BACKGROUND: Cervical cancer is listed as one of high-incidence endemic diseases in Xinjiang. Our study aimed to evaluate the expression of TLR9 in uterine cervical tissues of Uyghur women and examine associations with clinicopathological variables. We further characterized the direct effects of TLR9 upon the selective silencing of human papillomavirus (HPV) E6 and E7 oncoprotein expression in HPV 16-positive human cervical carcinoma cells treated with siRNA in vitro. MATERIALS AND METHODS: Immunohistochemistry was applied to evaluate TLR9 expression in 97 formalin-fixed paraffin-embedded cervical samples from Uyghur women; 32 diagnosed with cervical squamous cell carcinomas (CSCC) , 14 with low-grade cervical intraepithelial neoplasias (CINI), 10 medium-grade (CINII), 24 high-grade (CINIII), and 17 chronic cervicitis. BLOCK-iTTM U6 RNAi Entry Vector pENTRTM/U6-E6 and E7 was constructed and transfected the entry clone directly into the mammalian cell line 293FT. Then the HPV 16-positive SiHa human cervical carcinoma cell line was infected with RNAi recombinant lentivirus. RT-PCR and Western blotting were used to determine the expression of TLR9 in both SiHa and HPV 16 E6 and E7 silenced SiHa cells. RESULTS: Immunohistochemical staining showed that TLR9 expression was undetectable (88.2%) or weak (11.8%) in chronic cervicitis tissues. However, variable staining was observed in the basal layer of all normal endocervical glands. TLR9 expression, which was mainly observed as cytoplasmic staining, gradually increased in accordance with the histopathological grade in the following order: chronic cervicitis (2/17, 11.8%) <CINI (4/19, 28.6%) <CINII (3/10, 30.0%) <CINIII (12/24, 50.0%) <CSCC (17/32, 53.1%) (p<0.05), but not with tumor differentiation. RT-PCR and Western blotting showed that TLR9 expression was up-regulated in HPV16 E6 and E7 silenced SiHa cells at both mRNA and protein levels. CONCLUSIONS: TLR9 expression increases according to the histopathological grade of cervical pathological process. HPV E6 and E7 oncoprotein have negative effects on the expression and function of TLR9.
BACKGROUND:Cervical cancer is listed as one of high-incidence endemic diseases in Xinjiang. Our study aimed to evaluate the expression of TLR9 in uterine cervical tissues of Uyghur women and examine associations with clinicopathological variables. We further characterized the direct effects of TLR9 upon the selective silencing of human papillomavirus (HPV) E6 and E7 oncoprotein expression in HPV 16-positive human cervical carcinoma cells treated with siRNA in vitro. MATERIALS AND METHODS: Immunohistochemistry was applied to evaluate TLR9 expression in 97 formalin-fixed paraffin-embedded cervical samples from Uyghur women; 32 diagnosed with cervical squamous cell carcinomas (CSCC) , 14 with low-grade cervical intraepithelial neoplasias (CINI), 10 medium-grade (CINII), 24 high-grade (CINIII), and 17 chronic cervicitis. BLOCK-iTTM U6 RNAi Entry Vector pENTRTM/U6-E6 and E7 was constructed and transfected the entry clone directly into the mammalian cell line 293FT. Then the HPV 16-positive SiHa human cervical carcinoma cell line was infected with RNAi recombinant lentivirus. RT-PCR and Western blotting were used to determine the expression of TLR9 in both SiHa and HPV 16 E6 and E7 silenced SiHa cells. RESULTS: Immunohistochemical staining showed that TLR9 expression was undetectable (88.2%) or weak (11.8%) in chronic cervicitis tissues. However, variable staining was observed in the basal layer of all normal endocervical glands. TLR9 expression, which was mainly observed as cytoplasmic staining, gradually increased in accordance with the histopathological grade in the following order: chronic cervicitis (2/17, 11.8%) <CINI (4/19, 28.6%) <CINII (3/10, 30.0%) <CINIII (12/24, 50.0%) <CSCC (17/32, 53.1%) (p<0.05), but not with tumor differentiation. RT-PCR and Western blotting showed that TLR9 expression was up-regulated in HPV16 E6 and E7 silenced SiHa cells at both mRNA and protein levels. CONCLUSIONS:TLR9 expression increases according to the histopathological grade of cervical pathological process. HPV E6 and E7 oncoprotein have negative effects on the expression and function of TLR9.