The expression of Oct3/4A mRNA and not its isoforms is upregulated by the HPV16 E7 oncoprotein.

PURPOSE: Oct3/4 a transcription factor is involved in maintaining the characteristics of cancer stem cells. Oct3/4 can be expressed differentially with respect to the progression of cervical cancer (CC). In addition, Oct3/4 can give rise to three isoforms by alternative splicing of the mRNA Oct3/4A, Oct3/4B and Oct3/4B1. The aim of this study was to evaluate the mRNA expression from Oct3/4A, Oct3/4B and Oct3/4B1 in low-grade squamous intraepithelial lesion (LSIL), high-grade squamous intraepithelial lesion (HSIL), CC samples, and measure the effect of the HPV16 E7 oncoprotein on the mRNA expression from Oct3/4 isoforms in the C-33A cell line. METHODS: The expression levels of Oct3/4A, Oct3/4B and Oct3/4B1 mRNA were analyzed by reverse transcription quantitative polymerase chain reaction (RT-qPCR) in patients with LSILs, HSILs and CC. Additionally, C-33A cells that expressed the HPV16 E7 oncoprotein were established to evaluate the effect of E7 on the expression of Oct3/4 mRNA isoforms. RESULTS: Oct3/4A (p = 0.02), Oct3/4B (p = 0. 001) and Oct3/4B1 (p < 0. 0001) expression is significantly higher in patients with LSIL, HSIL and CC than in woman with non-IL. In the C-33A cell line, the expression of Oct3/4A mRNA in the presence of the E7 oncoprotein increased compared to that in nontransfected C-33A cells. CONCLUSION: Oct3/4B and Oct3/4B1 mRNA were expressed at similar levels among the different groups. These data indicate that only the mRNA of Oct3/4A is upregulated by the HPV16 E7 oncoprotein.

Women with chronic follicular gastritis positive for Helicobacter pylori express lower levels of GKN1.

In women, serum levels of CTSB, GKN2, LIPF, LIPFG, AZGP1, TOP2A and PGA4 are proposed as predictive markers of gastric cancer. It is unknown whether GKN1 expression varies with the sex of patients with chronic gastritis or gastric cancer. We studied 36 patients with histopathological diagnosis of chronic gastritis from the state of Guerrero, Mexico. PCR was performed for H. pylori detection and GKN1 expression was determined by RT-qPCR and western blot. GKN1 mRNA expression was significantly lower in patients with chronic follicular gastritis than in those with chronic chemical gastritis (p = 0.00071). The mRNA and protein level of expression of GKN1 were significantly lower in women with chronic follicular gastritis than in men with the same condition (p = 0.0279 and p = 0.0014, respectively); the lowest levels of GKN1 were detected in women with H. pylori-positive follicular gastritis (p = 0.0175 and p = 0.0111, respectively). Through a bioinformatic analysis, estrogen response elements were identified in the GKN1 promoter.

Differential proteins among normal cervix cells and cervical cancer cells with HPV-16 infection, through mass spectrometry-based Proteomics (2D-DIGE) in women from Southern México.

BACKGROUND: Cervical cancer (CC) is the fourth most common cancer in women worldwide with an estimated 528,000 new cases in 2012. The same year México had an incidence of 13,960 and a mortality of 4769 cases. There are several diagnosis methods of CC; among the most frequents are the conventional Pap cytology (Pap), colposcopy, and visual inspection with acetic acid (VIA), histopathological examination, tests of imaging and detection of high-risk papilloma virus (HR-HPV) with molecular tests (PCR, hybridization, sequencing). Proteomics is a tool for the detection of new biomarkers that can be associated with clinical stage, histological type, prognosis, and/or response to treatment. In this study we performed a comparative analysis of CC cells with normal cervical cells. The proteomic analysis was carried out with the fluorescent two-dimensional electrophoresis (2D-DIGE) technique to subsequently identify differential protein profiles using Decyder Software, and the selected proteins were identified by Mass Spectrometry (MALDI-TOF). RESULTS: The proteins that showed an increased expression in cervical cancer in comparison with normal cervix cells were: Mimecan, Actin from aortic smooth muscle and Lumican. While Keratin, type II cytoskeletal 5, Peroxiredoxin-1 and 14-3-3 protein sigma showed a decrease in their protein expression level in cervical cancer in comparison with normal cervix cells. CONCLUSIONS: Thus, this study was successful in identifying biomarker signatures for cervical cancer, and might provide new insights into the mechanism of CC progression.

Association of human papillomavirus 16 E6 variants with cervical carcinoma and precursor lesions in women from Southern Mexico.

BACKGROUND: HPV 16 is the cause of cervical carcinoma, but only a small fraction of women with HPV infection progress to this pathology. Besides persistent infection and HPV integration, several studies have suggested that HPV intratype variants may contribute to the development of cancer. The purpose of this study was to investigate the nucleotide variability and phylogenetically classify HPV 16 E6 variants circulating over a period of 16 years in women from Southern Mexico, and to analyze its association with precursor lesions and cervical carcinoma. METHODS: This study was conducted in 330 cervical DNA samples with HPV 16 from women who were residents of the State of Guerrero, located in Southern Mexico. According of cytological and/or histological diagnosis, samples were divided into the following four groups: no intraepithelial lesion (n = 97), low-grade squamous intraepithelial lesion (n = 123), high-grade squamous intraepithelial lesion (n = 19) and cervical carcinoma (n = 91). HPV 16 E6 gene was amplified, sequenced and aligned with reference sequence (HPV 16R) and a phylogenetic tree was constructed to identify and classify HPV 16 variants. Chi squared was used and data analysis and statistics were done with SPSS Statistics and STATA softwares. RESULTS: Twenty seven HPV 16 E6 variants were detected in women from Southern Mexico, 82.12% belonged to the EUR, 17.58% to AA1 and 0.3% to Afr2a sublineages. The most common was E-G350 (40%), followed by E-prototype (13.03%), E-C188/G350 (11.82%), AA-a (10.61%), AA-c (6.07%) and E-A176/G350 (5.15%). Eight new E6 variants were found and 2 of them lead to amino acid change: E-C183/G350 (I27T) and E-C306/G350 (K68T). The HPV 16 variant that showed the greatest risk of leading to the development of CC was AA-a (OR = 69.01, CI = 7.57-628.96), followed by E-A176/G350 (OR = 39.82, CI = 4.11-386.04), AA-c (OR = 21.16, CI 2.59-172.56), E-G350 (OR = 13.25, CI = 2.02-87.12) and E-C188/G350 (OR = 10.48, CI = 1.39-78.92). CONCLUSIONS: The variants more frequently found in women with cervical carcinoma are E-G350, AA-a, AA-c, E-C188/G350 and E-A176/G350. All of them are associated with the development of cervical carcinoma, however, AA-a showed the highest association. This study reinforces the proposal that HPV 16 AA-a is an oncogenic risk for cervical carcinoma progression in Mexico.

Schwannoma and Post-vaccine Changes: A Case Report.

Schwannomas are benign sheaths of Schwann cells that can present with degenerative and morphological changes; necrosis or hemorrhage are rare findings in these tumors. We present the case of a 28-year-old man with a C2-C4 cervical Schwannoma who experienced upper limb paresthesia in 2020 while presenting with COVID-19 symptoms. The patient later recovered and came to our institution, where surgery was scheduled one year after the initial diagnosis. One week before surgery, the patient received the first dose of the Moderna vaccine. Despite being asymptomatic, the patient underwent successful total resection of the schwannoma, which was confirmed histologically. However, extensive necrosis with abundant foamy macrophages was observed, suggesting a possible link to post-vaccine effects.

Plasma levels of YKL-40 as a prognostic factor in childhood acute lymphoblastic leukemia.

YKL-40, also known as chitinase-3-like protein 1 (CHI3L1), is an inflammatory glycoprotein secreted by different types of cells, such as inflammatory cells. The levels of this protein are elevated in the serum or plasma of patients with different types of cancer, and high concentrations are associated with poor prognosis and short survival in patients with liver, breast, lung, bladder and endometrial cancers. In Mexico, acute lymphoblastic leukemia (ALL) is the most common type of cancer affecting the pediatric population. The prognosis of patients with ALL is difficult to establish. Hence, the objective of the present study was to analyze the plasma levels of YKL-40 in Mexican children with ALL and investigate its role as a prognostic factor. A case-control study was performed in a population of 90 children aged 1-18 years, among whom 45 had ALL and 45 were hematologically healthy. The levels of YKL-40 in plasma samples were measured using ELISA and were found to be significantly higher in children with ALL compared with those in controls (P<0.0001). Children with ALL who had high plasma levels of YKL-40 (≥36.34 ng/ml) had shorter survival compared with those with low levels (<36.34 ng/ml; P<0.05). The findings of the present study revealed that the YKL-40 plasma level, age/initial leukocyte count and central nervous system invasion were associated with the prognosis of children with ALL [odds ratio (OR)=6.06, 95% confidence interval (CI): 1.1-31.6, P=0.03; OR=8.53, 95% CI: 1.2-58.2, P=0.03; and OR=6.45, 95% CI: 1.01-41.2, P=0.04, respectively]. Therefore, YKL-40 plasma levels may serve as a prognostic biomarker in pediatric patients with ALL.

E6/E7 Variants of Human Papillomavirus 16 Associated with Cervical Carcinoma in Women in Southern Mexico.

Persistent infection with the human papillomavirus 16 (HPV 16) is the cause of half of all cervical carcinomas (CC) cases. Moreover, mutations in the oncoproteins E6 and E7 are associated with CC development. In this study, E6/E7 variants circulating in southern Mexico and their association with CC and its precursor lesions were evaluated. In total, 190 DNA samples were obtained from scrapes and cervical biopsies of women with HPV 16 out of which 61 are from patients with CC, 6 from patients with high-grade squamous intraepithelial lesions (HSIL), 68 from patients with low-grade squamous intraepithelial lesions (LSIL), and 55 from patients without intraepithelial lesions. For all E7 variants found, the E7-C732/C789/G795 variant (with three silent mutations) was associated with the highest risk of CC (odd ratio (OR) = 3.79, 95% confidence interval (CI) = 1.46-9.85). The analysis of E6/E7 bicistron conferred to AA-a*E7-C732/C789/G795 variants revealed the greatest increased risk of CC (OR = 110, 95% CI = 6.04-2001.3), followed by AA-c*E7-C732/C789/G795 and A176/G350*E7-p. These results highlight the importance of analyzing the combinations of E6/E7 variants in HPV 16 infection and suggest that AA-a*E7-C732/C789/G795, AA-c*E7-C732/C789/G795, and A176/G350*E7-p can be useful markers for predicting CC development.

MiR-23b-3p reduces the proliferation, migration and invasion of cervical cancer cell lines via the reduction of c-Met expression.

Malignant transformation and progression in cancer is associated with the altered expression of multiple miRNAs, which are considered as post-transcriptional regulators of genes participating in various cellular processes. Although, it has been proposed that miR-23b-3p acts as a tumor suppressor in cervical cancer (CC), not all the pathways through which it alters the cellular processes have been described. The present study examines whether miR-23b-3p directly represses the c-Met expression and that consequently modifies the proliferation, migration and invasion of C33A and CaSki cells. c-Met has five microRNA response elements (MREs) for miR-23b-3p in the 3'-UTR region. The ectopic overexpression of miR-23b-3p significantly reduces c-Met expression in C33A and CaSki cells. The overexpression of miR-23b-3p reduces proliferation, migration and invasion of CaSki cells and the proliferation and invasion in C33A cells. In CaSki cells, the activation of Gab1 and Fak, downstream of c-Met, is reduced in response to the overexpression of miR-23b-3p. Together, the results in the present study indicate that miR-23b-3p is a tumor suppressor that modulates the progression of CC via post-transcriptional regulation of the c-Met oncogene.

Deregulation of folate pathway gene expression correlates with poor prognosis in acute leukemia.

The present study analyzed the mRNA expression levels of genes involved in the transport and metabolism of methotrexate (MTX) (RFC1, ABCC1, ABCB1, GGH, FPGS, ATIC, TS, MTHFR, MTRR, MS and MTHFD1) in patients with acute leukemia (AL). The expression levels of the examined genes were analyzed by reverse transcription quantitative polymerase chain reaction (RT-qPCR) in patients with AL (ALL:50/AML:19) and 66 healthy individuals. The mRNA expression levels of RFC1, MS, MTRR, MTHFR and ABCB1 were decreased (P<0.05), while those of GGH, FPGS, TS and MTHFD1 (P<0.05) were overexpressed in patients with AL. Patients with high mRNA levels of GGH (OR=4.28, 95% CI=1.29-14.14), TS (OR=7.14, 95% CI 1.84-27.81), MTHFR (OR=4.81, 95% CI=1.31-17.64), ABCB1 (OR=4.61, 95% CI=1.33-15.97) and ABCC1 (OR=5.50, 95% CI=1.12-27.06) had a higher chance of relapse. Interestingly, high mRNA levels of RFC1 are a protective factor in the risk of AL relapse (OR=0.22, 95% 0.06-0.80). The results of the present study indicated that deregulation of folate pathway gene expression is associated with poor prognosis in AL and that the expression levels of these markers could serve as novel molecular targets for the treatment of patients with AL.

Association Between the 5,10-MTHFR 677C>T and RFC1 80G>A Polymorphisms and Acute Lymphoblastic Leukemia.

BACKGROUND: Polymorphisms in folate-related genes are closely related to the development of cancer. The 5,10-MTHFR 677C>T and RFC1 80G>A polymorphisms are associated with an increased risk of susceptibility to pediatric ALL. OBJECTIVE: The aim of this study was to illustrate the association between 5,10-MTHFR 677C>T and RFC1 80G>A polymorphisms and ALL in a Mexican population. MATERIALS AND METHODS: This study was conducted in 60 pediatric ALL patients and 60 healthy individuals. The 5,10-MTHFR 677C>T and RFC1 80G>A polymorphisms were detected by the PCR-RFLP method. RESULTS: Our investigation revealed that the 5,10-MTHFR 677 C/T and 5,10-MTHFR 677T/T genotypes are associated with susceptibility to pediatric ALL (OR = 1.9, 95% IC = 1.36-12.09, p = 0.012 and OR = 2.8, 95% CI = 1.49-22.82, p = 0.011, respectively). Likewise, the G/A genotype from the RFC1 80G>A polymorphism showed an increased ALL risk compared to RFC1 G80G genotype (OR = 3. 3, 95% CI = 1.75-8.87, p = 0.002). CONCLUSION: Therefore, our results suggest that the 5,10-MTHFR 677C>T and RFC1 80G>A polymorphisms are factors involved in the susceptibility to ALL in Mexican population.

Methylation and expression of miRNAs in precancerous lesions and cervical cancer with HPV16 infection.

Abnormal expression and promoter methylation of microRNAs (miRNAs) are common events during cervical carcinogenesis. Worldwide, infection by types 18 and 16 of human papillomaviruses (HPVs) is considered the major risk factor for cervical cancer development. It has been reported that expression of the miRNAs can be deregulated by specific HPV genotypes. In this study we analyzed the promoter methylation of 22 miRNAs and the expression of three miRNAs in 10 non-squamous intraepithelial lesions (Non-SIL) without HPV16 infection, and 7 Non-SIL, 16 low-grade SIL (LSIL) and 16 cervical cancer samples, all with HPV16 infection. The methylation status was determined using Human Cancer miRNA EpiTect Methyl II Signature PCR Array® and the expression of miR-124, miR-218 and miR-193b was determined by qRT-PCR using individual TaqMan assays. Comparisons of groups defined were performed using the Fisher exact test for categorical variables and Mann-Whitney test for continuous variables. A p-value of <0.05 was considered statistically significant. The methylation levels of miR-124-2, miR-218-1, miR-218-2 and miR-34b/c promoters were significantly higher in cervical cancer than in LSIL samples. The methylation levels of miR-193b promoter were significantly lower in cervical cancer than in LSIL samples. The expression of miR-124 and miR-218 was significantly lower in cervical cancer than in LSIL samples. The expression of miR-193b was significantly higher in cervical cancer than in LSIL and Non-SIL samples. Our results suggest that the abnormal promoter methylation and expression of miR-124, miR-218 and miR-193b are common events during cervical carcinogenesis.

miR-23b as a potential tumor suppressor and its regulation by DNA methylation in cervical cancer.

BACKGROUND: The aberrant expression of miR-23b is involved in the development and progression of cancer. The aim of this study was to evaluate the potential role of methylation in the silencing of miR-23b in cervical cancer cell lines and to determine its expression in stages of malignant progression and in cervical cancer tissues HPV16-positive. METHODS: The methylation of the miR-23b promoter was determined in HeLa, SiHa, CaSki and C33A cells using a Human Cancer miRNA EpiTectMethyl II Signature PCR Array®. The cells were treated with 5-Aza-2'-deoxycytidine, and the expression of miR-23b, uPa, c-Met and Zeb1 was determined by qRT-PCR. miR-92a and GAPDH were used as controls. The expression of miR-23b was determined in cervical scrapes and biopsies of women without squamous intraepithelial lesions, with precursor lesions and with cervical cancer, all were HPV16-positive. The Fisher exact and Mann-Whitney tests were used to compare the differences of the expression of miR-23b, uPa, c-Met and Zeb1 among cell groups, and the difference among patients, respectively. The association between the expression of miR-23b and cervical cancer was determined by logistic regression with a confidence level of 95 %. A value of p < 0.05 was considered statistically significant. RESULTS: In C33A, HeLa and CaSki cells, methylation was associated with decreased expression of miR-23b. After treatment with 5-Aza-CdR, the expression of miR-23b increased in all cell lines and the expression of c-Met decreased in HeLa cells, while uPa and Zeb1 decreased in C33A and CaSki cells. In SiHa cells the expression of uPa, c-Met and Zeb1 increased. The expression of miR-23b decreased in relation to the increase in the severity of the lesion and was significantly lower in cervical cancer. In women with premalignant lesions HPV16-positive, decreased levels of miR-23b increased the risk of cervical cancer (OR = 36, 95 % CI = 6.7-192.6, p < 0.05). CONCLUSIONS: The results suggest that the expression of miR-23b is regulated by the methylation of its promoter and is possible that this microRNA influence the expression of uPa, c-Met and Zeb1 in cervical cancer cells lines. In women with premalignant lesions and cervical cancer infected with HPV16, the expression level of miR-23b agree with a tumor suppressor gene.

Identificación molecular de helicobacter pylori en tejidos gástricos con neoplasias malignas embebidos en parafina / Molecular identification of helicobacter pylori in paraffin embedded gastric tissues with malignant neoplasm

Introducción: Helicobacter pylori es considerado uno de los principales agentes causales de gastritis crónica, úlcera péptica y neoplasias gástricas malignas en humanos. Objetivo: evaluar el uso de la reacción en cadena de la polimerasa para la identificación de H. pylori y sus genotipos en tejidos gástricos con neoplasias malignas embebidos en parafina. Material y Métodos: se estudiaron secciones de 5 bloques de parafina procedentes de 5 pacientes mexicanos con neoplasias gástricas malignas. Se realizaron coloraciones de rutina y especiales de anatomía patológica, así como la técnica de la reacción en cadena de la polimerasa para la detección del microorganismo y sus genotipos. Resultados: la técnica de la reacción en cadena de la polimerasa identificó a este agente infeccioso en todos los bloques analizados en correspondencia con su detección a través de las técnicas histológicas. Esta metodología permitió demostrar una variabilidad genética del patógeno en las muestras analizadas según los genotipos vacA y cagA. Conclusiones: la reacción en cadena de la polimerasa podría ser un método eficaz en la identificación del H. pylori en tejidos gástricos con neoplasias malignas embebidos en parafina. Esta se perfila como una estrategia atractiva para realizar estudios de epidemiología molecular y permitirá establecer posibles asociaciones de genotipos/subtipos del microorganismo con variables clínicas, epidemiológicas y de manejo del paciente.

Introduction: Helicobacter pylori is considered one of the main causal agents of chronic gastritis, peptic ulcer and gastric malignant neoplasms in humans. Objective: to evaluate polymerase chain reaction for identification of Helicobacter pylori and its genotypes in paraffin embedded gastric tissues with malignant neoplasms. Material and Methods: sections of five paraffin blocks from five patients with gastric malignant neoplasms were studied. They were analyzed through routine and special stains of pathological anatomy, as well as the polymerase chain reaction technic for microorganism and genotypes detection. Results: the infectious agent was identified in all of the analyzed blocks through the polymerase chain reaction technic in correspondence with its detection through histologic techniques. This methodology showed a genetic variability of the pathogen in the analyzed samples in respect to vacA and cagA genotypes. Conclusions: the polymerase chain reaction could be an efficacious method for the identification of H. pylori in paraffin embedded gastric tissues with malignant neoplasms. It is projected as an attractive strategy for performing studies of molecular epidemiology and the establishment of possible associations between genotypes/subtypes of the microorganism and clinic or epidemiologic variables, and patient handling.