Epigallocatechin-3-gallate increases autophagy signaling in resting and unloaded plantaris muscles but selectively suppresses autophagy protein abundance in reloaded muscles of aged rats

Abstract

We have previously found that Epigallocatechin-3-gallate (EGCg), an abundant catechin in green tea, reduced apoptotic signaling and improved muscle recovery in response to reloading after hindlimb suspension (HS). In this study, we investigated if EGCg altered autophagy signaling in skeletal muscle of old rats in response to HS or reloading after HS. Fischer 344 x Brown Norway inbred rats (age 34 mo.) were given 1 ml/day of purified EGCg (50 mg/kg body weight), or the same sample volume of the vehicle by gavage. One group of animals received HS for 14 days and the second group of rats received 14 days of HS, then the HS was removed and they were allowed to recover by ambulating normally around the cage for two weeks. EGCg decreased a small number of autophagy genes in control muscles, but it increased the expression of other autophagy genes (e.g., ATG16L2, SNCA, TM9SF1, Pink1, PIM-2) and HS did not attenuate these increases. HS increased Beclin1, ATG7 and LC3-II/I protein abundance in hindlimb muscles. Relative to vehicle treatment, EGCg treatment had greater ATG12 protein abundance (35.8%, P<0.05), but decreased Beclin1 protein levels (−101.1%, P<0.05) after HS. However, in reloaded muscles, EGCg suppressed Beclin1 and LC3-II/I protein abundance as compared to vehicle treated muscles. EGCg appeared to “prime” autophagy signaling before and enhance autophagy gene expression and protein levels during unloading in muscles of aged rats, perhaps to improve the clearance of damaged organelles. However, EGCg suppressed autophagy signaling after reloading, potentially to increase the recovery of hindlimb muscles mass and function after loading is restored.

Coordinated regulation of skeletal muscle mass and metabolic plasticity during recovery from disuse

“…We showed an acute but transient increase in the autophagosome formation markers Map1lc3b-I, Gabarapl1, and Sqstm1 protein, and a decrease and subsequent increase in the Map1lc3b-II:I ratio that parallels the regulation of late stages of myogenesis. In line with our data, previous studies showed an increase in Sqstm1 protein expression (27) and an increase in Map1lc3b-II:I ratio (31). In addition, White et al (63) reported an unaltered Map1lc3b-II:I ratio after 14 d of RL, potentially reflecting a normalization of autophagy signaling upon completed recovery.…”

The role of SIRT1 in skeletal muscle function and repair of older mice

“…Skeletal muscle lysates were analysed with Western immunoblots as previously described, with slight modifications. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis was run on 4–12% gradient NuPAGE Bis‐Tris gels (ThermoFisher Scientific, Pittsburgh PA, USA) at 120 V for 2 h in NuPAGE MES SDS buffer (ThermoFisher Scientific, Pittsburgh PA, USA).…”

“…Protein concentrations of samples were determined by the Lowry method, and the purity of each fraction was confirmed as reported previously . Forty micrograms of protein were loaded into each well of a 4–12% gradient polyacrylamide gel (Invitrogen, Carlsbad, CA) and separated by routine sodium dodecyl sulfate‐polyacrylamide gel electrophoresis for 1 h at 120 V. The proteins were transferred to a nitrocellulose membrane for 1.5 h at 25 V. Non‐specific protein binding was blocked by incubating the membranes in 5% nonfat milk in Tris‐buffered saline containing 0.05% Tween 20 …”

Experimental Models of Sarcopenia: Bridging Molecular Mechanism and Therapeutic Strategy

“…Herein, we discuss animal models that have been used to study sarcopenia based on the molecular mechanisms of each model (Table 2). [150][151][152] Animal models of sarcopenia with chronic diseases (e.g., cancer cachexia, chronic pulmonary diseases) are not included, we summarize animal models that have been widely used as in vivo models associated sarcopenia. ↑; increase, ↓; decrease, ↔; no change.…”

“…The elevated animal's hindlimbs are adjusted and suspended from the ground, while the forelimbs remain free [149]. HLS can contribute to muscle atrophy by disrupting several cellular processes such as inducing oxidative imbalance [149], mitochondrial dysfunction [163], autophagy [150], and improper protein synthesis/degradation [164]. Hindlimb unloading disrupts the redox balance by inducing oxidative stress and decreasing antioxidant production (e.g., catalase and glutathione peroxidase) [149,165].…”