Besides playing a role in platelet

Besides playing a role in platelet aggregation (Chang et al., 2010; Kerrigan et al., 2009; Mourao-Sa et al., 2011), a function for CLEC2 on other immune flt3 has yet to be defined. Here we show that soluble CLEC2 regulates Kupffer cell polarization in the liver and improves glucose and lipid parameters in diabetic animals, thus revealing a novel physiological role for CLEC2 in both inflammation and metabolism. Our results demonstrate a previously unknown connection between CLEC2 and glucose and lipid metabolism, and support CLEC2 as a potential target for treating diabetes.
Materials and Methods
Results
Discussion Chronic inflammation, a hallmark feature of obesity and diabetes (Hotamisligil, 2006; Shoelson et al., 2006), is in part characterized by increased macrophage infiltration into adipose tissue, which in turn exacerbates insulin resistance (Weisberg et al., 2003; Xu et al., 2003). However, two distinct pathways for macrophage activation have been defined: pro-inflammatory M1 macrophages and anti-inflammatory M2 macrophages. Whereas M1 macrophages drive adipose inflammation in obese mice, macrophages residing in the adipose tissue of lean mice are predominantly of the M2 type and work to actively suppress inflammation (Lumeng et al., 2007a; Odegaard et al., 2007). Activation of liver-specific Kupffer cells contributes to liver steatosis and is also implicated in insulin resistance (Huang et al., 2010; Lanthier et al., 2010). Similar to adipose resident macrophages, Kupffer cells exhibit flexibility in that they can be activated toward either a classical pro-inflammatory state or an alternative anti-inflammatory state (Odegaard et al., 2008). Accordingly, alternatively activated M2-polarized Kupffer cells have been shown to ameliorate obesity-induced insulin resistance (Odegaard et al., 2008). Although a role for the C-type lectin-like receptor, CLEC2, in platelet activation has been well described (Chang et al., 2010; Kerrigan et al., 2009; Mourao-Sa et al., 2011), a role for CLEC2 in immune cell function had yet to be elucidated. Among all tissues examined, we observed the highest expression of Clec2 in the liver, presumably due to expression on the surface of Kupffer cells (Fig. 1C, Tang et al., 2010). Due to the lack of known endogenous ligands for CLEC2, we chose to focus on a loss-of-function study using the soluble CLEC2 ECD as a potential dominant negative factor to block endogenous CLEC2 receptor function. Using this approach, our results were consistent across all models tested and revealed that Fc-CLEC2(ECD) treatment can reproducibly induce markers of alternatively activated Kupffer cells, including Clec7a, Retnla, and Chi313, and significantly improved glucose and lipid metabolism. These results suggest that the endogenous CLEC2 receptor may play a pro-inflammatory role under disease conditions which is consistent with the observation that Clec2 expression in macrophages of diabetic animals is dramatically increased. Gene expression profiling further showed that Fc-CLEC2(ECD) stimulates a phagocytic activity in the liver. The liver has a highly developed filtering mechanism to maintain homeostasis in the circulation and Kupffer cells are key players in this process (Toth and Thomas, 1992). The Kupffer cells form a protective barrier for the systemic circulation and remove many harmful materials such as endotoxins and cellular debris through phagocytosis (Toth and Thomas, 1992). It is known that under diabetic and metabolic syndrome conditions, macrophages display altered immune activities such as decreased phagocytosis due to insulin resistance (Liang et al., 2007; Plotkin et al., 1996), and that dysfunction of the phagocytic activity can result in the accumulation of materials that cause inflammatory responses and exacerbate insulin resistance. Our results suggest that Fc-CLEC2(ECD) improves the deficiency in Kupffer cell phagocytosis present under diseased conditions, reduces pro-inflammatory cytokine release, and increases anti-inflammatory cytokine production. Given the important role played by inflammation in insulin resistance and metabolic disorders, our data suggests that the anti-inflammatory activity of Fc-CLEC2(ECD) contributes to the improvement of glucose and lipid metabolism observed in our studies.