Oxidative metabolism of lindane and its isomers with microsomes from rat liver and house fly abdomen

Lindane and other hexachlorocyclohexane isomers produced 2,4,6-trichlorophenol as the major oxidative metabolite when incubated in the presence of NADPH under aerobic condition. A mechanism for the formation of 2,4,6-trichlorophenol is proposed, which includes direct oxygenation of the cyclohexane ring. The proposed mechanism is supported by data from studies of model chemical reactions of the pentachlorocyclohexanol isomers. Pathways leading to 1,2,4-trichlorobenzene, tetrachlorobenzene isomers, 2,4,5-trichlorophenol, and tetrachlorophenol isomers are discussed, and are considered to include the route through pentachlorocyclohexene and hexachlorocyclohexene. Reductive dechlorination of lindane under anaerobic condition was observed using microsomes and NADPH.     

Decreases in serum apolipoprotein C-III concentration in cows with ethionine-induced fatty liver

To monitor the serum concentration of apolipoprotein C-III (apoC-III), one of the functional apoproteins in lipid metabolism, in cows with ethionine-induced fatty liver, and to investigate the association of apoC-III with liver triglyceride (TG) content and serum biochemical variables, seven nonpregnant nonlactating Holstein cows (3 to 6 years old) were used. Five cows were treated with ethionine, an analogue of methionine, (days 0, 7 and 14). The remaining two controls received saline as the vehicle. Liver TG contents in the treated cows were increased markedly whenever administered, and significant increases were observed at days 14 (666.4%, 85.3 mg/g) and 21 (675.0%, 86.4 mg/g) compared with day 0. In controls, no significant changes in liver TG content and serum biochemical variables were observed during this experiment. The serum apoC-III concentration in the treated cows was decreased drastically after the first administration and fell to the lowest value at day 10 (76.2 microg/ml, 32% of day 0). The apoC-III was significantly (p<0.05) correlated with non-esterified fatty acids (r= -0.526), gamma-glutamyl transpeptidase (r= -0.407), total bilirubin (r= -0.464), positively with apolipoprotein B-100 (apoB-100, r=0.601) and cholesterol ester (r=0.449). Although apoB-100 concentrations were also reduced by the administrations, the concentrations tended to recover smoothly toward the next administration. The distinct difference in change between apoC-III and apoB-100 suggests that apoC-III may be regulated by other pathways, in addition to inhibiting the synthesis of apoproteins by ethionine.     

Inhibition by sulfatide of 21-kDa protein phosphorylation by protein kinase C in cow mammary gland and its reversal by phosphatidylserine

The effect of sulfatide, a sulfated sphingolipid, on phosphorylation of endogenous proteins by protein kinase C (PKC) was examined in cow mammary gland. Several proteins, including 21-kDa, 43-kDa and 56-kDa proteins in the cytosolic fraction, were found to be substrates for PKC by phosphorylation in the absence or presence of the cofactors 1-oleoyl-2-acetyl-sn-glycerol (OAG), phosphatidylserine (PS) and Ca2+. Sulfatide inhibited the 21-kDa phosphorylation, whereas it enhanced the 56-kDa and 43-kDa phosphorylation. Experiments were then conducted to examine whether other sphingolipids, including sphingosine, dihydrosphingosine, ceramides, galactocerebrosides, psychosine and sphingomyelin, modulated phosphorylation of the PKC substrates. Sphingosine, dihydrosphingosine and psychosine did not inhibit the 21-kDa phosphorylation; however, they enhanced the 56-kDa and 43-kDa phosphorylation. Ceramides, galactocerebrosides and sphingomyelin did not inhibit the 21-kDa or enhance the 56-kDa and 43-kDa phosphorylation. The inhibition by sulfatide of the 21-kDa phosphorylation was reversed by excess addition of PS, but not by OAG or Ca2+; whereas the enhancement by sulfatide, as well as sphingosine, dihydrosphingosine and psychosine, of 56-kDa and 43-kDa phosphorylation was not affected by PS, OAG or Ca2+. It is suggested that sulfatide is involved in the regulation of PKC-dependent phosphorylation by modulating the association of PKC substrates, in particular the 21-kDa protein, with membrane phospholipids in cow mammary gland.     

Oxidative metabolism of tetrachlorocyclohexenes,pentachlorocyclohexenes, and hexachlorocyclohexenes with microsomes from rat liver and house fly abdomen

Microsomal mixed-function oxidase systems from rat liver and house fly abdomen effectively metabolized isomers of 3,4,5,6-tetrachlorocyclohexene, 1,3,4,5,6-pentachlorocyclohexene, and 1,2,3,4,5,6-hexachlorocyclohexene to tetrachlorocyclohexenol isomers, 2,4,5,-trichlorophenol, and 2,3,4,6-tetrachlorophenol, respectively. The $\text{(}\text{346}\text{5}\text{)-isomer}$ of pentachlorocyclohexene gave also an abundant amount of pentachlorocyclohexenol isomers. As the metabolites of ($\text{36}\text{45}$)-, ($\text{35}\text{46}$)-, and $\text{(}\text{34}\text{56}\text{)-hexachlorocyclohexene}$, some compounds such as 1,2,4-trichlorobenzene, 1,2,3,4-tetrachlorobenzene, and pentachlorobenzene were more abundantly formed, respectively, than 2,3,4,6-tetrachlorophenol. These oxidative metabolic reactions were shown to mainly proceed via “ene-like” hydroxylation accompanied by double bond migration. Inhibition by CO, piperonyl butoxide, and SKF 525-A suggested that the “ene-like” hydroxylating enzyme was cytochrome P-450 dependent. The formation of an isomer of pentachlorocyclohexenol from $\text{(}\text{36}\text{45}\text{)-hexachlorocyclohexene}$ was also observed, and this reaction was activated by SKF 525-A.     

Outbreak of larval Echinococcus multilocularis infection in Japanese monkey (Macaca fuscata) in a zoo, Hokkaido: western blotting patterns in the infected monkeys

A high prevalence of larval Echinococcus multilocularis (Em) infection was found in zoo primates in Hokkaido, Japan. In October 1997, a Japanese monkey (Macaca fuscata) died and histopathologically diagnosed as alveolar hydatidosis. Serum samples were collected from the remaining Japanese monkeys and examined for antibodies against Em by enzyme-linked immunosorbent assay and western blotting. Serological tests showed 12 more animals of the remaining 57 monkeys were possibly infected. Ultrasonography revealed that nine of these 12 animals had a cystic lesion in the liver. The band patterns of western blotting in the monkeys were very similar to those in human.     

Molecular cloning of the canine c-Met/HGF receptor and its expression in normal and regenerated liver

The c-Met proto-oncogene is the receptor for hepatocyte growth factor (HGF), which is a member of the tyrosine kinase family. Activation of the HGF/c-Met signal pathway leads to cell proliferation, motility, regeneration, and morphogenesis. In this study, the complete nucleotide sequence of complementary DNA (cDNA) of canine c-Met was cloned, and its distribution was determined in tissues. The canine c-Met cDNA clone had an open reading frame of 4419 bp that encoded a putative polypeptide of 1383 amino acids. The c-Met mRNA was expressed in a variety of canine tissues including peripheral blood mononuclear cells (PBMC), bone marrow, liver, kidney, lung, stomach, uterus, testis, thymus, lymph node, small intestine, colon, adrenal gland, thyroid gland, heart, muscle, skin, pancreas, ovary, prostate, spleen, fat, cerebrum, and cerebellum. In addition, the c-Met mRNA expression in normal and regenerated liver was examined. The levels of the mRNA increased 2-fold in regenerated liver compared to that found in normal liver, indicating that c-Met is involved in various functions including remodeling of canine hepatocytes.     

Disulfide Proteome Analysis of Buckwheat Seeds to Screen Putative Allergens

Accumulating evidence suggests a correlation between disulfide bonding and the allergenicity of proteins. Also, a significant characteristic of most food allergens is that they are stable to proteases. We sought to identify putative allergens of buckwheat seed comprehensively by combining a disulfide proteome technique with an in vitro digestibility test. First, a dilute acetic acid fraction of buckwheat seed was found to be rich in disulfide proteins. Internal amino acid sequence analyses of these proteins showed that most of them were known allergens or putative allergens sharing high amino acid sequence similarities to known allergens. Next, the fraction was subjected to in vitro protease digestion, which revealed relatively large fragments that were resistant to prolonged enzymatic digestion. These protease‐resistant fragments contained disulfide bonds that should have protected the potential enzyme cleavage sites by forming compact structures. These results confirm and extend our knowledge of the correlations among the disulfide bonding of proteins, their protease stability, and their allergenicity. Also, these observations suggest a new strategy to identify putative allergens by proteomic approaches as well as to mitigate them.     

Consecutive changes in serum alkaline phosphatase isoenzyme 3 activities in Holstein heifers during the first 18 months of life

This study investigated consecutive fluctuations in serum activities of bone-specific alkaline phosphatase (ALP) isoenzyme 3 (ALP3) in 11 clinically healthy Holstein heifers during the first 18 months of life. ALP3 activities at the first sampling time point after weaning (3 months) were significantly lower than those at multiple time points during the pre-weaning period. Those activities increased from a minimum at 3 months to a peak at 6 months during the post-weaning period. In the anthropometric data, daily body weight and wither height gains appeared to be below the public data at 4 months and 4–5 months, respectively. The data suggested that serum ALP3 activity can be used to monitor skeletal growth of heifers at weaning.     

A Quantitative Trait Locus for Chlorophyll Content and its Association with Leaf Photosynthesis in Rice

Leaf photosynthesis, an important determinant of yield potential in rice, can be estimated from measurements of chlorophyll content. We searched for quantitative trait loci (QTLs) for Soil and Plant Analyzer Development (SPAD) value, an index of leaf chlorophyll content, and assessed their association with leaf photosynthesis. QTL analysis derived from a cross between japonica cultivar Sasanishiki and high-yielding indica cultivar Habataki detected a QTL for SPAD value on chromosome 4. This QTL explained 31% of the total phenotypic variance, and the Habataki allele increased the SPAD value. Chromosomal segment substitution line (CSSL) with the corresponding segment from Habataki had a higher leaf photosynthetic rate and SPAD value than Sasanishiki, suggesting an association between SPAD value and leaf photosynthesis. The CSSL also had a lower specific leaf area (SLA) than Sasanishiki, reflecting its thicker leaves. Substitution mapping under Sasanishiki genetic background demonstrated that QTLs for SPAD value and SLA were co-localized in the 1,798-kb interval. The results suggest that the phenotypes for SPAD value and SLA are controlled by a single locus or two tightly linked loci, and may play an important role in increasing leaf photosynthesis by increasing chlorophyll content or leaf thickness, or both.