ABSTRACT
Inflammation and infection play an important role in the pathogenesis of many cancers.Toll-like receptors (TLRs) are a class of pattern recognition receptors that recognize conserved components of microbes and trigger the immune response against invading microorganisms.Toll-like receptor 9 (TLR9) recognizes non-methylated cytosine-phosphateguanosine (CpG) DNA sequences which are the surrogate for viral DNA.TLR9 may react to tumor development and progression during chronic inflammation that involves the tumor microenvironment.In order to study the role of TLR9 in cervical cancer,we analyzed the TLR9 expression in different types of HPV infection cervical cancer cells.Then we detected if CpG sequences influenced the TLR9 expression and the sensitivity to cisplatin (DDP) of these cervical cancer cells in vitro.The expression of TLR9 mRNA and protein in SiHa,Hela and C33A cells was detected by RT-PCR and Western blotting.Real-time PCR was used to examine the TLR9 expression changes induced by CpG.Chemosensitivity of the cervical cancer cells to cisplatin (DDP) was measured by MTT.It was observed that the expression of TLR9 mRNA and protein was increased gradually in SiHa (HPV16+),Hela (HPV18+) and C33A (HPV-) cells.Low doses of CpG increased the TLR9 expression only in C33A (HPV-) cells,but not in SiHa (HPV16+) and Hela (HPV18+) cells.Furthermore,low dose of CpG significantly increased the sensitivity ofC33A (HPV-) cells,but not that of SiHa (HPV16+) and Hela (HPV18+) cells.These results indicated that TLR9 may serve as a protective agent in HPV negative cervical cancer cells.It was concluded that TLR9 could improve the sensitivity to DDP in HPV negative cervical cancer cells and might represent a potential therapeutic option in clinical practice.
ABSTRACT
The mRNA and protein expression of thymidylate synthase(TS),thymidine phosphorylase(TP)and dihydropyrimidine dehydrogenase(DPD)and their relationship with prognosis were investigated.Real-time quantitative RT-PCR(Taqman)was used to detect the mRNA expression of TS,TP and DPD in formalin-fixed and paraffin-embedded 106 samples of epithelial ovarian cancer and 29 normal ovaries.A TATA box-binding protein(TBP)was used as an endogenous reference gene.A relationship between TS,TE DPD expression and clinicopathologic features was investigated.The protein location and expression of TS,TP and DPD was examined in the same patients by an avidin-biotin-peroxidase immunohistochemistry.TS and TP mRNA expression levels were significantly higher in tumor group than in normal controls,with the average value of TS and TP mRNA being 6.14±0.62 and 0.59±0.06 in tumor tissue,and 0.71±0.14 and 0.16±0.04 in normal tissue,respectively.DPD mRNA expression levels were significantly lower in tumor group(0.11±0.02)than in normal controls(0.38±0.05).There was statistically significant difference in TS and TP mRNA expression levels among different pathological grades and clinical stages(P<0.05),but histological subtype was not significantly associated with TS and TP mRNA expression.DPD gene expression was not significantly associated with any clinicopathological parameters.Immunohistochemistry revealed that TP protein was mainly distributed in nucleus,and TS and DPD mainly in cytoplasm.The protein expression intensity of TS,TP and DPD was coincided with the mRNA expression levels.It was concluded that TS,TP mRNA and protein expression levels were significantly higher in epithelial ovarian cancer,and DPD mRNA and protein expression levels were significantly lower.The expression levels of TS and DPD were related to the patients' prognosis and survival.Combined gene expression levels of TS,TP and DPD represent a new variable to predict the clinical outcome in ovarian cancer.The association of TS,TP and DPD expression levels with survival suggests an importance of these genes for tumor occurrence and progression.