Insulin-containing lipogenic stimuli suppress mast cell degranulation potential and up-regulate lipid body biogenesis and eicosanoid secretion in a PPARγ-independent manner

Abstract

Lipid bodies are most studied in adipocytes, where the lipogenic action of insulin initiates their formation. Here, we test the hypothesis that insulin may regulate lipid body content in mast cells and hence, modify their proinflammatory potential. Our data show that insulin causes lipid body accumulation in RBL2H3 and BMMCs. Lipid body accumulation in mast cells is associated with enhanced levels of leukotriene-synthesizing enzymes (LTC4S and 5-LO). Increased basal and antigen-stimulated release of LTC4 is observed in insulin-treated mast cells. Concomitantly, the insulin-containing lipogenic stimulus induces a phenotypic change in mast cells, where this enhancement in leukotriene levels is accompanied by a marked down-regulation in secretory granule content and release in response to stimulus. Mast cells exposed to insulin exhibit altered scatter and fluorescence properties, accumulating in a SSC(lo)FSC(hi) population that exhibits decreased BS staining and degranulation responses and is enriched in NR-positive lipid bodies and eicosanoid synthesis enzymes. Lipid body accumulation in mast cells is mechanistically distinct from the process in adipocytes; for example, it is independent of PPARγ up-regulation and does not involve significant accumulation of conjugated glycerides. Thus, chronic exposure to metabolic stimuli, such as insulin, may be a determinant of the proinflammatory potential of the mast cell.

Lipid body accumulation alters calcium signaling dynamics in immune cells

“…The latter are an established context for the study of FcεRI-induced calcium signalling, exhibiting store release from diverse thapsigargin-sensitive and –insensitive stores, store-operated calcium entry via Orai family members and non selective cation channel-mediated calcium influx through TRP family members [32]. Chronic exposure to insulin is a convenient method to induce a steatosis-like accumulation of LB in the cytosol [13], which can be confirmed using Oil Red O (ORO) staining (Figure 1A-C). At the population level (Figure 1D), FcεRI-induced bulk calcium fluxes are suppressed in cells treated with chronic insulin to induce steatosis.…”

“…The best-characterized example of this is the accumulation of dietary lipids in foamy macrophages during atherosclerosis, with these lipid droplet pools being sensitive to hormones such as resistin and leptin [12]. Recently we described sensitivity of mast cell lipid body numbers, both in vitro and ex vivo , to insulin, with chronic insulin exposure driving steatosis and marked phenotypic changes in these cells ([13], WEG and HT, manuscript submitted).…”

“…LB in immunocytes have some commonalities in terms of protein content and lipidome with LD, but also manifest important differences [12, 13, 23-25]. Our studies suggest that even in response to an endocrine input (chronic hyperinsulinemia), the expansion in LB content in mast cells is predominantly comprised of LB that store inflammatory precursors, alongside energy substrates that may also function as precursors for bioactive lipids [13, 24]. This expansion is, unsurprisingly, linked to an elevated potency in antigen mediated de novo synthesis and release of bioactive lipids such as LTC4 [13].…”

“…Our studies suggest that even in response to an endocrine input (chronic hyperinsulinemia), the expansion in LB content in mast cells is predominantly comprised of LB that store inflammatory precursors, alongside energy substrates that may also function as precursors for bioactive lipids [13, 24]. This expansion is, unsurprisingly, linked to an elevated potency in antigen mediated de novo synthesis and release of bioactive lipids such as LTC4 [13]. However, other consequences of steatosis in mast cells are less predictable, notably the induction of ER stress and remodeling and the marked suppression of degranulation responses ([13], WEG and HT, unpublished data).…”

“…This expansion is, unsurprisingly, linked to an elevated potency in antigen mediated de novo synthesis and release of bioactive lipids such as LTC4 [13]. However, other consequences of steatosis in mast cells are less predictable, notably the induction of ER stress and remodeling and the marked suppression of degranulation responses ([13], WEG and HT, unpublished data). Steatotic mast cells are phenotypically different than their normal counterparts, creating a need to more fully understand the impact of steatosis on mast cell signaling.…”