[MAFbx expression after free muscle transplantation and its relationship with muscle function]

Abstract

Objective: To study the quantitative changes of ubiquitin ligase MAFbx mRNA and protein expression, muscle atrophy and muscle function following free muscle transplantation and to explore relationships among them.

Methods: Thirty-six female SD rats, SPF grade, weighing (250 +/- 25) g, were used. One hind limb of the rat was randomly selected as experimental side to receive in situ free gracilis muscle transplantation, and the counterlateral hind limb underwent no operation serving as control side. General condition of the rats was observed after operation. Muscle contractive capacity and muscle wet weight maintenance rate of the experimental and the control side were detected 1, 2, 4, 10, 15, and 30 weeks after operation, and 6 rats were killed at each time point. Meanwhile, HE staining was performed to observe muscle fibre cross-sectional area, real-time quantitative PCR was applied to detect relative expression of MAFbx/Atrogin-1 mRNA, and Western blot test was used to observe MAFbx protein expression.

Results: All rats survived till the end of the experiment, all incisions healed well, and no dysfunction occurred in the experimental sides. The value of muscle contractive capacity, muscle wet weight maintenance rate, muscle's maximal force of single contraction, and muscle's maximal force of tetanic contraction in the experimental sides dramatically decreased in the first 4 weeks after operation and increased gradually over 4 to 30 weeks. The MAFbx mRNA expression of the experimental sides peaked and was seven times greater than the control sides 2 weeks after operation, then the value gradually decreased over 15 to 30 weeks after operation and was 1.1 to 1.5 times greater than the control sides, and significant difference was evident between the experimental sides and the control sides at each time point (P < 0.05). Significant difference was evident between the experimental sides and the control sides in terms of MAFbx protein expression of the muscle 1 to 15 weeks after operation according to the Western blot result (P < 0.05), and no significant difference was noted at 30 weeks (P > 0.05). The correlation coefficient between muscle wet weight maintenance rate and muscle's maximal force of single contraction maintenance rate was 0.95, between muscle wet weight maintenance rate and muscle's maximal force of tetanic contraction maintenance rate was 0.75, between muscle fibre cross-sectional area recovery rate and muscle's maximal force of single contraction maintenance rate was 0.93, and between muscle fibre cross-sectional area recovery rate and muscle's maximal force of tetanic contraction maintenance rate was 0.68 (P < 0.05). The correlation coefficient between MAFbx mRNA expression and the parameter of muscle wet weight maintenance rate, muscle fibre cross-sectional area recovery rate, muscle's maximal force of single contraction maintenance rate, and muscle's maximal force of tetanic contraction maintenance rate was -0.62 (P < 0.05), -0.45 (P > 0.05), -0.72 (P < 0.05) and -0.78 (P < 0.05), respectively; the correlation coefficient between MAFbx protein relative expression and the parameter of muscle wet weight maintenance rate, muscle fibre cross-sectional area recovery rate, muscle's maximal force of single contraction maintenance rate, and muscle's maximal force of tetanic contraction maintenance rate was -0.95 (P < 0.05), -0.82 (P < 0.05), -0.89 (P < 0.05), and -0.54 (P > 0.05), respectively.

Conclusion: Decrease of muscle function after transplantation correlates closely with muscle atrophy. The high expression of MAFbx mRNA and protein, especially their persistent increases from 4 to 15 weeks after nerve reinnervation, is a junction between the muscle atrophy and the decrease of muscle function.