Two-staged pancreatoduodenectomy with external drainage of pancreatic juice and omental graft technique.
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Aged↗
Biomedical subjects
Y Seyama
Publications and source records attributed to Y Seyama.
Sexual differences in branched chain amino acid metabolism into fatty acids and cholesterol in Harderian gland of golden hamster.
The Harderian gland of golden hamster (Mesocricetus auratus) secretes copious lipids, most of which is 1-alkyl-2,3-diacylglycerol (ADG). We previously reported that the composition of ADG shows marked sexual dimorphism [Seyama et al. (1995) J. Biochem. 117, 661-670]. Male ADG contains only straight chain alkyl and acyl groups, but female ADG contains a lot of branched chain ones too. In this study, we investigated the metabolism of branched chain amino acids (BCAAs) and analyzed the incorporation of the metabolites into lipids in the Harderian gland. Golden hamsters were injected intraperitoneally with [U-14C]BCAAs, and Harderian glands were obtained at 3, 6, 9, and 24 h after injection. Lipids were then extracted from the glands and analyzed. Thin layer chromatography revealed that the ADG was labeled in both sexes, but the profile depended on the sex. The cholesterol fraction was labeled only in the male gland. The alkyl and acyl groups of ADG were subjected to radio-gas liquid chromatography. As for the alkyl groups, radioactivity was detected in straight-C16 and -C18 chains in males, while branched-C17 and -C19 chains were labeled in females. As for the acyl groups, straight-C14, -C15, and -C16 chains were labeled in males, while in females, branched-C17 and -C19 chains were labeled as well as a straight-C16 chain. These results suggest that the BCAA metabolism should be regulated as to the sex at the step of branched chain acyl-CoA degradation in the Harderian gland of golden hamster, which causes the sexual dimorphism in the lipid composition in this gland.
Amino Acids↗
Genetic analysis enables definite and rapid diagnosis of cerebrotendinous xanthomatosis.
Mutations in the sterol 27-hydroxylase gene (CYP27) cause cerebrotendinous xanthomatosis (CTX). Early diagnosis of CTX is crucial because treatment with chenodeoxycholic acid can prevent or reverse some of the neurologic disability associated with the disease. We report the identification of three types of mutations (Arg441Trp, Arg372Gln, and Arg441Gln) in the CYP27 gene in five patients with suspected CTX from four unrelated families by restriction endonuclease analysis.
Adult↗
Comparison of elastolytic activity between experimental aneurysm and experimental diabetes mellitus.
In order to clarify the degradation of elastin under abnormal conditions, we examined the aortic elastolytic activity in rat experimental diabetes mellitus induced by treatment with streptozotocin and in rat experimental aneurysm induced by treatment with an inhibitor of lysyloxidase (beta-aminopropionitrile: BAPN). Measurement of the aortic elastolytic activity used 14C-labeled elastin as the substrate, and the determined value was compared with the aortic lysosomal enzyme (acid phosphatase) activity. In the case of experimental diabetes, the aortic elastolytic activity was not changed, but the aortic acid phosphatase activity was significantly increased compared with the control. In the case of the experimental aneurysm, the aortic elastolytic activity measured after 2 and 3 weeks was increased compared with each control. There was a negative correlation (r=-0.435, n=36) between the elastolytic activity and the cross-linking (desmosine) content in the aorta. The ratio of elastolytic activity to desmosine content was significantly increased compared with the control. Therefore, the degradation of aortic elastin in the experimental aneurysm was caused by elastase, not by lysosomal enzymes. We concluded that an elastase-like enzyme mainly contributed to the degradation of elastin in the experimental aneurysm since the inhibitory pattern of the elastolytic activity in the experimental aneurysm was similar to that of pancreatic elastase.
Animals↗
Alternative pre-mRNA splicing of the sterol 27-hydroxylase gene (CYP 27) caused by a G to A mutation at the last nucleotide of exon 6 in a patient with cerebrotendinous xanthomatosis (CTX).
A recently identified G to A mutation at the last nucleotide of exon 6 of the sterol 27-hydroxylase gene (CYP 27) in a patient with cerebrotendinous xanthomatosis (CTX) was shown here to cause alternative pre-mRNA splicing of the gene. Northern blot analysis of the patient's RNA revealed a broadened band in the human CYP 27 mRNA region compared to that of the normal sample, indicating that there may exist differently spliced mRNA species in the patient. RT-PCR produced three fragments in the patient, one was full-length size and the other two were of smaller sizes. Sequence analysis confirmed that the nucleotide of the full-length size was identical to that of the normal full-length cDNA, except for the G to A mutation at codon 362, which corresponds to the last nucleotide of exon 6. One of the smaller size species lacked exon 6 and the other was absent from the 3' terminal 88 bp of exon 6 due to the use of an activated cryptic 5' splice site in exon 6. The correctly spliced mRNA harbouring the G to A mutation was responsible for the deficiency of the sterol 27-hydroxylase activity, as confirmed by transfection experiment. Transfection of constructed minigenes, with or without the mutation, showed that correctly spliced mRNA was observed in the normal minigene while the mutant minigene was differently spliced. This is the first report of a G to A substitution at the last nucleotide of an exon resulting in both normal and abnormal pre-mRNA splicings, including exon skipping and activating of a coding region cryptic 5' splice site. The results reveal a new molecular basis for the CTX and provide information on aberrant splicing of pre-mRNA in multi-exon genes.
Adult↗
CPP32 activation during dolichyl phosphate-induced apoptosis in U937 leukemia cells.
Treatment of U937 cells with dolichyl phosphate led to an increase in the activity of the ICE family protease CPP32, accompanied with cleavage of pre-CPP32 to generate p17. Peptide inhibitors YVAD-cmk and Z-Asp-CH2-DCB (specific to ICE) and DEVD-CHO (specific to CPP32) blocked the dolichyl phosphate-induced apoptosis. The dolichyl phosphate-induced increase of CPP32 activity was inhibited by adenylate cyclase inhibitors, SQ 22536 and 2',5'-dideoxyadenosine. Dolichyl phosphate caused a transient increase of intracellular cAMP concentration. The results suggest that modulation of cAMP synthesis due to the stimulation of adenylate cyclase by dolichyl phosphate plays a critical role in CPP32 activation and apoptosis.
Adenine↗
Ornithine decarboxylase overexpression in mouse 10T1/2 fibroblasts: cellular transformation and invasion.
BACKGROUND: Ornithine decarboxylase (ODC) plays a pivotal role in the synthesis of polyamines, a group of chemical compounds that are essential for cell growth. Recent reports have shown that ODC overexpression may be involved in malignant transformation of immortalized NIH 3T3 cells. We have demonstrated that ODC-overproducing mouse breast cancer cells are more invasive in vitro than control cells. However, little information is available concerning the relationship between ODC overexpression, tumor invasion, and metastasis and the signal transduction pathways involved in ODC-induced transformation and invasion. PURPOSE: Our purpose was twofold: 1) to determine whether ODC overexpression is directly involved in tumor cell invasion and 2) to determine whether ODC overexpression induces mitogen-activated protein (MAP) kinase activities that are associated with cell growth and transformation. METHODS: We transfected C3H clone 8 mouse 10T1/2 fibroblasts with an expression vector that carries a complementary DNA encoding rat ODC. Neomycin-resistant cells that overproduced ODC (4-6.5 times the control levels) were isolated. The transformed phenotype of these cells was determined by assessing colony formation and anchorage-independent growth in soft agar. The invasiveness of the cells was studied by means of an invasion assay that used Matrigel-coated filters in Boyden chambers. The MAP kinase activity of the cells was assayed by an in-gel kinase assay, using myelin basic protein as the substrate. RESULTS: Overexpression of ODC induced not only cell transformation and anchorage-independent growth in soft agar but also invasiveness through a Matrigel-coated filter. The ODC-overproducing transfectants showed enhanced MAP kinase activity that paralleled the magnitude of cell invasiveness. CONCLUSIONS: ODC plays a pivotal role not only in cell transformation but also in cancer cell invasion. ODC overexpression enhanced MAP kinase activity. IMPLICATIONS: Our results demonstrate a connection between the polyamine/ODC and the MAP kinase signal transduction pathways and suggest that MAP kinase may play a pivotal role in ODC-induced cell transformation and invasion.
Animals↗
Novel regulation of delta-aminolevulinate synthase in the rat harderian gland.
The mode of expression of delta-aminolevulinate synthase (ALAS), as well as that of mRNAs for other heme pathway enzymes, was examined in the rat Harderian gland. Northern blot and in situ hybridization analyses demonstrated that the non-specific ALAS (ALAS-N) mRNA is highly expressed in this tissue, whereas the erythroid-specific ALAS (ALAS-E) mRNA is not. Immunoblot analysis of ALAS also confirmed this finding at the protein level. ALAS-N mRNA was maximally induced in the Harderian gland and was not increased further by treatment of animals with 2-allyl-2-isopropylacetamide (AIA). The levels of mRNAs for other heme pathway enzymes, i.e., delta-aminolevulinate dehydratase, porphobilinogen deaminase, uroporphyrinogen decarboxylase, and coproporphyrinogen oxidase, also were increased markedly in the Harderian gland and not influenced by AIA treatment. The level of ferrochelatase (FeC) mRNA in the gland was, however, lower than that in the liver. The gland contained an extremely high level of protoporphyrin, while heme was undetectable. Microsomal heme oxygenase-1 (HO-1) mRNA levels were significantly higher in the Harderian gland than in the liver. When isolated glands were incubated with hemin in vitro in organ cultures, the level of HO-1 mRNA was increased, whereas the ALAS-N mRNA level was not. These findings indicate that markedly elevated levels of protoporphyrin and extremely low levels of heme in the Harderian gland are the results of both decreased expression of FeC and markedly increased expression of ALAS-N and HO-1. The constitutive expression of the ALAS-N gene in the Harderian gland suggests a novel transcriptional control mechanism of this gene.
5-Aminolevulinate Synthetase↗
Effect of the photoperiod in modulating the androgenic control of 1-alkyl-2,3-diacylglycerol composition in the harderian gland of the golden hamster, Mesocricetus auratus.
The golden hamster Harderian gland produces a lipid secretion consisting mainly of 1-alkyl-2,3-diacylglycerol. We investigated the composition of alkyl and acyl groups in male and female hamster Harderian alkyldiacylglycerol, in animals kept in long and short photoperiods. Female hamsters in long days have alkyl groups with long saturated straight chains (C18:0 and C20:0) and methyl-branched chains (even and odd chain length iso-branched and odd chain length anteiso-branched chains). Acyl groups in females in long days are mostly long straight chains (C16:0) and methyl-branched chains. In females, short photoperiods led to reductions in the proportions of methyl-branched chains and changes in the proportions of straight chain alkyl and acyl groups; these changes were prevented by pinealectomy. Male hamsters with intact gonads, maintained in long days, had no methyl-branched chain alkyl or acyl groups; saturated straight chains were generally shorter than those of females and the odd chain length saturated C15:0 acyl group was common. Short photoperiods did not significantly alter the composition of male alkyldiacylglycerol. Castrated male hamsters in long days showed a distinctively female phenotype, with long straight chains and methyl-branched alkyl and acyl groups. Castrated males in short days showed a mixture of male and female characteristics: shorter straight chain alkyl and acyl groups, a total absence of methyl-branched alkyl groups, and the presence of methyl-branched acyl groups. These results and those of other studies suggest that testosterone controls the enzymes isovaleryl acyl-CoA dehydrogenase and 2-methyl branched-chain acyl-CoA dehydrogenase; in the absence of these enzymes, the primers for the synthesis of methyl-branched chain fatty acids are produced. Our results indicate that this control is modulated by short photoperiods (perhaps due to reduced prolactin levels). It is also suggested that characteristics of male-type alkyldiacylglycerol are better adapted to conditions of autumn and winter than are those of female-type alkyldiacylglycerol.
Animals↗
Anti-alpha3 integrin antibody induces the activated form of matrix metalloprotease-2 (MMP-2) with concomitant stimulation of invasion through matrigel by human rhabdomyosarcoma cells.
Proteolytic enzymes, such as matrix metalloproteases (MMPs), play a pivotal role in cancer invasion and metastasis. Invasive human rhabdomyosarcoma cells (RD) secreted proMMP-2 (72-kDa progelatinase). We found that anti-alpha3 and -alpha2 integrin antibodies induced the activated form of MMP-2 and enhanced proMMP-2 secretion by RD cells. The effect of anti-alpha2 integrin antibody was less prominent than that seen with anti-alpha3 integrin antibody. Moreover, we have found that anti-alpha3 and -alpha2 integrin antibodies enhanced RD-cell invasion through matrigel (reconstituted basement membrane) by 2.6- and 2.0-fold respectively this process was abrogated by neutralizing antibody to MMP-2. These data suggest that signaling events induced by anti-alpha3 integrin antibody may be involved in RD-cell invasion as a result of modulation of matrix-metalloprotease expression.
Antibodies, Anti-Idiotypic↗
Enhancement of apoptosis in developing chick neural retina cells by basic fibroblast growth factor.
To evaluate the role of various growth factors in naturally occurring cell death during development of the neural retina, we examined the effects of such factors on the nuclear morphology and the size of DNA in cultured chick embryonic neural retina cells. Basic fibroblast growth factor (bFGF) increased internucleosomal cleavage of DNA and nuclear fragmentation in a time- and dose-dependent manner. The effect was inhibited by anti-bFGF antibody, suramin, and cycloheximide. Epidermal growth factor, platelet-derived growth factor, nerve growth factor, tumor necrosis factor-alpha, and dexamethasone had no effect. These results provide evidence that bFGF may eventually act as a lethal factor inducing apoptotic cell death during the development of the neural retina in chick embryo.
Animals↗
Effects of monensin on tropoelastin metabolism in vascular smooth muscle cells: monensin causes intracellular degradation of accumulated tropoelastin.
Treatment with 80 nM monensin for 48 h resulted in impairment of tropoelastin secretion during the initial 6 h and subsequent reduction of tropoelastin synthesis from 18 to 48 h to one-tenth. The steady state level of tropoelastin mRNA started to decrease at 6 h and reached one-fifth of the control level by 48 h. A pulse-chase experiment after 24 h monensin treatment demonstrated that half of the accumulated tropoelastin in the cells was rapidly degraded within 120 min. These results indicate that the marked reduction in tropoelastin synthesis from 6 to 48 h treatment may be caused by both a reduction in the tropoelastin mRNA level and accelerated intracellular degradation of tropoelastin. Thus, monensin modulates tropoelastin expression through pretranslational and posttranslational mechanisms.
Animals↗
Cultured human keratinocytes express tropoelastin.
We detected elastin mRNA in cultured normal human keratinocytes by RNase protection assay. The content of elastin mRNA was estimated at approximately one-twentieth of that of cultured skin fibroblasts. Tropoelastin polypeptide with a molecular weight of 68 kDa was detected in the preparation of culture medium of normal human keratinocytes by western blot assays using anti-tropoelastin antibody. Immunohistochemical studies also demonstrated positive staining in cultured normal human keratinocytes as well as in skin fibroblasts. The expression of elastin by normal human keratinocytes was found to reach a maximum level at the quiescent phase of keratinocyte growth. When normal human keratinocytes were cultured on tropoelastin-coated dishes, their growth potential was greatly suppressed compared with other matrix protein-coated dishes. These results suggest that cultured normal human keratinocytes can actively synthesize elastin and that keratinocyte elastin may act as a growth-regulator for keratinocytes.
Blotting, Northern↗
[Properties in elastin of different arteries: cross-links, hydrophobicity and fibronectin in elastin fraction].
In order to evaluate the quantitative and qualitative changes of elastin in different arteries, biochemical properties were measured; that is, elastin content, isodesmosine (ID) as cross-links content, free thiol content (free SH/total SH), hydrophobic properties and coexisting fibronectin (FN) content in elastin fraction from different arteries, correlation coefficients of these properties and these their effect with aging were also calculated. The results from different arteries in regard to changes in elastin can be summarized as follows: 1. The basilar artery in the quantitative changes of elastin. There was no correlation of these parameters in the basial artery from the other arteries. 2. The content of ID in the coronary artery was lower than that of the other arteries. The level of ID was negatively associated with aging, FN, elastin and free SH. On the other hand, the level of free SH was positively associated with aging, cholesterol and FN. 3. The contents of free SH, hydrohobicity and FN in elastin from the arch to the thoracic artery were lower than that of the other arteries. Hydrohobicity was negatively correlated with ID, and FN was positively correlated with cholesterol. 4. In the abdominal artery, FN content was higher than that of other arteries. There was a correlation between FN and free SH. However, free SH was inversely associated with elastin, ID and hydrohobicity.
Adult↗
[Functional morphology of harderian gland].
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Animals↗
Novel homozygous and compound heterozygous mutations of sterol 27-hydroxylase gene (CYP27) cause cerebrotendinous xanthomatosis in three Japanese patients from two unrelated families.
The autosomal recessively inherited cholesterol metabolic disease, cerebrotendinous xanthomatosis (CTX), is caused by mutations in the sterol 27-hydroxylase gene. Three Japanese CTX patients from two unrelated families were studied genetically. By DNA sequence analysis a novel mutation of A for G substitution at amino acid position 372 (CGG 372Arg to CAG 372Gln) was identified in one of the CTX families. The mutation was also found in two patients from the other family, with a compound heterozygous pattern of A for G substitution at amino acid position 441 (CGG 441Arg to CAG 441Gln). The latter mutation was the same as previously reported by our group (J. Lipid Res. 1994. 35: 1031-1039). As the two mutations changed the restriction enzyme sites, rapid screening methods were developed for the detection of the carriers. Transfection of the two mutant cDNAs into COS cells resulted in markedly reduced sterol 27-hydroxylase activity. These results indicate that the two mutations are responsible for the deficiency of the sterol 27-hydroxylase activity in these patients. The features of mutations identified till now in Japanese CTX patients are also reviewed.
Cholestanetriol 26-Monooxygenase↗
[Calpain super family and its interacting-proteins].
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Amino Acid Sequence↗
Genetic analysis of a Japanese cerebrotendinous xanthomatosis family: identification of a novel mutation in the adrenodoxin binding region of the CYP 27 gene.
Cerebrotendinous xanthomatosis (CTX), an autosomal recessive lipid-storage hereditary disorder, is caused by mutations in the sterol 27-hydroxylase gene (CYP 27). A 24-year-old female Japanese CTX patient and her parents were studied for a CYP 27 mutation. Multiple xanthomas were the main complaint of the patient and plasma cholestanol level was markedly elevated. Sterol analysis of a xanthoma biopsy confirmed cholesterol and cholestanol deposition, and the cholestanol accounted for 8.1% of the total sterols. Sterol 27-hydroxylase activity in fibroblasts derived from the patient was undetectable, while the activities in fibroblasts from her mother and father were 54% and 41% of the normal level, respectively. Direct sequence analysis showed a missense mutation of A for G substitution in the CYP 27 gene at codon 362 (CGT 362Arg to CAT 362His) with a homozygous pattern in the patient, and a heterozygous pattern in the parents. The mutation, which eliminates a normal HgaI endonuclease site at position 1195 of the cDNA and is located at the adrenodoxin binding region of the gene, is most probably responsible for the decreased sterol 27-hydroxylase activity in this Japanese CTX family. The combined data strongly support that the primary enzymatic defect in CTX is the disruption of sterol 27-hydroxylase and that the disease is inherited in an autosomal recessive trait.
Adrenodoxin↗