Bone morphogenetic protein 7 (BMP7) reverses obesity and regulates appetite through a central mTOR pathway

Abstract

Body weight is regulated by coordinating energy intake and energy expenditure. Transforming growth factor β (TGFβ)/bone morphogenetic protein (BMP) signaling has been shown to regulate energy balance in lower organisms, but whether a similar pathway exists in mammals is unknown. We have previously demonstrated that BMP7 can regulate brown adipogenesis and energy expenditure. In the current study, we have uncovered a novel role for BMP7 in appetite regulation. Systemic treatment of diet-induced obese mice with BMP7 resulted in increased energy expenditure and decreased food intake, leading to a significant reduction in body weight and improvement of metabolic syndrome. Similar degrees of weight loss with reduced appetite were also observed in BMP7-treated ob/ob mice, suggesting a leptin-independent mechanism utilized by BMP7. Intracerebroventricular administration of BMP7 to mice led to an acute decrease in food intake, which was mediated, at least in part, by a central rapamycin-sensitive mTOR-p70S6 kinase pathway. Together, these results underscore the importance of BMP7 in regulating both food intake and energy expenditure, and suggest new therapeutic approaches for obesity and its comorbidities.

Figure 1.

Systemic administration of BMP7 reduces appetite and body weight in DIO mice. A) Body weight monitoring of DIO mice receiving BMP7 or LacZ adenovirus (control), showing a weight-lowering effect of BMP7. B) DIO mice treated with BMP7 adenovirus had increased oxygen consumption as measured by CLAMS, compared to mice treated with LacZ adenovirus. C) DIO mice treated with BMP7 adenovirus consumed less food during a 12-d period, compared to mice treated with LacZ adenovirus. D) DIO mice treated with BMP7 adenovirus lost more weight compared to pair-fed animals, indicating that the effects of BMP7 on body weight reflect both increased energy expenditure and reduced food intake. E) BMP7-mediated weight loss in DIO mice was due to a decrease in fat mass, not lean mass, as measured by DEXA scanning. F) BMP7-treated DIO mice had lower serum leptin levels at the end of the study, consistent with reduced adiposity in these mice, compared to DIO mice treated with LacZ adenovirus. G, H) DIO mice treated with BMP7 adenovirus had improved glucose tolerance and increased insulin sensitivity as evaluated by glucose (G) and insulin (H) tolerance tests. I) H&E-stained liver sections of DIO mice treated with BMP7 or LacZ for 12 d. Mice treated with BMP7 (right panel) had reduced fat accumulation in their liver compared to control mice (left panel). *P < 0.05, **P < 0.01, ***P < 0.001.

Figure 2.

Systemic effects of BMP7 are leptin-independent. A, B) Treatment of leptin-deficient ob/ob mice with BMP7 adenovirus resulted in significant weight loss (A) and reduced food consumption (B) compared to ob/ob mice that received LacZ adenovirus (control) during a 13-d study. C–E) BMP7 adenovirus-treated mice had lower plasma insulin levels (C), higher adiponectin levels (D), and no change in glucose levels (E), indicating increased insulin sensitivity compared to LacZ-adenovirus-treated ob/ob mice. N.S., not significant. *P < 0.05, **P < 0.01, ***P < 0.001 vs. control.

Figure 3.

Expression of BMP7 ligand and receptors in the hypothalamus. A) In situ hybridization analysis of hypothalamic sections for BMPRII (pink), showing caudal hypothalamus (left panel) and rostral hypothalamus (right panel). Antisense represents specific staining (top panel), while sense staining represents background (bottom panel). BMPRII, a type 2 BMP receptor, was highly expressed throughout the hypothalamus. B, C) Immunofluorescence staining (green FITC signal) for the type 1 BMP receptors ALK3 (B) and ALK2 (C) in the mouse hypothalamus, along the third ventricle. ALK3 staining was most robust in the arcuate and paraventricular nuclei, while ALK2 staining was most robust in the dorsomedial and paraventricular nuclei. D) LacZ staining of mouse hypothalamus (top panel) and dorsal midbrain (including ventricular system and choroid plexus; bottom panel) showing BMP7-positive cells (blue) in BMP7 reporter mice. Arrowheads indicate choroid plexus surrounding the ventricles (bottom panel).

Figure 4.

Intracerebroventricular administration of BMP7 suppresses food intake. A) Intracerebroventricular administration of rhBMP7 acutely reduced food intake in C57BL/6 mice. Results are presented from a representative of 7 independent experiments. Leptin served as a positive control. B) Food intake 4 h after i.c.v. rhBMP7 or vehicle treatment of ob/ob mice, showing that leptin-deficient mice maintain an anorectic response to BMP7 administration. C) Hypothalamic staining for c-Fos immunoreactivity (brown) at 60 min after i.c.v. injection of mice with either vehicle (left panel), rhBMP7 (middle panel), or leptin (right panel), showing increased c-Fos expression in response to both leptin and BMP7 treatments in the arcuate and ventromedial nuclei. D) mRNA expression of hypothalamic neuropeptides at 4 h after i.c.v. rhBMP7, leptin, or vehicle as measured by Q-RT-PCR, showing a significant increase in POMC expression by BMP7 treatment. E) α-MSH immunostaining (brown) of hypothalamic sections 60 min after i.c.v. injection of mice with either vehicle (left panel), rhBMP7 (middle panel), or leptin (right panel), showing increased α-MSH expression in response to both leptin and BMP7 treatments, in the arcuate nucleus. F) Food intake 4 h after i.c.v. BMP7 or vehicle treatment in Agouti mice (KK, Cg-A^y/J), showing that Agouti mice maintain an anorectic response to BMP7 administration. *P < 0.05, **P < 0.01, ***P < 0.001 vs. control.

Figure 5.

BMP7 utilizes the mTOR pathway to regulate food intake. A) Western blot analysis demonstrating that BMP7, when injected i.c.v. in mice, activated multiple signaling pathways in the hypothalamus, most notably the p70S6K and pSTAT3 pathways. Cyclophilin A serves as a loading control. B) Intracerebroventricular rhBMP7-activated phos-p70S6K (brown) in the ARC (left panels) and PVN (right panels), as detected by immunohistochemical staining of hypothalamic sections using a specific antibody against phospho-p70S6K at 30 min following injection of BMP7. Arrows indicate cells that are positive for phos-p70S6K. C) Activation of SMAD and p70S6K signaling pathways by BMP7 in the hypothalamic neuronal cell line N25/2, as evaluated by Western blot analysis using phospho-specific antibodies. D) Western blot analysis of phosphorylated p70S6K in response to 30 min of BMP7 or insulin stimulation in the hypothalamic neuronal cell line N25/2. The 70 kDa-band corresponding to p70S6K is completely blocked by 1 h rapamycin pretreatment. β-Tubulin serves as a loading control. E) Rapamycin, an mTOR inhibitor, reversed the anorectic effect of rhBMP7 injected i.c.v. into mice. Average food consumption at 4 h and 16 h post-treatment are presented. *P < 0.05.