Screening of extracts of Japanese woods for melanin biosynthesis inhibition

The effects of 120 methanol extracts prepared from bark and heartwood of 69 types of Japanese wood on the melanin production of B16 melanoma cells were examined. The melanin content of B16 melanoma cells was determined spectrophotometrically at 405nm. The extracts were also examined for their effects on cell viability. We found that the methanol extracts of Fagus crenata (buna, wood, 100μg/ml), Sapium sebiferum (Nankinhaze, wood, bark, 10μg/ml), and Zelkova serrata (keyaki, wood, 10μg/ml) greatly inhibited the melanin production of B16 melanoma cells without significant cytotoxicity. However, these extracts did not inhibit tyrosinase activity at the concentration of 100μg/ml. These findings indicate that the depigmenting mechanism of these extracts involves the suppression of some pigmenting signals in stimulating melanogenesis rather than the inhibition of tyrosinase activity. Part of this study was presented at the 53rd Annual Meeting of the Japan Wood Research Society, Fukuoka, Japan, March 2003     

Nickel-catalyzed carbonization of wood for coproduction of functional carbon and fluid fuels I: production of crystallized mesoporous carbon

Japanese larch wood loaded with nickel (1%–4%) alone or with nickel and calcium (0.25%–1.5%) was carbonized at 800°–900°C for 0–120min with a heating rate of 5°–20°C min⁻¹ in a helium flow of 5.8−46.4 ml STP cm⁻² min⁻¹ to examine the influence of these variables on the crystallization of carbon (the formation of T component) and the development of mesoporosity. From the obtained results, reaction conditions suitable for effective production of carbon with the dual functions of adequate electroconductivity and adsorption capacity in liquid phase were established, thereby explaining the factors that govern the process. It was also confirmed that mesopore having a diameter of about 4 nm was selectively produced at the cost of specific (BET) surface area in parallel with the formation of T component. This result provided good insight into how the simultaneous dual function could be realized.     

Combination of serum phosphorylated neurofilament heavy subunit and hyperintensity of intramedullary T2W on magnetic resonance imaging provides better prognostic value of canine thoracolumbar intervertebral disc herniation

The aim of this study was to evaluate the prognostic value of concurrent measurement of serum phosphorylated neurofilament heavy subunit (pNF-H) concentration and intramedullary T2W hyperintensity in paraplegic to paraplegic dogs. Our hypothesis was that concurrent measurement of these would provide a more accurate prediction of functional outcome in dogs with thoracolumbar intervertebral disc herniation (IVDH). A prospective case-control clinical study was designed using 94 dogs with acute onset of thoracolumbar IVDH. The association of serum pNF-H concentration, T2W hyperintensity on sagittal MRI (T2H/L2), deep pain perception and surgical outcome were evaluated with logistic regression analysis after three months for all 94 surgically treated dogs. Sensitivity to predict non-ambulatory outcome was compared among pNF-H and T2H/L2 and combination of both. Logistic regression analysis indicated that serum pNF-H concentration and T2H/L2 were significantly correlated with surgical outcome (P<0.05); however, deep pain perception was not (P=0.41). The results of logistic regression analysis indicated that the odds ratios of unsuccessful long-term outcome were 2.6 for serum pNF-H concentration, 1.9 for T2H/L2 and 2.3 for deep pain sensation. The sensitivity and specificity to predict non-ambulatory outcome for using serum parameter pNF-H>2.6 ng/ml, using T2H/L2 value of>0.84 and using both serum pNF-H and T2H/L2, were 95% and 75.7%, 65% and 86.5%, and 90.0% and 97.5%, respectively. Therefore, combined measurements of serum pNF-H and T2H/L2 might be useful for predicting long-term outcome in dogs with thoracolumbar IVDH.     

Trehalose in glycerol-free freezing extender enhances post-thaw survival of boar spermatozoa

Cryopreservation of boar semen is still considered suboptimal due to lower fertility as compared with fresh samples when glycerol, a permeating cryoprotectant, is used. Trehalose is a non-permeable cryoprotectant and nonreducing disaccharide known to stabilize proteins and biologic membranes. The aim of this study was to evaluate the cryosurvival and in vitro penetrability of boar spermatozoa when glycerol was replaced with trehalose in a freezing extender. Ejaculated Berkshire semen samples were diluted in egg yolk-based freezing extender containing glycerol (100 mM) or trehalose (0, 50, 100, 150, 200 and 250 mM) and cryopreserved using a straw freezing procedure. Thawed samples were analyzed for motility, viability, mitochondrial membrane potential (MMP), and acrosome integrity. In experiment 2, penetrability of spermatozoa cryopreserved with 100 mM glycerol or trehalose was examined. Replacement of cryoprotectant glycerol (100 mM) with trehalose had no effect on sperm viability, but replacing it with 100 mM trehalose improved motility, MMP and acrosome integrity significantly. Sperm motility and MMP were considerably higher in 100 mM trehalose, whereas the acrosome integrity was substantially higher in 100–250 mM trehalose. The in vitro penetration rate was also significantly higher in spermatozoa cryopreserved with trehalose (61.3%) than in those cryopreserved with glycerol (43.6%). In conclusion, 100 mM non-permeable trehalose can be used to replace glycerol, a permeating cryoprotectant, for maintenance of better post-thaw quality of boar spermatozoa.     

Influence of phosphate-buffered sucrose solution on early graft function in feline renal autotransplantation

Graft perfusion with cold heparinized saline has known to induce ischemia and reperfusion injury in feline kidney transplantation. In this study, the effects of phosphate-buffered sucrose solution and heparinized saline solution on early kidney graft function were compared in feline kidney autotransplantation. Perfusion of grafts with or without hypothermic storage with chilled phosphate-buffered sucrose solution prevented ischemia and reperfusion injury despite a very short ischemic time. The results of our study suggest that phosphate-buffered sucrose perfusion and storage solution should be effective to reduce ischemia and reperfusion injury despite a very short ischemic time in feline kidney transplantation.     

Expression Pattern of Vascular Endothelial Growth Factor in Canine Folliculogenesis and its Effect on the Growth and Development of Follicles after Ovarian Organ Culture

In this study, the expressions of VEGF in dog follicles were detected by immunohistochemistry and the effects of VEGF treatment on the primordial to primary follicle transition and on subsequent follicle progression were examined using a dog ovary organ culture system. The frozen‐thawed canine ovarian follicles within slices of ovarian cortical tissue were cultured for 7 and 14 days in presence or absence of VEGF. After culture, the ovaries were fixed, sectioned, stained and counted for morphologic analysis. The results showed that VEGF was expressed in the theca cells of antral follicles and in the granulosa cells nearest the oocyte in preantral follicle but not in granulosa cells of primordial and primary follicles; however, the VEGF protein was expressed in CL. After in vitro culture, VEGF caused a decrease in the number of primordial follicles and concomitant increase in the number of primary follicles that showed growth initiation and reached the secondary and preantral stages of development after 7 and 14 days. Follicular viability was also improved in the presence of VEGF after 7 and 14 days in culture. In conclusion, treatment with VEGF was found to promote the activation of primordial follicle development that could provide an alternative approach to stimulate early follicle development in dogs.