Metabolic Regulation of Collagen I in Fibroblasts Isolated from Normal Peritoneum and Adhesions by Dichloroacetic Acid (DCA)
Drs. Michael P. Diamond, Eslam El-Hammady, Rona Wang, Ghassan Saed

Wayne State University

Detroit, Michigan

Introduction: We have previously demonstrated that collagen I, a major component of postoperative adhesions, is differentially regulated in fibroblasts isolated from normal human peritoneum (HPF) and adhesions (ADF). Collagen I mRNA levels are significantly higher in ADF as compared to HBF; this variation is further accentuated by hypoxia.
Purpose: Since adhesions provide a means of supplying oxygen and nutrients to postsurgical ischemic tissue, we sought to examine the role of aerobic metabolism in the differential expression of collagen I. To examine this issue we have utilized a compound, DCA, which stimulates pyruvate dehydrogenase, causing pyruvate to be converted in the Kreb’s cycle rather than into lactate, thereby converting anaerobic to aerobic metabolism. Specifically we have exposed human HPF and ADF cultures to DCA (0,50, and 100 mg/ml) for 24 h under normal and hypoxic (2% O₂) conditions
Methods: Multiplex RT/PCR of collagen I, and b-actin was performed using mRNA extracted from all treatment points (N=3). Analysis of PCR-amplified products was performed by fractionation over a 2% agarose gel followed by ethidium bromide staining of DNA bands. A scanning densometer was used to determine the ratio of intensity of each band relative to b-actin. Densometric analysis of gel bands was performed using the NIH image analysis program.  

Results:                    Collagen mRNA Percent Increase Over Control

Summary: In summary these findings confirm that fibroblasts from adhesions are characterized by excessive collagen I production, which is further accentuated by hypoxia. These observations are extended to show the stimulation of oxidative metabolism by DCA increases collagen I production; in contrast DCA inhibits collagen I production by adhesion fibroblasts as well as under hypoxic conditions in both types of fibroblasts. Thus regulation of metabolic activity of peritoneal cells may provide a target for future interventions for reduction of postoperative adhesions.