T. S. Lefcourt1, C. Zgheib1, J. Hu1, J. Deacon1, J. Zuk1, J. Xu1, K. W. Liecthy1 1Laboratory For Fetal And Regenerative Biology, Department Of Surgery, School Of Medicine, University Of Colorado Denver Anschutz Medical Campus And Childrens Hospital Colorado,Surgery,Aurora, CO, USA
Introduction: Compared to the adult response to injury, the inflammatory response and scar formation in the neonates is decreased. We have previously shown that increased fibroblast age is associated with increased production of proinflammatory cytokines. Recent evidence suggests that one of the most important regulatory factors of protein production are microRNAs. One such microRNA, microRNA-146a (miR-146a), regulates the inflammatory response by suppressing interleukin-8 (IL-8). Epigenetic regulation such as histone modification has been implicated as a modulatory factor in microRNA expression. We hypothesize that differentially expressed miR-146a in different age groups is under epigenetic control, and that the increased production of proinflammatory cytokines by fibroblasts with increasing age may be due to decreased expression of miR-146a.
Methods: With IRB approval, we isolated fibroblasts from the discarded foreskin of young (<1 month of age) or old (>6 months) males undergoing elective circumcision. Fibroblasts were isolated and expanded no more than 5 passages. Young or old fibroblasts were then cultured for 24 hours with or without two different pan Histone Deacetylase inhibitor (HDACi) conditions: suberanilohydroxamic acid (SAHA), or Apicidin. Total cellular RNA was isolated. Expression of IL-8 and miR-146a was analyzed by quantitative RT-PCR analysis. Expression at baseline and following HDACi treatment was compared to vehicle (DMSO) control.
Results: Young fibroblasts had significantly decreased IL-8 gene expression compared to the older fibroblasts. Young fibroblasts also had significantly increased miR-146a expression compared to older fibroblasts. Treatment with an HDACi resulted in an increase in miR-146a expression in both young and old fibroblasts. In addition, expression of IL-8 was significantly decreased in the older fibroblasts under both HDACi treatment conditions (SAHA p < 0.01, Apicidin p < 0.05).
Conclusion: These findings identify age specific molecular mechanism(s) that modulate the inflammatory response. Increased inflammation and subsequent scar tissue formation with increasing age may be due to decreased expression of miR-146a. Our results also indicated that anti-inflammatory miR-146a is differentially expressed with age, and that the mechanism governing this differential expression may be under epigenetic control. Further studies are warranted to examine miR-146a as a potential therapeutic target to modulate the inflammatory response, and decrease the complications of fibrosis and scar formation.