Effects Of Ionizing Radiation On ICOS/ICOSL And Related Signaling Pathway In Immune Cells Of Mice

Study on the immune effects of ionizing radiation has been the subject ofRadiation Medicine. The mechanisms of radiation immune effects have established amore comprehensive,rigorous and systematic theory,which relates to the overallregulation,cell function,molecular expression,signal transduction,gene regulation andmany other fields as well.With the development of basic immunology,there are muchdeeper understandings about the immune cells of co-stimulatory molecules,of whichthe functions have been revealed continually and the roles in the mechanisms ofradiation immune effects remain to be thoroughly investigated.There have been a few reports about ICOS/ICOSL of T lymphocyte and antigenPresenting cell surface.In our study,a preliminary study on immune cell ICOS/ICOSL,CD40/CD40L and the related molecules NF-κB, PI3K and cotykine IL-21exposed toionizing radiation was carried out in different observation levels including protein andcells to explore the expression of time-and dose-effect changes,The experimentalresults will be how the immune effect of radiation mechanism to add newexperimental evidence.1The effects of ionizing radiation on ICOS/ICOSL of mice thymocytes andsplenocytesFloweytometry(FCM) was used to detect the expression changes of ICOS andICOSL of mice thymocytes after0.075Gy and2.0Gy X-ray. The results showedthat:0.075Gy X-rays caused down-regulation trend of ICOS on Thymocyte（sP<0.01）;but2.0Gy X-rays ICOS expression was incresing with the time. The similar changetrend appear in the ICOSL expression.The expression changes of ICOS and ICOSL of mice splenocytes after0.075Gy and2.0Gy X-ray. The results showed that: The expression of ICOS on mice increasedtransient after0.075Gy X-rays radiation in4h, then decreased until72h significantlybelow sham-irradiation group (P<0.01).2.0Gy X-ray irradiation cause the ICOSexpression significant increased compared with sham-irradiation group (P<0.05,P<0.01).Expression of ICOS/ICOSL double positive cells decresed after8h andreaehed its valley at72h(P<0.01). Then increased slightly, but it still below comparewith the control group;2.0Gy was not similared with after0.075Gy X-rays,ICOS/ICOSL expression appear to decrease in the trend.This analysis of the thymocytes in the low dose radiation exposure, the immunesystem through has a positive effect on the immune response of the co-stimulatorymolecule CD28/B7activated T cells, this time not express negative synergistic co-stimulatory molecule ICOS/ICOSL, causing the enhancement of immune functionand immune cells; high dose radiation effects not only the expression of CTLA-4,while ICOS/ICOSL expression the final result, immune suppression. Similar trendswere associated with thymus these results above, changes in spleen of each index islikely to depend on the corresponding indexes of thymocytes.2The effects of ionizing radiation on CD40/CD40L of mice splenocytesFCM was used to detect the expression changes of CD40and CD40L of micesplenocytes after0.075Gy and2.0Gy X-ray. The results showed that: Compared withsham-irradiation group，0.075Gy X-rays caused significant down-regulation of CD40on splenocytes(P<0.01).While2.0Gy X-rays post-irradiation caused the CD40expression on splenocytes up with the increasing doses, except16h compared withsham-irradiation group had significant difference (P<0.05, P<0.01);Compared withsham-irradiation group,the expression of CD40L on splenocytes was significantlyincreased after0.075Gy X-rays, and the expression significant decreased from4h to72h after2.0Gy X-rays radiation compared with sham-irradiation group(P<0.01).The data indicated that0.075Gy X-ray CD40expression was below,butCD40L expression was increased. The results showed that auxiliary function ofCD40/CD40L does not play a key role for LDR effect. 3The effect of NF-κB protein expression in thymocytes and splenocytes afterdifferent doses X-raysThe expressions of time-course changes of NF-κB protein of mice splenocytesand thymocytes were detected by Western Blot after low and high doses X-rays. Theresults showed that the thymocytes NF-κB protein level decreased after low doseX-rays, and the high dose of X radiation increased expression; and the similar changetrend in the splenocytes. The results showed that the expression level of thymocytesNF-κB protein increased after low dose radiation group,high dose radiation group ofprotein level decreased significantly; but the expression leve of splenocytes NF-κBprotein of dose-response change was opposite compare with thymocytes. The resultsshowed: low doses radiation cannot activate the immune cell nuclear factor NF-κB,while high doses radiation can activate NF-κB, which may relate to low and highdoses radiation induce expression of JAK-SOCS signaling pathway moleculesdifference. Eventually lead to low doses radiation to reduce the NF-κB activity,andhigh dose radiation enhanced NF-κB activity.4The effects of PI3K protein expression in thymocytes and splenocytes afterdifferent doses X-rays.The expressions of time-course changes of PI3K protein of mice splenocytes andthymocytes were detected by Western Blot after low and high doses X-rays. Theresults showed that the elevated levels of thymocytes PI3K protein after low doseradiation, and the high dose radiation decreased expression level. It is different withthe thymocytes that the high dose radiation on splenocytes had no obvious trend.Dose-effect experiments of PI3K protein expression in thymocytes and splenocytesafter irradiation show: both in low and high doses radiation for the expression changesare similar changes, increased in varying degrees.5The effects of IL-21in thymocytes and splenocytes after different dosesX-rays The expressions of time-course changes of IL-21of mice spleen and thymocyteswere detected by ELISA after low and high doses X-rays. Splenocytes andthymocytes after0.075Gy X-rays, IL-21secretion caused a down-regulation trend,but was increased in the2.0Gy X-rays. Dose-effect experiments in thymocytes andsplenocytes after irradiation show: Low dose radiation group IL-21secretion decline,high dose group increased secretion. It is similar that IL-21secretion withICOS/ICOSL expression.In short, it can be drawn from the in vivo experiments, low dose irradiation caninduce the costimulatory molecules CD40,ICOS and its ligand ICOSL expressiondecreased, then NF-κB, cytokine IL-21secretion decreased, but PI3K molecules inthe signal path expression increased; High-dose immune suppression may related toincreased of ICOS/ICOSL expression in immune activition period.