Effect Of Ageing Stromal Microenvironment On The Development Of Murine Colitis-associated Cancer

Background: Although the incidence of the major types of cancers increases exponentially with ageing(after age 50 years,middle-aged stage),the age-related patterns of cancer incidence are not straight forward.It is known that accumulated mutations with ageing,especially in epithelium stem cells,provide seeds for cancer formation.But except the accumulated mutations with ageing,the change of tissue microenvironment with ageing also plays important roles in cancer development.Even now,some researches have revealed that ageing immune system could impair the development of cancer,the relationship between ageing tissue stromal microenvironment and carcinogenesis of spontaneous cancer is poorly understood.Aims: Using young and ageing mice,we investigated how ageing process impair sporadic CRC and inflammation-driven CRC to provide some new insights on the relationship between ageing and CRC.Methods: Given the feasibility of experimental models in ageing body and the increased cancer incidence with ageing(after age 50 years,middle-aged stage),we chose young(~3 months)and ageing(~14 months)mice,to represent the young and ageing(~50 years)populations in humans,respectively.Firstly,through Coloscopy,HE Staining,Intestinal organoids culture and Gene expression profile,we assessed the differences on tissue structure and colon stem cells in homeostatic colon between young and ageing mice.Then we used young and ageing mice to induce AOM-induced sporadic CRC and AOM/DSS-induced colitis-associated cancer(CAC),which usually occurs in the distal part of the colon and resembles the location of CRC at the distal colon and rectum in human.Subsequently,through Coloscopy,Gene expression profile,Immunofluorescent staining,Immunohistochemistry,Flowcytometry,conditional lineage tracing and conditional deletion system,we dynamically monitored and analyzed how ageing impairs development of sporadic CRC and CAC.At last,we also collected clinical tissue samples of inflammatory bowel disease(IBD)and CRC patients to confirm the mechanism we found in mice.Results: We did not discover any obvious difference on colon length,tissue structure,proliferation,whole transcripts expression,number of colon stem cells and intestinal organoids between young and ageing mice in homeostasis.In AOM-induced sporadic CRC model,we observed that tumor initiation and development were almost synchronous in young and ageing mice.In both DSS-induced colitis model and the early phase of CAC,we found that young and ageing mice showed the same severity of tissue damage,but ageing mice exhibited the inhibition in weight recovery,organizational reconstruction,as well as re-epithelization during intestinal wound healing.As a result,compared to young mice,the initiation and formation of CAC were significantly inhibited in ageing mice.Altogether,these results indicate that ageing process significantly inhibits both intestinal wound healing and CAC initiation and formation.Mechanistically,we found both the ratio of total immune cells and the ratio of main subtypes of immune cells in the intestinal microenvironment after DSS-induced tissue damage were similar between young and ageing mice,but reduced fibroblasts in stromal microenvironment during intestinal wound healing were observed in ageing mice.Through conditional lineage tracing,an important source of fibroblasts transformed from intestinal smooth muscle cells(ISMCs,nearly intestinal epithelium) was identified orchestrating intestinal wound healing and CAC in young mice.However,we observed reduced ISMCs-derived fibroblasts during intestinal wound healing in ageing mice,accompanied with inhibited intestinal wound healing and inhibited CAC initiation.To further explore why ageing inhibits ISMCs-fibroblasts transformation during intestinal wound healing,we found that activation of YAP/TAZ in ISMCs is required for their transformation into fibroblasts.Compared to its rare expression in ISMCs under homeostatic conditions,activation of YAP/TAZ and nuclear location in ISMCs during intestinal wound healing were observed in young mice.At the same time,the activation of YAP in ISMCs was positively correlated with the number of fibroblasts around ISMCs.However,the failure of YAP/TAZ activation in ISMCs and reduced fibroblasts were observed in ageing mice.Our study further showed that the conditionally deletion of YAP/TAZ in ISMCs of young mice led to reduced fibroblasts in response to DSS-induced intestinal damage in the early phase of CAC,which in turn inhibited both intestinal wound healing and simultaneous CAC initiation.Besides,we also found that the failure of YAP/TAZ activation in ISMCs was accompanied with the reduced fibroblasts in ageing intestinal microenvironment in both DSS-induced acute murine colitis and TNBS-induced acute murine colitis.Finally,we found that the activation of YAP/TAZ in ISMCs also exist in clinical samples of IBD and CRC patients.Conclusions: Our work reveals that ageing intestinal stromal microenvironment could significantly inhibit intestinal wound healing and simultaneously inhibit the initiation of CAC.Mechanistically,an important source of fibroblasts,transformed from ISMCs,was identified during intestinal wound healing and CAC development only in young mice,but not in ageing mice.This transformation was depended on the activation of YAP/TAZ in ISMCs.Our novel findings may offer a new perspective to understand the mechanism of ageing process and cancer development in relation to tissue stromal microenvironment.