RESUMEN
OBJECTIVE: To screen interleukin (IL)-1ß secretion-related membrane transporters by macrophage experiment in vitro and conventional knockout mice. METHODS: THP-1 cell line was differentiated to obtain human THP-1-derived macrophages, and the primary macrophages were obtained from human peripheral blood. FVB wild-type mice with the same sex and age were used as the controls of MRP1 knockout mice. The macrophages in abdominal cavity and bone marrow of mice were cultivated. The cells were treated with ABCC1/MRP1, ABCG2/BCRP, ABCB1/P-gp, OATP1B1, and MATE transporter inhibitors, then stimulated by lipopolysaccharide and adenosine triphosphate. The secretion level of IL-1ß was detected by ELISA, Western blot, and immunofluorescence. RESULTS: After inhibiting ABCC1/MRP1 transporter, the secretion of IL-1ß decreased significantly, while inhibition of the other 4 transporters had no effect. In animal experiment, the level of IL-1ß secreted by macrophages in bone marrow of MRP1 knockout mice was significantly lower than control group (P < 0.05). CONCLUSION: ABCC1/MRP1 transporter is a newly discovered IL-1ß secretion pathway, which is expected to become a new target for solving clinical problems such as cytokine release syndrome.
Asunto(s)
Regulación hacia Abajo , Interleucina-1beta , Macrófagos , Ratones Noqueados , Proteínas Asociadas a Resistencia a Múltiples Medicamentos , Animales , Humanos , Ratones , Interleucina-1beta/metabolismo , Lipopolisacáridos , Macrófagos/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Células THP-1RESUMEN
Cerebral ischemic preconditioning (CIP) has been shown to improve brain ischemic tolerance against subsequent lethal ischemia. Reactive astrocytes play important roles in cerebral ischemia-reperfusion. Recent studies have shown that reactive astrocytes can be polarized into neurotoxic A1 phenotype (C3d) and neuroprotective A2 phenotype (S100A10). However, their role in CIP remains unclear. Here, we focused on the role of N-myc downstream-regulated gene 2 (NDRG2) in regulating the transformation of A1/A2 astrocytes and promoting to brain ischemic tolerance induced by CIP. A Sprague Dawley rat model of middle cerebral artery occlusion/reperfusion (MCAO/R) was used. Rats were divided into the following six groups: (1) sham group; (2) CIP group: left middle cerebral artery was blocked for 10 min; (3) MCAO/R group: left middle cerebral artery was blocked for 90 min; (4) CIP + MCAO/R group: CIP was performed 72 h before MCAO/R; (5) AAV-NDRG2 + CIP + MCAO/R group: adeno-associated virus (AAV) carrying NDRG2 was administered 14 days before CIP + MCAO/R; (6) AAV-Ctrl + CIP + MCAO/R group: empty control group. The rats were subjected to neurological evaluation 24 h after the above treatments, and then were sacrificed for 2, 3, 5-triphenyltetraolium chloride staining, thionin staining, immunofluorescence and western blot analysis. In CIP + MCAO/R group, the neurological deficit scores decreased, infarct volume reduced, and neuronal density increased compared with MCAO/R group. Notably, CIP significantly increased S100A10 expression and the number of S100A10+/GFAP+ cells, and also increased NDRG2 expression. MCAO/R significantly decreased S100A10 expression and the number of S100A10+/GFAP+ cells yet increased C3d expression and the number of C3d+/GFAP+ cells and NDRG2 expression, and these trends were reversed by CIP + MCAO/R. Furthermore, over-expression of NDRG2 before CIP + MCAO/R, the C3d expression and the number of C3d+/GFAP+ cells increased, while S100A10 expression and the number of S100A10+/GFAP+ cells decreased. Meanwhile, over-expression of NDRG2 blocked the CIP-induced brain ischemic tolerance. Taken together, these results suggest that CIP exerts neuroprotective effects against ischemic injury by suppressing A1 astrocyte polarization and promoting A2 astrocyte polarization via inhibiting NDRG2 expression.