Compositions and methods for regulating pancreatic beta cell function using adipsin

#The rising prevalence of obesity worldwide and its associated metabolic derangements, such as type 2 diabetes mellitus (T2DM), pose an enormous public health challenge (Van Gaal et al. (2006) Nature 444:875-880). Diabetes is a chronic and progressive disease characterized by insulin resistance and insulinopenia resulting from pancreatic beta cell failure and decreases in pancreatic beta cell mass (Ferrannini (2010) Cell Metabol. 11:349-352; Kasuga (2006) J. Clin. Invest. 116:1756-1760; Muoio and Newgard (2008) Nat. Rev. Mol. Cell. Biol. 9:193-205). Despite initial success in glycemic control with oral medications, patients with T2DM typically exhibit worsening glucose homeostasis over the span of a few years (Kahn et al. (2006) New Engl. J. Med. 355:2427-2443; Matthews et at (1998) Diab. Med. 15:297-303; Turner et al. (1996) Annal. Int. Med. 124:136-145). The natural history of T2DM includes a continuous decline in pancreatic beta cell function as illustrated by longitudinal follow up studies of patients (Festa et al. (2006) Diabetes 55:1114-1120). This deterioration leads to the need for additional therapies as well as the cumulative development of diabetic complications.

#Increasing adiposity is directly correlated with adipose inflammation and elaboration of proinflammatory cytokines, such as tumor necrosis factor-α, but whether this low-grade chronic inflammation is sufficient to trigger islet dysfunction is unknown (Hotamisligil (2006) Nature 444:860-867; Ouchi et al. (2011) Nat. Rev. Immunol. 11:85-97; Rosen and Spiegelman (2006) Nature 444:847-853; Shoelson et al. (2006) J. Clin. Invest. 116:1793-1801). Other factors, such as amyloidosis, glucolipotoxicity, failure of pancreatic beta cell expansion, apopotosis, and pancreatic beta cell dedifferentiation have also been posited (Kitamura (2013) Nat. Rev. Endocrinol. 9:615-623; Leroith and Accili (2008) Nat. Clin. Pract. Endocrinol. Metabol. 4:164-172; Muoio and Newgard (2008) Nat. Rev. Mol. Cell. Biol. 9:193-205; Prentki and Nolan (2006) J. Clin. Invest. 116:1802-1812; Weir et al. (2009) Diabetes Obes. Metabol. 11:82-90).

#Adipsin, also known as complement factor D, was the first adipokine described (Cook et al. (1987) Science 237:402-405). In fact, adipsin is one of the major proteins of adipose cells, but paradoxically circulating adipsin levels decline in many animal models of obesity and diabetes (Flier et al. (1987) Science 237:405-408). Adipsin was later identified to be complement factor D (Rosen et al. (1989) Science 244:1483-1487; White et al. (1992) J. Biol. Chem. 267:9210-9213), which catalyzes the rate-limiting step of the alternative pathway of complement activation (Xu et al. (2001) Proc. Natl. Acad. Sci. U.S.A. 98:14577-14582). Since then, adipsin has been shown to play pivotal roles in models of ischemia reperfusion (Stahl et al. (2003) Amer. J. Pathol. 162:449-455) and sepsis (Dahlke et al. (2011) J. Immunol. 186:3066-3075). Functions of this molecule include both the formation of the C5-C9 membrane attack complex and the generation of a number of signaling molecules, including the anaphylatoxins C3a and C5a (Ricklin et al. (2010) Nat. Immunol. 11:785-797). However, the function of adipsin in relation to energy homeostasis and systemic metabolism has been unknown.

#FIG. 1 includes 2 panels, identified as panels A and B, which show circulating adipsin levels wane with obesity. Panel A shows the results of adipsin protein analysis by Western blot of sera collected from WT mice placed on a regular chow or a high fat diet for the indicated number of weeks (w). Panel B shows the results of adipsin expression in WT and Adipsin−/− mice on a regular diet at the corresponding ages, in month (m). Sera were collected for analysis of adipsin protein by Western blot.

#FIG. 2 includes 2 panels, identified as panels A and B, which show the results of glucose homeostasis of adipsin-deficient mice on normal chow and short term high fat diets. A glucose tolerance test (GTT) was performed by intraperitoneal (i.p.) injection of glucose to WT and Adipsin−/− mice fed a regular diet (Panel A) or a HFD (Panel B) for 8 weeks with measurement of blood glucose concentrations at the indicated times.

#FIG. 3 includes 7 panels, identified as panels A, B, C, D, E, F, and G, which show that adipsin regulates adipose tissue inflammation and protects against diabetes. Panel A shows the results of WT and Adipsin−/− mice fed a high fat diet (HFD) and assayed for immune infiltration in visceral adipose tissues by qPCR. Panels B and C show the results of WT and Adipsin−/− adipose tissues stained for crown-like structures (CLS) with Mac2 antibody for immunohistochemistry. Representative pictures from each genotype are shown (Panel B) and numbers of CLS (Panel C) were quantitated. Paneled D and E show the results of WT and Adipsin−/− adipose tissues stained with toluidine blue for mast cells. Representative pictures from each genotype are shown (Panel D) and numbers of mast cells (Panel E) were quantitated. Panel F shows the results of a glucose tolerance test (GTT) performed by intraperitoneal (i.p.) injection of glucose to WT and Adipsin−/− mice fed a HFD with measurement of blood glucose concentrations at the indicated times. Panel G shows the results of an insulin tolerance test (ITT) performed by i.p. injection of insulin to WT and Adipsin−/− mice fed a HFD with measurement of blood glucose concentrations at the indicated times. For GTT and ITT experiments, n=12-16 per genotype. *P<0.05, **P<0.01, ***P<0.001. Also see FIGS. 1 and 2.

#For any figure showing a bar histogram, the bars from left to right for each indication correspond directly and in order to the boxes from top to bottom of the legend, as shown.

#The present invention is based in part on the discovery that adipsin is an adipokine secreted by fat cells that has a beneficial role in maintaining pancreatic beta cell function. Adipsin−/− animals were determined to exhibit exacerbated glucose intolerance due to insulinopenia. Furthermore, isolated islets from adipsin-deficient animals were determined to have reduced glucose-stimulated insulin secretion. Replenishment of adipsin to diabetic db/db mice effectively treated hyperglycemia by boosting insulin secretion. In addition, C3a, a peptide product generated in response to adipsin, was identified as a potent insulin secretagogue and the C3a receptor is shown to be required for optimal insulin secretion. C3a was determined to act on islets in part by enhancing cytosolic free Ca2+ in response to insulin secretagogues. In addition, type 2 diabetic patients with pancreatic beta cell failure were also determined to be deficient in adipsin. Thus, the adipsin pathway connects adipose cell function to pancreatic beta cell physiology and manipulation of this molecular switch can serve to therapeutically intervene in pancreatic beta cell-mediated disorders, such as diabetes.

#In order that the present invention may be more readily understood, certain terms are first defined. Additional definitions are set forth throughout the detailed description.