Presurgical miR‐675‐5p Associates With the Inflammatory Response Following Surgery and Increases Inflammatory Gene Expression In Vitro

Background

A loss of muscle mass and strength is a marker of poor outcomes following surgery. Changes in muscle gene expression following surgery are likely to contribute to postsurgical muscle loss, but the mechanisms controlling these changes are not known. We have previously shown that miRNAs associate with muscle loss in disease, including surgery, so may contribute to changes in gene expression. We analysed the change in muscle gene expression following surgery and compared it to pre‐ and postsurgical miRNA expression. We then analysed the effect of one of these miRNAs (miR‐675‐5p) on gene expression in myoblasts in the presence and absence of an inflammatory stimulus.

Methods

Differential gene expression analysis was performed on RNAseq data from rectus femoris biopsies ( n  = 18 male patients) pre‐ and postsurgery. Gene expression was compared with miRNA expression in the same samples to identify patterns of mRNA expression associated with physiology and miRNA expression. The effect of miR‐675‐5p on gene expression in the presence and absence of TNF‐ α was determined in LHCN myoblasts by RNAseq.

Results

Postsurgery muscle had increased expression of gene sets associated with TNF‐ α signalling (2.7‐fold) and MYC targets (2.7‐fold). Loss of muscle mass and strength associated with MYC target gene sets, but inflammatory gene sets only associated with loss of strength.

Comparing miRNA expression with postsurgery gene expression showed that presurgery miR‐675‐5p expression was most closely associated with postsurgical gene expression, indicating that the presurgery phenotype is important in determining the response to surgery. Presurgery miR‐675‐5p correlated with change in gene expression in response to surgery ( β  = 1.30, p  < 0.001), such that individuals with the highest presurgery miR‐675‐5p had the largest change in gene expression in response to surgery.

In LHCN myoblasts miR‐675‐5p increased CCL2 protein release approximately 1.5‐fold ( p  < 0.001). RNAseq showed that, in the absence of TNF‐ α , transfection with miR‐675‐5p increased the expression of 3422 genes with greatest enrichment of the hypoxia, myogenesis and TNF‐ α signalling via NF‐κB gene sets. In the presence of TNF‐ α , miR‐675‐5p increased the expression of 3403 genes, and the most enriched gene sets were the same suggesting that miR‐675‐5p increases the inflammatory response. miR‐675‐5p also increased the expression of MuRF1 in the presence and absence of TNF‐ α .

Conclusions

Male patients with higher presurgical expression of miR‐675‐5p experienced a larger increase in inflammatory gene expression following surgery. MiR‐675‐5p increased inflammatory gene expression in myoblasts suggesting that it may contribute to the size of the inflammatory response.