A very low carbohydrate ketogenic diet improves glucose tolerance in ob/ob mice independently of weight loss

Abstract

In mice of normal weight and with diet-induced obesity, a high-fat, low-carbohydrate ketogenic diet (KD) causes weight loss, reduced circulating glucose and lipids, and dramatic changes in hepatic gene expression. Many of the effects of KD are mediated by fibroblast growth factor 21 (FGF21). We tested the effects of KD feeding on ob/ob mice to determine if metabolic effects would occur in obesity secondarily to leptin deficiency. We evaluated the effect of prolonged KD feeding on weight, energy homeostasis, circulating metabolites, glucose homeostasis, and gene expression. Subsequently, we evaluated the effects of leptin and fasting on FGF21 expression in ob/ob mice. KD feeding of ob/ob mice normalized fasting glycemia and substantially reduced insulin and lipid levels in the absence of weight loss. KD feeding was associated with significant increases in lipid oxidative genes and reduced expression of lipid synthetic genes, including stearoyl-coenzyme A desaturase 1, but no change in expression of inflammatory markers. In chow-fed ob/ob mice, FGF21 mRNA was elevated 10-fold compared with wild-type animals, and no increase from this elevated baseline was seen with KD feeding. Administration of leptin to chow-fed ob/ob mice led to a 24-fold induction of FGF21. Fasting also induced hepatic FGF21 in ob/ob mice. Thus, KD feeding improved ob/ob mouse glucose homeostasis without weight loss or altered caloric intake. These data demonstrate that manipulation of dietary macronutrient composition can lead to marked improvements in metabolic profile of leptin-deficient obese mice in the absence of weight loss.

Fig. 1.

Effects of ketogenic diet (KD) feeding on weight and energy expenditure. A: 60 days of KD feeding led to sustained weight loss in wild-type (wt) but not ob/ob mice. Filled circles, chow-fed wt; open circles, KD-fed wt; filled squares, chow-fed ob/ob mice; open squares, KD-fed ob/ob mice. B: caloric intake was greater in chow-fed ob/ob mice compared with wt controls. Filled bars, chow fed; open bars, KD fed. *P < 0.001 wt vs. ob/ob. C: core temperature was lower in ob/ob mice but was not different in chow (filled bars) and KD (open bars)-fed groups. *P < 0.001 wt vs. ob/ob. D: indirect calorimety revealed increased V̇o₂ in KD-fed wt groups that was apparent in the dark phase (filled bars) or light phase (open bars). This effect was absent in ob/ob mouse groups. E: heat production during 48-h period of indirect calorimetry revealed no difference between chow-fed and KD-fed wt mice or chow-fed and KD-fed ob/ob mice. F: respiratory exchange ratio (RER) was markedly decreased in KD-fed groups in both the dark phase and light phase (*P < 0.001 chow vs. KD).

Fig. 2.

Effects of KD feeding on glucose metabolism. A: KD feeding normalized fasting glucose levels and reduced glucose excursions in ob/ob mice. Open circles, chow-fed wt; filled circles, KD-fed wt; open squares, chow-fed ob/ob mice; filled squares, KD. *P < 0.001 chow- vs. KD-fed ob/ob. B: KD feeding reduced fasting insulin levels. Filled bars, fasted; open bars, 45 min insulin. +P < 0.05 chow- vs. KD-fed wt mice. *P < 0.001 chow- vs. KD-fed ob/ob mice. Increment in circulating insulin in response to glucose challenge returned in KD-fed ob/ob mice. #P < 0.01 wt chow-fed basal vs. 45 min and ob/ob KD-fed basal vs. 45 min. C: insulin tolerance test revealed increased sensitivity in KD-fed ob/ob mice. Filled circles, chow-fed wt; open circles, KD-fed wt; filled squares, chow-fed ob/ob mice; open squares, KD. *P < 0.001 chow- vs. KD-fed ob/ob mice. D: quantitative insulin sensitivity check index (QUICKI) metric was greatly improved by KD feeding of ob/ob mice. Filled bars, chow; open bars, KD. #P < 0.01 chow- vs. KD-fed wt mice. *P < 0.001 chow- vs. KD-fed ob/ob mice.

Fig. 3.

Effects of KD feeding on expression of enzymes of hepatic metabolism. A and B: enzymes of lipid synthesis [fatty acid synthase (FAS) and stearoyl-coenzyme A desaturase 1 (SCD1)]. C–E: genes required for fatty acid transport and oxidation [fatty acid translocase (CD36) and long-chain acyl-coenzyme A dehydrogenase (ACADL) and 3-hydroxyacyl-coenzyme A dehydrogenase (HADH)]. F: peroxisome proliferator-activated receptor (PPAR)α target gene uncoupling protein-2 (UCP2). G and H: genes of ketosis [3-hydroxy-3-methylglutaryl-coenzyme A synthase 2 (HMGCS2) and 3-hydroxybutyrate dehydrogenase (HBDH)]. Filled bars, chow fed; open bars, KD fed. +P < 0.05, #P < 0.01, and *P < 0.001 chow vs. KD fed.

Fig. 4.

Effects of KD feeding on hepatic regulatory gene expression. A: fibroblast growth factor 21 (FGF21). B: PPARα. C: PPARγ. D: PPARγ coactivator-1α (PGC1α). E: PPARγ coactivator-1β (PGC1β). F: sterol-regulatory element-binding protein-1c (SREBP1c). Filled bars, chow fed; open bars, KD fed. +P < 0.05, #P < 0.01, and *P < 0.001 wt vs. ob/ob mice.

Fig. 5.

Leptin treatment and fasting induces FGF21 in ob/ob mice. A: weight loss following 72-h administration of saline (filled bars) or leptin (open bars) to wt or ob/ob mice. *P < 0.001 leptin treatment vs. saline control. B: induction of FGF21 gene expression resulting from leptin-mediated anorexia. +P < 0.05. C: FGF21 is induced by fasting in ob/ob mice. Filled bars, fed; open bars, fasted. +P < 0.05 and *P < 0.001 fasted vs. fed control. D: circulating levels of FGF21 reflect FGF21 gene expression in ob/ob mice fed (filled bars) and fasted (open bars). +P < 0.05 and *P < 0.001 fasted vs. fed control.