Insulin Sensitivity Initially Worsens but Later Improves With Aging in Male C57BL/6N Mice.

Aging is believed to induce insulin resistance in humans. However, when and how insulin sensitivity changes with aging remains unclear in both humans and mice. In this study, groups of male C57BL/6N mice at 9-19 weeks (young), 34-67 weeks (mature adult), 84-85 weeks (presenile), and 107-121 weeks of age underwent hyperinsulinemic-euglycemic clamp studies with somatostatin infusion under awake and nonrestrained conditions. The glucose infusion rates for maintaining euglycemia were 18.4 ± 2.9, 5.9 ± 1.3, 20.3 ± 7.2, and 25.3 ± 4.4 mg/kg/min in young, mature adult, presenile, and aged mice, respectively. Thus, compared with young mice, mature adult mice exhibited the expected insulin resistance. In contrast, presenile and aged mice showed significantly higher insulin sensitivity than mature adult mice. These age-related changes were mainly observed in glucose uptake into adipose tissue and skeletal muscle (rates of glucose disappearance were 24.3 ± 2.0, 17.1 ± 1.0, 25.5 ± 5.2, and 31.8 ± 2.9 mg/kg/min in young, mature adult, presenile, and aged mice, respectively). Epididymal fat weight and hepatic triglyceride levels were higher in mature adult mice than those in young and aged mice. Our observations indicate that, in male C57BL/6N mice, insulin resistance appears at the mature adult stage of life but subsequently improves markedly. These alterations in insulin sensitivity are attributable to changes in visceral fat accumulations and age-related factors.

Efficient Detection and Purification of Cell Populations Using Synthetic MicroRNA Switches.

Isolation of specific cell types, including pluripotent stem cell (PSC)-derived populations, is frequently accomplished using cell surface antigens expressed by the cells of interest. However, specific antigens for many cell types have not been identified, making their isolation difficult. Here, we describe an efficient method for purifying cells based on endogenous miRNA activity. We designed synthetic mRNAs encoding a fluorescent protein tagged with sequences targeted by miRNAs expressed by the cells of interest. These miRNA switches control their translation levels by sensing miRNA activities. Several miRNA switches (miR-1-, miR-208a-, and miR-499a-5p-switches) efficiently purified cardiomyocytes differentiated from human PSCs, and switches encoding the apoptosis inducer Bim enriched for cardiomyocytes without cell sorting. This approach is generally applicable, as miR-126-, miR-122-5p-, and miR-375-switches purified endothelial cells, hepatocytes, and insulin-producing cells differentiated from hPSCs, respectively. Thus, miRNA switches can purify cell populations for which other isolation strategies are unavailable.

A novel efficient feeder-free culture system for the derivation of human induced pluripotent stem cells.

In order to apply human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs) to regenerative medicine, the cells should be produced under restricted conditions conforming to GMP guidelines. Since the conventional culture system has some issues that need to be addressed to achieve this goal, we developed a novel culture system. We found that recombinant laminin-511 E8 fragments are useful matrices for maintaining hESCs and hiPSCs when used in combination with a completely xeno-free (Xf) medium, StemFit™. Using this system, hESCs and hiPSCs can be easily and stably passaged by dissociating the cells into single cells for long periods, without any karyotype abnormalities. Human iPSCs could be generated under feeder-free (Ff) and Xf culture systems from human primary fibroblasts and blood cells, and they possessed differentiation abilities. These results indicate that hiPSCs can be generated and maintained under this novel Ff and Xf culture system.