Cervical Intraepithelial Neoplasia grade 2 biopsy: Do p16INK4a and Ki-67 biomarkers contribute to the decision to treat? A cross-sectional study

ABSTRACT BACKGROUND: Managing cervical intraepithelial neoplasia grade 2 (CIN2) is challenging, considering the CIN2 regression rate, perinatal risks associated with excisional procedures, and insufficient well-established risk factors to predict progression. OBJECTIVES: To determine the ability of p16INK4a and Ki-67 staining in biopsies diagnosed with CIN2 to identify patients with higher-grade lesions (CIN3 or carcinoma). DESIGN AND SETTING: Cross-sectional study conducted at a referral center for treating uterine cervical lesions. METHODS: In 79 women, we analyzed the correlation of p16INK4a and Ki-67 expression in CIN2 biopsies with the presence of a higher-grade lesions, as determined via histopathology in surgical specimens from treated women or via two colposcopies and two cytological tests during follow-up for untreated women with at least a 6-month interval. The expression of these two biomarkers was verified by at least two independent pathologists and quantified using digital algorithms. RESULTS: Thirteen (16.8%) women with CIN2 biopsy exhibited higher-grade lesions on the surgical excision specimen or during follow-up. p16INK4a expression positively and negatively predicted the presence of higher-grade lesions in 17.19% and 86.67% patients, respectively. Ki-67 expression positively and negatively predicted the presence of higher-grade lesions in 40% and 88.24% patients, respectively. CONCLUSIONS: Negative p16INK4a and Ki67 immunohistochemical staining can assure absence of a higher-grade lesion in more than 85% of patients with CIN2 biopsies and can be used to prevent overtreatment of these patients. Positive IHC staining for p16INK4a and Ki-67 did not predict CIN3 in patients with CIN2 biopsies.

Comparison of values of immunocytochemical P16/Ki-67 double staining, P16 INK4α single staining and high-risk human papillomavirus testing in screening of high-grade cervical lesions / 肿瘤研究与临床

Objective:To investigate the screening values of immunocytochemical P16/Ki-67 double staining, P16 INK4α single staining and high-risk human papillomavirus (HR-HPV) testing for high-grade cervical lesions. Methods:The clinical data of 622 patients who underwent cervical thin-layer liquid-based cytology (TCT) and HR-HPV testing in General Hospital of Taiyuan Iron and Steel (Group) Co., Ltd. from March 2019 to July 2021 were retrospectively analyzed. The remaining cytological specimens were detected by P16/Ki-67 double staining and P16 INK4α single staining. Among them, 334 patients with TCT results suggesting atypical squamous cell of undetermined significance (ASCUS) and above and HPV-positive underwent colposcopy pathological biopsy. Using pathological results as reference, the positive predictive value, sensitivity, specificity and accuracy of P16/Ki-67 double staining, P16 INK4α single staining and HR-HPV testing for screening of high-grade squamous intraepithelial neoplasia (HSIL) and cervical cancer were compared. Results:Taking the results of histopathology as references, combined with the results of TCT, 31 of 622 patients were HSIL, of which 22 (71.0%) were positive for P16/Ki-67 double staining, 23 (74.2%) were positive for P16 INK4α single staining, and 25 (80.6%) were positive for HR-HPV testing; 4 cases were cervical cancer, and the positive rates of the three detection methods were all 100.0% (4/4). Among 622 patients, the positive rates of P16/Ki-67 double staining, P16 INK4α single staining and HR-HPV testing for screening of HSIL and cervical cancer were 13.99% (87/622), 25.40% (158/622) and 21.38% (133/622); the positive predictive values were 29.89%, 17.09% and 21.08%; the accuracies were 91.19%, 78.94% and 83.28%; the specificities were 89.77%, 77.98% and 82.46%; the sensitivities were 74.29%, 77.14% and 82.86%. The positive rate, positive predictive value, specificity and accuracy of P16/Ki-67 double staining were higher than those of P16 INK4α single staining and HR-HPV testing, and the differences were statistically significant ( z values were -5.062 and -3.418, 2.328 and 2.450, 5.436 and 3.570, 6.043 and 4.161, all P < 0.05); the sensitivity of HR-HPV testing was higher than that of P16/Ki-67 double staining and P16 INK4α single staining, but the differences were not statistically significant ( z values were -0.890 and 1.017, both P > 0.05). Conclusions:HR-HPV testing is more suitable for primary cervical lesion screening; P16/Ki-67 double staining can be used as a potential combined cell screening tool or an effective triage tool; P16 INK4α single staining has certain limitations.

Evaluation of the P16INK4a Expression in Cervical Biopsy Specimen SBMCH

In early 1980 human papillomavirus (HPV) were the risk factor and most commonly affects younger women. Many test have developed since then and among that a biomarker test system have developed and clinically evaluated. P16INK4a is used as an important marker for indicating neoplastic transformation for cervical dyplasia. This study was done to evaluate the P16INK4a expression in cervical biopsy in 50 cases. Two cases were identified a P16INK4a positive and remaining 48 didn’t show P16INK4a expression proving the hypothesis that p16INK4a is capable of showing the dysplasia positive cases

Guilu Erxian Glue () Inhibits Chemotherapy-Induced Bone Marrow Hematopoietic Stem Cell Senescence in Mice May via p16 / 中国结合医学杂志

OBJECTIVE@#To evaluate the effect of Guilu Erxian Glue (, GEG) on cyclophosphamide (CTX)-induced bone marrow hematopoietic stem cells (HSCs) senescence in mice and explore the underlying mechanism.@*METHODS@#The H@*RESULTS@#Compared with the model group, GEG increased cell viability as well as proliferation (P<0.05 or P<0.01) and reduced β -gal expression. Furthermore, GEG significantly decreased the expressions of p16@*CONCLUSION@#GEG can alleviate CTX-induced HSCs senescence in mice, and the p16

Hypermethylation Analysis of p16INK4a and p15INK4b Promoters in Chronic Lymphocytic Leukaemia Patients and Normal Individuals

@#Introduction: Chronic Lymphocytic Leukaemia (CLL) is a common type of leukaemia in persons of predominantly European descent but is rare in the Asian population. Disparities in CLL incidence among people of Asian and European descent may be related to the genetic make-up of the two different populations. Hypermethylation event might be one of the silencing mechanisms that inactivate the tumour suppressor genes in CLL. The aim of this study was to determine the hypermethylation status of p16INK4a and p15INK4b among CLL patients and normal individuals. Materials & Methods: A total of 25 CLL patients and 25 normal individuals were recruited for this study and their genomic DNA were extracted from the peripheral blood. The hypermethylation status of p16INK4a and p15INK4b were determined using Methylation Specific-PCR (MS-PCR) whereas DNA sequencing method was applied to selected samples for validation of the MS-PCR results. We also evaluated the association between hypermethylation of these genes with the clinical and demographic characteristics of each group of subjects. Results: Among the CLL patients, p15INK4b partialmethylation occurred in 6 (24%) subjects while methylation occurred in 1 (4%) subject. All the remaining patients were unmethylated at p15INK4b. All the samples showed unmethylation at p16INK4a. Statistically significant associations were found between p15INK4b hypermethylation with the presence of CLL (p=0.01) and with race (p=0.02). Conclusion: Further study using a larger sample size is warranted to explore the significance of DNA methylation incidence among the CLL patients of the Malaysian population. Hence, we suggest that hypermethylation at p15INK4b has a huge influence that kick-starts CLL disease among Malaysians and MS-PCR technique is applicable to be used in methylation study.

Expression of Bcl2 inhibitor of transcription 1 and E-cadherin in different regions of cervical squamous cell carcinoma and their relationship with expression of P16INK4a / 第二军医大学学报

Objective: To determine the expression of anoikis factor Bcl2 inhibitor of transcription 1 (Bit1), epithelial-mesenchymal transformation (EMT) marker E-cadherin and P16INK4a in tumor budding and central tumor of cervical squamous cell carcinoma, and to explore the significance of Bit1 and E-cadherin expression in the process of obtaining high invasiveness of cervical cancer and their relationship with P16INK4a expression. Methods: A total of 77 paraffin-embedded specimens of cervical squamous cell carcinoma were collected from the Department of Pathology of Gansu Provincial Cancer Hospital between 2014 and 2018. The expression levels of Bit1, E-cadherin and P16INK4a in tumor budding and central tumor of these specimens were detected by immunohistochemistry. Taking the median scores of protein expression in the central tumor and tumor budding as dividing points, the specimens were divided into high expression group and low expression group. The differences of Bit1 and E-cadherin expression under different p16INK4a expression and their relationship with the clinicopathological characteristics of the patients were analyzed. The correlation between Bit1 and E-cadherin expression in central tumor and tumor budding was explored. The χ2 test, continuous correction χ2 test and Spearman rank correlation analysis were used for statistical analysis. Results: In 77 cases of paraffin-embedded specimens of cervical squamous cell carcinoma, the high expression rates of P16INK4a, E-cadherin and Bit1 in central tumor and tumor budding were 32.5% (25/77), 67.5% (52/77) and 63.6% (49/77), and 67.5% (52/77), 33.8% (26/77) and 37.7% (29/77), respectively, and the differences were significant (χ2 18.935, 17.561 and 10.391, all P < 0.01). Both in central tumor and in tumor budding, there were no significant differences in Bit1 or E-cadherin expression between high and low P16INK4a expression regions (all P < 0.05). In central tumor, the low expression of Bit-1 was related to lymphovascular invasion and lymph node metastasis (χ2 5.053 and 4.400, both P < 0.05). In tumor budding, the low expression levels of E-cadherin and Bit-1 were both associated with lymph node metastasis (χ2 5.580 and 7.573, both P < 0.05). Spearman rank correlation analysis showed that there was positive correlation between E-cadherin and Bit1 expression in central tumor and tumor budding (r 0.287, P = 0.011; r 0.236, P < 0.039). Conclusion: The increased invasiveness of cervical cancer may be related to the decreased expression of Bit1 and E-cadherin and the increased expression of P16INK4a. Cervical cancer cells may acquire high invasiveness by inhibiting Bit1 to obtain anoikis resistance and affecting the EMT, but P16INK4a is not involved in this process.

Methylation of p16 ink4a promoter is independent of human papillomavirus DNA physical state: a comparison between cervical pre-neoplastic and neoplastic samples

BACKGROUND Epigenetic modifications in host cells, like p16 ink4a methylation, have been considered as putative complementary mechanisms for cancer development. Because only a small proportion of infected women develop cervical cancer, other factors might be involved in carcinogenesis, either independently or in association with high-risk human papillomavirus (HR-HPV) infections, including epigenetic factors. OBJECTIVES We hypothesised that p16 ink4a methylation might have a role in cancer development driven by HPV16, mainly in the presence of intact E1/E2 genes. Thus, our objectives were to assess the status of p16 ink4a methylation and the HPV16 E1/E2 integrity in samples in different stages of cervical diseases. METHODS Presence of HPV16 was determined by E6 type-specific polymerase chain reaction (PCR). Methylation status of the p16 ink4a promoter was assessed by methylation-specific PCR in 87 cervical specimens comprising 29 low-grade (LSIL), 41 high-grade (HSIL) lesions, and 17 cervical cancers (CC). Characterisation of E1 and E2 disruption (as an indirect indicator of the presence of episomal viral DNA) was performed by PCR amplifications. FINDINGS We observed a significantly increased trend (nptrend = 0.0320) in the proportion of methylated p16 ink4a in cervical samples during cancer development. Concomitant E1 and E2 disruptions were the most frequent pattern found in all groups: CC (76%), HSIL (54%), and LSIL (73%). No statistically significant differences between p16 ink4a methylation and E1/E2 integrity, in histological groups, was observed. MAIN CONCLUSIONS There was an increase in methylation of the p16 ink4a promoter from pre-neoplastic lesions to cancer. Additionally, a high frequency of E1/E2 disruptions in LSIL/HSIL suggested that viral DNA integration was an early event in cervical disease. Moreover, the methylation status was apparently independent of HPV16 integrity.

Mutations of Pl6INK4a/CDK4/CCND1 genes in 134 Chinese melanoma patients and their clinical significance / 基础医学与临床

Objective To investigate the frequency of P16INK4a, CDK4 and CCND1 gene mutations in Chinese melanoma patients and to find out the potential clinical significance. Methods The samples in this study were collected from 134 melanoma patients(37 acral melanomas, 87 mucosal melanomas, 10 nonacral skin melanomas), hospitalized in Beijing Cancer Hospital from January 2010 to December 2014.The mutation status of P1 6INK4a, CDK4 and CCNDl was detected by PCR amplification and Sanger sequence. Statistical analyses were used to investigate the correlation between gene mutation and prognosis. Results Among 134 samples, the mutation frequency of P16INK4a, CDK4, CCNDl was 8.2％(11/134), 0.75％(1/134), 0％(0/134) respectively.81.8％ (9/11) of the P16INK4a gene mutation may affect protein function.The median survival time of melanoma patients with P16INK4a mutations was significantly shorter than the patients without Pl6INK4a mutations (X2 = 8.872, P＜0.01).P16INK4a gene mutation was an independent prognostic factor for melanoma (P＜0.05). Conclusions P16INK4a may be a breaking point of targeted therapy for melanoma.

Expression and significance of MCM5 and p16INK4A in cervical squamous cell carcinoma / 临床与实验病理学杂志

Purpose To investigate the expression and significance of MCM5 and p16INK4A in cervical carcinoma and cervical intraepithelial neoplasia (CIN 1,CIN 2 ～ 3) with different degrees of HPV 16 infection.Method RT-PCR and immunohistochemistryof SABC method were used to detect the expression of mRNA and protein in HPV 16 infected normal cervix,CIN 1 tissue,CIN 2 ～3 tissue and cervical squamous cell carcinoma and analyzed the clinical significance.Result The expression of p16INK4A and MCM5 mRNA in cervical cancer tissues were significantly higher than that in normal tissues (x2 =-6.589,P ＜0.001,x2 =-4.349,P ＜0.001).The degree of cervical lesions increased gradually (x2 =57.141,P ＜ 0.001,x2 =47.628,P ＜0.01).Expression of mRNA MCM5 was correlated with pathological grade and clinical stage.Ⅰ a-Ⅰ b period was significantly lower than that of Ⅱ a-Ⅱ b (x2 =-4.93,P ＜0.01),the expression decreased with the decrease of pathological grade (x2 =-4.017,P ＜0.01).The expression of p16INK4A mRNA in cervical cancer decreased with the decrease of pathological grade (x2 =8.560,P ＜ 0.01).The most obvious expression of p16INK4A and p16INK4A mRNA in squamous cell carcinoma.Expression of MCM5 protein and p16INK4A protein in cervical squamous cell carcinoma was positively correlated (r =0.497).Conclusion There is high expression of MCM5 and p16INK4A in cervical carcinoma.MCM5 can be a better reaction of cervical malignant hyperplasia,and p16INK4A joint detection for the improvement of CIN classification and prognosis of the significance of the judgment.

Expression of polycomb protein SUZ12 predicts poor prognosis and response of patients with intrahepatic cholangiocarcinoma to adjuvant gemcitabine-based chemotherapy / 实用肿瘤学杂志

Objective This study explored the expression of polyclonal protein SUZ12 in patients with intrahepatic cholangiocarcinoma(ICC),and its role in predicting the survival and treatment of ICC patients.Methods The expression of SUZ12 and p16INK4a was detected by immunohistochemical assay in 207 liver tissue samples including ICC patients,BilIN-1,-2,-3 and non-tumor-like cholangiocarcinoma.The expression of these proteins was assessed to be related to the pathological characteristics of the ICC patients receiving chemotherapy and the outcome of survival as well as the subsequent chemotherapy response.Results The expression level of SUZ12 was gradually increased from non-neoplastic bile duct tissue to BilIN-1,-2,-3 and ICC.The expression of p16INK4a protein was expressed in non-neoplastic-like cholangiocarcinoma,but it decreased gradually in BilIN-1,-2,-3 and ICC tissues.SUZ12 expression was associated with undifferentiated ICC,lymph node metastasis and advanced cancer.Kaplan-Meier curve analysis showed that ICC patients with high expression of SUZ12 had a significant reduction in overall survival and disease-free survival in comparison with ICC patients with the low expression of SUZ12.SUZ12 expression was significantly associated with overall survival of patients receiving adjuvant gemcitabine-based chemotherapy(AGC).Conclusion SUZ12 expression is able to predict the overall survival and disease-free survival of ICC patients with adjuvant gemcitabine-based chemotherapy.

Detection p16INK4a protein in cervical tissues with time-resolved fluoroimmunoassay / 临床与实验病理学杂志

Purpose To establish a new method for detecting p16INK4a in cervical tissues with time-resolved fluoroimmunoassay (TRFIA).Methods 126 cases of paraffin imbedding tissues of cervix were selected for immunohistochemistry (IHC) of EnVision two-step and TRFIA.Results There were 20 cases of no intraepithelial lesion or malignancy,24 cases of low-grade squamous intraepithelial lesion (LSIL),53 cases of high-grade squamous intraepithelial lesion (HSIL) and 29 cases of squamous cell carcinoma (SCC).In the groups of no intraepithelial lesion or malignancy,LSIL,HSIL and SCC,p161NK4a positive was seen in 1,19,53 and 28,respectively.TRFIA test results displayed p16INK4a positive in 3,17,50 and 27 cases,respectively.Positive of p16 using by TRFIA in no intraepithelial lesion or malignancy,LSIL,above HSIL was 15.00％,70.83％ and 93.90％,respectively (P ＜ 0.01).Conclusion TRFIA is suitable for detecting of p16INK4a protein and demand low detection equipment,p16INK4a expression detected by TRFIA may helpful for large scale detection in various clinical institution.

Expression of ASPP2 and P16INK4a and its relationship with apoptosis in esophageal carcinoma / 实用医学杂志

Objective To study the expression of ASPP2 and P16INK4a in esophageal carcinoma and their relationship to the apoptosis and related clinicopathological characteristics. Methods Immunohistochemistry S-P method was used to examine the expression of ASPP2 and P16INK4a in the pathological specimens of 112 esophageal carcinoma cases and 31 cases of normal esophageal mucosa. TUNEL was also employed to detect the rate of apopto-sis in 37 esophageal carcinoma cases and 12 cases of normal esophageal mucosa. Results The difference between the expression of ASPP2 and P16INK4a in esophageal carcinoma and normal esophageal mucosa was statistically signif-icant(P < 0.05). Their abnormal expressions were all related to lymph node metastasis and differentiation degree (P < 0.05). The difference between the positive rate of apoptosis in esophageal carcinoma and normal esophageal mucosa was statistically significant(P < 0.05). The abnormal expression of ASPP2 and P16INK4a was all related to apoptosis in esophageal carcinoma. Conclusions The different expression of ASPP2 and P16INK4a may cooperatively play a role in differentiation degree ,lymph nodes metastasis and apoptosis in esophageal carcinoma. Co-examina-tion of them may be useful for the diagnosis and guiding the clinical treatment in esophageal carcinoma.

Expression of ASPP2 and P16INK4a and its relationship with apoptosis in esophageal carcinoma / 实用医学杂志

Objective To study the expression of ASPP2 and P16INK4a in esophageal carcinoma and their relationship to the apoptosis and related clinicopathological characteristics. Methods Immunohistochemistry S-P method was used to examine the expression of ASPP2 and P16INK4a in the pathological specimens of 112 esophageal carcinoma cases and 31 cases of normal esophageal mucosa. TUNEL was also employed to detect the rate of apopto-sis in 37 esophageal carcinoma cases and 12 cases of normal esophageal mucosa. Results The difference between the expression of ASPP2 and P16INK4a in esophageal carcinoma and normal esophageal mucosa was statistically signif-icant(P < 0.05). Their abnormal expressions were all related to lymph node metastasis and differentiation degree (P < 0.05). The difference between the positive rate of apoptosis in esophageal carcinoma and normal esophageal mucosa was statistically significant(P < 0.05). The abnormal expression of ASPP2 and P16INK4a was all related to apoptosis in esophageal carcinoma. Conclusions The different expression of ASPP2 and P16INK4a may cooperatively play a role in differentiation degree ,lymph nodes metastasis and apoptosis in esophageal carcinoma. Co-examina-tion of them may be useful for the diagnosis and guiding the clinical treatment in esophageal carcinoma.

Expressions of p16INK4a protein in cervical lesion tissues of Mongolian patients and their significances / 吉林大学学报(医学版)

Objective:To detect the expressions of p16INK4a protein in the cervical lesion tissues of the Mongolian patients, and to explore the relationship between its expression and the occurrence and development of cervical cancer in the Mongolian patients.Methods:A total of 100 cases of paraffin sections of cervical cancer, cervical intraepithelial neoplasia(CIN),chronic cervicitis and uterine leiomyoma were divided into 25 cases of cervical cancer, 35 cases of CIN, 20 cases of chronic cervicitis, and 20 cases of uterine leiomyoma groups. The expressions of p16INK4a protein in different cervical tissues were detected by immunohistochemical SP method.Results:The positive rates of p16INK4a protern in cervical cancer, CIN, chronic cervicitis and uterine leiomyoma tissnes were 100.0%, 74.3%, 25.0%,and 10.0%, respectively.The results of K-W H rank sum test for multiple sample comparisons showed that the positive expression rate of p16INK4a protein in cervical cancer tissue was significantly higher than those in CIN, chronic cervicitis and uterine leiomyoma tissues(P<0.05).Conclusion:p16INK4a protein can be used as a indicator to screen the Mongolian patients with early cervical cancer.

P16INK4a: A Potential Diagnostic Adjunct For Prediction Of Highgrade Cervical Lesions In Liquid-Based Cytology: With HPV Testing And Histological Correlation

Human papillomavirus (HPV) is a necessary cause of cervical cancer and its precursors. Increased expression of high-risk hrHPV viral oncogenes in abnormal cells might increase the expression of p16INK4a. We aimed to determine the role of p16INK4a in detecting hrHPV-transformed epithelial cells in liquid-based cervical cytology, and compared the results with hrHPV DNA testing by realtime polymerase chain reaction (RT-PCR). Fifty-seven cytological samples were tested for p16INK4a immunomarker and hrHPV DNA. Test performance of both tests was determined by comparing sensitivity, specificity and predictive values using available histological follow-up data as gold standard. Of 57 samples, 36 (63.2%) showed immunoreactivity for p16INK4a and 43 (75.4%) were hrHPV-infected. A fairly low concordance rate (k = 0.504) between p16INK4a immunolabelling and hrHPV DNA status was noted. For prediction of cervical intraepithelial neoplasia (CIN) II and worse lesions, p16INK4a had a sensitivity and specificity of 93.5% and 60%; whereas hrHPV DNA testing had a sensitivity and specificity of 100% and 20%. Dual testing by combining p16INK4a and hrHPV showed sensitivity and specificity of 100% and 33.3%. In conclusion, p16INK4a is useful in predicting severity of the cytological abnormalities. Although p16INK4a is more specific but less sensitive than hrHPV in detecting high-grade cervical lesions, a combination of both tests failed to demonstrate significant improvement in diagnostic sensitivity, specificity and predictive value. Larger-scale prospective studies are required to assess further whether this biomarker should be routinely used as primary screening tool independently or in combination with hrHPV testing to improve diagnostic accuracy in cervical cytology.

Correlation of p16INK4a immunoexpression and human papillomavirus (HPV) detected by in-situ hybridization in cervical squamous neoplasia

Persistence and eventual integration of high-risk HPV (hrHPV) into the cervical cell is crucial to the progression of cervical neoplasia and it would be beneficial to morphologically identify this transformation in routine surgical pathology practice. Increased p16INK4a (p16) expression is a downstream event following HPV E7 binding to pRB. A study was conducted to assess the correlation between hrHPV detection using a commercial in-situ hybridization assay (Ventana INFORM HPV ISH) and p16 immunoexpression (CINtec Histology Kit) in cervical squamous intraepithelial lesions and squamous carcinoma. 27 formalin-fixed, paraffin-embedded cervical low-grade squamous intraepithelial lesions (LSIL), 21 high-grade squamous intraepithelial lesions (HSIL) and 51 squamous carcinoma (SCC) were interrogated. hrHPV was significantly more frequent in HSIL (76.2%) and SCC (88.2%) compared to LSIL(37.0%). p16 expression was similarly more frequent in HSIL (95.2%) and SCC (90.2%) compared to LSIL(3.7%). That the rates of hrHPV when compared with p16 expression were almost equivalent in HSIL and SCC while p16 was expressed in only 1 of the 10 LSIL with hrHPV, are expected considering the likelihood that transformation has occurred in HSIL and SCC but does not occur in majority of LSIL.

Conventional cytology, visual tests and evaluation of P16INK4A as a biomarker in cervical intraepithelial neoplasia.

OBJECTIVES: (1) To detect cervical intraepithelial neoplasia (CIN) using Papanicolaou test (PAP test), visual tests (visual inspection after the application of acetic acid [VIA], visual inspection after the application of Lugol’s iodine [VILI]), colposcopy, and biopsy. (2) To study the biomarker p16INK4A expression by immunostaining. MATERIALS AND METHODS: Experimental study was conducted from November 2009 to April 2011. 1500 women were screened for cancer cervix using conventional PAP test, VIA, and VILI. Sensitivity, specificity, positive, and negative predictive values of these tests were calculated individually, sequentially, and in parallel. Women having positive results underwent colposcopy and biopsy if required. p16INK4Aexpression in biopsy samples was studied using immunohistochemistry. RESULTS: All test positive cases (n = 235) underwent colposcopy. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of PAP with atypical squamous cells of undetermined significance (ASCUS) as cut‑off was 40%, 99.25%, 35.25%, and 99.39%; VIA was 60%, 93.06%, 8.03%, and 99.56% and VILI was 80%, 86.06%, 5.4%, and 99.76%, respectively. When PAP, VIA, and VILI were used in parallel sensitivity, specificity, PPV, and NPV improved to 100%, 85.18%, 6.38%, and 100%, respectively. Colposcopic abnormalities were detected in 83 and biopsy proven CIN in 15. p16INK4A expression was seen in eight of 15 CIN cases. CONCLUSIONS: (1) PAP test and visual techniques are complementary. (2) p16INK4Aexpression was seen in majority of CIN 2 lesions suggesting a higher grade lesion.

p16INK 4a and Ki-67 expression in human papilloma virus-related head and neck mucosal lesions / Expresión de p16INK 4a y Ki-67 en las lesiones de cabeza y cuello relacionados con el virus del papilloma humano

Human papilloma virus (HPV) is postulated as a risk factor in the etiology of some specific mucosal pathologies in the head and neck regions. Despite the frequent use of p16INK4a as a surrogate marker for HPV-infection, there is still controversy with respect to its reliability. This study has been undertaken to assess the potential role of p16INK 4a and Ki-67 expression in HPV-related lesions. The study was conducted on 71 specimens of oral, tonsillar and laryngeal lesions which comprised 25 dysplasia and 46 papilloma specimens. Specimens were immunohistochemically stained for p16INK4A and Ki-67 proteins. HPV DNA was determined by one step multiplex polymerase chain reaction. HPV DNA was detected in 33.8% of all lesions. Tonsil and larynx lesions showed significant differences with oral lesions for HPV positivity (p<0.001). p16INK 4a over-expression was seen in 56.5% of papilloma and 60% of dysplasia specimens. HPV status showed a positive correlation with p16INK 4a expression in tonsillar dysplasias (p<0.001). p16INK 4a expression may have a value as a marker in high risk HPV induced dysplasias, but not in low risk infected lesions. The proliferation index is not related to HPV-induced lesions and may be evaluated as an independent marker in head and neck premalignant lesions.

El virus del papiloma humano (VPH) se postula como un factor de riesgo en la etiología de algunas patologías de la mucosa, específicas en las regiones de cabeza y cuello. A pesar de usar con frecuencia el p16INK4A como un marcador sustituto para la infección por VPH, todavía existe controversia con respecto a su fiabilidad. Este estudio se ha llevado a cabo para evaluar el papel potencial de la expresión de p16INK 4a y de Ki-67 en las lesiones relacionadas con el VPH. El estudio se realizó en 71 muestras de lesiones orales, tonsilares y laríngeas que comprendían 25 displasias y 46 especímenes de papiloma. Los especímenes fueron teñidos inmunohistoquímicamente para p16INK4a y Ki-67. El ADN del VPH se determinó mediante una PCR multiplex de un paso. ADN del VPH se detectó en el 33,8% de todas las lesiones. Las lesiones de la amígdala y laringe mostraron diferencias significativas con lesiones orales para la positividad de VPH (p <0,001). Sobre-expresión de p16INK 4a se observó en 56,5% de las muestras de papiloma y 60% de las muestras de displasia. El estatus del VPH mostró una correlación positiva con la expresión de p16INK4a en displasias tonsilares (p <0,001). La expresión de p16INK4a puede tener valor como marcador en las displasias inducidas por VPH de alto riesgo, pero no en las lesiones infectadas de bajo riesgo. El índice de proliferación no está relacionado con las lesiones inducidas por VPH y puede ser evaluado como un marcador independiente en las lesiones premalignas de la cabeza y del cuello.

Inhibitory effect of chlorogenic acid on senescence of human skin fibroblasts / 中华皮肤科杂志

[Abstract ] Objective To evaluate the inhibitory effect of chlorogenic acid on senescence of human skin fibroblasts (HSFs). Methods Fibroblasts isolated from human foreskin were treated with 1 mmol/L glyoxal in vitro to develop a model for cellular senescence. In order to select effective concentrations of chlorogenic acid, some HSFs were treated with 1 mmol/L glyoxal alone or in combination with chlorogenic acid at different concentrations (5, 10, 20, 40, 80 μmol/L)for 3 days, with those receiving no treatment serving as the blank control group. Then, methyl thiazolyl tetrazolium (MTT)assay was performed to evaluate the proliferative activity of HSFs. Some HSFs were divided into 5 groups to be cultured alone(blank control group), or treated with 1 mmol/L glyoxal(glyoxal group)or the combination of 1 mmol/L glyoxal and chlorogenic acid at effective concentrations of 10, 20 and 40 μmol/L (glyoxal + chlorogenic acid groups). Senescence associated β-galactosidase (SA-β-gal)staining and real-time fluorescence-based quantitative PCR were conducted to determine the percentage of senescent cells and expression level of p16INK4a mRNA respectively. Statistical analysis was carried out by one-way analysis of variance followed by the least significant difference(LSD)-t test. Results Compared with the blank control group, the glyoxal group showed significantly decreased cellular proliferative activity of HSFs (55.65% ± 2.00% vs. 100% ± 6.90%, P 0.05). Therefore, 10 - 40 μmol/L was selected as the effective concentrations of chlorogenic acid. The glyoxal group showed significant increases in the percentage of senescent (SA-β-gal-positive)cells (35.65% ± 2.24% vs. 13.00% ± 2.22%, P < 0.01)and expression level of p16INK4a mRNA (2-ΔΔCt: 1.00 ± 0.06 vs. 0.26 ± 0.05, P <0.01)compared with the blank control group, while the glyoxal + 10-, 20-, 40-μmol/L chlorogenic acid groups showed significantly decreased percentage of senescent cells (31.50% ± 2.13% , 22.31% ± 3.11% and 19.32% ± 3.01%respectively)and expression level of p16INK4a mRNA (2-ΔΔCt: 0.88 ± 0.08, 0.73 ± 0.06 and 0.68 ± 0.04 respectively) compared with the glyoxal group (all P < 0.05). Additionally, the percentage of senescent cells decreased with the increase in chlorogenic acid concentrations in the glyoxal + chlorogenic acid groups. Conclusion Chlorogenic acid can protect HSFs from glyoxal-induced senescence.

Expression and significance of P16INK4,P57kip2 and Ki67 in hepatocellular carcinoma / 重庆医学

Objective Analysis the expression and significance of P16INK4 ,P57kip2 and Ki67 in hepatocellular carcinoma .Meth-ods 21 normal liver tissue ,48 hepatocellular carcinoma tissues and 48 corresponding adjacent tissue were collected for this study . Immunohistochemical analysis the expression levels of P16INK4 ,P57kip2 and Ki67 .Results P16INK4 and P57kip2 in hepatocellular carci-noma tissue were significantly lower ,the positive rates were 49 .3% and 34 .2% respectively ,Ki67 was significantly higher than the normal liver tissue and corresponding adjacent tissue ,the difference was statistically significant(P<0 .05) ,the results were consist-ent with the Western blot ;different differentiated group p57kip2 ,Ki67 expression and P16INK4 were significant differences(P<0 .05) . Conclusion Low expression of P16INK4 and P57kip2 ,the higher expression of Ki67 play an important role in the development of hepa-tocellular carcinoma .