Targeting Of Integrin ?v Ameliorates Radiation-induced Pulmonary Fibrosis

Part ? Conditional knockout Itgav from myofibroblasts prevent radiation-induced pulmonary fibrosis in miceObjective Radiotherapy is a main approach for treating thoracic tumor that may induce pulmonary fibrosis.TGF-?1 is a key factor regulating pulmonary fibrosis.Studies have shown that integrin?v-mediated TGF-?1 activation plays an important role in liver fibrosis,and the role of integrin?v in radiation-induced pulmonary fibrosis has not been reported.The current study will explore the feasibility of using integrin?v as a target to interfere with radiation-induced pulmonary fibrosis.Methods Itgav^loxP/loxP;Pdgfrb-Cre mice were established to conditionally knockout Itgav in myofibroblasts.All mice received 16 Gy radiation or sham radiation.Degree of lung fibrosis was measured by modified Ashcroft scale and microCT.Protein expression of collagen and?-SMA in lung tissue were determined using Western blotting.Results Conditional knockout Itgav from myofibroblasts in mice significantly attenuated radiation-induced pulmonary fibrosis measured by modified Ashcroft scale?p<0.01?.Lung images were detected using microCT and hounsifield units were reduced in knockout mice compared with control group?p<0.001?.Protein expression of collagen and?-SMA in lung tissue was significantly reduced in transgentic mice compared with wildtypes.Conclusion The current study found that conditional knockout Itgav from myofibroblasts could significantly attenuate radiation-induced pulmonary fibrosis in micePart ? Integrin ?v promotes radiation-induced pulmonary fibrosis by activating TGF-?1 and its downstream pathwayObjective By establishing Itgavlox P/lox P;Pdgfrb-Cre conditional knockout mice,we found that integrin ?v expressed by myofibroblasts played key roles in the pathological process of radiation-induced pulmonary fibrosis.Studies have shown that integrin ?v is critical for the activation of TGF-?1.However,it is unclear whether integrin ?v promotes the occurrence of radiation-induced pulmonary fibrosis by activating TGF-?1.The current study will further explore the mechanism of integrin ?v in radiation-induced pulmonary fibrosis.Methods ELISA determined the plasma levels of TGF-?1 in knockout mice and wild type mice,and the difference in expression levels was compared.MRC-5 cells were irradiated with 8Gy,and the expression changes of integrin ?v,p-Smad2/3,collagen and ?-SMA were compared before and after irradiation.Using exogenous RGD peptide,exogenous latent or active TGF-?1,and gene knockdown technology,the activation of TGF-?1 by integrin ?v and its mechanism were clarified.Results Itgav knockout resulted in a decrease in secreted active TGF?1 in vivo,while it had no effect on the expression of latent TGF?1.Irradiation resulted an up-regulated expression of integrin ?v,phosphorylated Smad2/3,?-SMA and collagen in MRC-5.Exogenous RGD polypeptide treatment or Itgav knockdown in MRC-5 resulted in down-regulation of fibrosis marker protein expression.The addition of exogenous active TGF?1 reversed that reduction,while latent TGF?1 did not have the effect.Conclusion The current study found that integrin ?v activated latent TGF-?1 by interacting with the RGD sequence,thereby promoting the development of radiation-induced pulmonary fibrosis.Part ? Association between single nucleotide polymorphisms of integrin gene and radiation-induced lung injuryObjective Radiation therapy plays an important role in the treatment of lung cancer.As a radiation-induced normal tissue complication,radiation-induced lung injury is a major limiting factor in the dose of chest radiation.However,even if the dosimetric parameters of radiotherapy are the same,there is a large individual difference in the probability and extent of radiation-induced lung injury in patients,which means that genetic factors may play a role in the pathogenesis of radiation-induced lung injury.Based on the first two parts of the study,this section will further study the correlation between integrin gene single nucleotide polymorphisms and radiation-induced lung injury,and establish a predictive model of radiation-induced lung injury.Methods Mass Array system and Sanger sequencing techniques were used to determine integrin single nucleotide polymorphisms in lung cancer patients received thoracic radiotherapy.Cox regression survival analysis was used to study the correlation between integrin genotype and radiation-induced lung injury.The Kaplan-Meier curve was used to describe the relationship between the risk and time of radiation-induced lung injury in patients with different rs4665162 genotypes.Immunohistochemical analysis was used to compare the differences in integrin expression levels in patients with different genotypes.Based on the correlation between integrin genotype,dosimetric parameters and radiation-induced lung injury,a predictive model of radiation-induced lung injury was established,and the sensitivity and specificity of each model were compared using ROC curve.Results Cox regression survival analysis found that patients' age,IMRT,V20,MLD and rs4665162 genotype were associated with the risk of radiation-induced lung injury.The rs4665162 AG/GG genotype combined with dosimetric parameters MLD? 13.5Gy or V20? 24.1% and patients who did not receive IMRT had the highest risk of radiation-induced lung injury.Immunohistochemical staining revealed higher levels of integrin expression in lung tissue of patients with AG/GG genotype.Compared with the simple dose parameters to predict the risk of onset,the prediction model established by the combination of patient age,MLD,IMRT,and rs4665162 genotype has the largest area under the ROC curve,which has the best sensitivity and specificity.Conclusion The current study found that rs4665162 genotype was associated with the risk of radiation-induced lung injury,and the prediction model established by the combination of patient age,MLD,IMRT,rs4665162 genotype has good sensitivity and specificity.