Preliminary Study The Influence Of Colorectal Cancer Associated Fibroblasts Irradiation Effects Interacting With Colorectal Cancer Cells

Objective To preliminary study the influence of colorectal cancer cells on the colorectal cancer associated fibroblasts(CCAFs) irradiation effects and the effects of irradiated CCAFs on the colorectal cancer cells.Method Using tissue culture, digestion the fibroblasts which climb out from the tissue, cultured to 3-4 generations, then CCAFs were identified by immunofluorescence,then co-cultured with the CCAFs and divided into four groups, followed by①non-co-culture 0Gy group ②co-culture 0Gy group ③ co-culture 4Gy group ④non-co-culture 4Gy group; after treatment each group, testing CCAFs’ growth, apoptosis,autophagy and senescence;Colorectal cancer CCL-224 cells were cultured using CCAFs/CCL-224 cocultrue conditioned medium(CCAFs group), irradiated CCAFs/CCL-224 coculture conditioned medium(RCCAFs group) and serum-free medium(blank control group). The colony formation, proliferation, growth and migration abilities of CCL-244 were compared.Result CCAFs and CNFs were all expressing fibroblast marker Vimentin;The CCAFs had specific immuno-stain with the α—SMA,while the CNFs was no; After irradiated the co-culture CCAFs with 4Gy Cobalt 60 irradiation, it’s growth was inhibited,apoptosis rate was significantly increased, the level of autophagy was increased and there were also occur the senescence response. However, these effects were more pronounced increased when irradiated the CCAFs alone;The colony-forming efficiency of RCCAFs group was(0.6589 ± 0.009493),higher than CCAFs group(0.6378 ± 0.01365)(P> 0.05) and control. The absorbance values(OD) of CCL-244 cells in MTT assay from the RCCAFs group was 0.3502 ±0.1278 on day three and 0.6571 ± 0.1351 on day five, whereas, the OD values in theCCAFs group were 0.3234 ± 0.00986 and 0.7116 ± 0.1259 respectively. The difference of OD value on day 3 between RCCAFs group and CCAFs group was not statistically significant(P> 0.05) but there was statistically significant difference on day 5(P = 0.007).Cell proliferation index(PI) in the RCCAFs group was 0.4203 ± 0.01308 and cell mobility rates in 48 h were( 39.33 ± 3.80)%. No statistically significant difference was observed between RCCAFs group and CCAFs group(P> 0.05). The colony-forming efficiency,OD value in MTT assay, proliferation index and migration abilities in the RCCAFs and CCAFs groups were significantly enhanced when compared to blank control.Conclusion CCAFs did not have the ability of strong endurance with the radiation,4Gy single dose irradiation could produce the effects of irradiation on the CCAFs obviously; it may have a relatively protective effect to the CCAFs radiation damage under the condition of existence of tumor cells;The tumor promoting effects of CCAFs on the proliferation and migration abilities of colorectal cancer CCL-224 cells were not influenced by 4Gy single dose irradiation.