Formation of a glutathione conjugate from butylated hydroxytoluene by rat liver microsomes

Butylated hydroxytoluene (BHT) was converted to S-(3,5-di-tert-butyl-4-hydroxybenzyl)-glutathione (BHT-glutathione) by rat liver microsomes in the presence of NADPH, molecular oxygen, and glutathione. NADH was far less effective than NADPH and exhibited little synergistic effect when used together with NADPH. Cytochrome P-450 inhibitors, such as SKF 525-A, alpha-naphthoflavone, metyrapone, and carbon monoxide, significantly inhibited BHT-glutathione formation. Liver microsomes from phenobarbital-treated rats catalyzed the formation of BHT-glutathione at a rate that was nine times the rate of adduct formation by control microsomes. No stimulation of BHT-glutathione formation was observed with the addition of liver cytosol fraction to the microsomal incubation mixtures even at low glutathione concentrations. These results support the view that BHT is converted by the cytochrome P-450 monooxygenases to a chemically reactive metabolite, possibly BHT-quinone methide, which forms BHT-glutathione by nonenzymatic conjugation with glutathione.     

A quantitative anatomic characterization of the quadrilateral surface for periacetabular osteotomy

The periacetabular osteotomy described by Ganz et al is used widely, and includes an outward osteotomy from the quadrilateral surface. Because intraarticular extension of the osteotomy can complicate the Ganz osteotomy, it is important to image the margin of the hip. To prevent this complication, and to do this procedure more safely, 32 hemipelves from cadavers were used in the current study. Some landmarks were selected that can be clarified on the quadrilateral surface during the periacetabular osteotomy. The acetabulum was hollowed out using an acetabular reamer of the same size as each femoral head, and the margin of the penetrated hole through the acetabulum was determined using these landmarks. The posterior margin of the hip is located approximately 2 cm anterior to the sciatic notch. The anatomic guidepoint for the osteotomy of the ischium averaged 14 mm inferior to the distal margin of the hip. By clarifying the margin of the hip presumed on the quadrilateral surface in this way, the periacetabular osteotomy can be done more safely, without causing complications such as intraarticular chisel penetration.     

The genetic and antigenic diversity of avian influenza viruses isolated from domestic ducks, muscovy ducks, and chickens in northern and southern Vietnam, 2010–2012

To estimate the prevalence of avian influenza virus infection in Vietnam, surveillance was conducted in domestic and wild birds from households, live-bird markets, slaughtering sites, and bird sanctuaries in Vietnam between October 2010 and October 2012. Of the 4,550 samples collected, 226 influenza A virus isolates were obtained from domestic ducks, muscovy ducks, and chickens. Of these, 25 and 22 H5N1 highly pathogenic avian influenza viruses (HPAIVs) were isolated from apparently healthy domestic ducks in live-bird markets and slaughtering sites in northern and southern Vietnam, respectively. The HA genes of H5 viruses isolated from birds in northern Vietnam phylogenetically belonged to the genetic clade 2.3.2.1 and those in southern Vietnam belonged to the genetic clade 1.1. In addition, 39 H3, 12 H4, 1 H5, 93 H6, 2 H7, 18 H9, 3 H10, and 11 H11 viruses were isolated. Phylogenetic and antigenic analyses of the H6 and H9 viruses revealed that they were closely related to the isolates obtained from domestic poultry in China. Phylogenetic analyses of internal gene segments of these isolates revealed that these viruses were circulating in both domestic and wild birds in Asia and reassortment events had occurred frequently. Therefore, it will be important to continue the surveillance and strict controls over the movement and trade of poultry and poultry products in order to eradicate H5N1 HPAIV from Asia.     

Lateral thrust of anterior cruciate ligament-insufficient knees and posterior cruciate ligament-insufficient knees

Leaving anterior cruciate ligament (ACL) insufficiency and posterior cruciate ligament (PCL) insufficiency untreated frequently leads to osteoarthritis (OA). The purpose of this study was to evaluate dynamically the lateral thrust of ACL-insufficient knees and PCL-insufficient knees, and from the findings investigate the relationship between cruciate ligament insufficiency and OA occurrence. An acceleration sensor was attached to the affected and control anterior tibial tubercles, acting in medial-lateral and perpendicular directions. The lateral thrust immediately after heel strike was measured continuously by a telemeter under stabilised walking conditions. When compared to the contralateral healthy knee, the peak value of lateral acceleration immediately after heel strike was significantly larger in the ACL-insufficient knee; and lateral thrust was increased, but not significantly, in the PCL-insufficient knee. Given that lateral thrust of the knee during walking increases due to ACL or PCL injury, it may be a principal contributor to OA progression.     

Age-dependent change in activities of lysosomal enzymes in rat brain

The age-dependent change in activities of seven lysosomal enzymes (cathepsin D, β-glucuronidase, acid phosphatase, acid/alkaline DNases and acid/alkaline RNases) was studied in four brain regions (cerebrum, hippocampus, pons and cerebellum) of Wistar rats. The activity of cathepsin D was significantly increased with aging in the four regions. The age-dependent change in activities of acid and alkaline DNases showed the characteristic regional difference, and the ratio of acid to alkaline DNases was increased with aging in all regions. Acid RNase showed the lowest activity in 18-month-old rats, and alkaline RNase activity was decreased with aging. The activity of β-glucuronidase was higher in 2-month-old rats in all of the regions studied. Acid phophatase showed no significant age-dependent change except in pons. The study demonstrated that all of the lysosomal enzyme activities do not change in parallel with aging, and that the age-dependent change showed the characteristic regional difference.     

CD4+ T-cells from peripheral blood of a patient with psoriasis recognize keratin 14 peptide but not 'homologous' streptococcal M-protein epitope

Psoriasis has been recognized as an immunologically mediated inflammatory skin disease that has been associated with group A, β-haemolytic streptococcal infections. Notably cross-reactive autoimmune mechanism, which is mediated by T cells reacting to epitopes that are common to streptococcal M-protein and keratin, has been proposed in psoriasis. In order to investigate this possibility, peptides corresponding to M-protein and human epidermal keratin, which share some amino acid sequence between them, were synthesized and tested for their ability to stimulate T-cells of patients with psoriasis. Among five cases examined, we isolated a CD4⁺ T-cell line that recognized the type I keratin (K14)_p168–181 when it was presented by the patient's HLA-DR molecules from a single psoriatic patient, whose MHC allele was HLA-A2/A26, -B27/B16, -DR4/DR8, -DQ8. Further analysis disclosed that the critical peptide recognized by the T-cell line was 10-mer keratin_p171–180 (DLRNKILTAT). However, corresponding M6 protein with homology to K14 did not stimulate the T-cell response and no evidence for cross-reactivity was obtained. The K14-responsive T cell line produced IFN-γ, but little IL-4 when stimulated with irradiated autologous PBMC pulsed with this peptide. Thus, the finding that human epidermal keratin peptide is immunogenic in a psoriasis patient may provide the evidence that T lymphocytes play an important role in the pathogenesis of psoriasis as an autoimmune disorder participated with Th1 like cells. However, the keratin-responsive T cell line was detected in only one of five cases of psoriasis examined, suggesting that such T cell line appears to be not so popular in psoriatic patients. No evidence for cross-reactivity to streptococcal M protein also suggests that the contribution of streptococci may simply be inducing proliferation of various repertoire of T cells (including K14-responsive T cells) possibly through a superantigen-dependent process.     

Gait analysis by floor reaction force in double support period

Eight characteristic points in the double support period were determined by digitalization of floor reaction curves including vertical and fore-aft components. Phases, vertical forces and differentials of the characteristic points in variable step lengths and cadences were studied in normal gaits and pathological gaits of patients with hip, knee or ankle disorders. In normal gait, the order of appearance of characteristic points was constant in spite of changes in step length and cadence. In pathological gait, on the other hand, a reversed order or wide variation was observed in the appearance of characteristic points and differentials. Characteristic point Z1, the maximum of the differential of vertical force in the forelimb, was especially valuable in the study of pathological gait.     

Molecular studies of chronic myelogenous leukemia using the polymerase chain reaction

Thirty-two cases of chronic myelogenous leukemia (CML) were studied to determine whether there was a correlation between the position of the chromosome breakpoint within the breakpoint cluster region (bcr) on chromosome 22 and the type of chimeric mRNA expression. One case with the chromosome breakpoint in zone 2 of the major bcr (Mbcr) and six cases with breakpoints in zone 3 expressed Mbcr exon 2-abl (b2-a) mRNA, and they were in distinguishable at the level of mRNA expression. The remaining ten cases with breakpoints in zone 3 and all ten cases with breakpoints in zone 4 expressed Mbcr exon 3-abl (b3-a) mRNA with or without b2-a mRNA. Three cases with breakpoints in zone 5 expressed b3-a mRNA, and none of these expressed Mbcr exon 4-abl(b4-a) mRNA. The cases with breakpoints in zones 4 or 5 had b3-a mRNA expression indistinguishable from those with breakpoints in zone 3. In two patients, the breakpoint in the bcr could not be determined by Southern hybridization using the 3' bcr probe or the large bcr probe. However, when analyzed for chimeric mRNA expression, both of them exhibited b3-a chimeric mRNA, suggesting the possibility that the entire Mbcr is deleted in the majority of leukemic cells in these patients. These studies indicate that Southern hybridization analysis combined with the polymerase chain reaction assay is a useful approach to understanding the pathologic role of bcr-abl gene recombination and expression in the development of CML.     

Evaluation of adenovirus type 4 and type 7 recombinant hepatitis B vaccines in dogs.

Recombinant hepatitis B virus vaccines based on adenovirus (Ad) vectors Ad4 and Ad7 have been prepared. However, immunogenicity testing of such vaccines in experimental animals is difficult because these human adenoviruses exhibit a highly restricted host range. In this study, the dog was evaluated as a model for screening Ad4- and Ad7-vectored vaccines. Intratracheal inoculation of dogs with Ad4 and Ad7 induced substantial type-specific humoral immune responses that were significantly higher than responses obtained following pharyngeal or oral inoculations. Inoculation of dogs with recombinant Ad7 and Ad4 vaccines expressing hepatitis B surface antigen (HBsAg) elicited large antibody responses to HBsAg (anti-HBs). Substantial secondary anti-HBs responses were produced upon sequential immunizations with heterotypic Ad7 and Ad4 recombinant vaccines. These data thus indicate that the dog is a useful model for evaluating immune responses to vaccines based on Ad4 and Ad7 vectors.