| With changes in lifestyle and dietary structure,incidence of colorectal cancer has ranked third worldwide.Folate have a certain preventive effects in a variety of cancers including colorectal cancer,it could also inhibit mucosa cell proliferation in postoperative colorectal cancer patients.Folate receptor ?(FR?,as FOLR1 protein)encoded by FOLR1 gene,have a crucial role in folate intake decided by the affinity to folate and expression level of FR?.FOLR1 protein could specifically bind folate and transport it into cells.Folate transport into cells can through one-carbon metabolism involve in DNA synthesis,damage repair and methylation.Folate deficiency and/or metabolic abnormalities usually cause chromosome instability that including abnormal DNA structure and gene expression,then the risk of abnormal nervous system development,a variety of degenerative diseases and carcinogenesis would increase.To explore the effects of different status of folate deficiency on the expression of FOLR1 and the chromosome instability of different physiological conditions colonic cell,our study analysised the effect of expression of FOLR1 mRNA,protein and chromosome instability(CIN)in colonic normal and cancer cells after intervening with folic acid(FA)and 5-methyltetrahydrofolate(5-MeTHF).Concentration of folate in fixed RPMI-1640 medium(2266nmol/L)was as control in our works.Concentration of folate deficiency(22.66 nmol/L)and maintaining genomic stability(226.6 nmol/L)were as the experimental concentrations to intervene colorectal adenocarcinoma cells COLO205 and human normal colonic epithelium cells CCD-841-CoN(CCD)for 28 days.Extracted cells total RNA and total protein in intervention trials for 14 and 28 days.The expression of FOLR1 gene was detected by RT-PCR and 2-??CT analysis and the expression of FOLR1 protein was detected by Western Blotting at above two time points;the rate of CIN in two test cell lines were examined at 9th,18 th and 27 th day of the intervening culture by Cytokinesis-Block Micronucleus Cytome assay(CBMN-Cyt).Our results showed that:(1)The expression of FOLR1 protein in CCD cells was significantly lower than that in COLO205 cells(P <0.001)in natural condition.(2)In COLO205 cells,the trends in expression of FOLR1 mRNA and protein were consistently after FA and 5-MeTHF were intervened for 14 and 28 days,with concentration of FA and 5-MeTHF decreased expression of FOLR1 m RNA(P <0.001 ~ 0.01)and protein(P < 0.01 ~ 0.05)were significantly increased;besides,the CIN rate was significantly higher than that of the control(2266nmol/L,P < 0.01)in folate deficiency concentration(22.66 nmol/L).In CCD cells,the expression of FOLR1 mRNA(P < 0.001)and protein(P < 0.01 ~ 0.05)were significantly higher than those in the control at 14 th day,while the expression of FOLR1 m RNA(P <0.001)and protein(P < 0.01 ~ 0.05)was significantly lower than that in the control at28 th day;simultaneously,the CIN rate was significantly higher than that of the control(P < 0.01)in folate deficiency concentration.(3)The degree of effects in different status of folate on the transcription of FOLR1 in COLO205 and CCD cells was different: the expression of FOLR1 in5-MeTHF group was significantly higher than that in FA group at 22.66 nmol/L(P <0.01).The results suggest that folate deficiency could cause expression of FOLR1 mRNA and protein up-regulation in normal and colonic adenocarcinoma cells,and the rate of CIN is increase;however,in normal cells,sustained shortage of folate would lead to expression of FOLR1 m RNA and protein down-regulation in late period,which conjecture that folate deficiency may induced damage of chromosome and eventually cause growth arrest or apoptosis in cells.The study also indicated that folate deficiency could cause expression of FOLR1 increase,the effects of 5-MeTHF are stronger than FA in up-regulation of gene transcription. |
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