Ghassan M Saed1, Adnan R Munkarah, Michael P Diamond. 1. Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Detroit, Michigan 48201, USA. g.saed@wayne.edu
Abstract
OBJECTIVE: To determine whether the COX-2 gene is expressed in human fibroblasts isolated from normal peritoneal and adhesion tissues. DESIGN: Prospective experimental study. SETTING: University medical center. PATIENT(S): Five patients undergoing laparotomy for pelvic pain. Primary cultures of fibroblasts were taken from both peritoneum and adhesion tissues. INTERVENTION(S): Hypoxia treatment of the primary cultured fibroblasts. MAIN OUTCOME MEASURES: We used the multiplex reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry techniques to determine whether COX-2 mRNA and its protein were present in normal peritoneal and adhesion fibroblasts from the same patients. Total RNA was extracted from cultured fibroblasts and subjected to multiplex RT-PCR to detect the presence of COX-2 mRNA in these cells. Cultured fibroblasts from all tissues were also fixed on slides and stained with COX-2 monoclonal antibody labeled with immunofluorescence. RESULT(S): COX-2 mRNA and its protein were absent in normal peritoneal fibroblasts from all five subjects but were present in adhesion fibroblasts from the same patients, as indicated by the multiplex RT-PCR and immunohistochemistry techniques. Hypoxia treatment significantly induced the mRNA and COX-2 protein levels in normal peritoneal fibroblasts to levels seen in adhesion fibroblasts under normoxic conditions. However, hypoxia had no effects on COX-2 expression by adhesion fibroblasts. CONCLUSION(S): Adhesion fibroblasts develop a specific phenotype, an adhesion phenotype, which is in part characterized by the expression of COX-2. The expression of COX-2 mRNA in adhesion fibroblasts and the induction of COX-2 in peritoneal fibroblasts in response to hypoxia indicate a possible inflammatory response. Regulation of COX-2 may alter peritoneal healing and may provide the opportunity to reduce postoperative adhesion development.
OBJECTIVE: To determine whether the COX-2 gene is expressed in human fibroblasts isolated from normal peritoneal and adhesion tissues. DESIGN: Prospective experimental study. SETTING: University medical center. PATIENT(S): Five patients undergoing laparotomy for pelvic pain. Primary cultures of fibroblasts were taken from both peritoneum and adhesion tissues. INTERVENTION(S): Hypoxia treatment of the primary cultured fibroblasts. MAIN OUTCOME MEASURES: We used the multiplex reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry techniques to determine whether COX-2 mRNA and its protein were present in normal peritoneal and adhesion fibroblasts from the same patients. Total RNA was extracted from cultured fibroblasts and subjected to multiplex RT-PCR to detect the presence of COX-2 mRNA in these cells. Cultured fibroblasts from all tissues were also fixed on slides and stained with COX-2 monoclonal antibody labeled with immunofluorescence. RESULT(S): COX-2 mRNA and its protein were absent in normal peritoneal fibroblasts from all five subjects but were present in adhesion fibroblasts from the same patients, as indicated by the multiplex RT-PCR and immunohistochemistry techniques. Hypoxia treatment significantly induced the mRNA and COX-2 protein levels in normal peritoneal fibroblasts to levels seen in adhesion fibroblasts under normoxic conditions. However, hypoxia had no effects on COX-2 expression by adhesion fibroblasts. CONCLUSION(S): Adhesion fibroblasts develop a specific phenotype, an adhesion phenotype, which is in part characterized by the expression of COX-2. The expression of COX-2 mRNA in adhesion fibroblasts and the induction of COX-2 in peritoneal fibroblasts in response to hypoxia indicate a possible inflammatory response. Regulation of COX-2 may alter peritoneal healing and may provide the opportunity to reduce postoperative adhesion development.
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