RESUMEN

Type 1 diabetes (T1D) is a consequence of autoimmune destruction of ß-cells and macrophages (MΦ) have a central role in initiating processes that lead to ß-cell demise. We reported that Ca2+-independent phospholipase A2ß (iPLA2ß)-derived lipid (iDL) signaling contributes to ß-cell death. As MΦ express iPLA2ß, we assessed its role in T1D development. We find that selective reduction of myeloid-iPLA2ß in spontaneously diabetes-prone nonobese diabetic (NOD) mice (a) deceases proinflammatory eicosanoid production by MΦ, (b) favors anti-inflammatory (M2-like) MΦ phenotype, and (c) diminishes activated CD4+ and CD8+ T-cells phenotype in the pancreatic infiltrate, prior to T1D onset. These outcomes are associated with a significant reduction in T1D. Further, inhibition of select proinflammatory lipid signaling pathways reduces M1-like MΦ polarization and adoptive transfer of M2-like MΦ reduces NOD T1D incidence, suggesting a mechanism by which iDLs impact T1D development. These findings identify MΦ-iPLA2ß as a critical contributor to TID development and potential target to counter T1D onset.

RESUMEN

Leukocytes migrate through the blood and extravasate into organs to surveil the host for infection or cancer. Recently, we demonstrated that intravenous (IV) anti-CD45.2 antibody labeling allowed for precise tracking of leukocyte migration. However, the narrow labeling window can make this approach challenging for tracking rare migration events. Here, we show that altering antibody administration route and fluorophore can significantly extend the antibody active labeling time. We found that while both IV and intraperitoneal (IP) anti-CD45.2 antibody labeled circulating leukocytes after injection, they had different kinetic properties that impacted labeling time and intensity. Quantification of circulating antibody revealed that while unbound IV anti-CD45.2 antibody rapidly decreased, unbound IP anti-CD45.2 antibody increased over one hour. Using in vitro and in vivo serial dilution assays, we found that Alexa Fluor 647 (AF647) and Brilliant Blue 700 (BB700) dyes had the greatest labeling sensitivity compared to other fluorophores. However, IP antibody injection with anti-CD45.2 BB700, but not AF647, resulted in continuous blood leukocyte labeling for over 6 hours. Finally, we leveraged IP anti-CD45.2 BB700 antibody to track slower migrating leukocytes into tumors. We found that IP anti-CD45.2 antibody injection allowed for the identification of ~seven times as many tumor-specific CD8+ T cells that had recently migrated from blood into tumors. Our results demonstrate how different injection routes and fluorophores affect anti-CD45.2 antibody leukocyte labeling and highlight the utility of this approach for defining leukocyte migration in the context of homeostasis and cancer.

RESUMEN

The Ca2+-independent phospholipase A2ß (iPLA2ß) is a member of the PLA2 family that has been proposed to have roles in multiple biological processes including membrane remodeling, cell proliferation, bone formation, male fertility, cell death, and signaling. Such involvement has led to the identification of iPLA2ß activation in several diseases such as cancer, cardiovascular abnormalities, glaucoma, periodontitis, neurological disorders, diabetes, and other metabolic disorders. More recently, there has been heightened interest in the role that iPLA2ß plays in promoting inflammation. Recognizing the potential contribution of iPLA2ß in the development of autoimmune diseases, we review this issue in the context of an iPLA2ß link with macrophages and T-cells.

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RESUMEN

Phospholipases A2 (PLA2s) catalyze hydrolysis of the sn-2 substituent from glycerophospholipids to yield a free fatty acid (i.e., arachidonic acid), which can be metabolized to pro- or anti-inflammatory eicosanoids. Macrophages modulate inflammatory responses and are affected by Ca2+-independent phospholipase A2 (PLA2)ß (iPLA2ß). Here, we assessed the link between iPLA2ß-derived lipids (iDLs) and macrophage polarization. Macrophages from WT and KO (iPLA2ß-/-) mice were classically M1 pro-inflammatory phenotype activated or alternatively M2 anti-inflammatory phenotype activated, and eicosanoid production was determined by ultra-performance LC ESI-MS/MS. As a genotypic control, we performed similar analyses on macrophages from RIP.iPLA2ß.Tg mice with selective iPLA2ß overexpression in ß-cells. Compared with WT, generation of select pro-inflammatory prostaglandins (PGs) was lower in iPLA2ß-/- , and that of a specialized pro-resolving lipid mediator (SPM), resolvin D2, was higher; both changes are consistent with the M2 phenotype. Conversely, macrophages from RIP.iPLA2ß.Tg mice exhibited an opposite landscape, one associated with the M1 phenotype: namely, increased production of pro-inflammatory eicosanoids (6-keto PGF1α, PGE2, leukotriene B4) and decreased ability to generate resolvin D2. These changes were not linked with secretory PLA2 or cytosolic PLA2α or with leakage of the transgene. Thus, we report previously unidentified links between select iPLA2ß-derived eicosanoids, an SPM, and macrophage polarization. Importantly, our findings reveal for the first time that ß-cell iPLA2ß-derived signaling can predispose macrophage responses. These findings suggest that iDLs play critical roles in macrophage polarization, and we posit that they could be targeted therapeutically to counter inflammation-based disorders.

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RESUMEN

The Ca2+-independent phospholipases, designated as group VI iPLA2s, also referred to as PNPLAs due to their shared homology with patatin, include the ß, γ, δ, ε, ζ, and η forms of the enzyme. The iPLA2s are ubiquitously expressed, share a consensus GXSXG catalytic motif, and exhibit organelle/cell-specific localization. Among the iPLA2s, iPLA2ß has received wide attention as it is recognized to be involved in membrane remodeling, cell proliferation, cell death, and signal transduction. Ongoing studies implicate participation of iPLA2ß in a variety of disease processes including cancer, cardiovascular abnormalities, glaucoma, and peridonditis. This review will focus on iPLA2ß and its links to male fertility, neurological disorders, metabolic disorders, and inflammation.

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