Calcineurin and heat-shock proteins modulation in clenbuterol-induced hypertrophied rat skeletal muscles

Abstract

To examine the changes in heat shock proteins (HSPs) and calcineurin (CaN), a calcium/calmodulin regulated protein phosphatase, in hypertrophied rat skeletal muscles, adult male Wistar rats were administered clenbuterol (CLB, 30 mg l(-1) in drinking water), a beta 2-agonist, for 4 weeks. Compared to controls, CLB-treated rats had significantly larger body (10%) and relative (to body weight) soleus (Sol, 16%), plantaris (Plt, 32%) and gastrocnemius (Gast, 27%) weights. Immunohistochemically classified fast fibers were hypertrophied in the Sol (64%), Plt (62%), and deep (d, 70%) and superficial (s, 44%) regions of the Gast, whereas slow fibers were hypertrophied only in the Plt (47%). The percentage of fast fibers in the Sol increased from 10% to 21%. The myosin heavy chain (MHC) isoform composition shifted from slow to fast in the Sol (increase in the percentage of type IIa MHC and de novo synthesis of type IIx MHC) and Gast-d (de novo synthesis of type IIb MHC) and to the faster isoforms in the Plt (increase in the percentage of type IIb MHC). Hsp72 and Hsp90 levels in CLB-treated rats were 52% and 50% lower in the Sol and 44 and 41% lower in the Gast-d, respectively, than in control rats. In control rats, the relative content of CaN was: Sol>Gast-d>Plt>Gast-s, and CLB treatment enhanced the CaN content by 1.4-, 1.2-, 5.0-, and 3.3-fold, respectively. These results indicate that the down-regulation of HSPs in the Sol and Gast-d was closely related to a decrease in the slow phenotype, and that the relative up-regulation of CaN among the muscles/regions was closely related to the selective hypertrophy of fast fibers in the CLB-treated rats.

Slower skeletal muscle phenotypes are critical for constitutive expression of Hsp70 in overloaded rat plantaris muscle

“…1B). Thus these data support previous suggestions (22,24,25) that Hsp70 appears to play another yet-to-be-determined role in skeletal muscle in response to contractile activity with normal recruitment patterns.…”

“…For instance, fast-to-slow type remodeling occurs following the increased contractile activity associated with compensatory overload, as evidenced by the increase in relative expression of MHC I (12,27). This observation is noteworthy since expression of Hsp70 not only tends to parallel slower, more oxidative fiber-type gene expression in unstressed muscles (13,22) but also during changes in expression of this gene program as a consequence of compensatory overload (12,21,23), clenbuterol-induced hypertrophy (24), or altered thyroid hormone status (12). More recently, it has been observed (22,25) that the sudden and sustained increase of contractile activity in the developing rat is not accompanied by an immediate increase in Hsp70.…”

Clenbuterol, a β2-adrenergic agonist, reciprocally alters PGC-1 alpha and RIP140 and reduces fatty acid and pyruvate oxidation in rat skeletal muscle

“…In addition, reductions in mitochondrial proteins (CS, PDHE1␣, ␤-HAD, COXIV) concur with previously reported, clenbuterol-induced changes (14,49,50,53,66), which suggest that skeletal muscle mitochondrial content was reduced. However, this interpretation requires some care since several of these marker proteins can also be reduced within mitochondria (57,68), as was observed for CS and ␤-HAD activity, and for PDHE1␣ protein in the present study.…”

“…This is the first evidence that clenbuterol appears to exert its metabolic effects via simultaneous and reciprocal changes in the nuclear receptor coactivator PGC-1␣ and the nuclear corepressor RIP140. mitochondria; oxidative capacity CLENBUTEROL, A ␤2-ADRENERGIC agonist, induces skeletal and cardiac muscle hypertrophy, due to increased protein synthesis and a concomitant decrease in protein degradation in this tissue (2,46,50). This ␤2-adrenergic agonist is a commonly prescribed bronchodilator treatment for asthmatic patients, while its anabolic properties have led to its illegal use in animal meat production and doping in sports.…”

“…Following a 3-to 7-day acclimation period, rats were randomly divided into two treatment groups. The control group (n ϭ 16) was provided with standard chow and water ad libitum for 3 wk, while the clenbuterol group (n ϭ 16) was provided with standard chow and treated for 3 wk with clenbuterol (Sigma-Aldrich, St. Louis, MO) administered via the drinking water (30 mg clenbuterol/l), as has been done previously by others (50,51,54,61). Body weight and the amount of water consumed were recorded twice each week.…”