The Research On Destruction Of Paracellular Pathway In Rat Submandibular Glands Mediated By Inflammatory Response Post-Irradiation

Objective: To investigate the change of tight junction(TJ)proteins claudin-1 and claudin-3 in submandibular glands(SMGs)post-irradiation,and the relationship between the inflammatory response induced by radiation and the destruction of paracellular pathway.Methods: The rats were randomly divided into control group,1week,6weeks,and 12 weeks post-irradiation groups.The right SMGs of rats in each group was exposed to a single dose of 20 Gy radiation.At different time points,the resting saliva was measured.The pathological changes of SMGs were observed by HE staining,the positive expression of tumor necrosis factor alpha(TNF-α)was observed by IHC staining.The levels of TNF-α,interleukin(IL)-1β and IL-6 in SMGs were detected by ELISA,and the protein levels and m RNA expression of claudin-1 and claudin-3 in SMGs were detected by WB and RT-q PCR,respectively.The distribution of claudin-1 and claudin-3 in SMGs were observed by IF staining.In addition,the level of NF-κB protein in SMGs post-irradiation were detected by WB.Results: After SMGs was exposed to 20 Gy X-ray radiation,the resting salivation decreased gradually.At 1week post-irradiation,the pathological changes of the SMGs mainly were minor interstitial edema and different degrees of vacuolation.At 6weeks post-irradiation,the increased degree of interstitial edema and vacuolation were observed.At 12 weeks post-irradiation,the degree and number of vacuolation of glandular epithelial cells were further increased,and inflammatory cell infiltration was observed.However,the interstitial edema was somewhat reduced.In addition,the levels of TNF-α,IL-1β and IL-6 in SMGs increased gradually in each groups post-irradiation.The protein levels and m RNA expressions of claudin-1 and claudin-3 were down-regulated in a time-dependent manner,and the immunofluorescence intensity showed decreasing trends.However,the phosphorylation NF-κB protein level was increased by western blot.Conclusion: Radiation of rats SMGs induces increased levels of inflammatory cytokines including TNF-α,IL-1β and IL-6.The increased inflammatory response may result in time-dependent down-regulation of claudin-1 and claudin-3 expressions by influencing the NF-κB protein phosphorylation level,resulting in SMGs radiation-induced dysfunction.