| Colorectal cancer is one of the most common human cancer. With the improvementof living standards and diet changes, the incidence of colorectal cancer in China isincreasing rapidly. The incidence of the disease in economically developed areas isclose to that of the Western developed countries (ranking the second of all themalignancies). The mortality of colorectal cancer ranks the fifth among all themalignancies in China and has been increasing year after year. The pathogenesis ofcolorectal cancer is a long and progressing period from normal tissue, polyps, adenomas,high-level neoplasia (CIN) lesions, and eventually the malignant tumor.Although radiotherapy is among the most effective treatment of malignant, surgeryand chemotherapy are two major therapeutics on colorectal cancer patients.Radiotherapy could only be applied before and after operation as adjuvant therapy inpatients with locally advanced tumor or recurrent tumor. Because at present, even bymedium-dose irradiation on abdomen and pelvic, extensive degeneration, necrosis andshedding of the small intestine mucosa usually lead to diarrhea and malabsorptiondigestion of nutrients and water which limited the effects and application ofradiotherapy on colorectal cancer.To resolve contradiction between the effectiveness of the therapy and susceptivityof small intestine by irradiation, a new method that could synergies and increase theeffects of radiotherapy and reduce radiation injury should be developed. In this study,the BABL/c mice subcutaneous tumor model was established to study the effects ofToll-like receptor2(TLR2) agonist peptidoglycan in immune regulation and radiationinjury repair. Part1：Effects of peptidoglycan on post-irradiation immuneregulation and damage repairPurpose: To study the immunomodulatory effects of PGN in the process oftumor radiotherapy and intestinal damage repair.Method：The model mice were established and divided into4groups, namelynon-treatment group (untreated), single administration group (PGN), a single irradiation(15Gy), irradiation plus administration group the (15Gy+PGN), n=30. Abdominalirradiation of15Gy was given7d after tumor cell inoculation.1d after irradiation, themice of15Gy+PGN group and the PGN group were injected i.p. PGN (1.5mg/kg).Survival ratio and tumor volume of10mice of each group were observed and recorded.Another20of each group were scarified at day1.25d,3.5d and9d to investigatepathological changes of intestinal tissues such as length of intestinal villi, number ofcrypt and Ki67cells.Results：(1) Compared with the15Gy group, the weight loss rate, the smallintestine pathological, villus height, crypt number and intestinal crypt proliferation ofthe15Gy+PGN group are significantly different, indicating a lesser degree of intestinaldamage.(2) The tumor proliferation of15Gy+PGN group was inhibited in comparisonwith15Gy group.Conclusion：(1) PGN can promote intestinal radiation damage repair, promote therecovery of bowel function, improve the quality of radiotherapy the survival of mice.(2)PGN synergizes radiation treatment of tumors, inhibit tumor growth and delay tumorrecurrence.Part2：Mechnisms of peptidoglycan on post-irradiation immuneregulation and damage repairPurpose: To explore the mechanism of peptidoglycan on immune regulation anddamage repair.Methods：Tumor and small intestine were taken3.5d after irradiation from eachmice tumor model group, we used liquid nitrogen grinding method to extract RNA and protein, then used real-time PCR to detect different gene expression in cell survivalrelated pathway PI3K/Akt，immunoblotting method to detect related protein expression,through these methods to find which genes are regulated by PGN. We also usedimmunofluorescence method to detect the infiltration of CD8+T cells, macrophages, Bcells, NK cells, and mast cells in different tumor matrix. After grinding tumor tissue toextract protein，we used enzyme-linked immunosorbent assay (ELISA) to detect theexpression of the three cytokines IL-21, IL-1α, TNF-α.Results：(1) In15Gy+PGN group，Akt3expression level in small intestinalvillus epithelial and endothelial cells were higher than in15Gy group; But there was nodifference in the tumor tissue between the two groups (2) In15Gy+PGN group，theinfiltration of CD8+T cells， NK cells, B cells, mast cells, macrophage were more andthe expression of cytokine TNF-α, IL-1α,，IL-21were higher than in15Gy group，withstatistical difference.Conclusion：(1) PGN can stimulate the expression of Akt3, to regulate intestinalvillus cells after irradiation withered thereby regulating villus epithelial and endothelialcell apoptosis and survival, promote small intestine radiation damage repair, improvebowel function, improve radiotherapy mice quality of life.(2) Immune escape of tumorcells after irradiation reduced，then peptidoglycan can promote the infiltration ofimmune cells in the tumor microenvironment and cytokine expression, killing tumorcells, inhibition of tumor growth and recurrence.In conclusion, the research found that TLR2agonists have a synergistic effect ofradiation therapy. It could promote normal intestinal damage repair, improve theefficacy of radiotherapy and reduce side effects. The mechanism of PGN in this processlies on stimulating the expression of Akt3and promoting the intestinal villi cell survivalafter irradiation and inhibit tumor growth and recurrence through promoting immunecell infiltration in the tumor stroma and cytokine secretion. |
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