Regular ArticleAssociation between Cervical Neoplasia and Apoptosis as Detected byin SituNuclear Labeling
Abstract
Invasive cervical cancer is thought to arise from the progression of precancerous lesions. How these lesions proceed from precancers to cancer remains unknown. Data regarding other tissues indicate that altered programmed cell death (apoptosis), in addition to cellular proliferation, is associated with the development of neoplasia. Therefore, in order to better understand the development of cervical neoplasia, we investigated the rate of apoptosis in cervical precancer and cancer. Archival cervical samples from normal epithelium (n= 11), low-grade squamous intraepithelial lesions (LSIL,n= 11), high-grade squamous intraepithelial lesions (HSIL,n= 10), and squamous cancers (n= 10) were evaluated for chromatin cleavage, a hallmark of programmed cell death. We usedin situend-labeling of DNA strand breaks by terminal deoxynucleotidyltransferase incorporation of biotinylated deoxyuridine to 3′-OH ends of DNA, identified by a nickel–avidin–peroxidase. The apoptotic index (sum of apoptotic bodies divided by the total nuclei times 100) significantly decreased (P< 0.001) as the degree of neoplasia increased: 3.5% (±0.4) in normal cervical epithelium (4.8 ± 0.4) in LSIL, 1.4% (±0.4) in HSIL, and 0.4% (±0.1) in squamous cancers. Compared to normal epithelium, the total cell number per 10 mm2increased significantly (P< 0.001): 124 (±12) in normal epithelium, 162 (±9.7) in LSIL, 315 (±31) in HSIL, and 413 (±32) in squamous cancers. We conclude that increasing cervical atypicality is associated with a decrease in apoptosis. We hypothesize from our data that one factor involved in the progression of neoplasia in the uterine cervix is a decrease in the rate of normal cellular deletion.
References (0)
Cited by (22)
Evaluation of p53 and Bcl-2 expression as prognostic markers in invasive cervical carcinoma stage IIb/III patients treated by radiotherapy
2003, Gynecologic OncologyObjective. The objective was to evaluate the immunohistochemical expression of p53 and Bcl-2 proteins as prognostic markers in locally advanced [FIGO Stage IIb/III] invasive squamous cell carcinoma of the uterine cervix. It was also our aim to determine if they had any relationship to each other and to the pretreatment levels of apoptosis determined morphologically.
Methods. Seventy-six consecutive cases of invasive squamous cell carcinoma of the cervix diagnosed in 1995 were analyzed retrospectively for the apoptotic index and p53 and Bcl-2 expression determined immunohistochemically and correlated to the patient outcome at the end of a 5-year follow-up period.
Results. The overall cumulative 5-year survival was 52.63%. p53 immunoreactivity was seen in 53.9% cases with variable levels of expression. By univariate analysis, p53 positivity correlated with poor survival [chi-square P = 0.029; log-rank test P = 0.0468] but not upon multivariate analysis. The apoptotic index which ranged from 0 to 40 and Bcl-2 expression seen in 38.1% cases showed no correlation to survival. Neither p53 nor Bcl-2 expression correlated with the apoptotic index or with each other. Seven cases with the immunophenotype p53-/Bcl-2+ had an excellent survival.
Conclusions. Neither p53 nor Bcl-2 expression are independent predictors of prognosis in locally advanced cervical squamous cancers. However, evaluation of their combined expression may affect clinical outcome and needs further investigation.
Apotosis-related proteins in cervical intraepithelial neoplasia and squamous cell carcinoma of the cervix
2002, Gynecologic OncologyObjectives. A loss of balance between proliferation and apoptosis leads to tumor formation. Normal cervical epithelium becomes dysplastic before potentially developing into carcinoma. This study was conducted to delineate the role of apoptosis-related proteins in various stages of development in cervical neoplasia. Both regulator and effector proteins were examined.
Methods. The expression of apoptosis-related proteins, including five members of the Bcl-2 family and two members of the caspase family, was evaluated in 26 low-grade cervical intraepithelial neoplasias (CIN-L), 37 high-grade cervical intraepithelial neoplasias (CIN-H), and 43 cervical squamous cell carcinomas, using immunohistochemistry. Specimens showing cytosolic immunoreactivity in 10% or more of the neoplastic cells were considered immunopositive.
Results. One hundred six subjects were studied. All seven apoptotic regulators examined were positive in a proportion of these neoplasms. The expression of Caspase 3 was significantly higher in CIN-H than in CIN-L (P = 0.016). The expression of Bak, Caspase 3, and Caspase 6 was reduced in cervical carcinoma compared to CIN-H (P < 0.01, P = 0.026, P < 0.01, respectively).
Conclusions. There is an increase in expression of Caspase 3 in CIN-H compared to CIN-L and the increase is thought to be related to the increased proliferative activity in dysplastic cells. The reduction of Bax, Caspase 3, and Caspase 6 expression in carcinoma indicates that the apoptotic mechanism has become defective in the process of malignant transformation.
Human papillomavirus infection among Egyptian females with cervical carcinoma: Relationship to spontaneous apoptosis and TNF-α
2001, Clinical BiochemistryObjectives: This study was designed to detect HPV type-16 in Cervical carcinoma (CC) tissue specimens. The results were correlated with clinicopathological parameters of the carcinoma, with spontaneous apoptosis and with immunoreactivity to TNF-α antibodies
Methods: Fresh frozen tissue specimens representing 30 cases of cervical carcinoma as well as 20 normal cervical tissues (NCT) were the subjects of this study. HPV-16 DNA was detected by Polymerase Chain Reaction (PCR). The occurrence of spontaneous apoptotic cell death was analyzed by the apoptosis assay. Apoptotic cells were also counted by light microscopy and the apoptotic index (AI) was calculated. Electron microscopy was used to confirm the morphology of apoptotic cells. TNF-α was quantified using EIA kit.
Results: HPV-16 DNA was more frequent in CC than in NCT. No correlation was observed between HPV infection and grade, stage or pathologic type of CC. The occurrence of spontaneous apoptosis was significantly higher in CC than in NCT, where it was correlated to advanced tumor stage and tumor pathology being more in adenocarcinoma (AC) than in squamous cell carcinoma (SCC). Moreover, AI was negatively correlated to HPV-16 infection. TNF-α levels were significantly higher in CC vs. NCT, where they were positively correlated to advanced tumor stage. TNF-α levels were correlated to DNA fragmentation and AI (r = 0.47 and 0.57 respectively). A cut-off value for TNF-α was calculated to be 9.1 pg/mg protein (using ROC curve). At the determined cut-off point the sensitivity was 70% and the specificity was 80%.
Conclusion: HPV infection, high levels of TNF-α and spontaneous apoptosis were strongly associated with malignant phenotype of cervical tissues. Rate of spontaneous apoptosis was higher in AC compared to SCC. On the other hand, HPV negativity was correlated with AI. Moreover, TNF-α and apoptotic cell death were correlated to each other as well as to tumor progression. No correlation was detected between TNF-α and HPV-16 infection.
Decreased programmed cell death in the uterine cervix associated with high risk human papillomavirus infection
1999, Pathology and Oncology ResearchThe relationship between apoptosis, apoptosis regulatory proteins, cell proliferation and human papillomavirus infection during various phases of tumor progression in the uterine cervix was studied. Apoptosis was defined by morphological criteria and the TUNEL assay. Expression of p53, bcl-2, bax, cyclin D1, Ki 67 and E6 protein was evaluated by immunocytochemistry. Presence of mutant p53 was detected using a mutant specific ELISA. Type of HPV infection was determined by PCR using type specific primers. Apoptosis showed significant negative correlation with increasing histological abnormality (p=0.0005). Higher tumor cell proliferation was associated with increasing histological abnormality (p=0.001 for Ki 67 and cyclin D1). There was significant correlation between histological grade and immunoreactivity of p53 (p=0.0001) and bcl-2 (p=0.0002). However, mutant p53 was expressed by only 12 of the 230 samples. Expression of bax and the bax/bcl-2 ratio showed an inverse correlation to histological grade (p=0.0003 and 0.0001, respectively). There was also an inverse correlation between extent of apoptosis and immunoreactivity of p53 (p=0.0001) and bcl-2 (p=0.0001). A significant positive correlation between expression of the bax protein and apoptosis was evident (p=0.0001). HPV infection significantly correlated to the extent of histological abnormality (p=0.0001). High risk HPV-E6 protein also showed this significant correlation (p=0.0002). There was an inverse correlation between apoptosis and HPV infection (p=0.0002). High risk HPV infection was associated with decreased apoptosis and also increased human cell proliferation. Lowest levels of bax/bcl-2 ratio was also associated with HPV 16 and 18 infection (p=0.0001). Modulation of apoptosis and apoptotic regulatory proteins by high risk HPV infection may be an important factor in the development of cervical cancer. Copyright 1999 W.B. Saunders Company Ltd on behalf of the Arányi Lajos Foundation
Altered apoptosis levels in hearts of human fetuses with Down syndrome
1998, American Journal of Obstetrics and GynecologyObjective: We sought to test the hypothesis that apoptosis is a method of remodeling in second-trimester fetal heart development. We also hypothesized that hearts from fetuses with Down syndrome would have different levels of apoptosis than would control hearts, associated with their abnormal heart development. Study Design: We obtained hearts from fetuses between 14 and 23 weeks’ gestation with Down syndrome without anomalies (n = 10) and with the atrioventricular canal defect (n = 5). These hearts were compared with control hearts without anomalies (n = 5 ). Hearts were subjected to in situ end-labeling of deoxyribonucleic acid to test for evidence of apoptosis. The apoptotic indices were compared by anatomic location. Results: Apoptotic nuclei were observed in each anatomic location in every category. The apoptotic indices were significantly lower in the atrial myocardial tissues of fetuses with Down syndrome than in control preparations (P < .05). The apoptotic index did not differ significantly in the atrial septum, ventricular myocardium, or ventricular septum. Conclusions: Apoptosis occurs in human fetal hearts during the second trimester. The different levels observed in our study suggest a different remodeling process in hearts of fetuses with Down syndrome than in hearts of control fetuses. Further study is needed to determine whether the different level of apoptosis associated with Down syndrome is due to the abnormal karyotype or to the presence of an anomaly. (Am J Obstet Gynecol 1998;179:962-5.)
Quick assessment of DNA damage in cervical epithelial cells using a chromatin dispersion test
2021, Archives of Gynecology and Obstetrics
- 1
To whom reprint requests should be addressed at Department of Obstetrics and Gynecology, Brigham and Women's Hospital, 75 Francis St., Boston, MA 02115. Fax: (617) 738-5124.