Effects of exercise in the heat on thermoregulation of Japanese and Malaysian males

The effect of low-intensity exercise in the heat on thermoregulation and certain biochemical changes in temperate and tropical subjects under poorly and well-hydrated states was examined. Two VO2max matched groups of subjects consisting of 8 Japanese (JS) and 8 Malaysians (MS) participated in this study under two conditions: poorly-hydrated (no water was given) and well-hydrated (3 mL x Kg(-1) body weight of water was provided at onset of exercise, and the 15th, 35th and 55th min of exercise). The experimental room in both countries was adjusted to a constant level (Ta: 31.6+/-0.03 degrees C, rh: 72.3+/-0.13%). Subjects spent an initial 10 min rest, 60 min of cycling at 40% VO2max and then 40 min recovery in the experimental room. Rectal temperatures (Tre) skin temperatures (Tsk), heart rate (HR), heat-activated sweat glands density (HASG), local sweat rate (M sw-back) and percent dehydration were recorded during the test. Blood samples were analysed for plasma glucose and lactate levels.The extent of dehydration was significantly higher in the combined groups of JS (1.43+/-0.08%) compared to MS (1.15+/-0.05%). During exercise M sw-back was significantly higher in JS compared to MS in the well-hydrated condition. The HASG was significantly more in JS compared to MS at rest and recovery. Tre was higher in MS during the test. Tsk was significantly higher starting at the 5th min of exercise until the end of the recovery period in MS compared to JS.In conclusion, tropical natives have lower M sw-back associated with higher Tsk and Tre during the rest, exercise and recovery periods. However, temperate natives have higher M sw-back and lower Tsk and Tre during experiments in a hot environment. This phenomenon occurs in both poorly-hydrated and well-hydrated states with low intensity exercise. The differences in M sw-back, Tsk and Tre are probably due to a setting of the core temperature at a higher level and enhancement of dry heat loss, which occurred during passive heat exposure.     

Blood group A glycosphingolipid accumulation in the hair of patients with alpha-N-acetylgalactosaminidase deficiency

In the hair of individuals with blood group AB, the level of blood group A glycosphingolipids is much lower than that of blood group B. We hypothesized that in hair, blood group A determinants are converted by alpha-N-acetylgalactosaminidase (alpha-NAGA, E.C.3.2.1.49) to H determinants. To address our hypothesis, the relative amount of ABH glycosphingolipids in hairs and nails of normal subjects, patients with Kanzaki disease, and heterozygous carriers of alpha-NAGA deficiency were analyzed by dot-blotting and enzyme-linked immunosorbent assay. In hair from normal subjects with blood group B, ABH glycosphingolipids consisted of 88% blood group B- and 12% blood group H glycosphingolipids. In blood group A subjects, 14% were group A- and 86% were group H glycosphingolipids. In Kanzaki patients, 81% were blood group A- and 19% were blood group H glycosphingolipids. In 2 alpha-NAGA deficiency carriers, the ABH glycosphingolipids consisted of 67% blood group A- and 33% blood group H glycosphingolipids. These results indicate that blood group A glycosphingolipids are catabolized to H glycosphingolipids by alpha-NAGA, resulting in lower levels of blood group A glycosphingolipids in the hair of normal subjects, and alpha-NAGA deficiency causes accumulation of blood group A glycosphingolipids in the hair of Kanzaki patients. This finding is of clinical relevance because it suggests that hair may be used to diagnose and assess the alpha-NAGA status of individuals.     

Anti-vascular endothelial growth factor gene therapy attenuates lung injury and fibrosis in mice

Vascular endothelial growth factor (VEGF) is an angiogenesis factor with proinflammatory roles. Flt-1 is one of the specific receptors for VEGF, and soluble flt-1 (sflt-1) binds to VEGF and competitively inhibits it from binding to the receptors. We examined the role of VEGF in the pathophysiology of bleomycin-induced pneumopathy in mice, using a new therapeutic strategy that comprises transfection of the sflt-1 gene into skeletal muscles as a biofactory for anti-VEGF therapy. The serum levels of sflt-1 were significantly increased at 3-14 days after the gene transfer. Transfection of the sflt-1 gene at 3 days before or 7 days after the intratracheal instillation of bleomycin decreased the number of inflammatory cells, the protein concentration in the bronchoalveolar lavage fluid and with von Willebrand factor expression at 14 days. Transfection of the sflt-1 gene also attenuated pulmonary fibrosis and apoptosis at 14 days. Since the inflammatory cell infiltration begins at 3 days and is followed by interstitial fibrosis, it is likely that VEGF has important roles as a proinflammatory, a permeability-inducing, and an angiogenesis factor not only in the early inflammatory phase but also in the late fibrotic phase. Furthermore, this method may be beneficial for treating lung injury and fibrosis from the viewpoint of clinical application, since it does not require the use of a viral vector or neutralizing Ab.     

In vivo conversion of cellular prion protein to pathogenic isoforms, as monitored by conformation-specific antibodies

The central event in prion disease is thought to be conformational conversion of the cellular isoform of prion protein (PrP(C)) to the insoluble isoform PrP(Sc). We generated polyclonal and monoclonal antibodies by immunizing PrP(C)-null mice with native PrP(C). All seven monoclonal antibodies (mAbs) immunoprecipitated PrP(C), but they immunoprecipitated PrP(Sc) weakly or not at all, thereby indicating preferential reactivities to PrP(C) in solution. Immunoprecipitation using these mAbs revealed a marked loss of PrP(C) in brains at the terminal stage of illness. Histoblot analyses using these polyclonal antibodies in combination of pretreatment of blots dissociated PrP(C) and PrP(Sc) in situ and consistently demonstrated the decrease of PrP(C) at regions where PrP(Sc) accumulated. Interestingly, same mAbs showed immunohistochemical reactivities to abnormal isoforms. One group of mAbs showed reactivity to materials that accumulated in astrocytes, while the other group did so to amorphous plaques in neuropil. Epitope mapping indicated that single mAbs have reactivities to multiple epitopes, thus implying dual specificities. This suggests the importance of octarepeats as a part of PrP(C)-specific conformation. Our observations support the notion that loss of function of PrP(C) may partly underlie the pathogenesis of prion diseases. The conversion of PrP(C) to PrP(Sc) may involve multiple steps at different sites.     

Purification and characterization of a new fungalysin-like metallopeptidase from the culture filtrate of a plant worm,Nomuraea atypicola

A new protease was purified from the culture filtrate of a plant worm, Nomuraea atypicola. The activity of the protease was suppressed by metalloprotease inhibitors such as EDTA and 1,10-phenanthroline, suggesting that it might be a metalloprotease. Its molecular mass was estimated to be 48 kDa by SDS-PAGE, and its optimal pH and temperature were pH 8.5–9.0 and 40 °C, respectively. The N-terminal amino acid sequence of the metalloprotease was similar to those of fungalysin metallopeptidases of the M36 family from fungi such as Coccidioides posadasii, Pyrenophora tritici-repentis, and Arthroderma gypseum, supporting the idea that it is a fungalysin-like metallopeptidase.     

The Three-Dimensional Human Skin Reconstruct Model: a Tool to Study Normal Skin and Melanoma Progression

Most in vitro studies in experimental skin biology have been done in 2-dimensional (2D) monocultures, while accumulating evidence suggests that cells behave differently when they are grown within a 3D extra-cellular matrix and also interact with other cells (1-5). Mouse models have been broadly utilized to study tissue morphogenesis in vivo. However mouse and human skin have significant differences in cellular architecture and physiology, which makes it difficult to extrapolate mouse studies to humans. Since melanocytes in mouse skin are mostly localized in hair follicles, they have distinct biological properties from those of humans, which locate primarily at the basal layer of the epidermis. The recent development of 3D human skin reconstruct models has enabled the field to investigate cell-matrix and cell-cell interactions between different cell types. The reconstructs consist of a "dermis" with fibroblasts embedded in a collagen I matrix, an "epidermis", which is comprised of stratified, differentiated keratinocytes and a functional basement membrane, which separates epidermis from dermis. Collagen provides scaffolding, nutrient delivery, and potential for cell-to-cell interaction. The 3D skin models incorporating melanocytic cells recapitulate natural features of melanocyte homeostasis and melanoma progression in human skin. As in vivo, melanocytes in reconstructed skin are localized at the basement membrane interspersed with basal layer keratinocytes. Melanoma cells exhibit the same characteristics reflecting the original tumor stage (RGP, VGP and metastatic melanoma cells) in vivo. Recently, dermal stem cells have been identified in the human dermis (6). These multi-potent stem cells can migrate to the epidermis and differentiate to melanocytes.     

An anti-sulfatide antibody O4 immunoprecipitates sulfatide rafts including Fyn, Lyn and the G protein α subunit in rat primary immature oligodendrocytes

The association of sulfatide with specific proteins in oligodendrocytes was examined by co-immunoprecipitation with an anti-sulfatide antibody. Protein kinase activity was detected in precipitates with a monoclonal antibody to sulfatide (O4) from the rat primary immature oligodendrocytes. We conducted in vitro kinase assay of tyrosine phosphorylated proteins of 80, 59, 56, 53 and 40 kDa by gel electrophoresis. Of these proteins, the proteins of 59 kDa and 53/56 kDa were identified as the Src family tyrosine kinases Fyn and Lyn on the basis of their sequential immunoprecipitation with anti-Fyn and anti-Lyn antibodies, respectively. The 40 kDa protein was identified as the α subunit of the heterotrimeric G protein. These observations suggest that O4 immunoprecipitates sulfatide rafts including Fyn, Lyn and the α subunit of the heterotrimeric G protein.