Molecular regulation of protein catabolism in trauma patients

Abstract

In skeletal muscle, protein degradation is accomplished by several proteolytic systems located at different parts of the cell (1,2). In the lysosomes, cathepsin B, D, E, H and L are mainly involved in the degradation of non-myofibrillar proteins such as membrane-associated proteins and endocytosed proteins. In the cytoplasm, the ubiquitin proteasome system degrades the bulk of myofibrillar protein and many other cellular proteins. Proteins to be degraded are covalently bound to multiple ubiquitin molecules by ubiquitinconjugating enzymes, and the degradation of such ubiquitinprotein complex occurs in the proteosome. Calpains m and ~t are also cytoplasmic proteases with unclear functions. Distinct proteolytic enzymes are also located in the mitochondria. Critically ill patients are characterized by accelerated degradation of both myofibrillar and non-myofibrillar muscle proteins. Recent evidence shows that in patients with severe trauma gene expression for cathepsin D, calpain m and critical components of the ubiquitin proteolytic pathway are simultaneously increased in skeletal muscle (3). However, the relative importance and the regulatory mechanisms of the different muscle proteolytic systems after trauma are not completely understood. Molecular control of proteolytic enzymes involves gene transcription, mRNA maturation, mRNA translation and subsequent protein activation (4). The aim of this study was to develop a methodology to simultaneously evaluate in human skeletal muscle gene expression of critical components of different proteolytic systems and their enzymatic activity. We used a competitive polymerase chain reaction (PCR) methodology (5) to accurately quantitate mRNA levels for cathepsin B and ubiquitin in skeletal muscle of severely traumatized patients. Ubiquitin is encoded in the human genome as a multigene family (6). Among the different ubiquitin genes we chose the UbB polyubiquitin gene which codes for three direct repeats of the ubiquitin sequence (6). Enzymatic activity of cathepsin B was also determined in the same muscle specimens.