JAMDA jo u rn a l h o m e p a g e : w w w. j a m d a. c o m

BackgroundA pharmacologic strategy for age‐related muscle weakness is desired to improve mortality and disability in the elderly. Angiotensin‐converting enzyme 2 (ACE2) cleaves angiotensin II into angiotensin 1‐7, a peptide known to protect against acute and chronic skeletal muscle injury in rodents. Since physiological aging induces muscle weakness via mechanisms distinct from other muscle disorders, the role of ACE2‐angiotensin 1‐7 in age‐related muscle weakness remains undetermined. Here, we investigated whether deletion of ACE2 alters the development of muscle weakness by aging and whether angiotensin 1‐7 reverses muscle weakness in older mice.MethodsAfter periodic measurement of grip strength and running distance in male ACE2KO and wild‐type mice until 24 months of age, we infused angiotensin 1‐7 or vehicle for 4 weeks, and measured grip strength, and excised tissues. Tissues were also excised from younger (3‐month‐old) and middle‐aged (15‐month‐old) mice. Microarray analysis of RNA was performed using tibialis anterior (TA) muscles from middle‐aged mice, and some genes were further tested using RT‐PCR.ResultsGrip strength of ACE2KO mice was reduced at 6 months and was persistently lower than that of wild‐type mice (p < 0.01 at 6, 12, 18, and 24‐month‐old). Running distance of ACE2KO mice was shorter than that of wild‐type mice only at 24 months of age [371 ± 26 vs. 479 ± 24 (m), p < 0.01]. Angiotensin 1‐7 improved grip strength in both types of older mice, with larger effects observed in ACE2KO mice (% increase, 3.8 ± 1.5 and 13.3 ± 3.1 in wild type and ACE2KO mice, respectively). Older, but not middle‐aged ACE2KO mice had higher oxygen consumption assessed by a metabolic cage than age‐matched wild‐type mice. Angiotensin 1‐7 infusion modestly increased oxygen consumption in older mice. There was no difference in a wheel‐running activity or glucose tolerance between ACE2KO and wild‐type mice and between mice with vehicle and angiotensin 1‐7 infusion. Analysis of TA muscles revealed that p16INK4a, a senescence‐associated gene, and central nuclei of myofibers increased in middle‐aged, but not younger ACE2KO mice. p16INK4a and central nuclei increased in TA muscles of older wild‐type mice, but the differences between ACE2KO and wild‐type mice remained significant (p < 0.01). Angiotensin 1‐7 did not alter the expression of p16INK4a or central nuclei in TA muscles of both types of mice. Muscle ACE2 expression of wild‐type mice was the lowest at middle age (2.6 times lower than younger age, p < 0.05).ConclusionsDeletion of ACE2 induced the early manifestation of muscle weakness with signatures of muscle senescence. Angiotensin 1‐7 improved muscle function in older mice, supporting future application of the peptide or its analogues in the treatment of muscle weakness in the elderly population.

The angiotensin-converting enzyme 2 (ACE2)-angiotensin 1-7 (A1-7)-A1-7 receptor (Mas) axis plays a protective role in the renin–angiotensin system (RAS). We recently found that ACE2 knockout (ACE2KO) mice exhibit earlier aging-associated muscle weakness, and that A1-7 alleviates muscle weakness in aging mice. In the present study, we investigated the role of the A1-7-Mas pathway in the effect of ACE2 on physiological aging. Male wild-type, ACE2KO, and Mas knockout (MasKO) mice were subjected to periodical grip strength measurement, followed by administration of A1-7 or vehicle for 4 weeks at 24 months of age. ACE2KO mice exhibited decreased grip strength after 6 months of age, while grip strength of MasKO mice was similar to that of wild-type mice. A1-7 improved grip strength in ACE2KO and wild-type mice, but not in MasKO mice. Muscle fibre size was smaller in ACE2KO mice than that in wild-type and MasKO mice, and increased with A1-7 in ACE2KO and WT mice, but not in MasKO mice. Centrally nucleated fibres (CNFs) and expression of the senescence-associated gene p16INK4a in skeletal muscles were enhanced only in ACE2KO mice and were not altered by A1-7. ACE2KO mice, but not MasKO mice, exhibited thinning of peripheral fat along with increased adipose expression of p16INK4a. A1-7 significantly increased bone volume in wild-type and ACE2KO mice, but not in MasKO mice. Our findings suggest that the impact of ACE2 on physiological aging does not depend on the endogenous production of A1-7 by ACE2, while overactivation of the A1-7-Mas pathway could alleviate sarcopenia and osteoporosis in aged mice.

We hypothesized that low‐intensity endurance exercise might be more effective in preventing cancer cachexia–induced muscle atrophy through both an increase in protein synthesis and a decrease in protein degradation. The purpose of present study was to evaluate the effects and to clarify the mechanism of low‐intensity endurance exercise on cancer cachexia–induced muscle atrophy. Twenty‐four male Wistar rats were randomly divided into 4 groups: control (Cont), Cont plus exercise (Ex), AH130‐induced cancer cachexia (AH130), and AH130 plus Ex. Cancer cachexia was induced by intraperitoneal injections with AH130 Yoshida ascites hepatoma cells; we analyzed the changes in muscle mass and the gene and protein expression levels of major regulators or indicators of skeletal muscle protein degradation and synthesis pathway in the soleus muscles. Low‐intensity exercise inhibited the muscle mass loss through a suppression of the ubiquitin‐proteasome pathway, increased hypoxia‐inducible factor‐1α and phosphorylated AMPK, and inhibited the deactivation of mammalian target of rapamycin pathway in the soleus muscle, which contributed to the prevention of cancer cachexia–induced muscle atrophy. These results suggest that low‐intensity exercise has the potential to become an effective therapeutic intervention for the prevention of cancer cachexia–induced muscle atrophy.—Tanaka, M., Sugimoto, K., Fujimoto, T., Xie, K., Takahashi, T., Akasaka, H., Kurinami, H., Yasunobe, Y., Matsumoto, T., Fujino, H., Rakugi, H. Preventive effects of low‐intensity exercise on cancer cachexia–induced muscle atrophy. FASEB J. 33, 7852–7862 (2019). http://www.fasebj.org

Summary Arrestin-dependent activation of a G-protein-coupled receptor (GPCR) triggers endocytotic internalization of the receptor complex. We analyzed the interaction between the pattern recognition receptor (PRR) lectin-like oxidized low-density lipoprotein (oxLDL) receptor (LOX-1) and the GPCR angiotensin II type 1 receptor (AT1) to report a hitherto unidentified mechanism whereby internalization of the GPCR mediates cellular endocytosis of the PRR ligand. Using genetically modified Chinese hamster ovary cells, we found that oxLDL activates Gαi but not the Gαq pathway of AT1 in the presence of LOX-1. Endocytosis of the oxLDL-LOX-1 complex through the AT1-β-arrestin pathway was demonstrated by real-time imaging of the membrane dynamics of LOX-1 and visualization of endocytosis of oxLDL. Finally, this endocytotic pathway involving GPCR kinases (GRKs), β-arrestin, and clathrin is relevant in accumulating oxLDL in human vascular endothelial cells. Together, our findings indicate that oxLDL activates selective G proteins and β-arrestin-dependent internalization of AT1, whereby the oxLDL-LOX-1 complex undergoes endocytosis.