Detection Of The Relationship Between The Expression Of P16 Protein In The Differential Cervical Lesion And HPV16/18 Infection

Objective: Cervical cancer is one of the most common cancer and a second leading cause of deaths among women, Incidence was younger and constantly on the rise. There were about 500,000 new cases every year all over the world, secondly to the breast cancer. The majority of cervical cancer gradually developed from precancerous lesions-cervical intraepithelial neoplasia, Its occurrence results of a multi-factor and is closely related to the infection of HPV. Early diagnosis and treatment can effectively prevent progress and transformation of the disease. For some precancerous lesions, due to the observers subjective factors, it is difficult to make accurate diagnosis. Therefore, it is very important to find an aided diagnosis for cervical precancerous lesions.The domestic and foreign researches indicate that the disease is caused due to persistent infection of high-risk human papillomaviruses(HR-HPVs), most commonly by HPV types 16/18. The carcinogenic process of HPV has often accompanied with proto-oncogene mutation or antioncogene inactivation. p16 gene, also known as multiple tumor suppressor gene. It is found the first gene which directly play role in the cell cycle, inhibiting cell division, Previous studies confirmed that there exist abnormal expression of P16 protein, but it is uncertain that its expression is increased or decreased; In recently years, There are more research about P16 and cervical cancer, but the role and significance of P16 protein expression in CIN histopathologic diagnosis and classification has not been clear.This issue will detect the expression of P16 protein and the infection of HPV in normal cervical squamous epithelium, CIN, cervical squamous cell carcinoma through self-made tissue microarray, to study the role of the virus in cervical carcinogenesis, to observe the expression of P16 protein in different lesions and discuss the significance of P16 overexpress in cervical carcinogenesis. To investigate the closely correlations between human papillomavirus (HPV) and P16 in cervical squamous cell carcinoma and CIN tissues, and analyze the influention of HPV16/18. So as to find a more objective and reliable indicators for monitoring the progress of disease, as well as to provide an evidence for further revealing the mechanism of HPV carcinogenic and synergy factors .There is an important practical significance for early prevention of cervical cancer early detection, and early treatment .Method1 Collected 168 cases of cervical tissue samples, of which 44 cases of cervical squamous cell carcinoma, 96 cases of CIN, and 28 cases of normal cervix.2 By manual tissue chip technique, this is done by using a needle to biopsy a standard histologic section and placing the core into an array on a recipient paraffin block, then fusing, slicing, gaining and baking.3 Immunohistochemistry SP method was used to examine the expression of P16 in cervical carcinoma, CIN and normal cervical squamous epithelia.4 In situ hybridization (ISH) was used to detect HPV16/18 in cervical carcinoma, CIN and normal cervical squamous epithelia.5 Analysis the data use SPSS13.0, significance was set at P<0.05.Result:1 Preparation of tissue chipTissue chips (6×5, 7×5) were made. Dots of tissue chip lined up in order, uniformed in size and no shifting. 11 tissue dots were absence or no significant tissue structures. The significant tissue structures were watched in the other dots. The rate of complete tissue dots was more than 90%. The haematoxylin-eosin (HE) staining was uniformity and no dropping, moving and wrinkling.2 Expression of P16 in different cervical intraepithelial lesionsUsing immunohistochemical methods to detect the expression of P16 in 41 cases of cervical squamous cell carcinoma, 90 cases of CIN and 28 cases of normal cervical squamous epithelial tissue. There is no expression in normal cervical squamous epithelium, the expressions of P16 are still have obvious stratification, the positive cells are mainly restricted to the middle or lower layer in low-grade intraepithelial lesions,and high-grade of CIN may be more than the 2/3 or the full thickness. The P16 positive rate: 100%(41/41) in cervical squamous cell carcinoma, 96.67%(29/30) in CINⅢ, 96.30%(26/27) in CINⅡand 42.42%(14/33) in CINⅠ. There was significant difference between the normal cervix and CIN, so was the CINⅠand CINⅡ(P<0.05). There was no significant difference between CINⅡand CINⅢ, CINⅢand cervical squamous cell carcinoma(P>0.05). The intensity of positive expression in the normal cervix, CIN and cervical squamous cell carcinoma is significantly different(P<0.05), As the lesions increasing, the positive expression is mainly on over-expression(++~+++).3 Expression of HPV16/18 in differentcervical intraepithelial lesionsHPV-DNA16/18 in 41 cases of cervical squamous cell carcinoma, 90 cases of CIN(including 30 cases of CINⅢ, 27 Example CINⅡ, 33 Example CINⅠ)and 28 cases of normal cervical squamous epithelial tissues were detectd by In situ hybridization. There is no expression of HPV protein in normal cervical epithelia, the positive expression rates of HPV in CINⅠ, CINⅡ, CINⅢand cervical squamous cell carcinoma were 15.2%(5/33), 44.4%(12/27), 60%(18/30), 53.7%(22/41), respectively, The expression of HPV-DNA was significantly higher in cervical squamous cell carcinoma compared to normal controls, CINⅠ, CINⅡ, CINⅢand cervical squamous cell carcinoma. There was no significant difference between CINⅡand CINⅢ, CINⅢand cervical squamous cell carcinoma (P>0.05).4 Correlation of p16 and HPV16/18 in cervical lesions There was a positive correlation between expression of P16 and HPV16/18 infection (r=0.36, P<0.05).Conclusion:1 Immunohistochemical P16 protein, rate of positive expression in CIN and cervical squamous cell carcinoma is higher than normal cervical tissue, It can help to distinguish the normal cervical squamous intraepithelial and the cervical lesions.2 Immunohistochemical P16 protein, from the normal cervical squamous epithelium to cervical squamous cell carcinoma, As the lesions increasing, the positive expression is mainly on over-expression(++~+++). the positive degree can help to determine the extent of lesions.3 There is no expression in normal cervical squamous epithelium, the expressions of P16 are still have obvious stratification, the positive cells are mainly restricted to the middle or lower layer in low-grade intraepithelial lesions, and high-grade of CIN may be more than the 2/3 or the full thickness. It can help to determine the level of CIN. 4 In situ hybridization detect HPV-DNA16/18, positive rate of cervical lesions is higher than the normal cervix group, It confirmed that cervical squamous cell carcinoma is closely related with HPV16/18 infection.