The Role And Mechanism Of Neutrophils In The Pathogenesis Of Rosacea

Background:Rosacea is a common chronic inflammatory skin disease that mainly occurs on the convexities of central face.It has a high incidence and is most common in young and middle-aged women.There is a lack of effective treatment and it is prone to recurrence,seriously affecting patients’physical and mental health and quality of life.The pathogenesis of rosacea is still unclear,but the basic histopathological change is an increased infiltration of inflammatory cells(e.g.,neutrophils and CD4~+T lymphocytes)surrounding the pilosebaceous unit.Neutrophils display plasticity and heterogeneity with the ability to adapt their function in different disease microenvironment.To date,the specific role of neutrophils in the rosacea pathogenesis involved remains unknown.Objective:To explore the role and mechanism of neutrophils in the pathogenesis of rosacea,and to provide new directions and targets for the development of new therapeutic approaches.Methods:In this study,we first confirmed the infiltration of neutrophils in rosacea lesions by H&E staining and the levels of neutrophil-associated chemokines in the serum of rosacea patients by Bio-Plex protein microarray analysis.We then constructed LL37-induced rosacea-like dermatitis using two different neutropenia mouse models(Genista mice and mice treated with anti-Gr-1 neutralizing antibody)to observe and assess the severity of skin lesions,compare and analyze the histopathological manifestations,and detect the expression levels of rosacea-related genes at the lesions,so as to clarify the role of neutrophils in the pathogenesis of rosacea.Further immunohistochemistry and flow cytometry confirmed different polarization states of neutrohphils in rosacea patients and mouse model.Subsequently,we successfully induced the polarization of human and mouse N1 and N2 in vitro and explored their effect on rosacea pathogenesis by localized replenishment of N2neutrophils in mouse skin lesions.Next,we screened significantly differentially expressed genes by RNA-seq analysis of mouse skin lesions and in vitro induced N1 and N2 type neutrophils.The effect of N2neutrophils on the proliferation of CD4~+T lymphocytes was explored using cell co-culture.Finally,we explored the potential upstream regulatory mechanisms that may affect neutrophil polarization through in vitro sh RNA knockdown techniques.Results:(1)Infiltration of neutrophils in rosacea lesions is increased.(2)The neutrophil-associated chemokine CXCL8 is elevated in the serum of rosacea patients,and its level is positively correlated with disease severity.(3)LL37-induced rosacea-like dermatitis is significantly exaggerated in neutropenia mouse models,thus suggesting that neutrophils play a major anti-inflammatory role in rosacea.(4)Neutrophils polarize into different subtypes in the lesions of rosacea patients and mouse model.(5)Localized replenishment of N2-polarized neutrophils significantly ameliorates the rosacea-like erythema and inflammation in mice,and N2-polarized neutrophils exert anti-inflammatory effects in rosacea by regulating vascular factors and inhibiting the proliferation of CD4~+T cells.(6)Transcription factor Id3 may be one of potential upstream regulator mediating neutrophil polarizationConclusion:Neutrophils are capable of reprogramming into different polarization states in rosacea lesions,in which N2-polarized neutrophils exert anti-inflammatory effects in rosacea by regulating blood circulation-related factors and inhibiting the proliferation of CD4~+T cells.