NGF Induces Changes In The Phenotype And Function Of Adrenal Medullary Chromaffin Cells In Mice With Allergic Rhinitis Through JAK1/STAT1, P38 And ERK Pathways

Background:Allergic rhinitis(AR),or allergic rhinitis,is currently a global health problem.According to the statistics of the World Health Organization(WHO),allergic diseases affect about 40% of the world’s population.With the increase of disease severity and the expansion of the allergy spectrum to unrelated allergens,their incidence rate continues to increase.AR is an inflammation of the upper respiratory tract.If AR is not properly treated,it may transform into other diseases such as bronchial asthma,chronic sinusitis,tympanitis,nasal polyposis,serous otitis media,allergic conjunctivitis,and so on.Bronchial asthma(BA)is a respiratory system disease caused by the crosstalk between innate and adaptive immunity;It is essentially a heterogeneous disease characterized by respiratory airway inflammation driven by immune cells such as lymphocytes,T cells,eosinophils,and cytokines such as interleukin,and is associated with bronchial hyperresponsiveness.Pathological characteristics of AR and BA are similar,with prominent manifestations of airway inflammation and hyperresponsiveness.Previous studies have shown a close relationship between AR and BA.In the absence of BA,AR patients may also have significant lower airway dysfunction,including histological,physiological,and biochemical changes that are very similar to BA.If both the upper and lower respiratory tracts are involved,AR usually precedes asthma,and it can be inferred that AR may be an independent risk factor for the development of BA.However,the pathogenesis of AR developing into BA has not yet been fully elucidated.Nerve growth factor(NGF)is originally discovered as a signaling molecule that participates in a variety of physiological processes,including neuronal survival,protection,differentiation,proliferation,neurotransmission,and axonal growth.Studies have shown that NGF was persistently and highly expressed in the nasal epithelium and submucosa of AR patients,and was mainly limited to inflammatory cells.The concentration of NGF in the serum of BA patients was significantly higher than the normal level.The above indicates that NGF may participate in the pathophysiological processes of allergic rhinitis and bronchial asthma.This study hypothesized that the significantly increased expression of NGF in AR mouse models can induce the transition of AMCC from an endocrine phenotype to a neuronal phenotype,leading to a decrease in AD secretion.Its pathogenesis may be related to the activation of JAK1-STAT1,p38,and ERK signaling pathways,ultimately leading to bronchial diastolic dysfunction,which leads to the development of BA.This hypothesis not only supplements and enriches the theoretical theory of "one airway,one disease",but also may provide unique insights for the early prevention and treatment of allergic rhinitis.Objective:To explore the molecular mechanism of insufficient adrenaline release from chromaffin cells in the adrenal medulla during the development of allergic rhinitis,and to clarify the role and pathogenesis of nerve growth factor(NGF)in inducing the transformation of allergic rhinitis into bronchial asthma.Methods:1.A mouse model of allergic rhinitis was established by referring to the literature.Male mice(22±2 g)were randomly divided into a control group and a NGF model group.The control group was sensitized and stimulated with normal saline;The NGF model group was sensitized and challenged with ovalbumin(OVA)in mice.Finally,adrenal glands were collected and the model of allergic rhinitis was identified.2.Primary isolated chromaffin cells from the adrenal medulla were identified using Giemsa staining and transmission electron microscopy(TEM),respectively.3.The ultrastructural changes of chromaffin cells in the adrenal medulla were observed by transmission electron microscopy after treatment with NGF and K252 a,respectively.Real-time quantitative polymerase chain reaction(RT-q PCR),Western blot(WB),and immunohistochemistry were used to detect the expression of synaptophysin(SYN).4.The STAT1 sh RNA interference lentivirus vector was constructed,and the transfection efficiency was verified by q PCR and Western blot.5.Cells were divided into the following groups: 1)control group;2)NGF stimulation group;3)NGF stimulation+K252a group;4)NGF stimulation+PD98059group;5)NGF stimulation+Sh-STAT1 group;6)NGF stimulation+Sh-STAT1 NC group.RT-q PCR and Western blot were used to detect the m RNA and protein expression of JAK/STAT1,P38 and ERK pathways.The concentration changes of epinephrine and norepinephrine in the supernatant of cell culture in each group were detected by enzyme-linked immunosorbent assay(ELISA).Results:1.Allergic rhinitis mice sensitized by OVA exhibit symptoms such as sneezing,runny nose,and scratching the nose.2.Sensitization of OVA in allergic rhinitis mice resulted in histopathological changes such as disordered arrangement of epithelial cells and cilia,inflammatory cells infiltration(lymphocytes and eosinophils)and interstitial swelling,compared with control mice.3.Compared with the control group,there were villous synaptic changes in the chromaffin cell membrane of the adrenal medulla,swelling of mitochondria,reduction of chromaffin particles in the cytoplasm,and significant upregulation of the expression level of SYN in the adrenal medulla after NGF intervention;After K252 a intervention,the degree of cell damage could be reduced,thereby the expression level of SYN was inhibited.4.Compared with the control group,NGF treatment could inhibit the expression level of adrenalin concentration in the supernatant and promoted the secretion of norepinephrine;Compared with the NGF model group,K252 a,PD98059,and STAT1 sh RNA treatment could promote the expression level of epinephrine concentration in the supernatant and inhibited the secretion of norepinephrine.5.Compared with the control group,NGF stimulation significantly upregulated the expression levels of JAK1,STAT1,p38,and ERK proteins and m RNA;However,K252 a,PD98059,and STAT1 sh RNA treatment significantly downregulated the expression of JAK1,STAT1,p38,and ERK proteins and m RNA,thereby affecting the JAK1/STAT1,p38,and ERK signaling pathways.Conclusion: NGF could regulate the phenotypic and functional changes of chromaffin cells in the adrenal medulla,and its mediated pathogenesis be involved in inducing allergic rhinitis to evolve into bronchial asthma by reducing the release of epinephrine through JAK1/STAT1,p38,and ERK signaling pathways.