ABSTRACT
Multivesicular bodies (MVB) are endocytic compartments that enclose intraluminal vesicles (ILVs) formed by inward budding from the limiting membrane of endosomes. In T lymphocytes, ILVs are secreted as Fas ligand-bearing, pro-apoptotic exosomes following T cell receptor (TCR)-induced fusion of MVB with the plasma membrane at the immune synapse (IS). In this study we show that protein kinase C δ (PKCδ), a novel PKC isotype activated by diacylglycerol (DAG), regulates TCR-controlled MVB polarization toward the IS and exosome secretion. Concomitantly, we demonstrate that PKCδ-interfered T lymphocytes are defective in activation-induced cell death. Using a DAG sensor based on the C1 DAG-binding domain of PKCδ and a GFP-PKCδ chimera, we reveal that T lymphocyte activation enhances DAG levels at the MVB endomembranes which mediates the association of PKCδ to MVB. Spatiotemporal reorganization of F-actin at the IS is inhibited in PKCδ-interfered T lymphocytes. Therefore, we propose PKCδ as a DAG effector that regulates the actin reorganization necessary for MVB traffic and exosome secretion.
Subject(s)
Actins/metabolism , Exosomes/metabolism , Multivesicular Bodies/metabolism , Protein Kinase C-delta/metabolism , T-Lymphocytes/immunology , Apoptosis/immunology , Cell Line, Tumor , Cell Membrane/metabolism , Humans , Jurkat Cells , Lymphocyte Activation/immunology , Protein Kinase C-delta/genetics , RNA Interference , RNA, Small Interfering/genetics , Receptors, Antigen, T-Cell/metabolismABSTRACT
BACKGROUND: Cardiovascular disease is the leading comorbidity in renal patients and has been related to impaired nitric oxide signaling. Estrogens exert protective effects on the vascular system. This study investigates the effects of biological sex and nitric oxide-independent soluble guanylate cyclase (sGC) stimulator BAY 41-8543 on aortic remodeling in experimental mild uremia. METHOD: Age-matched male and female Wistar rats were assigned for 18 weeks into sham-operated, subtotally nephrectomized (SNX), SNXâ+âBAY 41-8543 and SNXâ+âhydralazine. Analysis involved functional, histological, and molecular kidney and thoracic aorta parameters. RESULTS: SNX significantly increased SBP, which was comparably reduced to control levels by BAY 41-8543 and hydralazine. In SNX males, uremic aortic remodeling was characterized by marked media thickening and increased media-to-lumen ratio (Pâ<â0.01), vascular smooth muscle cell (VSMC) proliferation, macrophage infiltration, extracellular matrix turnover, decreased aortic elastin-to-collagen ratio (Pâ<â0.01) and endothelial nitric oxide-synthase (eNOS) mRNA expression (Pâ<â0.05). No significant alterations of aortic media-to-lumen ratio, VSMC proliferation, macrophage infiltration, matrix metalloproteinase-2, and eNOS mRNA expressions were seen in female uremic animals. BAY 41-8543 significantly ameliorated uremic aortic remodeling and stiffening involving reduced VSMC proliferation, collagen I-deposition, extracellular matrix turnover, and increased elastin content and eNOS mRNA expression. Hydralazine treatment did not substantially alter aortic remodeling. CONCLUSION: Experimental mild uremia leads to pronounced aortic hypertrophic remodeling and stiffening with sex-dependent alternations, and these are more severe in male rats. BAY 41-8543 ameliorates uremic aortic remodeling in a blood pressure-independent manner. The results suggest that sGC-stimulators may offer a novel treatment mode for pathological arterial wall remodeling in patients with impaired renal function.
