Pathogenesis Of Neutrophil Extracellular Traps Inducing Pulmonary Microvascular Endothelial-to-Mesenchymal Transition In Dermatomyositis-associated Interstitial Lung Disease

Background: Interstitial lung disease(ILD)is a frequent and severe manifestation of dermatomyositis(DM)associated with poor outcome.The previous research group found that abnormal regulation of neutrophil extracellular traps(NETs)may be involved in the pathogenesis of DM and could be one of the factors that initiate and aggravate ILD,TLR9-mi R-7-Smad2 signalling pathway is involved in the proliferation of pulmonary fibroblasts and their differentiation into myofibroblast.At the same time,the study found that NETs can also induce Endothelial-to-esenchymal Transition(Endo MT).These studies suggest that NETs induced endothelial cell damage in the DM-ILD,especially in anti-melanoma differentiation-associated gene 5(MDA5)-positive DM with rapidly progressive interstitial lung disease(RP-ILD).whether the specific mechanism remains unknown.Objective: To explore the role and possible mechanism of NETs in DM with ILD,to provide new insights into the occurrence of this disease,and to provide potential targets for the treatment of DM-ILD.Methods: The concentrations of plasma NETs markers,endothelial injury markers and anti-MDA5 antibody-positive patients were measured and correlation analysis was performed,the ability of plasma in different populations to induce NETs formation in vitro was observed,the extracted NETs were isolated in vitro to stimulate immortalized HPMECs(HPMEC-ST),CCK8 was used to detect cell viability,WB and immunofluorescence were used to detect Endo MT markers,transcriptome sequencing was used to detect differential genes,and WB and q PCR verification were performed with sequencing results.Finally,the experimental autoimmune myositis(EAM)model was used for pathological verification.Results: NETs were abnormally formed in DM with ILD patients with endothelial damage,anti-MDA5 antibody titers were positively correlated with v WF level and cf DNA concentration,and plasma of anti-MDA5 antibody positive patients could induce the formation of NETs in vitro.WB and immunofluorescence experiments proved that NETs could induce HPMEC-ST to produce Endo MT,and transcriptome sequencing showed that the expression of NLRP3,AIM2,IL-1β,TGF-βand other genes was significantly increased,which was consistent with the q-PCR and WB verification results.Infiltration of NETs and occurrence of Endo MT have also been found in EAM mouse models and in lung tissues of DM with ILD patients.Conclusion: NETs can promote the occurrence of Endo MT by activating NLRP3 inflammasomes in DM with ILD.Therefore,inhibiting NETs formation and blocking NETs from activating NLRP3 inflammasomes may be potential therapeutic targets for DM with ILD.