Effect And Mechanism Of Dectin-1 In Secondary Brain Injury After Intracerebral Hemorrhage

Background:Intracerebral hemorrhage(ICH)is a severe subtype of stroke.Most patients have some residual neurological dysfunction.Secondary Brain Injury(SBI)after ICH is the main cause of neurological impairment,and neuroinflammation is the important mechanism of SBI after ICH.However,our current understanding of the regulatory mechanism of neuroinflammation after ICH is still incomplete.Dendritic cell-associated C-type lectin 1(Dectin-1)is involved in inflammatory responses in a variety of diseases.Although a recent study indicated that Dectin-1 regulated microglia M1/M2 polarization after ICH and participated in inflammatory response,the role and specific mechanism of Dectin-1 in SBI after ICH are still not fully understood.Objective:To investigate the effect and possible mechanism of Dectin-1 in secondary brain injury after intracerebral hemorrhage.Methods:A mouse model of collagenase intracranial hemorrhage was established in vivo.Western blot(WB)was used to observe the expression of Dectin-1 at different time points after ICH.The localization of Dectin-1 in microglia,astrocytes and neurons after ICH was determined by double immunofluorescence.Laminarin,an inhibitor of dectin-1,was intraperitoneally injected after ICH.Neurobehavioral tests were used to assess neurological impairment;Brain edema was assessed by brain water content;Microglia activation and neutrophil infiltration were assessed by Immunofluorescence(IF)and WB;Dectin-1 and its downstream factors were detected by WB,qRT-PCR and ELISA.In vitro experiments,LPS were used to stimulate BV2 microglia to establish ICH inflammatory model in vitro,and WB was used to assess the expression of Dectin-1 at different time points.Then Laminarin was used for intervention,and expression of dectin-1 and downstream factors was detected by WB.Results:The protein expression of Dectin-1 in the brain tissue surrounding hematoma after ICH in mice showed a trend of first increasing and then decreasing,significantly increasing at 12 h,reaching the peak at day 3 and decreasing at day 7.Dectin-1 was expressed in microglia but not in astrocytes and neurons.After stimulated by LPS,the expression of dectin-1 in BV2 microglia showed a trend of first increasing and then decreasing,significantly increasing at 12 h,peaking at 24h and beginning to decrease at 48 h.Intraperitoneal injection of dectin-1 inhibitor Laminarin(450mg/kg)after ICH effectively improved neurological deficits,and reduce brain edema and brain injury.Inhibition of Dectin-1 reduced microglia activation and neutrophil infiltration in brain tissues surrounding hematoma after ICH,and decreased the expression of Iba-1 and MPO.Inhibition of Dectin-1 reduced the expression of pyroptosis executor protein GSDMD-N,inflammatory cytokines IL-1βand IL-18,alleviating pyroptosis and neuroinflammation after ICH.On the mechanism,Dectin-1 signaling was activated after ICH in vivo.Inhibition of Dectin-1 inhibited NLRP3 inflammasome activation,as well as reduced IL-1β and IL-18 production.In vitro,LPS-stimulated dectin-1 signaling was activated in microglia.Inhibition of Dectin-1 down-regulated the expression of NLRP3 inflammasome components and reduced the production of IL-1β and GSDMD-N.Conclusion:Dectin-1 plays an important role in SBI after ICH in mice by regulating NLRP3 signaling pathway.Inhibition of Dectin-1 alleviates microglia pyroptosis and neuroinflammation by attenuating NLRP3,reducing SBI after ICH.