SummaryChanges to innate cells, such as macrophages and myeloidderived suppressor cells (MDSCs), during aging in healthy or tumor-bearing hosts are not well understood. We compared macrophage subpopulations and MDSCs from healthy young (6-8 weeks) C57BL/6J mice to those from healthy geriatric (24-28 months) mice. Spleens, lymph nodes, and bone marrow of geriatric hosts contained significantly more M2 macrophages and MDSCs than their younger counterparts. Peritoneal macrophages from geriatric, but not young, mice co-expressed CD40 and CX3CR1 that are usually mutually exclusively expressed by M1 or M2 macrophages. Nonetheless, macrophages from geriatric mice responded to M1 or M2 stimuli similarly to macrophages from young mice, although they secreted higher levels of TGF-b in response to IL-4. We mimicked conditions that may occur within tumors by exposing macrophages from young vs. geriatric mice to mesothelioma or lung carcinoma tumor cell-derived supernatants. While both supernatants skewed macrophages toward the M2-phenotype regardless of age, only geriatric-derived macrophages produced IL-4, suggesting a more immunosuppressive tumor microenvironment will be established in the elderly. Both geriatric-and young-derived macrophages induced allogeneic T-cell proliferation, regardless of the stimuli used, including tumor supernatant. However, only macrophages from young mice induced T-cell IFN-c production. We examined the potential of an IL-2/agonist anti-CD40 antibody immunotherapy that eradicates large tumors in young hosts to activate macrophages from geriatric mice. IL-2-/CD40-activated macrophages rescued T-cell production of IFN-c in geriatric mice. Therefore, targeting macrophages with IL-2/anti-CD40 antibody may improve innate and T-cell immunity in aging hosts.

This study compared the capacity of young and old male C57Bl/6J mice to exercise with increasing resistance over 10 weeks, and its impact on muscle mass. Young mice (aged 15-25 weeks) were subjected to low (LR) and high (HR) resistance exercise, whereas only LR was used for old mice (107-117 weeks). Weekly patterns of voluntary wheel activity, food consumption and body weights were measured. Running patterns changed over time and with age, with two peaks of activity detected for young, but only one for old mice: speed and distance run was also less for old mice. The mass for six limb muscles was measured at the end of the experiment. The most pronounced increase in mass in response to exercise was for the soleus in young and old mice, and also quadriceps and gastrocnemius in young mice. Soleus and quadriceps muscles were analyzed histologically for myofiber number and size. A striking feature was the many small myofibers in response to exercise in young (but not old) soleus, whereas these were not present after exercise in young or old quadriceps. Overall, there was a striking difference in response to exercise between muscles and this was influenced by age.

Shavlakadze T, Anwari T, Soffe Z, Cozens G, Mark PJ, Gondro C, Grounds MD. Impact of fasting on the rhythmic expression of myogenic and metabolic factors in skeletal muscle of adult mice. Am J Physiol Cell Physiol 305: C26 -C35, 2013. First published April 17, 2013 doi:10.1152/ajpcell.00027.2013.-Circadian rhythms and metabolism are tightly integrated, and rhythmic expression of metabolic factors is common in homeostatic processes. We measured the temporal changes in the expression of myogenic regulatory factors and expression and activity level of molecules involved in protein metabolism in skeletal muscles and livers in mice and examined the impact of fasting. Tissues were collected over 24 h (at zeitgeber times ZT1, ZT5, ZT9, ZT13, ZT17, ZT21, and ZT1 the following day) from adult male C57Bl/6J mice that had been either freely fed or fasted for 24 h. In skeletal muscle, there was a robust rise in the mRNA expression of the myogenic regulatory factors MyoD and myogenin during dark hours which was strongly suppressed by fasting. Circadian pattern was observed for mRNA of MuRF1, Akt1, and ribosomal protein S6 in muscles in fed and fasted mice and for Fbxo32 in fed mice. Activity (phosphorylation) levels of Akt(Ser473) displayed temporal regulation in fasted (but not fed) mice and were high at ZT9. Fasting caused significant reductions in phosphorylation for both Akt and S6 in muscles, indicative of inactivation. Hepatic phosphorylated Akt(Ser473) and S6(Ser235/236) proteins did not exhibit daily rhythms. Fasting significantly reduced hepatic Akt(473) phosphorylation compared with fed levels, although (unlike in muscle) it did not affect S6(Ser235/236) phosphorylation. This in vivo circadian study addresses for the first time the signaling activities of key molecules related to protein turnover and their possible cross-regulation of expression of genes related to protein degradation.Akt; MuRF1; Fbxo32; MyoD; circadian rhythm THE CIRCADIAN CLOCK INFLUENCES most physiological processes in mammals (6, 20, 21) since it synchronizes daily cyclical changes in metabolism, endocrine activity, and behavior with the 24-h cycle of the external environment (8). The principal orchestrator of the circadian clock in mammals is the suprachiasmatic nucleus in the central nervous system that drives behavioral rhythms (i.e., sleeping and feeding) (22). Apart from the central clock, peripheral tissues such as liver, kidney, heart, and skeletal muscle possess local circadian clocks (33,34,50,57,63) that can influence diverse processes including many aspects of metabolism and homeostasis (8, 50).The central clock is primarily entrained by a light-dark cycle, whereas the peripheral clocks can be synchronized by a wide range of factors such as rhythmically secreted hormones (12, 49), changes in the feeding schedule, food metabolites (23), and exercise (61). Central and peripheral circadian clocks are sustained by a rhythmic expression of clock genes through the interaction of positive and negative transcriptional-translational feedback...

Analysis of protein kinase B (AKT) and S6 kinase1 (p70S6K) activity is widely used to assess the efficacy of interventions designed to increase or maintain skeletal muscle mass; these studies are often performed on feed-deprived mice. One problem associated with feed deprivation is that it promotes catabolism, and young or metabolically compromised mice may have less tolerance. The aim of our study was to determine the effect of various times of feed deprivation on the activity of AKT and p70S6K signaling and markers of protein catabolism in young, growing mice compared with adult mice. Young 23-d-old and adult 3-mo-old mice were feed deprived for 8, 10, and 12 h starting at 0700 h. In addition, adult mice were feed deprived for 24 h. AKT(Ser473) phosphorylation decreased by 50 and 76% from fed amounts by 10 and 12 h of feed deprivation, respectively, in young but not adult muscles. In adult muscles, feed deprivation for 24 h reduced AKT(Ser473) phosphorylation by 70%. Significant de-phosphorylation of p70S6K(Thr389) occurred in all feed-deprived young and adult mice. There was an increase in muscle RING-finger protein-1 (Murf1; 133-1245%) and muscle atrophy F-box protein or Atrogin-1 (Fbxo32; 210-2420%) mRNA in all young but not adult groups deprived of feed for 8-12 h, and there was a trend (P = 0.08) toward increased MURF1 associated with the contractile protein-enriched fraction isolated from young muscles of mice feed deprived for 12 h. This study demonstrates that skeletal muscles of young mice respond rapidly to feed deprivation by decreasing AKT activity and upregulating the protein degradation program.

The age-related loss of skeletal muscle mass and function is termed sarcopenia and has been attributed to a decline in concentrations of insulin-like growth factor-1 (IGF-1). We hypothesized that constitutively expressed IGF-1 within skeletal muscles with or without exercise would prevent sarcopenia. Male transgenic mice that overexpress IGF-1 Ea in skeletal muscles were compared with wild-type littermates. Four-month-old mice were assigned to be sedentary, or had access to free-running wheels, until 18 or 28 months of age. In wild-type mice, the mass of the quadriceps muscles was reduced at 28 months and exercise prevented such loss, without affecting the diameter of myofibers. Conversely, increased IGF-1 alone was ineffective, whereas the combination of exercise and IGF-1 was additive in maintaining the diameter of myofibers in the quadriceps muscles. For other muscles, the combination of IGF-1 and exercise was variable and either increased or decreased the mass at 18 months of age, but was ineffective thereafter. Despite an increase in the diameter of myofibers, grip strength was not improved. In conclusion, our data show that exercise and IGF-1 have a modest effect on reducing aged-related wasting of skeletal muscle, but that there is no improvement in muscle function when assessed by grip strength.