylase protein abundance was up regulated by AI in a time‐dependent manner. This was associated with increased 14C‐palmitate oxidation under EG (267%, P<0.001) or HG (200%, P<0.02) vs. the controls. However, the AMPK inhibitor, adenosine 9‐beta‐d‐arabinofuranoside (araA), attenuated the AI‐induced increase in fatty acid oxidation (P<0.01) under both conditions. HG results in insulin resistance and impaired glucose uptake in adipocytes. Compared to the control, AI improved this glucose uptake by 210% (P<0.01), even in the presence of araA (180%, P<0.05). These data indicate that intracellular Ca++ is a significant determinate of fatty acid oxidation via the control of AMPK activity. In contrast, the ability of increased intracellular Ca++ to enhance glucose uptake in HG conditions is independent of the activation of AMPK.

Dietary calcium attenuation of body fat gain during high-fat feeding in mice

Human epidemiological studies have supported the hypothesis that a dairy food-rich diet is associated with lower fat accumulation, although prospective studies and intervention trials are not so conclusive and contradictory data exist in animal models. The purpose of this study was to assess the effects on body weight and fat depots of dairy calcium (12 g/kg diet) in wild-type mice under ad libitum high-fat (43%) and normal-fat (12%) diets and to gain comprehension on the underlying mechanism of dairy calcium effects. Our results show that calcium intake decreases body weight and body fat depot gain under high-fat diet and accelerates weight loss under normal-fat diet, without differences in food intake. No differences in gene or protein expression of UCP1 in brown adipose tissue or UCP2 in white adipose tissue were found that could be related with calcium feeding, suggesting that calcium intake contributed to modulate body weight in wild-type mice by a mechanism that is not associated with activation of brown adipose tissue thermogenesis. UCP3 protein but not gene expression increased in muscle due to calcium feeding. In white adipose tissue there were effects of calcium intake decreasing the expression of proteins related to calcium signalling, in particular of stanniocalcin 2. CaSR levels could play a role in decreasing cytosolic calcium in adipocytes and, therefore, contribute to the diminution of fat accretion. Results support the anti-obesity effect of dietary calcium in male mice and indicate that, at least at the time-point studied, activation of thermogenesis is not involved.

Do adipocytes contain high affinity sulfonylurea receptors.

Sulfonylureas interact with specific, high affinity receptors on the pancreatic beta-cell to close ATP-sensitive K+ channels, depolarize the cell, activate Ca2+ influx through voltage-dependent Ca2+ channels, and trigger insulin secretion. We tested the hypothesis that sulfonylureas promote glucose uptake into 3T3-L1 cells or isolated rat adipocytes by similar mechanisms. Using 125I-labeled 5-iodo-2-hydroxyglyburide and either equilibrium binding or photoaffinity labeling, a high affinity sulfonylurea receptor was not found on plasma membranes of either the 3T3-L1 cells or rat adipocytes. Furthermore, glyburide did not inhibit 86Rb+ efflux (a marker for ATP-sensitive K+ channel conductance), increase free cytosolic calcium in adipocytes or 3T3-L1 cells, or increase basal or insulin-stimulated glucose uptake into 3T3-L1 cells or rat adipocytes. Parallel studies using a hamster insulin-secreting tumor cell line (HIT cells) easily demonstrated both the receptor and biological effects of glyburide on free cytosolic calcium and insulin secretion. Thus, rat adipocytes and 3T3-L1 cells do not possess the high affinity sulfonylurea receptor or respond to glyburide alone. We conclude that the antidiabetogenic effects of sulfonylureas are not mediated by a direct action of sulfonylureas to increase glucose uptake into adipose tissue and suggest that the major locus for sulfonylurea action is the beta-cell.

Regulation of GLUT-4 phosphorylation by intracellular calcium in adipocytes.

Sustained elevations in cytosolic calcium concentrations ([Ca2+]i) have been shown to render insulin target cells resistant to insulin action. In this study we examined the mechanisms of the detrimental effect of high levels of [Ca2+]i on insulin-induced 2-deoxyglucose (2-DOG) uptake. To elevate [Ca2+]i, we incubated rat adipocytes with either 40 mM potassium (K+) or 20 ng/ml PTH for 1 h for in vitro experiments and injected rats with PTH (injections of 50 micrograms, ip, every hour for 3 h) for in vivo studies. Adipocytes with elevated [Ca2+]i demonstrated a 30% decrease in insulin-stimulated 2-DOG uptake. A calcium channel blocker (nitrendipine) and a cAMP antagonist (RpcAMP) each partially restored insulin-stimulated glucose transport, but together they completely restored 2-DOG uptake. Concomitantly, we found a significant increase in phosphorylation of GLUT-4 in adipocytes with elevated [Ca2+]i. This change in GLUT-4 phosphorylation was also attenuated by nitrendipine and RpcAMP. These observations confirm that elevated [Ca2+]i diminishes insulin-stimulated glucose transport and suggest that increased phosphorylation of GLUT-4 in adipocytes with high [Ca2+]i may alter its intrinsic activity.

Effect of hormones on cytosolic free calcium in adipocytes

Some studies have indicated that insulin was able to increase the level of free cytosolic calcium in adipocytes [e.g. 7]. The present study was designed to examine this phenomenon. Insulin did not increase free cytosolic calcium, however oxytocin, vasopressin, α-adrenergic agonists and ATP did increase free cytosolic calcium in adipocytes. Other agonists which also did not alter calcium were epidermal growth factor, angiotensin II, glucagon, and β-adrenergic agonists. The effect of oxytocin at increasing free cytosolic calcium was inhibited by activation of protein kinase C with phorbol 12-myristate 13-acetate and by ADP ribosylation of a G i like protein with islet activating protein. The hormones that did increase cytosolic free calcium did so by mobilizing internal calcium and by promoting calcium influx. Even though insulin did not increase free cytosolic calcium, it was able to attenuate the α-adrenergic mediated increase in cytosolic free calcium. The fact that certain hormones can increase the level of the second messenger calcium in adipocytes implies that it may be a key intracellular regulator of adipocyte function as it is in many other tissues.

Cytosolic Free Calcium in Adipocytes: Distinct mechanisms of regulation and effects on insulin action

It has been proposed that an elevation in cytosolic free Ca2+ may play a role in either mediating or antagonizing the ability of insulin to stimulate glucose uptake in adipocytes. This question has been addressed in the present studies using isolated fura-2-loaded rat adipocytes stimulated with a variety of agonists. The effects of insulin, oxytocin, norepinephrine, ATP, and ionomycin on cytosolic free Ca2+ levels were assessed and compared with their effects on transport-limited glucose oxidation. Oxytocin and ionomycin at concentrations which caused 3-5-fold increases in cytosolic Ca2+, by releasing Ca2+ from internal stores, had no effect on insulin-stimulated glucose oxidation. ATP and norepinephrine which caused more modest increases in Ca2+, by mechanisms at least partially dependent on external stores, inhibited insulin-stimulated glucose oxidation. Insulin had no effect on basal Ca2+ levels nor did it modulate the Ca2+ elevation caused by other agonists. These data suggest that insulin-stimulated glucose transport is not associated with an increase in cytosolic Ca2+. In addition, although there appears to be a correlation between inhibition of insulin-stimulated glucose transport and the effect of certain agonists to promote Ca2+ influx, there is not a general obligatory relationship between an elevation in cytosolic Ca2+ and antagonism of this insulin action.