Role Of Nuclear Factor-κB On Airway Smooth Muscle Cells Phenotype Modulation And The Effect Of PDTC Intervention

Objective: Phenotype modulation of airway smooth muscle cell is a unique characteristic of airway remodeling, our preliminary in vitro study has found that NF-k B signaling pathway plays a crucial role in phenotypic modulation. On this basis, the current study aimed to explicit the effect of in vivo inhibition of NF-k B on airway smooth muscle phenotype modulation. Further, we will discuss the impact on airway remodeling, airway hyper-responsiveness and chronic inflammation, thus provide theoretical and experimental basis for the search for new asthma treatments. Methods: The models of acute and chronic asthma were established by intraperitoneal injection and repeated inhalation of ovalbumin. PDTC, a NF-k B inhibitor, was deliverd by intraperitoneal injection 2hr before OVA inhalation. Airway hyper-responsiveness was measured by using BUXCO WBP system. General histological changes were examined by HE staining tissue. Collagen deposition and airway smooth muscle mass were detected by Masson trichrome staining and IHC respectively. Collagen and airway smooth muscle area were quantified by morphologic analysis. Phenotypic marker of airway smooth muscle cells and COL1A1 m RNA Level were measured by RT-PCR. protein expression of phosphorylated P65 and α-SMA were detected by Western-Blot.The expression of inflammatory cytokines in serum protein was quantified by Ray Biotech ELISA array. α-SMA and PCNA immunofluorescence were performed to evaluate proliferating cells. Results:(1) Penh values were significantly increased in asthmatic mouse compared with normal control group, while PDTC intervention decreased Penh values in both acute and chronic model.(2) The smooth muscle area was significantly increased in chronic but not acute model, and was mitigated by administration of PDTC.(3) Airway collagen area was significantly increased in chronic but not acute model, and was mitigated by administration of PDTC.(4) Co-express of PCNA and α-SMA was not detected, while the number of PCNA+ cell was increased significantly in acute model compared to control group, mainly located around blood vessels and airways. PCNA+ cell were undetected in chronic asthma mice.(5) The decrease in m RNA level of α-smooth muscle actin, calponin, and SM-MHC was observed in chronic model, while only calponin was found to be lowered down in acute model. PDTC intervention partially reversed phenotypic modulation in chronic and acute asthma. Calponin decline in chronic model cannot be reversed PDTC. Western blot showed that after PDTC intervention phospho-P65 expression was significantly reduced, and α-SMA protein was increased.(6) In the acute model, G-CSF, IL-6, and IL-17 were significantly higher in OVA+vehicle mice compared with naive controls, while PDTC treatment significantly reduced G-CSF, IL-6 levels but not IL-17. In the chronic model, however, only IL-17 and TNF-α were significantly higher in OVA+vehicle mice and TNF-α was reduced in OVA+PDTC mice. Though IL-6 was higher in the chronic model and was reduced by PDTC treatment, the statistical analysis showed no significance. Conclusion:(1) The in vivo progression of ASMC phenotype modulation, accompany with cell function alteration, is a dynamic procedure with the contractile protein began to reduce in the acute phase, and later decrease to a much greater extent in the chronic phase.(2) Intervention of NF-k B pathway reversed the airway smooth muscle cell phenotype modulation to some extent.(3) Intervention of NF-k B signaling pathway alleviate the asthmatic airway remodeling, airway hyper responsiveness and chronic inflammation to some extent, therefore can serve as an intervention target for asthma therapy.