Detection Of The Relationship Between The Amplification Of TERC Gene In The Differential Cervical Lesion And The Expression Of TERT Protein

Objective:Cervical cancer, one of the the female reproductive organs malignant tumors, is harmful to women's lives and health. The majority of cervical cancer gradually developed from precancerous lesions-cervical intraepithelial neoplasia. Pathogenesis of cervical cancer is associated with many factors and phases, but it is still unclear enough. Studying the molecular mechanism from gene level to protein level contributes to explore the biomarker as diagnosis precancerous lesions, As well as expound the pathogenesis of cervical cancer.Telomeres are the termini of chromosomes. They are critical for maintaining chromosome integrity and stability. In normal cell, the telomeres are shortening with split. When telomeres become critically short, cell apoptosis occurs. Telomerase is a special reverse transcriptase which can extend telomere sequence. Activited telomerase make cell unlimited proliferation by maintaining the lenghth of telomere. It is bound up with the tumor occurrence and development. Telomerase consists of the telomerase reverse transcriptase (TERT), the telomerase RNA (TERC), and telomere-related proteins. In recent years, The study demonstrated that the amplifications of TERC gene is the frequent genetic changes in the process of Cervical squamous epithelium into cervical carcinoma. TERC gene detection in cervical liquid-based cytological specimens exfoliated contributes to diagnosis for low-grade squamous intraepithelial lesions and high-grade squamous intraepithelial lesions. However, It is unclear that the value of detection of TERC gene in cervical paraffin sections for grade cervical intraepithelial neoplasia diagnosis, as well as the relationship between TERC gene amplification andTERT protein expression is unclear. This issue will detect the amplification of TERC gene in normal cervical squamous epithelium, CIN, cervical squamous cell carcinoma to evaluate the clinical value of detection of TERC gene as a biomarker in paraffin sections by self-made tissue microarray and fluorescence in situ hybridization. Detect the expression of TERT protein by self-made tissue microarray and Immunohistochemistry to study the role of TERT protein expression for precancerous lesions diagnosis. Investigate the closely correlations between the amplification of TERC gene and the expression of TERT protein in cervical squamous cell carcinoma and CIN tissues, to analyze the effect of the amplification of TERC gene for expression of TERT protein, so as to provide an evidence for further revealing the mechanism of the amplification of TERC gene carcinogenicMethods:1 24 cases of normal cervical squamous epithelia,104 cases of CIN and 48 cases of cervical carcinoma were collected.2 Put 176 cases cervical tissues by manual tissue chip technique into a recipient paraffin block, then fusing, slicing, gaining and baking, and made into tissue chip.3 Fluorescence In situ hybridization (FISH) was used to detect TERC gene amplification in cervical carcinoma, CIN and normal cervical squamous epithelia.4 Immunohistochemistry SP method was used to examine the expression of TERT protein in cervical carcinoma, CIN and normal cervical squamous epithelia.5 Analysis the data use spss 16.0, significance was set at p<0.05.Results:1 The result of tissue chip4 tissue chips (7 X 7,6 X 5) were made. Dots of tissue chip lined up in order, uniformed in size and no shifting.3 tissue dots were absence,14 tissue no significant tissue structures. The significant tissue structures were watched in the other dots. The rate of complete tissue dots was 90.3%. The haematoxylin-eosin (HE) staining was uniformity and no dropping, moving and wrinkling.The immunohistochemical and FISH results show that the positive location was accurate and the background was clear. 2 The amplification of TERC gene in different cervical intraepitheliallesionsDetect the amplification of TERC gene in 23 cases of normal cervical,91 cases of CIN and 45 cases of cervical squamous cell carcinoma by fluorescence in situ hybridization. In normal cervical epithelia, the Amplification of TERC was absence, from CIN I 13.8%, CINâ…¡45.2%, CINâ…¢61.3% to SCC 84.4%. There was significant difference between normal cervical squamous epithelia, CIN I and CIN II, so was the CIN III and cervical invasive squamous cell carcinoma (P<0.05). There was no significant difference between normal cervical epithelia and CINâ… ,CINâ…¡and CINâ…¢(P>0.05).3 The expression of TERT protein in different cervical intraepithelial lesionsDetect the expression of TERT protein in 45 cases of cervical squamous cell carcinoma,91cases of CIN and 23 cases of normal cervical squamous epithelial tissue by immunohistochemical. There was hardly any expression in normal cervical squamous epithelium, the expressions of TERT was still have obvious stratification, the positive cells were mainly restricted to the middle or lower layer in low-grade CIN, and high-grade of CIN may be more than the 2/3 or the full thickness. The TERT protein positive rate:93.3%in cervical squamous cell carcinoma,71.0% in CINâ…¢,64.5% in CINâ…¡and 55.2% in CIN I. There was significant difference between the normal cervix and CIN I, so was the CINIII and cervical squamous cell carcinoma (P<0.05). There was no significant difference between CINâ… and CINâ…¡, CINâ…¡and CINâ…¢(P>0.05).4 The relation of the amplification of TERC and the expression of TERTproteinThere was a positive correlation between the amplification of TERC and the expression of TERT protein (r=0.345, P<0.05). Conclusions:1 Fluorescence in situ hybridization detect TERC gene, the rate of positive amplification in CINâ…¡, CINâ…¢and cervical cancinoma was significant higher than those in CIN I and normal cervical squamous epithelium There was significant clinical practical significance in antidiastole CINâ… and CINâ…¡.2 Immunohistochemical detect TERT protein, the rate of positive expression in CIN and cervical squamous cell carcinoma was significant higher than those in normal cervical squamous epithelium, TERT protein could be used as an biomarker to screening cervical precancerous lesions.3 The amplification of TERC gene was positive correlated with the expression of TERT protein,which indicates the amplification of TERC gene can precipitate occurrence and development in cervical squamous cancer by improving the expression of TERT protein.4 Mannual intensive tissue chips have the advantages of low cost, high efficience, which are more beneficial to improve the quality of experiment.