Skip to main page content
U.S. flag
See More

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2016 Sep;22(9):1023-32.
doi: 10.1038/nm.4145. Epub 2016 Aug 1.

Withaferin A is a leptin sensitizer with strong antidiabetic properties in mice

Jaemin Lee  1 , Junli Liu  1 , Xudong Feng  1 , Mario Andrés Salazar Hernández  1 , Patrick Mucka  1 , Dorina Ibi  1 , Jae Won Choi  1 , Umut Ozcan  1
Affiliations

Withaferin A is a leptin sensitizer with strong antidiabetic properties in mice

Jaemin Lee et al. Nat Med. 2016 Sep.

Abstract

The increasing global prevalence of obesity and its associated disorders points to an urgent need for the development of novel and effective therapeutic strategies that induce healthy weight loss. Obesity is characterized by hyperleptinemia and central leptin resistance. In an attempt to identify compounds that could reverse leptin resistance and thus promote weight loss, we analyzed a library of small molecules that have mRNA expression profiles similar to that of celastrol, a naturally occurring compound that we previously identified as a leptin sensitizer. Through this process, we identified another naturally occurring compound, withaferin A, that also acts as a leptin sensitizer. We found that withaferin-A treatment of mice with diet-induced obesity (DIO) resulted in a 20-25% reduction of body weight, while also decreasing obesity-associated abnormalities, including hepatic steatosis. Withaferin-A treatment marginally affected the body weight of ob/ob and db/db mice, both of which are deficient in leptin signaling. In addition, withaferin A, unlike celastrol, has beneficial effects on glucose metabolism that occur independently of its leptin-sensitizing effect. Our results show that the metabolic abnormalities of DIO can be mitigated by sensitizing animals to endogenous leptin, and they indicate that withaferin A is a potential leptin sensitizer with additional antidiabetic actions.

PubMed Disclaimer

Conflict of interest statement

COMPETING FINANCIAL INTERESTS

U. Ozcan is a scientific founder, shareholder, and scientific advisory board and board of directors member of ERX Pharmaceuticals Inc.

Figures

Figure 1
Figure 1
Identification of withaferin A with similar gene expression profile as celastrol. (a) Summary flow chart showing the identification of withaferin A as a leptin sensitizer and anti-obesity candidate. (b) Heat map representing the celastrol-regulated genes (20 most upregulated, blue; 20 most downregulated, red) obtained from MEFs treated with vehicle or celastrol (250 nM, 7 h). See also Supplementary Fig. 1a. (c) Distribution of absolute enrichment scores of the individual compounds in the CMAP database obtained using celastrol gene expression signature (Supplementary Fig. 1a). Also indicated is the chemical structure of withaferin A. (d) The heat maps generated from the CMAP database representing the celastrol regulated genes (50 most upregulated, blue; 50 most downregulated, red) and the corresponding changes in the same genes induced by withaferin A. (e) The heat maps representing the most upregulated (blue) and the most downregulated (red) 50 genes in the hypothalamus of celastrol-treated DIO mice (100 μg/kg, 4 d) and their corresponding gene expression changes in the hypothalamus of withaferin A-treated DIO mice (2 mg/kg, 4 d). The color scales in b,d,e represent the celastrol- and withaferin A-induced logarithmic fold changes in gene expressions. (f,g) Distribution of the celastrol- versus withaferin A-induced logarithmic fold changes in the expression of the genes in d,e, respectively. Line of best fit, and Pearson product-moment correlation coefficients (r) are shown.
Figure 2
Figure 2
Withaferin A reduces body weight and food intake of diet-induced obese mice but not that of lean mice. (a–e) DIO mice received vehicle or withaferin A (1.25 mg/kg) for 21 d. (a) Body weight and (b) percent change in body weight (n = 9, vehicle; n = 8, withaferin A). (c) Daily food intake during the first week treatment. The experiments in a–c were repeated in five cohorts (total n = 42, vehicle; n = 40, withaferin A). (d) Plasma leptin concentrations of DIO mice after 1, 2, and 3 weeks of treatment (n = 10 per group). (e) Fat percentage (left panel) and lean mass (right panel) after 3-week treatment (n = 9, vehicle; n = 8, withaferin A), which were repeated in five cohorts (total n = 46, vehicle; n = 44, withaferin A). (f–j) Lean mice received vehicle or withaferin A (1.25 mg/kg) for 21 d. (f) Body weight and (g) percent change in body weight (n = 9, vehicle; n = 10, withaferin A). (h) Daily food intake during the first week treatment. The experiments in f–h were repeated in three cohorts (total n = 21 per group). (i) Circulating leptin concentrations after 3-week treatment (n = 9, vehicle; n = 10, withaferin A), which were repeated in three cohorts (total n = 19, vehicle; n = 18, withaferin A). (j) Fat percentage (left) and lean mass (right) of lean mice after 3-week treatment (n = 9, vehicle; n = 10, withaferin A). Values are averages ± s.e.m. P values are determined by two-way ANOVA (a,b,f,g) or Student’s t test (c,d,e,h,i,j). ***P < 0.001. n.s.; not significant.
Figure 3
Figure 3
Leptin signaling-deficient mice are resistant to the weight reducing effect of withaferin A. (a–e) db/db mice received vehicle or withaferin A (1.25 mg/kg) for 21 d. (a) Body weight and (b) percent change in bodyweight during the treatment (n = 10, vehicle; n = 8, withaferin A). The experiments in a,b were repeated in seven cohorts (total n = 53, vehicle; n = 47, withaferin A). (c) Daily food intake during the first week treatment. (d) Plasma leptin concentrations after 3-week treatment period (n = 10, vehicle; n = 7, withaferin A). (e) Fat percentage (left) and lean mass (right) after 21-d treatment (n = 10, vehicle; n = 9, withaferin A). (f–i) ob/ob mice received vehicle or withaferin A (1.25 mg/kg) for 20 d. (f) Body weight and (g) percent change in bodyweight (n = 10 per group). The experiments in f,g were repeated in three cohorts (total n = 29, vehicle; n = 22, withaferin A). (h) Daily food intake during the first week treatment. (i) Fat percentage (left) and lean mass (right) of ob/ob mice after 20 d of treatment (n = 10 per group). Values are averages ± s.e.m. P values are determined two-way ANOVA (a,b,f,g) or by Student’s t test (c,d,e,h,i). ** P < 0.01, *** P < 0.001. n.s.; not significant.
Figure 4
Figure 4
Withaferin A increases leptin sensitivity in the hypothalamus of DIO mice. (a) Representative immunoblots of p-STAT3Tyr705, total STAT3 and tubulin from whole hypothalamus (left), and the quantified ratio between p-STAT3Tyr705 and total STAT3 (right). DIO mice received vehicle or withaferin A (5 mg/kg, 15 h), and then were injected with saline or leptin (1 mg/kg, 30 min). The experiments in a,b were repeated three times with similar outcomes (total n = 9, Veh + Sal; n = 10, Veh + Lep; n = 9, Wit A + Sal; n = 10, Wit A + Lep). (b–g) DIO mice were administered with vehicle or withaferin A (2 mg/kg) for 3 d and subsequently received saline or leptin (1 mg/kg). The hypothalamic samples were analyzed by immunohistofluorescence staining using p-STAT3Tyr705 specific antibodies. (b,d,f) Representative images of p-STAT3Tyr705 positive cells from (b) dorsomedial hypothalamus (DMH), (d) ventromedial hypothalamus (VMH) and (f) arcuate nucleus (ARC) (total numbers of analyzed images are provided in Online Methods). (c,e,g) The quantified results of total p-STAT3Tyr705 positive cell numbers (TCN, top panel) and fluorescence intensities (TFI, bottom panel) of the groups in b,d,f. Bar graphs in c,e,g represent the average of three experiments (total n = 12 mice per group). Values are averages ± s.e.m. P values are determined by one-way ANOVA with Bonferroni (a) or Tukey post hoc test (c,e,g). See Online Methods for the details of statistical analysis. *P < 0.05, **P < 0.01, ***P < 0.001. n.s.; not significant. Scale bars, 100 μm.
Figure 5
Figure 5
Withaferin A reduces ER stress. (a–c) DIO mice received vehicle or withaferin A (2 mg/kg) for 3 d. (a) Agrp, Npy, Pomc and Socs3 mRNA expression in the hypothalamus. (b) The quantified ratio of the signals between p-PERKThr980 and total PERK of the hypothalamic samples from five independent experiments in Supplementary Fig. 5c (total n = 23, vehicle; n = 26, withaferin A). (c) mRNA expression levels of spliced Xbp1s, Ddit3 and endoplasmic reticulum chaperones in the hypothalamus. Bar graphs in a,c represent the average of three independent experiments (total n = 16 per group). Values are represented as mean ± s.e.m. P values were determined by Student’s t test (a,c) or two-way ANOVA (b). *P < 0.05, **P < 0.01, ***P < 0.001. See also Supplementary Fig. 5.
Figure 6
Figure 6
Withaferin A’s beneficial effect on metabolic homeostasis. Individually caged (a–c) DIO (total n = 16 per group from two cohorts), (d–e) ob/ob (n = 5 per group) and (g–i) db/db mice (n = 6 per group) were placed in metabolic cages and received vehicle or withaferin A (1.5 mg/kg) for 3 d. Energy expenditure (kcal/h) of (a) DIO, (d) ob/ob and (g) db/db mice. Respiratory exchange ratios (RER) (VCO2/VO2) of (b) DIO, (e) ob/ob and (h) db/db mice. Ambulatory physical activity of (c) DIO, (f) ob/ob and (i) db/db mice. Bar graphs represent average of two dark (24–36 h and 48–60 h) and two light cycles (12–24 h and 36–48 h). Results in a–c are the average of two independent cohorts. Values are averages ± s.e.m. P values are determined by Student’s t test (a–i). **P < 0.01, ***P < 0.001.
See this image and copyright information in PMC

Comment in

References

    1. Wang Y, Beydoun MA, Liang L, Caballero B, Kumanyika SK. Will all Americans become overweight or obese? estimating the progression and cost of the US obesity epidemic. Obesity. 2008;16:2323–2330. - PubMed
    1. Olshansky SJ, et al. A potential decline in life expectancy in the United States in the 21st century. The New England journal of medicine. 2005;352:1138–1145. - PubMed
    1. Berrington de Gonzalez A, et al. Body-mass index and mortality among 1.46 million white adults. The New England journal of medicine. 2010;363:2211–2219. - PMC - PubMed
    1. Rodgers RJ, Tschop MH, Wilding JP. Anti-obesity drugs: past, present and future. Disease models & mechanisms. 2012;5:621–626. - PMC - PubMed
    1. Dalamaga M, et al. Leptin at the intersection of neuroendocrinology and metabolism: current evidence and therapeutic perspectives. Cell metabolism. 2013;18:29–42. - PubMed

MeSH terms