De Mexico (approval number DIC/03/311/04/051) and was performed in accordance with

De Mexico (approval number DIC/03/311/04/051) and was performed in accordance with the ethical principles described in the 1964 Declaration of Helsinki. Informed written consent was obtained from all participants prior to their inclusion in the study.epithelium evaluated in the Department of Obstetrics and Gynecology at the Hospital General de Mexico in Mexico City. ?The CC samples were a subset selected from a total of 462 patients with CC who were recruited sequentially from November 2003 through July 2007, which represented BMS-200475 manufacturer approximately 80 of the patients newly diagnosed with CC during this period due to the restrictive inclusion criteria (no previous treatment, incident case, born in Mexico with Mexican ancestry for 2 generations). The selection criteria for the CC subset were based on the availability of a fresh tumor biopsy for RNA extraction with more than 70 tumor cells in the morphological analysis (see below), mostly FIGO stages I/II, and viral type. This subset included 47 samples positive for HPV16 and 22 samples positive for other virus types, including HPV18, 31, 33, 45, 51, 58, and 59. Among them, 54 samples were of squamous cell carcinomas, 14 samples were of adenocarcinomas, and 1 sample was of an adenosquamous carcinoma. The average age of patients with cancer was 48 years (range, 23?8 years; Table S1). All patients received complete clinical evaluations. The tumors of CC patients were staged according to the last international revised protocol for gynecologic cancer [22]. One or two biopsies, conducted under colposcopy examination, were taken from tumors. One biopsy was divided in 2 equal parts, 1 part was fixed in buffered formol for morphological analysis and the other part, together with the second biopsy, was snap-frozen on dry ice and stored at 280uC until analysis. All CC patients were referred for surgery, radiation, chemotherapy, or a combination of these treatments according to the guidelines of the American Cancer Society (see below). Control cervical specimens were obtained from patients undergoing hysterectomy due to myomatosis at the Gynecology Service of the Hospital General de Mexico. They were previously diagnosed with a normal cervix by cytology and colposcopy. Immediately after receiving a cervix SQ 34676 fragment from the operating room, the exocervical and endocervical epitheliums were dissected under a stereoscopic microscope to avoid the stromal cells. The tissues were then snap frozen in liquid nitrogen and stored at 280uC until use. For HPV detection and typing, a scrape from the endocervix and ectocervix was collected with a cytobrush from the patients and controls, the cells were suspended in a vial with extraction buffer, and then stored at 220uC until analysis. Analysis of global gene expression (8,638 genes) was performed in RNAs extracted from 43 fresh tumor biopsies positive for HPV16 and from 12 samples of normal cervical epithelium using the HG-Focus microarray. Global gene expression was validated in 24 samples, including 19 CCs and 5 cervical epithelium controls, by a second high throughput microarray (HG-ST1.0). The 23 genes that showed the greatest deregulation were validated by real time PCR (qRT-PCR) in 44 HPV16-positive CC and 25 control samples. The 6 most differentially expressed genes (CCNB2, CDC20, PRC1, SYCP2, NUSAP1, and CDKN3) were further explored in 29 low-grade cervical intraepithelial neoplasias (CIN1) and 21 high-grade CIN (CIN2/3) to investigate whether they could differe.De Mexico (approval number DIC/03/311/04/051) and was performed in accordance with the ethical principles described in the 1964 Declaration of Helsinki. Informed written consent was obtained from all participants prior to their inclusion in the study.epithelium evaluated in the Department of Obstetrics and Gynecology at the Hospital General de Mexico in Mexico City. ?The CC samples were a subset selected from a total of 462 patients with CC who were recruited sequentially from November 2003 through July 2007, which represented approximately 80 of the patients newly diagnosed with CC during this period due to the restrictive inclusion criteria (no previous treatment, incident case, born in Mexico with Mexican ancestry for 2 generations). The selection criteria for the CC subset were based on the availability of a fresh tumor biopsy for RNA extraction with more than 70 tumor cells in the morphological analysis (see below), mostly FIGO stages I/II, and viral type. This subset included 47 samples positive for HPV16 and 22 samples positive for other virus types, including HPV18, 31, 33, 45, 51, 58, and 59. Among them, 54 samples were of squamous cell carcinomas, 14 samples were of adenocarcinomas, and 1 sample was of an adenosquamous carcinoma. The average age of patients with cancer was 48 years (range, 23?8 years; Table S1). All patients received complete clinical evaluations. The tumors of CC patients were staged according to the last international revised protocol for gynecologic cancer [22]. One or two biopsies, conducted under colposcopy examination, were taken from tumors. One biopsy was divided in 2 equal parts, 1 part was fixed in buffered formol for morphological analysis and the other part, together with the second biopsy, was snap-frozen on dry ice and stored at 280uC until analysis. All CC patients were referred for surgery, radiation, chemotherapy, or a combination of these treatments according to the guidelines of the American Cancer Society (see below). Control cervical specimens were obtained from patients undergoing hysterectomy due to myomatosis at the Gynecology Service of the Hospital General de Mexico. They were previously diagnosed with a normal cervix by cytology and colposcopy. Immediately after receiving a cervix fragment from the operating room, the exocervical and endocervical epitheliums were dissected under a stereoscopic microscope to avoid the stromal cells. The tissues were then snap frozen in liquid nitrogen and stored at 280uC until use. For HPV detection and typing, a scrape from the endocervix and ectocervix was collected with a cytobrush from the patients and controls, the cells were suspended in a vial with extraction buffer, and then stored at 220uC until analysis. Analysis of global gene expression (8,638 genes) was performed in RNAs extracted from 43 fresh tumor biopsies positive for HPV16 and from 12 samples of normal cervical epithelium using the HG-Focus microarray. Global gene expression was validated in 24 samples, including 19 CCs and 5 cervical epithelium controls, by a second high throughput microarray (HG-ST1.0). The 23 genes that showed the greatest deregulation were validated by real time PCR (qRT-PCR) in 44 HPV16-positive CC and 25 control samples. The 6 most differentially expressed genes (CCNB2, CDC20, PRC1, SYCP2, NUSAP1, and CDKN3) were further explored in 29 low-grade cervical intraepithelial neoplasias (CIN1) and 21 high-grade CIN (CIN2/3) to investigate whether they could differe.

Leave a Reply