Uncoupling Protein-3 (UCP3) mRNA Expression in Reconstituted Human Muscle after Myoblast Transplantation in RAG2−/−/γc/C5− Immunodeficient Mice

Abstract

Uncoupling protein-3 (UCP3), which is expressed abundantly in skeletal muscle, is one of the carrier proteins dissipating the transmitochondrial electrochemical gradient as heat and has therefore been implicated in the regulation of energy metabolism. Myoblasts or differentiated muscle cells in vitro expressed little if any UCP3, compared with the levels detected in biopsies of skeletal muscle. In the present report, we sought to investigate UCP3 mRNA expression in human muscle generated by myoblast transplantation in the skeletal muscle of an immunodeficient mouse model. Time course experiments demonstrated that 7-8 weeks following transplantation fully differentiated human muscle fibers were formed. The presence of differentiated human muscle fibers was assessed by quantitative PCR measurement of the human ␣-actin mRNA together with immunohistochemical staining using specific antibodies for spectrin and the slow adult myosin heavy chain. Interestingly, we found that the expression of UCP3 mRNA was dependant on human muscle differentiation and that the UCP3 mRNA level was comparable with that found in human muscle biopsies. Moreover, the human UCP3 (hUCP3) promoter seems to be fully functional, since triiodothyronine treatment of the mice not only stimulated the mouse UCP3 (mUCP3) mRNA expression but also strongly stimulated the hUCP3 mRNA expression in human fibers formed after myoblast transplantation. To our knowledge, this is the first time that primary myoblasts could be induced to express the UCP3 gene at a level comparable of that found in human muscle fibers.Uncoupling protein-3 (UCP3) 1 is a mitochondrial membrane protein that is predominantly expressed in human and rodent skeletal muscle and brown fat. The gene encoding UCP3 is located on human chromosome 11 and mouse chromosome 7 (1). The UCP3 gene encodes for a protein with 60% homology to the brown fat-specific mitochondrial uncoupling protein UCP1 (2, 3). The sequence similarity between UCP1 and UCP3 suggested to us that UCP3, like UCP1, might be a mitochondrial uncoupling protein involved in adaptive thermogenesis and energy expenditure in muscle. Moreover, biochemical experiments based on the overexpression of UCP3 in yeast or reconstituted transport activity of UCP3 in liposomes have confirmed the uncoupling activity of the UCP3 protein (4, 5). Furthermore, a correlation between the increase in UCP3 protein in skeletal muscle and the nonphosphorylating mitochondrial respiration rates was demonstrated in rats by modulating their metabolic status via triiodothyronine (T3) levels.Despite accumulating evidence that UCP3 plays a role as a mitochondrial uncoupling protein, UCP3 does not appear to be involved in adaptive thermogenesis in rodents in response to cold exposure and diet. Moreover, the role of UCP3 in energy expenditure and uncoupling effect is subject to controversy. First, homozygous UCP3 Ϫ/Ϫ mice lacked the obese phenotype and displayed an unchanged metabolic rate in response to cold exposure, fasting, stress, or T3 (6, 7). Second, tran...

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