Yasuto Suzuki

The Role of Elastases Secreted by Fibroblasts in Wrinkle Formation: Implication Through Selective Inhibition of Elastase Activity ¶

Abstract We have previously demonstrated that decreases in skin elasticity, accompanied by increases in the tortuosity of elastic fibers, are important early events in wrinkle formation. In order to study the role of elastases in the degeneration of elastic fibers during wrinkle formation we examined the effects of an inhibitor of skin fibroblast elastase, N-phenethylphosphonyl-l-leucyl-l-tryptophane (NPLT), on wrinkle formation in hairless mice skin following UV irradiation. Dorsal skins of hairless mice were exposed daily to UV light for 18 weeks at doses of 65–95 mJ/cm2 and treated topically with 100 μL of 1 mM NPLT immediately after each UV irradiation. Wrinkles on dorsal skins were evaluated from week 6 through week 18. The daily exposure of mouse skin to UV light with less than 1 minimal erythemal dose significantly enhanced the activity of elastase in the exposed skin by week 4, and the elevated levels of elastase activity were significantly reduced by the in vitro incubation with NPLT in a dose-dependent manner to a level similar to that in unexposed mice skin, indicating that NPLT can efficiently inhibit the UV-inducible elastase activity. Topical application of NPLT significantly suppressed wrinkle formation when compared with vehicle controls by week 15 of treatment (P < 0.05). Histochemistry of elastic fibers with Orcein staining demonstrated that there were no obvious decreases of the fine elastic fibers in UV-exposed NPLT-treated skin in contrast to their marked decreases in the UV-exposed vehicle-treated skin. These findings suggest that skin fibroblast elastase plays a decisive role in wrinkle formation through the degeneration of elastic fiber.

Coffee polyphenols suppress diet-induced body fat accumulation by downregulating SREBP-1c and related molecules in C57BL/6J mice

The prevalence of obesity is increasing globally, and obesity is a major risk factor for type 2 diabetes and cardiovascular disease. We investigated the effects of coffee polyphenols (CPP), which are abundant in coffee and consumed worldwide, on diet-induced body fat accumulation. C57BL/6J mice were fed either a control diet, a high-fat diet, or a high-fat diet supplemented with 0.5 to 1.0% CPP for 2-15 wk. Supplementation with CPP significantly reduced body weight gain, abdominal and liver fat accumulation, and infiltration of macrophages into adipose tissues. Energy expenditure evaluated by indirect calorimetry was significantly increased in CPP-fed mice. The mRNA levels of sterol regulatory element-binding protein (SREBP)-1c, acetyl-CoA carboxylase-1 and -2, stearoyl-CoA desaturase-1, and pyruvate dehydrogenase kinase-4 in the liver were significantly lower in CPP-fed mice than in high-fat control mice. Similarly, CPP suppressed the expression of these molecules in Hepa 1-6 cells, concomitant with an increase in microRNA-122. Structure-activity relationship studies of nine quinic acid derivatives isolated from CPP in Hepa 1-6 cells suggested that mono- or di-caffeoyl quinic acids (CQA) are active substances in the beneficial effects of CPP. Furthermore, CPP and 5-CQA decreased the nuclear active form of SREBP-1, acetyl-CoA carboxylase activity, and cellular malonyl-CoA levels. These findings indicate that CPP enhances energy metabolism and reduces lipogenesis by downregulating SREBP-1c and related molecules, which leads to the suppression of body fat accumulation.

Coffee polyphenols modulate whole-body substrate oxidation and suppress postprandial hyperglycaemia, hyperinsulinaemia and hyperlipidaemia

Postprandial energy metabolism, including postprandial hyperglycaemia, hyperinsulinaemia and hyperlipidaemia, is related to the risk for developing obesity and CVD. In the present study, we examined the effects of polyphenols purified from coffee (coffee polyphenols (CPP)) on postprandial carbohydrate and lipid metabolism, and whole-body substrate oxidation in C57BL/6J mice. In mice that co-ingested CPP with a lipid-carbohydrate (sucrose or starch)-mixed emulsion, the respiratory quotient determined by indirect calorimetry was significantly lower than that in control mice, whereas there was no difference in VO2 (energy expenditure), indicating that CPP modulates postprandial energy partitioning. CPP also suppressed postprandial increases in plasma glucose, insulin, glucose-dependent insulinotropic polypeptide and TAG levels. Inhibition experiments on digestive enzymes revealed that CPP inhibits maltase and sucrase, and, to a lesser extent, pancreatic lipase in a concentration-dependent manner. Among the nine kinds of polyphenols (caffeoyl quinic acids (CQA), di-CQA, feruloyl quinic acids (FQA)) contained in CPP, di-CQA showed more potent inhibitory activity than CQA or FQA on these digestive enzymes, suggesting a predominant role of di-CQA in the regulation of postprandial energy metabolism. These results suggest that CPP modulates whole-body substrate oxidation by suppressing postprandial hyperglycaemia and hyperinsulinaemia, and these effects are mediated by inhibiting digestive enzymes.

Structure-based design of dipeptide derivatives for the human neutral endopeptidase.

Neutral endopeptidase (NEP) plays a key role in the metabolic inactivation of various bioactive peptides such as atrial natriuretic peptide (ANP), endothelins, and enkephalins. Furthermore, NEP is known to work as elastase in skin fibroblast. Therefore, effective inhibitors of NEP offer significant therapeutic interest as antihypertensives, analgesics, and skin anti-aging agents. Recently, the X-ray crystal structure of human NEP complexed with phosphoramidon has been reported and provided insights into the active site specificity of NEP. Here, we designed new inhibitors by using in silico molecular modeling and synthesized them by short steps. Expectedly, we found highly effective inhibitors with sub-nanomolar levels of IC(50) values. These results indicate that our structure-based molecular designing program is useful for obtaining novel NEP inhibitors. Furthermore, these inhibitors may be attractive leads for the generation of new pharmaceuticals for NEP-related diseases.