We examined the effect of oral intake of pure glucosylceramide derived from konjac extract on skin barrier function evaluated by transepidermal water loss (TEWL) in hairless mice with sodium dodecyl sulfate (SDS)-induced skin roughness. The difference of TEWL between SDS-treated site and untreated sites in the pure glucosylceramide-fed group was significantly lower than that in control group on day 14 of ingestion. We investigated interleukin-1α (IL-1α) production in the hairless mouse skin, and it was significantly lower in the glucosylceramide-fed group than that of control animals. This reduced IL-1α production should contribute to improvement of skin barrier function. To investigate the effect of oral intake of glucosylceramide in human, we conducted a randomized double-blind placebo-controlled study including 100 healthy subjects whose TEWL in cheek was relatively high. As a result, cheek TEWL was significantly lower in the test product group as compared with the control group in weeks 8 and 12 of ingestion (p = 0.023 and p = 0.002 respectively).

RASopathies are autosomal dominant disorders caused by mutations in more than 10 known genes that regulate the RAS/MAPK pathway. Noonan syndrome (NS) is a RASopathy characterized by a distinctive facial appearance, musculoskeletal abnormalities, and congenital heart defects. We have recently identified mutations in RIT1 in patients with NS. To delineate the clinical manifestations in RIT1 mutation-positive patients, we further performed a RIT1 analysis in RASopathy patients and identified 7 RIT1 mutations, including two novel mutations, p.A77S and p.A77T, in 14 of 186 patients. Perinatal abnormalities, including nuchal translucency, fetal hydrops, pleural effusion, or chylothorax and congenital heart defects, are observed in all RIT1 mutation-positive patients. Luciferase assays in NIH 3T3 cells demonstrated that the newly identified RIT1 mutants, including p.A77S and p.A77T, and the previously identified p.F82V, p.T83P, p.Y89H, and p.M90I, enhanced Elk1 transactivation. Genotype-phenotype correlation analyses of previously reported NS patients harboring RIT1, PTPN11, SOS1, RAF1, and KRAS revealed that hypertrophic cardiomyopathy (56 %) was more frequent in patients harboring a RIT1 mutation than in patients harboring PTPN11 (9 %) and SOS1 mutations (10 %). The rates of hypertrophic cardiomyopathy were similar between patients harboring RIT1 mutations and patients harboring RAF1 mutations (75 %). Short stature (52 %) was less prevalent in patients harboring RIT1 mutations than in patients harboring PTPN11 (71 %) and RAF1 (83 %) mutations. These results delineate the clinical manifestations of RIT1 mutation-positive NS patients: high frequencies of hypertrophic cardiomyopathy, atrial septal defects, and pulmonary stenosis; and lower frequencies of ptosis and short stature.

Accumulating evidence shows that post-ischemic inflammation originated by Toll-like receptors (TLR) plays critical roles in ischemic stroke. However, the functions of other innate immune receptors are poorly understood in cerebral ischemia. Macrophage-inducible C-type lectin, Mincle, is one of the innate immune receptor C-type lectin-like receptor (CLR) to response against dying cells. In the present study, we showed that Mincle, its ligand SAP130, and its downstream phospho-Syk/Syk were upregulated after ischemia, and that Mincle is expressed in immune and non-immune cells in the ischemic brains of mice and human. We treated mice with piceatannol, a Syk inhibitor, and consequently the infarct volume and swelling were suppressed by piceatannol. The levels of phospho-Syk, MMP9 and ICAM-1 were downregulated, and the level of Claudin5 was uplegurated in piceatannol-treated groups. These data indicate that innate immune system, such as Mincle and Syk plays a pivotal role in the pathogenesis after the ischemia and reperfusion.

Circulating insulin-like growth factor binding proteins (IGFBPs) play pivotal roles in stabilizing IGFs and regulating their availability to target tissues. In the teleost circulation, three major IGFBPs are typically detected by ligand blotting with molecular masses around 20-25, 28-32 and 40-45 kDa. However, their identity is poorly established and often confused. We previously identified salmon 22-and 41-kDa forms as IGFBP-1 and-2b, respectively. In the present study, we cloned the cDNA of 28-kDa IGFBP from Chinook salmon (Oncorhynchus tshawytscha) as well as rainbow trout (O. mykiss) based on the partial N-terminal amino acid sequence of purified protein and identified it as an ortholog of IGFBP-1. Structural and phylogenetic analyses revealed that the 28-kDa IGFBP is more closely related to human IGFBP-1 and zebrafish IGFBP-1a than the previously identified salmon IGFBP-1 (i.e. 22-kDa IGFBP). We thus named salmon 28-and 22-kDa forms as IGFBP-1a and-1b, respectively. Salmon IGFBP-1a contains a potential PEST region involved in rapid protein turnover and phospholylation sites typically found in mammalian IGFBP-1, although the PEST and phospholylation scores are not as high as those of human IGFBP-1. There was a striking difference in tissue distribution patterns between subtypes; Salmon igfbp-1a was expressed in a variety of tissues while igfbp-1b was almost exclusively expressed in the liver, suggesting that IGFBP-1a has more local actions. Direct seawater exposure (osmotic stress) of Chinook salmon parr caused increases in both IGFBP-1s in plasma, while IGFBP-1b appeared to be more sensitive. The presence of two co-orthologs of IGFBP-1 in the circulation in salmon, and most likely in other telesots, provides a good opportunity to investigate subfunction partitioning of duplicated IGFBP-1 during postnatal growth. Keywords insulin-like growth factor binding protein; salmon; identification; gene duplication; subfunction partitioning circulating 28-32-kDa IGFBP in salmonids. 2. Materials and methods 2.1. Serum collection Serum was collected from spawning male Chinook salmon (Oncorhynchus tshawytscha) in the adult return pond on the University of Washington campus, Seattle, WA, in late October and early November. Fish were anesthetized in MS-222. Blood was withdrawn by syringe from the caudal veins, allowed to clot overnight at 4°C and then centrifuged at 1350 g for 30 min. Serum was stored at-80°C until use. One-year-old rainbow trout (Oncorhynchus mykiss) reared at Nanae Freshwater Experimental Station, Hokkaido University (Kameda, Hokkaido, Japan) were injected with cortisol (Sigma-Aldrich, St. Louis, MO) at a dose of 50 µg/g body weight and blood was withdrawn 8 h after injection. Serum was collected as described above and stored at-30°C until use. The experiment was carried out in accordance with the guidelines of Hokkaido University Field Science Center Animal Care and Use Committee. 2.2. Electrophoresis and Western ligand blotting Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) with a ...

Responses of insulin-like growth factor (IGF)- 12University, 2-9-1 Sakura, Nanae, Kameda-gun, Hokkaido 041-1105, Japan. 32Fasting/re-feeding also affected their mRNA levels in the liver. These results suggest that 33 circulating IGF-I and IGFBP-1b could serve as positive and negative indices of growth, 34respectively, in masu salmon. Different sensitivities of IGBP-1a and IGFBP-1b may be useful to 35assess a broad range of catabolic conditions when they are combined.