Stages Of Cervical Carcinogenesis Research Articles

Metabolomics variation profiling of vaginal discharge identifies potential targets for cervical cancer early warning.

Cervical cancer (CC) continues being the second leading cause of cancer death in women aged 20 to 39 years[1]. Recent evidence suggests that cervical microbiota differs in different stages of cervical carcinogenesis [2]. Previous research revealed that gut microbiota is linked to host metabolism and thus influence disease progression [3]. Can cervicovaginal flora affect host metabolism and therefore the progression of cervical cancer? To solve this problem, metabolomics has been implemented to detect small molecular differential metabolites in cancer progression. Metabolomics is the study of the thousands of low-molecular-weight molecules found in biological fluids and tissues of different individuals, whether normal or afflicted with disease, which reflect changes in biological functions.

Gene expression profiling of HPV-associated cervical carcinogenesis in formalin-fixed paraffin-embedded (FFPE) tissues using the NanoString nCounterTM platform.

Infection by high-risk human papillomavirus (HPV) causes genetic alterations in host cervical cells with consequent changes in gene expression affecting downstream molecular pathways, leading to the development of cervical cancer. In this exploratory study, we aimed to identify the perturbed cellular pathways during the various stages of cervical carcinogenesis. Total RNA was extracted from three formalin-fixed paraffin-embedded (FFPE) samples each of normal cervix, HPV-infected low-grade squamous intraepithelial lesion (LSIL), high-grade SIL (HSIL) and squamous cell carcinoma (SCC). Gene expression profiling was performed using the 770-gene panel from NanoString nCounter® PanCancer Pathways Panel to identify differentially expressed genes (DEGs) and significantly associated pathways in each stage of cervical cancer development. We identified 121 DEGs involved in cervical carcinogenesis. In the transformation from normal cells to LSIL, the MAPK, transcriptional misregulation and JAK-STAT pathways are implicated, while IL1B may promote inflammation and indirectly activates MMP9, resulting in collagen breakdown and cell migration. The cell cycle - apoptosis pathway with upregulation of E2F1 and MCM2, and DNA repair genes BRCA2-BRIP1 and FANCA are crucial during the progression from LSIL to HSIL. In the final stage of progression to SCC, the cell cycle and signaling pathways, as well as upregulation of c-MYC appear essential. In conclusion, archived FFPE-derived tissue samples are a valuable resource for gene expression profiling. The postulated dysregulated pathways and genes provide a guide of the molecular mechanisms that may be involved in the development of HPV-associated cervical cancer, for further investigation and validation studies.

The feature of cervical microbiota associated with the progression of cervical cancer among reproductive females

ObjectivesThe aim of this study was to characterize cervical microbiome feature of reproductive-age women in the progression of squamous intraepithelial lesions (SIL) to cervical cancer. MethodsWe characterized the 16S rDNA cervical mucus microbiome in 94 participants (age from 18 to 52), including 13 cervical cancer (CA), 31 high-grade SIL (HSIL), 10 low-grade SIL (LSIL), 12 HPV-infected (NH) patients and 28 healthy controls (NN). Alpha (within sample) diversity was examined by Shannon and Simpson index, while Beta (between sample) diversity by principle coordinate analysis (PCoA) of weighted Unifrac distances. Relative abundance of microbial taxa was compared using Linear Discriminant Analysis Effect Size (LEfSe). Co-occurrence analysis was performed to identify correlation among marker genera, and Phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt) to explore functional features and pathways of cervical microbiota. ResultsAlpha diversity(p < 0.05) was higher in severer cervical pathology with lower relative abundance of Lactobacillus as well as higher of anaerobes. Beta diversity (p < 0.01) was significantly different. Marker genera were identified including Porphyromonas, Prevotella and Campylobacter of CA and Sneathia of HSIL. The correlation of differential functional pathways with Prevotella was opposite to that with Lactobacillus. ConclusionOur study suggests differences in cervical microbiota diversity and relative abundance of reproductive-age females in different stages of cervical carcinogenesis. Marker genera might participate in the lesion progression and will be helpful for diagnosis, prevention and treatment. These findings may lead the way to further study of the cervical microbiome in development of cervical cancer.

Host-cell DNA methylation patterns during high-risk HPV-induced carcinogenesis reveal a heterogeneous nature of cervical pre-cancer

ABSTRACTCervical cancer development following a persistent infection with high-risk human papillomavirus (hrHPV) is driven by additional host-cell changes, such as altered DNA methylation. In previous studies, we have identified 12 methylated host genes associated with cervical cancer and pre-cancer (CIN2/3). This study systematically analyzed the onset and DNA methylation pattern of these genes during hrHPV-induced carcinogenesis using a longitudinal in vitro model of hrHPV-transformed cell lines (n = 14) and hrHPV-positive cervical scrapings (n = 113) covering various stages of cervical carcinogenesis. DNA methylation analysis was performed by quantitative methylation-specific PCR (qMSP) and relative qMSP values were used to analyze the data. The majority of genes displayed a comparable DNA methylation pattern in both cell lines and clinical specimens. DNA methylation onset occurred at early or late immortal passage, and DNA methylation levels gradually increased towards tumorigenic cells. Subsequently, we defined a so-called cancer-like methylation-high pattern based on the DNA methylation levels observed in cervical scrapings from women with cervical cancer. This cancer-like methylation-high pattern was observed in 72% (38/53) of CIN3 and 55% (11/20) of CIN2, whereas it was virtually absent in hrHPV-positive controls (1/26). In conclusion, hrHPV-induced carcinogenesis is characterized by early onset of DNA methylation, typically occurring at the pre-tumorigenic stage and with highest DNA methylation levels at the cancer stage. Host-cell DNA methylation patterns in cervical scrapings from women with CIN2 and CIN3 are heterogeneous, with a subset displaying a cancer-like methylation-high pattern, suggestive for a higher cancer risk.

DNA methylation regulated microRNAs in human cervical cancer

Regulation of miRNA gene expression by DNA promoter methylation may represent a key mechanism to drive cervical cancer progression. In order to understand the impact of DNA promoter methylation on miRNAs at various stages of cervical carcinogenesis, we performed DNA methylation microarray on Normal Cervical Epithelium (NCE), Cervical Intraepithelial Neoplasia (CIN I-III) and Squamous Cell Carcinoma (SCC) tissues to identify differentially methylated miRNAs followed by validation by bisulfite sequencing. Further, expression of miRNAs was analyzed by qRT-PCR in clinical tissues and cervical cancer cell lines. Transcriptional activity was determined by luciferase assay. We identified a total of 69 hypermethylated and hypomethylated miRNA promoters encompassing 78 CpG islands in all except Y chromosome, among the three groups. The candidate DNA promoters of miR-424 were significantly hypermethylated and miR-200b and miR-34c were significantly hypomethylated in SCC compared to NCE (P < 0.05). Expression of miR-424, miR-200b, and miR-34c were inversely correlated with promoter DNA methylation in tissue samples. Treatment of cell lines with 5-aza-2'-deoxycytidine showed differential expression in all three miRNAs. We observed a decrease in miRNA promoter activity following in vitro SssI methylase treatment of miR-424, miR-200b, and miR-34c. Luciferase assay demonstrated that miR-200b and miR-424 functionally interacts with 3'-UTR of HIPK3 and RBBP6 respectively and decreased their activity in presence of miR-200b and miR-424 mimics transfected in SiHa cells. Taken together, we have identified deregulation of miRNAs by aberrant DNA promoter methylation, leading to its transcriptional silencing during cervical carcinogenesis, which can be potential targets for diagnosis and therapy.

E6-associated transcription patterns in human papilloma virus 16-positive cervical tissues.

The change in transcription pattern induced by post-transcriptional RNA splicing is an important mechanism in the regulation of the early gene expression of human papilloma virus (HPV). The present study was conducted to establish a method to specifically amplify HPV-16 E6-associated transcripts. The E6-related transcripts from 63 HPV-16-positive cervical tumor tissue samples were amplified, consisting of eight cases of low-risk intraepithelial lesions, 38 cases of high-risk intraepithelial lesions and 17 cases of cervical cancer (CxCa). The appropriate amplified segments were recovered following agarose gel electrophoresis, and subjected to further sequencing and sequence alignment analysis. Six groups of E6 transcription patterns were identified from HPV-16-positive cervical tumor tissue, including five newly-discovered transcripts. Different HPV-16 E6-associated transcription patterns were detected during the development of CxCa. Over the course of the progression of the low-grade squamous intraepithelial lesions to CxCa, the specific HPV-16 E6-associated transcription patterns and the dominant transcripts were all different. As indicated by this study, the transcription pattern of the E6 early gene of HPV-16 was closely associated with the stages of cervical carcinogenesis, and may also be involved in the development of CxCa.

Human Papillomavirus Infection and the Multistage Carcinogenesis of Cervical Cancer

This short review outlines our understanding of cervical cancer precursors, concentrating on the central etiologic role of persistent human papillomavirus infection. The stages of cervical carcinogenesis are better understood than for most other major cancers, providing a successful cancer etiology and prevention model.

Cervical intraepithelial neoplasia- predictive molecular growth factors in natural history.

There is considerable controversy regarding the possible over-treatment of patients with mild Cervical Intraepithelial Neoplasia (CIN), with lesions being often excised or ablated. Thus, identifying the markers of potentially malignant lesions would be of a great prognostic value. In the current study, we hypothesized that using colposcopic, cytological and histological findings together with assessing the expression of molecular growth factors can predict CIN outcome. The study group consisted of 285 women between 19 and 81 years of age (median age, 37,8 years). The follow up were 60 months and considered 138 women: 50 women with Subclinical Papillomavirus Infection (SPI), 50 women with CIN1 and 38 women with CIN2. All patients underwent cytology, colposcopy, and sampling for subsequent testing for HPV. In cases in which colposcopy suggested the presence of suspicious lesions, biopsy specimens were taken. HPV DNA was genotyped for HPV types 16, 18, 31, 33, and 45 by multiplex PCR. Transcripts of HR HPV types 16, 18, 31, 33, and 45 were detected by the NucliSens EasyQ HPV assay. The VEGF expression was analyzed with immunohistochemistry, RNA extraction, cDNA synthesis and RT-PCR analysis and Western blot. We found that so called lymphangiogenetic switch (over expression of VEGF C and VEGFR-2) appears already in CIN 2, which is a rare observation, Persistent HPV HR infection is not only a trigger but also a maintenance factor in the cervical carcinogenesis. CIN2/3 and cervical cancer is in high percentage associated with the presence of HR DNA HPV as well as E6/E7 DNA mRNA. In CIN2/3 and cervical cancer VEGF and its receptor expression correlate with the stage of cervical carcinogenesis. Progression of cervical intraepithelial neoplasia occurs when co expression of all: HR DNA HPV, E6/E7 HR HPV mRNA and VEGF is present.

The role of co-factors in the progression from human papillomavirus infection to cervical cancer

ObjectiveCo-factors for cervical cancer, including oral contraceptive (OC) use, smoking and multiparity have been identified; however, the stage at which they act in cervical carcinogenesis is not clear. We compared established risk factors among women with CIN2 and CIN3 to evaluate the heterogeneity of these factors in precancer and also assessed their role during cervical carcinogenesis. MethodsThe current analysis included 2783 women with various stages of cervical disease who were enrolled in the Study to Understand Cervical Cancer Early Endpoints and Determinants (SUCCEED) and the Biopsy Study. Associations of co-factors within cervical precancer and at different stages of cervical carcinogenesis were estimated using logistic regression. ResultsLong-term OC use (10+years vs. never: OR=2.42, 95% CI: [1.13–5.15]), multiparity (3+ births vs. nulliparous: OR=1.54 [1.04–2.28]), smoking (ever vs. never: OR=1.95 [1.48–2.58]), and no Pap test in the previous five years (2.05 [1.32–3.17]) were positively associated with CIN3 compared to CIN2. We observed that long-term OC use, parity and smoking were associated with an increased risk of CIN3 compared to <CIN2 (1.97 [1.12–3.46]; 2.23 [1.59–3.11]; 2.60 [2.04–3.30], respectively), whereas associations were not significantly different (OC use, parity) or showed decreased risk (smoking) when comparing cancer to CIN3. ConclusionsDifferences in established risk factors suggest that CIN3 is a more specific definition of precancer than CIN2. Hormonally-related factors and smoking play a role in the transition from human papillomavirus infection to precancer.

CCDB: a curated database of genes involved in cervix cancer

The Cervical Cancer gene DataBase (CCDB, http://crdd.osdd.net/raghava/ccdb) is a manually curated catalog of experimentally validated genes that are thought, or are known to be involved in the different stages of cervical carcinogenesis. In spite of the large women population that is presently affected from this malignancy still at present, no database exists that catalogs information on genes associated with cervical cancer. Therefore, we have compiled 537 genes in CCDB that are linked with cervical cancer causation processes such as methylation, gene amplification, mutation, polymorphism and change in expression level, as evident from published literature. Each record contains details related to gene like architecture (exon–intron structure), location, function, sequences (mRNA/CDS/protein), ontology, interacting partners, homology to other eukaryotic genomes, structure and links to other public databases, thus augmenting CCDB with external data. Also, manually curated literature references have been provided to support the inclusion of the gene in the database and establish its association with cervix cancer. In addition, CCDB provides information on microRNA altered in cervical cancer as well as search facility for querying, several browse options and an online tool for sequence similarity search, thereby providing researchers with easy access to the latest information on genes involved in cervix cancer.

Using Biomarkers as Objective Standards in the Diagnosis of Cervical Biopsies

Histopathologic diagnosis of cervical biopsies determines clinical management of patients with an abnormal cervical cancer-screening test yet is prone to poor interobserver reproducibility. Immunohistochemical staining for biomarkers related to the different stages of cervical carcinogenesis may provide objective standards to reduce diagnostic variability of cervical biopsy evaluations but systematic, rigorous evaluations of their potential clinical utility are lacking. To address diagnostic utility of human papillomavirus (HPV) L1, p16(INK4a), and Ki-67 immunohistochemical staining for improving diagnostic accuracy, we conducted a community-based and population-based evaluation using 1455 consecutive cervical biopsies submitted to the Department of Pathology at the University of Virginia during a period of 14 months. Thin-sections of each biopsy from 1451 of 1455 (99.7%) biopsies underwent evaluation of immunohistochemical stains for the 3 biomarkers, masked to the original diagnosis, and the results were compared with an adjudicated, consensus diagnosis by 3 pathologists. p16 immunostaining, using the strongest staining as the cutpoint, was 86.7% sensitive and 82.8% specific for cervical intraepithelial neoplasia (CIN) grade 2 or more severe (CIN2(+)) diagnoses. The performance of p16(INK4a) was more sensitive (P<0.001), less specific (P<0.001), and of similar overall accuracy for CIN2(+) compared with the combined performance of all pathologist reviews in routine clinical diagnostic service (sensitivity=68.9%, specificity=97.2%). Ki-67 immunostaining was also strongly associated with a CIN2(+) diagnosis but its performance at all staining intensities was inferior to p16 immunostaining, and did not increase the accuracy of CIN2(+) diagnosis when combined with p16(INK4a) immunostaining compared with p16(INK4a) immunostaining alone. We found no utility for L1 immunostaining in distinguishing between CIN and non-CIN. In conclusion, with a rigorous evaluation, we found immunohistochemical staining for p16 to be a useful and reliable diagnostic adjunct for distinguishing biopsies with and without CIN2(+).

36 Cytomegalovirus (CMV) retinitis post rituximab therapy: A case report

We wanted to identify the most promising methylation marker candidates for cervical cancer early detection.A systematic literature review was performed in Medline and weighted average frequencies for methylated genes stratified by tissue source and methods used were computed.51 studies were identified analyzing 68 different genes for methylation in 4376 specimens across all stages of cervical carcinogenesis. 15 genes, DAPK1, RASSF1, CDH1, CDKN2A, MGMT, RARB, APC, FHIT, MLH1, TIMP3, GSTP1, CADM1, CDH13, HIC1, and TERT have been analyzed in 5 or more studies. The published data on these genes is highly heterogeneous; 7 genes (CDH1, FHIT, TERT, CDH13, MGMT, TIMP3, and HIC1) had a reported range of methylation frequencies in cervical cancers of greater than 60% between studies. Stratification by analysis method did not resolve the heterogeneity. Three markers, DAPK1, CADM1, and RARB, showed elevated methylation in cervical cancers consistently across studies.There is currently no methylation marker that can be readily translated for use in cervical cancer screening or triage settings. Large, well-conducted methylation profiling studies of cervical carcinogenesis could yield new candidates that are more specific for HPV-related carcinogenesis. New candidate markers need to be thoroughly validated in highly standardized assays.

Correlation of inhibitor of differentiation 1 expression to tumor progression, poor differentiation and aggressive behaviors in cervical carcinoma

Objective To study the correlation of inhibitor of differentiation 1 (Id-1) to tumor invasion and metastasis by examining Id-1 expression levels in different stages of cervical carcinogenesis. Methods Id-1 mRNA and protein expression was detected in total of 171 cervical samples including precancerous and cancerous tissues by quantitative RT-PCR (qRT-PCR) and immunohistochemistry (IHC), respectively. Twenty-five normal cervical tissues were used as a normal control. Correlation between Id-1 positive rates and expression levels to cancer progression and clinicopathologic features was statistically analyzed. Results A gradual increase of Id-1 protein expression associated with cervical cancer progression was detected (4%, 16%, 50% and 75.9% in normal, low squamous intraepithelial lesion (LSIL), high squamous intraepithelial lesion (HSIL) and cancer tissue, respectively, p < 0.001). A similar trend of Id-1 mRNA expression was also observed (1.3, 3.4 and 10.4 fold higher than normal tissues in LSIL, HSIL and cancer tissue, respectively, p < 0.001). Furthermore, the Id-1 expression level was correlated to tumor grade ( p = 0.005), lymph node metastasis ( p = 0.001), interstitial invasive ( p < 0.001) and tumor size ( p < 0.001). These results suggest that high Id-1 expression is associated with tumor growth, invasion and metastasis. Conclusion Id-1 expression is correlated to progression and aggressive behaviors in cervical cancer, suggesting a tumor-promoting role for Id-1 in progression of this malignancy.

Interaction between inflammation and angiogenesis during different stages of cervical carcinogenesis

Objective. The primary aim of this study was to investigate the role of angiogenesis and inflammatory cell response in cervical carcinogenesis. Methods. Formalin-fixed tissue specimens from 58 uterine cervical specimens (8 CIN1, 14 CIN2, 28 CIN3, and 8 SCC), representing the different stages of cervical carcinogenesis, were immunohistochemically analyzed. Normal cervical tissue specimens were also included as controls. The present study assessed the expression of CD31 and CD105 to evaluate microvessel density (MVD), the macrophage marker CD68 and the panleukocyte marker CD45. In addition, expression of iNOS (inducible Nitric Oxide Synthase) was also evaluated. Results. MVD, measured by either CD31 or CD105, increased along the continuum from normal epithelium to squamous cell carcinoma, and a significant correlation between the CD105-MVD and the CD31-MVD was observed ( r = 0.8735; p < 0.0001). Furthermore, the number of infiltrating macrophages was significantly associated with progression to malignancy. Interestingly, there was a close positive correlation between macrophage counts and CD105-MVD ( r = 0.7525; p < 0.0001). In striking contrast to the other angiogenic and inflammatory markers tested, iNOS expression was significantly reduced as cervical lesion grade progressed from low to high. Conclusions. Our findings demonstrated a positive correlation between neovascularity and macrophage counts, whereas iNOS expression displayed an inverse relationship with macrophage density and tumor progression. Low iNOS expression may modify the function of tumor-infiltrating macrophages toward a malignant phenotype that promotes tumor progression rather than an anti-tumor response.

Inestabilidad microsatelital en lesiones preneoplásicas y neoplásicas del cuello uterino: Correlación con el genotipo del virus papiloma humano

The association between some specific human papilloma virus (HPV) types and cervix cancer is well known. However, the genetic conditions that favor the development of cervical cancer are less well known. To determine the presence of satellite instability (MSI) in preneoplastic and neoplastic lesions of the cervix and correlate these findings with HPV genotypes. Biopsy samples of cervical lesions were studied. Sixteen had low grade lesions, 22 had high grade lesions and 28 had an epidermoid cancer. Viral types were identified with polymerase chain reaction, dot-blot hybridization and restriction fragment length polymorphism. MSI was determined using a panel of eight highly informative microsatellites. Microsatellite instability in at least one locus was observed in 91, 56 and 69% of low grade lesions, high grade lesions and epidermoid carcinomas, respectively. MSI-High grade, MSI-Low grade instability and microsatellite stability were observed in 5, 60 and 46% of samples, respectively. Two of three samples with high grade instability had HPV 52 genotype. Other viral subtypes had frequencies that ranged from 78% to 100%, with the exception of HPV16 that was present in only 53% of samples with low grade instability. Two thirds of biopsy samples from cervical lesions had MSI, mechanism that can be involved in the first stages of cervical carcinogenesis. The low frequency of high grade instability, its association with HPV52 and the low frequency of HPV16 in samples with low grade instability, suggest different coadjutant mechanisms in cervical carcinogenesis.

Genetic Polymorphisms of Phase I and Phase II Xenobiotic Enzymes in Human Papillomavirus Related Lesion and Cancer of the Uterine Cervix

Objective: Other than a major role of human papillomavirus (HPV) infection, exposure to cigarette smoke, estrogen and possibly other carcinogens may be related to the development of cancer. The aim of this study was to investigate whether the polymorphism of CYP1A1, CYP2E1, mEH, and GSTM1 contributes to the risk of invasive cervical cancer (ICC) and the low-grade squamous intraepithelial lesions (LSIL) of the uterine cervix. Materials and Methods: In this study, genetic polymorphisms of xenobiotic metabolizing enzymes CYP1A1, CYP2E1, mEH, and GSTM1, as well as the status of HPV infection were determined in 113 cases with invasive cervical cancer (ICC), 80 cases of low grade squamous intraepithelial lesion (LSIL, the early lesion of HPV infection), as well as in 1:1 age-matched control subjects. Results: As expected, subjects with results positive for HPV had significant risks for LSIL and ICC, with OR (95% CI) of 19.6 (6.9-55.3) and 61.1 (25.3-147.9), respectively. In comparison with the A/A genotype, A/G or G/G genotype of the CYP1A1 gene showed a protection for ICC, but not for LSIL, with an OR of 0.3 (95% CI of 0.1-0.9) after adjustment with HPV status. Subjects carrying GSTM1 null genotype were at risk for LSIL (OR=2.5; 95% CI: 1.0-6.3) and ICC (OR: 2.4; 95% CI: 1.0-6.0) after HPV adjustment. The combination of CYP1A1 A/A and GSTM1 null genotype showed a higher risk for ICC (OR of 4.2, 95% CI of 1.4-12.3; p＜0.01). Conclusions: The results of this study suggest there is a significant role for the polymorphism of GSTM1 in early stage and that of CYP1A1 in the invasive stage of cervical carcinogenesis in a Taiwanese population.

The VEGF-C/Flt-4 axis promotes invasion and metastasis of cancer cells

Flt-4, a VEGF receptor, is activated by its specific ligand, VEGF-C. The resultant signaling pathway promotes angiogenesis and/or lymphangiogenesis. This report provides evidence that the VEGF-C/Flt-4 axis enhances cancer cell mobility and invasiveness and contributes to the promotion of cancer cell metastasis. VEGF-C/Flt-4-mediated invasion and metastasis of cancer cells were found to require upregulation of the neural cell adhesion molecule contactin-1 through activation of the Src-p38 MAPK-C/EBP-dependent pathway. Examination of tumor tissues from various types of cancers revealed high levels of Flt-4 and VEGF-C expression that correlated closely with clinical metastasis and patient survival. The VEGF-C/Flt-4 axis, through upregulation of contactin-1, may regulate the invasive capacity in different types of cancer cells.

Epigenetics of cervical cancer. An overview and therapeutic perspectives.

Cervical cancer remains one of the greatest killers of women worldwide. It is difficult to foresee a dramatic increase in cure rate even with the most optimal combination of cytotoxic drugs, surgery, and radiation; therefore, testing of molecular targeted therapies against this malignancy is highly desirable. A number of epigenetic alterations occur during all stages of cervical carcinogenesis in both human papillomavirus and host cellular genomes, which include global DNA hypomethylation, hypermetylation of key tumor suppressor genes, and histone modifications. The reversible nature of epigenetic changes constitutes a target for transcriptional therapies, namely DNA methylation and histone deacetylase inhibitors. To date, studies in patients with cervical cancer have demonstrated the feasibility of reactivating the expression of hypermethylated and silenced tumor suppressor genes as well as the hyperacetylating and inhibitory effect upon histone deacetylase activity in tumor tissues after treatment with demethylating and histone deacetylase inhibitors. In addition, detection of epigenetic changes in cytological smears, serum DNA, and peripheral blood are of potential interest for development of novel biomolecular markers for early detection, prediction of response, and prognosis.

Ex vivo study of MAPK profiles correlated with parameters of apoptosis during cervical carcinogenesis

Cervical cancer is a leading cause of death in developing countries and is the second highest occurring cancer in women all over the world. The progression of cancer is a multistep process affecting aspects of cellular function such as proliferation, differentiation and apoptosis. Mitogen activated protein kinases (MAPKs), which include p38-MAPK, c-Jun NH 2-terminal kinase (JNK) and extracellular signal-regulated kinases (ERKs) are closely associated with cell proliferation and apoptosis and the balance between them could determine a cell's fate. Despite the expanding research effort in vitro, little is known about MAPK activation in clinical specimens of cervical cancer. Therefore, the aim of this ex vivo study was to correlate the phosphorylation status (activity) of MAPKs (p38-MAPK, JNK and ERK), as well as poly (ADP-ribose) polymerase (PARP) and caspase-3 (two cellular markers of apoptosis), during the different stages of cervical carcinogenesis, to observe whether correlations between MAPK activities and apoptosis during the disease process exist. Decreased p38-MAPK phosphorylation was found in the carcinoma (Ca) group) compared to the normal tissues, as well when the low grade squamous intraepithelial lesion—LSIL) group and high grade squamous intraepithelial lesion—HSIL) group were compared with the Ca group. Interestingly, a significant decrease in ERK44 phosphorylation was observed in Ca when compared to LSIL and HSIL. There was also a significant decrease in JNK phosphorylation in Ca when compared with normal tissue and HSIL. As expected, caspase-3 activation and PARP cleavage was significantly lower in Ca when compared with normal tissue. Our results present the first evidence of in vivo involvement of MAPKs in cervical cancer and indicate a possible correlation between MAPK activities and apoptosis in the disease process.

Death receptors and ligands in cervical carcinogenesis: an immunohistochemical study

Objective Increasing imbalance between proliferation and apoptosis is important in cervical carcinogenesis. The death ligands FasL and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induce apoptosis by binding to their cognate cell-surface death receptors Fas or death receptor (DR) 4 and DR5. This study aims to examine if changes in death ligand and death receptor expression during different stages of cervical carcinogenesis are related to an imbalance between proliferation and apoptosis. Methods The immunohistochemical expression and localization of Fas/FasL and DR4/DR5/TRAIL were assessed in 11 normal cervices, 15 cervical intraepithelial neoplasia (CIN) grade I, 15 CIN II, 13 CIN III, and 25 (microinvasive) squamous cell cervical cancers. The number of apoptotic cells was determined by morphological criteria and the number of proliferating cells by counting Ki-67-positive cells. Results A marked increase in proliferation as well as apoptosis percentage was found with increasing severity of neoplasia. In normal cervix and CIN I samples, FasL, DR4, DR5, and TRAIL staining was mainly observed in the basal/parabasal layer, whereas Fas staining was localized in the superficial, more differentiated epithelial layer. Frequency of Fas-positive staining decreased with increasing severity of CIN. In contrast, homogeneous FasL, DR4, DR5, and TRAIL expression throughout the lesions was more frequently observed in CIN III and cervical cancer. FasL, DR4, DR5, and TRAIL staining patterns were correlated, although TRAIL expression was more intense in low-grade lesions. No association was found between death receptor or ligand expression with the percentage of apoptosis or proliferation. Conclusion The loss of Fas and the deregulation of FasL, DR4, DR5, and TRAIL in the CIN-cervical cancer sequence suggest a possible functional role of these death ligands and receptors during cervical carcinogenesis. The frequent expression of DR4 and DR5 presents these receptors as promising targets for innovative therapy modalities in cervical cancer.