ISCHAEMIC PRECONDDITIONING OF SKELETAL MUSCLE – EFFECT IN A KNEE ARTHROPLASTY POPULATION

Abstract

Introduction: Ischaemic preconditioning (IPC) is a well recognised and powerful phenomenon where a tissue becomes more tolerant to prolonged ischaemia when it is first subjected to short bursts of ischaemia/reperfusion. IPC has been most comprehensively studied in cardiothoracic surgery, to date there has been little use of this powerful phenomenon in orthopaedic surgery. In this study, we report on the first clinical trial of IPC on human skeletal muscle, and show the potential of IPC in orthopaedics using global gene expression analysis.

Methods: After local ethics committee approval and informed consent, patients undergoing primary knee arthroplasty were randomly assigned into an IPC group and a control group. Diabetic patients or patients with an ankle/brachial index of less than 1 were excluded.

The IPC consisted of three five-minute periods of tourniquet insufflation on the operative limb, interrupted by five minute periods of reperfusion. The tourniquet was again insufflated and the operation started. The control group simply had tourniquet insufflation as normal prior to the start of surgery.

Muscle samples were taken from the operative knee of all patients at the immediate onset of surgery (t=0), and again, at one hour into the surgery (t=1). Total RNA was extracted from the muscle samples, and the gene expression profiles were determined using microarray technology.

Results: Comparison of IPC and control samples identified 702 transcripts with differences of ≥1.5-fold in their expression. Of these, 137 were altered at t=0 while 565 were altered at t=1. Amongst these changes was an up-regulation in the expression of a number of heat shock proteins (HSPs) in the IPC group as compared to the control group. Notably, there was up-regulation of the well known cytoprotective/anti-apoptotic gene, HSP72, at one hour post IPC (1.5-fold, p=0.039). There was also up-regulation of important oxidative stress defense genes, such as glutathione-S-transferase (1.6-fold, p = 0.021) and superoxide dismutase 2 (3.6-fold, p= 0.048). Microarray analysis also revealed a down-regulation in the expression of genes involved in metabolism, down-regulation of pro-apoptotic genes and up-regulation of genes necessary for transformation to a hypoxia-tolerant state.

Discussion: We present convincing evidence that IPC is beneficial to human skeletal muscle and for the first time show that IPC of human skeletal muscle works in the clinical setting. In this study, the protective effect of IPC involved a down-regulation in the expression of genes associated with metabolism, and an up-regulation in the expression of genes that provide protection from cell stress, oxidative stress and apoptosis. HSPs, and especially HSP72, have well documented roles in cell stress protection. Their presence has been cited by other studies as an indicator of cell adaptation to stress.