Study Of The Mechanisms Involed In The Anti-inflammatory Function Of Interleukin-27

Part I Protective function of interleukin 27 in colitis associated Cancer via suppression of inflammatory cytokines in intestinal epithelial cellsThe relationship between inflammation and cancer is one of the key points in cancer immunology study.Previous studies indicated chronic inflammation accelerates the tumor formation.Colitis associated cancer(CAC)is a typical inflammation-related tumor,which results from the oncogenic mutation derived from chronic epithelium destruction and regeneration.Interluekin-27(IL-27)is composed by p28 and EBI3 and expressed by activated antigen-presenting cells.Its receptor is formed by WSX-1 and gp130 and expressed by various immune cells,epithelial cells and endothelial cells.Previous research implied IL-27 play both anti-or pro-inflammation role in inflammation associated diseases and regulated the pathogenesis progression.In colitis studies,the exact function of IL-27 is controversial,and so far the role of IL-27 in colitis-associated cancer is not unveiled.Herein,We utilized the AOM/DSS induced CAC murine model,to study the function of IL-27.We hope our study would provide new insight into the relationship between inflammation and cancer and raise more knowledge of cancer immunology.First we induced the AOM/DSS model on wildtype(WT)C57BL/6 mice,and separate the tumor tissue,adjacent tissue and healthy tissue from control mice.By Q-PCR and western blot analysis,we detected that the expression of IL-27 and its receptors both upregulated in tumor tissue,and IL-27 is mostly expressed by lamina propria cells.Then we use IL-27R(WSX-1)knock out mice(WSX-1 KO)and WT mice to induce AOM/DSS model.We found that the tumor load and tumor size both increased in WSX-1 KO mice.Inflammatory cytokines and cell proliferation are both upregulated in WSX-1 models.As we observed higher inflammation level in WSX-1 KO mice,to further explore the function of the results above,we analyzed the immune cell subunit in lamina propria and found myeloid-derived suppressor cells(MDSC)is accumulated in WSX-1 KO mice.And the chemokine receptor CXCR2 on MDSCs is not significantly changed compared to WT mice.Then we evaluated the cytokine expression in intestinal epithelial cells(IECs)and find CXCL1,the ligand of CXCR2 is significantly upregulated.Besides,inflammatory cytokines like IL-6,TNF-? and GM-CSF are also upregulated.These results indicated that the alteration of these cytokines might be the reason of MDSCs accumulation.The we proved that IECs-derived cytok:ine is crucial for MDSCs accumulation by in vitro experiments.After Blocking of IL-6,GM-CSF or CXCL1 in IECs supernatant,MDSCs accumulation is decreased.Then we evaluated the tight junction related protein in WT and WSX-1 mice by Q-PCR and immunofluorescence and found comparable expression.And LPS in serum is also comparable.These results indicated the variation of inflammatory cytokines is not the consequence of the alteration of PAMPSs quantity but the inflammation signaling in IECs itself.Hence,we use LPS or IL-27 stimulated the primary IECs in vitro.We found WSX-1 KO IECs expressed higher amount of inflammation cytokines;and IL-27 stimulation decreased inflammatory cytokines expression in WT IECs.At last we feed the mice with antibiotic cocktails to ablate the PAMPs by remove the colon microbiota,then AOM/DSS model is induced.We found the tumor load,MDSCs accumulation and cytokines expression is comparable between WT and WSX-1 KO antibiotic-treated AOM/DSS models.So the protective function of IL-27 is dependent on PAMPs induced inflammation signaling.To sum up,in our study,we find IL-27 protect the mice from CAC by decreasing the accumulation via suppression of PAMPs-induced inflammatory cytokines in IECs.We hope our studies would provide new evidence in tumor microenvironment regulation and shed new insights in colon cancer immunotherapy.Part II Deficiency of IL-27R(WSX-1)leads to increased obesity phenotypeObesity has become one of the most danger diseases worldwide.The metabolism disorders induced by obesity influence the health of people mostly in western countries.The function of adipose was once considered mainly to be energy storage,and nowadays the endocrine function of adipose has been paid more attention.Obesity is considered to be a chronic inflammation state,and the inflammatory cytokines in adipose tissue,such as IL-6,TNF-a and C-reactive protein(CRP),are significantly upregulated in the obesity individuals and leads to metabolism disease like insulin resistance.Recently Treg in visceral adipose tissue(VAT Treg)has been discovered.It plays a vital role to regulate the adipose metabolism and homeostasis via secreting IL-10 and TGF-?.In the homeostasis condition,VAT Treg accumlates in the adipose tissue and display higher ratio than that in other lymphoid tissue and non-lymphoid tissue.But in the obese mice,VAT Treg decrease sharply in the adipose tissue.Because we found that WSX-1 KO mice(older than 24 weeks)always gain more weight that their WT littermates,and previously the function of IL-27 in obesity is not studied yet.So we utilized normal fat diet(NFD)and high fat diet(HFD)mice to study the regulation function of IL-27.We discovered that WSX-1 KO mice gained more bodyweight and VAT weight both after NFD and HFD feeding.And VAT Treg and its transcription factors were downregulated in the adipose tissue,indicating the protective function of IL-27 in obesity via maintain the accumulation of VAT Treg.Furthermore,we also found that IL-27 elevated in the plasma of obese children,implied that IL-27 might be secreted in the obese condition and exerts the compensatory effects.Hence,in this part we found the enhancement of obese phenotype in WSX-1 KO mice,and discovered the anti-inflammatory function of IL-27 in the obesity model for the first time.We hope our research would provide a new perspective on obesity and metabolism disorders.