RNAseq Analysis of Treatment-dependent Signaling Changes During Inflammation in a Mouse Cutaneous Wound Healing Model.

Abstract

Background: Despite proven therapeutic effects in inflammatory conditions, the specific mechanisms of phytochemical therapies are not well understood. The transcriptome effects of Tr14 (Traumeel), a multicomponent natural product, and diclofenac, a non-selective cyclooxygenase (COX) inhibitor, were compared in a mouse cutaneous wound healing model to identify both known and novel pathways for the anti-inflammatory effect of plant-derived natural products. Methods: Skin samples from abraded mice were analyzed by single-molecule, amplification-free RNAseq transcript profiling at 7 points between 12-192 hours after injury. Immediately after injury, the wounds were treated with either diclofenac, Tr14, or placebo control (n=7 per group/time). RNAseq levels were compared between treatment and control at each time point using a systems biology approach. Results: At early time points (12-36 hours), both control and Tr14-treated wounds showed marked increase in the inducible COX2 enzyme mRNA, while diclofenac-treated wounds did not. Tr14, in contrast, modulated lipoxygenase transcripts, especially ALOX12/15, and phospholipases involved in arachidonate metabolism. Notably, Tr14 modulated a group of cell-type specific markers, including the T cell receptor, that could be explained by an overarching effect on the type of cells that were recruited into the wound tissue. Conclusions: Tr14 and diclofenac had very different effects on the COX/LOX synthetic pathway after cutaneous wounding. Tr14 allowed normal autoinduction of COX2 mRNA, but suppressed mRNA levels for key enzymes in the leukotriene synthetic pathway. Tr14 appeared to have a broad ‘phytocellular’ effect on the wound transcriptome by altering the balance of cell types present in the wound.