Hang Shi

Regulation of adiposity by dietary calcium

Recent data from this laboratory demonstrate that increasing adipocyte intracellular Ca2+ results in a coordinated stimulation of lipogenesis and inhibition of lipolysis. We have also noted that increasing dietary calcium of obese patients for 1 year resulted in a 4.9 kg loss of body fat (P<0.01). Accordingly, we tested the possibility that calcitro‐phic hormones may act on adipocytes to increase Ca2+ and lipid metabolism by measuring the effects of 1,25‐(OH)2‐D in primary cultures of human adipocytes, and found significant, sustained increases in intracellular Ca2+ and a corresponding marked inhibition of lipolysis (EC50 ~50 pM; P<0.001), suggesting that dietary calcium could reduce adipocyte mass by suppressing 1,25‐(OH)2‐D. To test this hypothesis, we placed transgenic mice expressing the agouti gene specifically in adipocytes on a low (0.4%) Ca/high fat/high sucrose diet either unsupplemented or with 25 or 50% of the protein replaced by non‐fat dry milk or supplemented to 1.2% Ca with CaCO3 for 6 wk. Weight gain and fat pad mass were reduced by 26–39% by the three high calcium diets (P<0.001). The high calcium diets exerted a corresponding 51% inhibition of adipocyte fatty acid synthase expression and activity (P< 0.002) and stimulation of lipolysis by 3.4‐ to 5.2‐fold (P< 0.015). This concept of calcium modulation of adiposity was further evaluated epi‐demiologically in the NHANES III data set. After controlling for energy intake, relative risk of being in the highest quartile of body fat was set to 1.00 for the lowest quartile of Ca intake and was reduced to 0.75, 0.40, and 0.16 for the second, third, and fourth quartiles, respectively, of calcium intake for women (n=380;P≪ 0.0009); a similar inverse relationship was also noted in men (n=7114; P<0.0006). Thus, increasing dietary calcium suppresses adipocyte in‐tracellular Ca2+ and thereby modulates energy metabolism and attenuates obesity risk.—Zemel, M. B., Shi, H., Greer, B., DiRienzo, D., Zemel, P. C. Regulation of adiposity by dietary calcium. FASEB J. 14, 1132–1138 (2000)

Effects of dietary calcium on adipocyte lipid metabolism and body weight regulation in energy-restricted aP2-agouti transgenic mice

We have demonstrated previously a regulatory role for intracellular Ca2+ ([Ca2+]i) in adipocyte lipogenesis and lipolysis and have recently reported that 1,25‐(OH)2‐D increases adipocyte [Ca2+]i, which causes increased lipogenesis and decreased lipolysis. We have now tested the hypothesis that suppressing 1,25‐(OH)2‐D by increasing dietary calcium will suppress adipocyte [Ca2+]i, thereby facilitating weight loss by stimulating lipolysis and inhibiting lipogenesis in calorically (Kcal)‐restricted (70% of ad lib) aP2‐agouti transgenic (aP2‐a) mice. Mice (aP2‐a) exhibiting a pattern of obesity gene expression similar to humans were fed a low‐Ca (0.4%)/high‐fat/high‐sucrose diet for six weeks, resulting in a 27% and twofold increase in body weight and total fat pad mass, respectively, with a twofold increase in adipocyte [Ca2+]i (p<0.001). Mice were then either maintained on the same low‐Ca basal diet ad lib or Kcal‐restricted (70% of ad lib) on this diet either unsupplemented (basal) or with 25% or 50% of the protein replaced by non‐fat dry milk (medium or high) dairy or supplemented with CaCO3 to 1.2% Ca for six weeks. Adipocyte [Ca2+]i was unaffected by Kcal restriction but was reduced markedly by all three high Ca diets (290 vs. 130 nM, p<0.001). Body weight was reduced by 11%, 19%, 25%, and 29% by the restricted basal, high‐Ca, medium‐dairy and high‐dairy diets, with corresponding decreases in fat pad mass of 8, 42, 60, and 69% vs. ad lib basal (p<0.001), respectively. The high‐Ca diets caused similar decreases in fatty acid synthase activity 35%‐63%, p<0.001), increases in lipolysis (two‐ to threefold, p<0.001). All three high‐Ca diets increased adipocyte uncoupling protein 2 expression approximately twofold (p<0.02), with a corresponding increase in core temperature (0.48‐0.67°C, p<0.05). Thus, high‐Ca diets suppress adipocyte [Ca2+]i and thereby reduce energy storage and increase thermogenesis during Kcal restriction.

1α,25-dihydroxyvitamin D3 inhibits uncoupling protein 2 expression in human adipocytes

We recently demonstrated that suppressing 1α,25‐(OH)2‐D3 by increasing dietary calcium decreases adipocyte intracellular Ca2+ ([Ca2+]i), stimulates lipolysis, and inhibits lipogenesis. High calcium diets also increase core temperature and white adipose tissue uncoupling protein 2 (UCP2) expression in aP2‐agouti transgenic mice. Accordingly, we have evaluated the role of 1α,25‐(OH)2‐D3 in regulating human adipocyte UCP2 expression. Treatment of human adipocytes for 48 h with 1 nM 1α,25‐(OH)2‐D3 inhibited UCP2 mRNA and protein levels by 50% (P<0.002) and completely blocked isoproterenol‐ or fatty acid‐stimulated two‐ to threefold increases in UCP2 expression. However, a specific agonist for the membrane vitamin D receptor (mVDR), 1α,25‐dihydroxylumisterol3, was unable to inhibit basal, isoproterenol‐stimulated, or fatty acid‐stimulated UCP2 expression, whereas a specific mVDR antagonist,1β,25dihydroxyvitamin D3, was unable to prevent the 1α,25‐(OH)2‐D3 inhibition of UCP2 expression. In contrast, nuclear vitamin D receptor (nVDR) knockout via antisense oligodeoxynucleotide (ODN) prevented the inhibitory effect of 1α,25‐(OH)2‐D3 on adipocyte UCP2 expression and protein levels. These data indicate that 1α,25‐(OH)2‐D3 exerts an inhibitory effect on adipocyte UCP2 expression via the nVDR. Thus, suppression of 1α,25‐(OH)2‐D3 and consequent up‐regulation of UCP2 may contribute to our previous observation of increased thermogenesis in mice fed with high calcium diets.

Role of the sulfonylurea receptor in regulating human adipocyte metabolism

A regulatory role for intracellular Ca2+ ([Ca2+]i) in adipocyte lipogenesis, lipolysis and triglyceride accumulation has been demonstrated. Compounds acting on the pancreatic sulfonylurea receptor (SUR) to increase (e.g., glibenclamide) or decrease (e.g., diazoxide) [Ca2+]i cause corresponding increases and decreases in weight gain. However, these weight gain and loss effects have been attributed to insulin release rather than to the primary effects of these compounds on the adipocyte SUR and its associated KATP channel. Accordingly, we have evaluated the direct role of the human adipocyte SUR in regulating adipocyte metabolism. We used RT‐PCR with primers for a highly conserved region of SUR1 to demonstrate that human adipocytes express SUR1. The PCR product was confirmed by sequence analysis and used as a probe to demonstrate adipocyte SUR1 expression by Northern blot analysis. Adipocytes exhibited glibenclamide dose‐responsive (0–20 μM) increases in [Ca2+]i (P<0.05). Similarly, glibenclamide (10 μM) caused a 67% increase in adipocyte fatty acid synthase activity (P<0.001), a 48% increase in glycerol‐3‐phosphate dehydrogenase activity (P<0.01) and a 68% inhibition in lipolysis (P<0.01), whereas diazoxide (10 μM) completely prevented each of these effects. These data demonstrate that human adipocytes express a SUR that regulates [Ca2+]i and, consequently, exerts coordinate control over lipogenesis and lipolysis. Accordingly, the adipocyte SUR1 may represent an important target for the development of therapeutic interventions in obesity.—Shi, H., Moustaid‐Moussa, N., Wilkison, W. O., Zemel, M. B. Role of the sulfonylurea receptor in regulating human adipocyte metabolism. FASEB J. 13, 1833–1838 (1999)