Establish Of An In Vitro Culture System Of Human Nasal Epithelial Stem/progenitor Cells And The Impact Of IL-18 To The Nasal Epithelial Barrier Functions

Purpose:Human nasal epithelial stem cells(h NESPCs)were cultured in vitro to study the effects of LPS on Tight junction(TJs)protein expression,epithelial remodeling and immune response of nasal epithelial cells.Method:1.Human nasal epithelial stem cells(h NESPCs)were constructed in vitro.In the future,stem cells,fibroblasts and blood cells were separated from nasal polyp mucosa and inferior turbinate mucosa by low temperature digestion with protease K and differential adhesion method,and h NESPCs with good activity and high purity were obtained.2.Human nasal epithelial cell line(HNEp C)was cultured,and the expression of IL-4,IL-5,IL-1B,IL-18,and IL-33 was determined after lipopolysaccharide LPS was added with100ug/ml.3.Six human nasal epithelial stem cell(h NESPCs)models were cultured in vitro at the airliquid interface and stimulated by LPS 100ug/ml to observe the changes of transmembrane resistance(TEER)and the expression of tight junction proteins Claudin-4,Occludin and ZO-1.Results:(1)Human nasal polyp mucosa and inferior turbinate mucosa could be extracted and cultured with good activity and high purity human nasal epithelial stem cells.(2)Human nasal epithelial stem/progenitor cells(h NESPCs)were co-cultured with mouse 3T3 fibroblast feeder cells,which could be expanded in vitro while maintaining cell dryness.(3)Self-modified growth medium and differentiation medium with reasonable composition and proportion can promote the proliferation and differentiation of human nasal epithelial stem cells.(4)HE staining of nasal polyps with chronic sinusitis showed epithelial cell defect,ciliary collapse,goblet cell metaplasia and other epithelial barrier dysfunction and abnormal tissue remodeling.(5)After LPS stimulation,the expressions of IL-1B,IL-18,IL-33 and TNF-γwere increased,while the expressions of IL-4 and IL-5 were decreased.Conclusion:Human nasal epithelial stem/progenitor cells(h NESPCs)with good activity and high purity could be isolated and purified by in vitro culture model construction.The primary cells could be amplified by co-culture with 3T3 feeder cells.The air-liquid interface culture was beneficial to stem cell differentiation,and the self-modified medium was beneficial to stem cell proliferation and differentiation.The establishment of human nasal epithelial stem/progenitor cells(h NESPCs)in vitro culture model is of great significance for the study of the pathogenesis of nasal polyps,the function of nasal mucosal epithelial barrier and tissue remodeling.Cell wall structure lipopolysaccharide can stimulate the secretion of inflammatory cytokines IL-1B,IL-18,IL-33 and IFN-γin nasal epithelial cell.