Roles Of Programmed Death Protein 1/Programmed Death-ligand 1 In Secondary Brain Injury After Intracerebral Hemorrhage In Rats: Selective Modulation Of Microglia Polarization To M2 Phenotype

Background: Recent reports have shown that microglial cells/macrophages are the main regulatory cells in the immune defense response of the central nervous system(CNS),which plays critical roles in intracerebral hemorrhage-induced secondary brain injury.Programmed death protein 1/programmed death-ligand 1 has been reported to regulate neuroimmune cell functions.Signal transducers and activators of transcription 1 participates in microglia polarization and programmed death protein 1/programmed death-ligand 1 could regulate the activation of signal transducers and activators of transcription 1.We herein show the critical role of programmed death protein 1/programmed death-ligand 1 in the polarization of microglia during intracerebral hemorrhage-induced secondary brain injury in rat models.Methods: An autologous blood intracerebral hemorrhage model was established in Sprague Dawley rats(weighing 250-300 g).First,to examine PD-1 and PD-Ls expression at different time points after ICH,rats were divided into 3 groups: normal control group(n = 6),sham group(n = 6),and ICH group(n = 36).Normal control group animals were not subjected to a procedure.Sham group animals underwent withdrawal artery blood.The ICH group was randomly divided into 6 groups: group A(6h),group B(12h),group C(24h),group D(48h),group E(72h),group F(1w),which were subjected artery blood.The brain tissue was got out from SD rats for detecting the expression of PD-1 and PD-Ls by Western blot and immunofluorescence.Then we chose the 24 h as the sample for the point of time.The samples in vivo or vitro were all randomly divided into 10 groups.The primary cultured microglial cells were exposed to oxyhemoglobin to mimic intracerebral hemorrhage in vitro.Specific siRNAs and pDNA for programmed death protein 1 and programmed death-ligand 1 were exploited both in vivo and in vitro.At last,brain tissue was tested using Western blot analysis,immunofluorescence analysis,TUNEL method,Fluoro-Jade B(FJB)staining.The microglial cells were detected by microglia polarization analysis with immunofluorescence analysis.Results: In the brain tissue around hematoma,the protein levels of programmed death protein 1 and programmed death-ligand 1 and the interaction between them,as well as the phosphorylation of signal transducers and activators of transcription 1 were higher than that of the sham group,and collectively peaked at 24 h after intracerebral hemorrhage.Overexpression of programmed death protein 1 and programmed death-ligand 1 ameliorated intracerebral hemorrhage-induced secondary brain injury,including brain cell death and neuronal degeneration,while their knockdown induced an opposite effect.In addition,overexpression of programmed death protein 1 and programmed death-ligand 1 selectively promoted microglia polarization to M2 phenotype after intracerebral hemorrhage and inhibited the phosphorylation of signal transducers and activators of transcription 1,suggesting that intracerebral hemorrhage-induced increases in programmed death protein 1 and programmed death-ligand 1 maybe a self-help.Conclusions: Enhancing the expressions of programmed death protein 1 and programmed death-ligand 1 may induce a selective modulation of microglia polarization to M2 phenotype for intracerebral hemorrhage treatment.