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Biomedical subjects

M Shin

Publications and source records attributed to M Shin.

At least 73 records · Page 4Linked to original sources
PubMed1998

NAD+ biosynthesis from tryptophan in the presence of nicotinic acid or vice versa by rat hepatocytes--effect of clofibrate-feeding.

NAD+ biosynthesis from tryptophan in the presence of nicotinic acid or vice versa by rat hepatocytes was investigated. In the control hepatocytes, NAD+ synthesis from tryptophan was not affected by nicotinic acid from 0.026 to 0.26 mM. NAD+ synthesis from nicotinic acid was slightly inhibited with varying concentrations of tryptophan from 0.1 to 1.0 mM. In the clofibrate-treated hepatocytes, NAD+ synthesis from tryptophan was greatly increased (234% of the control), while that from nicotinic acid was decreased (71.2% of the control). Both, NAD+ synthesis from tryptophan and that from nicotinic acid were decreased by the coexisting nicotinic acid or tryptophan. Total amount of NAD+ synthesized from tryptophan and nicotinic acid at their physiological concentrations was significantly higher than that in the control hepatocytes as a result of a large increase of NAD+ synthesized from tryptophan. When the metabolic flux of 0.1 or 0.5mM tryptophan was investigated, the glutarate pathway was suppressed in the clofibrate-treated hepatocytes, the quinolinic acid-NAD+ flux being elevated. Similarly to clofibrate, DEHP and CPP revealed an increase in NAD+ synthesis from tryptophan. Mutual relationship of NAD+ biosyntheses from tryptophan and nicotinic acid in rat hepatocytes is discussed and the relevance with peroxisomal proliferation is suggested.

Animals↗
PubMed1998

[Detection of receptor mRNAs using nonradioactive in situ hybridization].

To understand better the regulatory mechanism of cell function through bioactive substance such as hormones and cytokines, analysis of the gene expression of their receptors at the individual cell level is required. Since the presence of specific mRNA is a good index of gene expression, in situ hybridization (ISH) has been recognized as a powerful tool. Especially, nonradioactive ISH with synthetic oligodeoxynucleotide probe is a safe, quick and highly sensitive method. We have developed a unique method with thymine-thymine (T-T) dimer as a nonradioactive labeling, in which adjacent thymines are dimerized by ultraviolet light irradiation. The T-T dimers in hybrids are immunohistochemically recognized using an anti-T-T dimer antibody. In this article, we will present an example of ISH protocol, which works very well both T-T dimer and digoxigenin-labeled probes.

Animals↗
PubMed1998

Different patterns of p53 mutations in prostatic intraepithelial neoplasia and concurrent carcinoma: analysis of microdissected specimens.

Prostatic intraepithelial neoplasia (PIN) is characterized by intraluminal proliferation of epithelial cells and can be divided into high-grade (HGPIN) and low-grade (LGPIN) lesions. HGPIN is regarded as the most likely precursor of prostatic carcinoma (PCA). Microdissected DNA selectively extracted from paraffin-embedded sections of 29 cases of PCA and 1 benign prostatic hypertrophy were analyzed for p53 mutation by single-strand conformation polymorphism (SSCP) of polymerase chain reaction (PCR)-amplified DNA fragments followed by direct sequencing. These patients had received total prostatectomy (27 cases) or transurethral resection (3 cases). Under direct microscopic observation, DNA was microdissected from 108 lesions: 67 lesions from 22 cases of PIN (55 HGPIN and 12 LGPIN), 29 from 22 cases of PCA, and 12 from 11 cases of adjoining benign glands. Analysis revealed 13 mutations in 10 lesions from six cases. All 13 were point mutations: 7 missense, 5 silent, and 1 nonsense. Mutations were detected in 3 cases (14%) of PIN and 5 cases (25%) of PCA. PIN lesions with p53 mutations were all categorized as HGPIN. Neither LGPIN nor benign glands adjoining PIN and/or PCA had mutations. Two PIN and one PCA lesion in each of two cases had mutations that were different from each other. G-to-A transition was the commonest mutation pattern. The current findings showed that HGPIN, but not LGPIN, and PCA are similar with regard to p53 mutation. The diverse patterns of p53 mutation among HGPIN and PCA lesions suggested multiclonal development of prostatic precancerous lesions.

Amino Acid Substitution↗
PubMed1997

Incidence of prostatic intra-epithelial neoplasia in Osaka, Japan.

High-grade prostatic intra-epithelial neoplasia (HGPIN) is the most likely precancerous lesion for prostatic carcinoma. A high incidence of its association with cancer has been reported in Western countries. On the other hand, information regarding its incidence is limited in Japan, where the mortality due to prostate cancer is much lower. We reviewed 53 clinical stage T2 or T3 prostatic cancers of Japanese patients living in Osaka, Japan (mean age, 67.2 years). These cases were subdivided into a pre-operatively non-castrated group (34 cases) and a medically or surgically castrated group (19 cases). HGPIN was found in 27 cases. The incidence of HGPIN was significantly lower in the castrated group (21.0%) compared with the non-castrated group (67.6%). In the non-castrated group, patient age, pathological stage, Gleason score, tumor size and serum prostate-specific antigen showed no significant correlation with HGPIN. Advanced pathological stage and tumor size tended to decrease the incidence of HGPIN, although this was not statistically significant. When the study group was limited to stage T2 tumors of the non-castrated group, the incidence of HGPIN was 81.0%. HGPIN in Japan may also be clinically and etiologically significant as a precursor of clinical cancer.

Aged↗
PubMed1997

Thiocyanato and Iodo Complexation of Cadmium(II) Ions in Micellar Solutions of a Nonionic Surfactant Triton X-100

Complexation of the cadmium(II) ion with thiocyanate and iodide ions have been studied by precise titration calorimetry in micellar solutions of a nonionic surfactant Triton X-100 containing 0.4 mol dm-3 NaClO4 as a constant ionic medium at 298 K. With regard to the Cd(II)-SCN- system, the formation of [Cd(NCS)]+ and [Cd(NCS)(SCN)] in aqueous solution and [Cd(NCN)2(SCN)]- in micelles was established, though the complexation in micelles is not extensive. With the Cd(II)-I- system, the formation of [CdI]+, [CdI3]-, and [CdI4]2- was established in aqueous solution together with the formation of [CdI2], [CdI3]-, and [CdI4]2- in micelles. It is revealed that the complexation in micelles is accompanied by relatively large and negative enthalpies, which is ascribed to the enthalpies of transfer of [CdI3]- and [CdI4]2- from aqueous solution to micelles. This suggests that a hydrophobic complex with no capacity to form strong hydrogen bonds in water is favored in micelles.

Journal Article↗
PubMed1997

[Examination of scanning technique for lung cancer screening with helical CT].

As a new application of helical CT scanning we evaluated the parameters for lung cancer screening with an extremely low doses and large helical pitch. On the phantom studies, the image quality obtained with the low dose parameter was not inferior to that of the usual screening technique but artifacts were increased with the large helical pitch. A scanning technique using 120 kv, 40-60mA, 10mmth, 20mm/ sec, and a reconstruction pitch of 2 was used for lung cancer screening (screening parameters) (50 cases), and comparison was made between the detectability of the screening parameters and the routine parameters (120 kv, 200mA, 10mmth, 10-13mm/sec, and a reconstruction pitch of 1-1.3). Detectability with the screening parameters was as follows: nodular lesions (< 5mm in size: 76%. 5-10mm: 90-93% 10mm < 100%), linear lesions: 94-95%, infiltrations: 93-100%. There were no false negative lesions, when the reconstruction pitch of the screening parameters was changed from 1 to 0.25. In conclusion, reconstruction pitch had the most influence on lesion detectability.

Adult↗
PubMed1996

Mice with disrupted GM2/GD2 synthase gene lack complex gangliosides but exhibit only subtle defects in their nervous system.

Gangliosides, sialic acid-containing glycosphingolipids, are abundant in the vertebrate (mammalian) nervous system. Their composition is spatially and developmentally regulated, and gangliosides have been widely believed to lay essential roles in establishment of the nervous system, especially in neuritogenesis and synaptogenesis. However, this has never been tested directly. Here we report the generation of mice with a disrupted beta 1,4-N-acetylgalactosaminyltransferase (GM2/GD2 synthase; EC 2.4.1.92) gene. The mice lacked all complex gangliosides. Nevertheless, they did not show any major histological defects in their nervous systems or in gross behavior. Just a slight reduction in the neural conduction velocity from the tibial nerve to the somatosensory cortex, but not to the lumbar spine, was detected. These findings suggest that complex gangliosides are required in neuronal functions but not in the morphogenesis and organogenesis of the brain. The higher levels of GM3 and GD3 expressed in the brains of these mutant mice may be able to compensate for the lack of complex gangliosides.

Animals↗
PubMed1996

NAD+ biosynthesis and metabolic fluxes of tryptophan in hepatocytes isolated from rats fed a clofibrate-containing diet.

Hepatocytes were isolated from rats fed a diet with or without 0.25% clofibrate, and NAD+ synthesis by the hepatocytes was determined using either [carboxyl-14C]nicotinic acid or [5-3H]tryptophan. NAD+ and total pyridine nucleotides synthesized from [14C]nicotinic acid by the clofibrate-treated cells were not significantly different from those synthesized by the control cells when expressed on the basis of nanomoles per hour per milligram of DNA. On the contrary, NAD+ synthesized from [3H]tryptophan was significantly higher in the clofibrate-treated cells (158% of the control cells) on the basis of nanomoles per hour per milligram of DNA. Clofibrate was inhibitory to tryptophan metabolism as a whole, affecting the glutarate pathway more (decreased to 37% of control) than the kynureninase flux (decreased to 64% of control). As a result, the quinolinate-NAD flux, estimated as the difference in the amounts of tryptophan metabolized by the two metabolic pathways, increased in the clofibrate-treated hepatocytes. The increase in quinolinate during the incubation was 8 times more in the clofibrate-treated cells than in the control cells, which confirmed alteration in the metabolic fluxes of tryptophan in the clofibrate-treated cells. Hepatic quinolinate phosphoribosyltransferase (EC 2.4.2.19) activity increased with dietary clofibrate and returned to the control level 1 week after removing clofibrate from the diet. Nicotinate phosphoribosyltransferase (EC 2.4.2.11) and NAD+ glycohydrolase (EC 3.2.2.5) activities remained unchanged with dietary clofibrate.

Animals↗
PubMed1996

Chemotherapeutic drugs released from polymers: distribution of 1,3-bis(2-chloroethyl)-1-nitrosourea in the rat brain.

PURPOSE: The distribution of [(3)H]BCNU following release from polymer implants in the rat brain was measured and evaluated by using mathematical models. METHODS: [(3)H]BCNU was loaded into p(CPP:SA) pellets, which were subsequently implanted intracerebrally in rats; [(3)H]BCNU was also directly injected into the brains of normal rats and rats with intracranially transplanted 9L gliomas. Concentrations of [(3)H]BCNU on coronal sections of the brain were measured by autoradiography and image processing. For comparison, the kinetics of [(3)H]BCNU release from the p(CPP:SA) polymer discs into phosphate-buffered saline were also measured. RESULTS: High concentrations of BCNU (corresponding to 1 mM) were measured near the polymer for the entire 30-day experiment. The penetration distance, defined as the distance from the polymer surface to the point where the concentration of [(3)H]BCNU in the tissue had dropped to 10 percent of the maximum value, was determined: penetration distance was 5 mm at day 1 and 1 mm at days 3 through 14. Local concentration profiles were compared with a mathematical model for estimation of the modulus phi (2), an indicator of the relative rate of elimination to diffusion in the brain. From day 3 to 14, phi(2) was 7, indicating that BCNU elimination was rapid compared to the rate of diffusive penetration into tissue. The enhanced penetration observed on day 1 appears to be due to convection of extracellular fluid caused by transient, vasogenic edema, which disappears by day 3. CONCLUSIONS: Polymer implants produce very high levels of BCNU in the brain, but BCNU penetration into brain tissue is limited due to rapid elimination.

Animals↗
PubMed1996

Detection of cytokine mRNA-expressing cells in peripheral blood of patients with IgA nephropathy using non-radioactive in situ hybridization.

IgA nephropathy (IgA-N) is considered to be an immune-mediated disorder and several immunological abnormalities have been observed. In the present study, we optimized non-radioactive in situ hybridization and applied this technique to evaluate the degree of expression of various cytokine mRNAs in peripheral blood mononuclear cells (PBMC) taken from patients with IgA-N on cytospin preparation. Using this method, together with image analysis, we examined the expression of mRNA in cells which secrete cytokines, such as IL-2, interferon-gamma (IFN-gamma), IL-4, IL-5 and IL-6. The expression of these mRNAs was clearly observed in monocytes and lymphocytes at a single-cell level. Compared with healthy adults, the expression of IL-4 mRNA, IL-5 mRNA and IL-6 mRNA, but not IL-2 mRNA or IFN-gamma mRNA, was significantly enhanced. Our results indicate that non-radioactive in situ hybridization method is a powerful technique for analysis of cytokine mRNAs in PBMC at a cellular level. Our results also suggest that mRNA expression of IL-4, IL-5 and IL-6, produced by Th2 cells, is increased in patients with IgA-N. The abnormal regulatory process involved in cytokine expression may play an important immunopathologic role in IgA-N.

Adult↗
PubMed1995

Four ferredoxins from Japanese radish leaves.

We detected three ferredoxin components, a, b, and c, in green shoots of Japanese radish seedlings by hydrophobic HPLC analysis using a phenyl-5PW column. All components were also present in mature leaves. Component a was further separated into two components, a1 and a2, by reversed-phase HPLC after carboxymethylation. The existence of four ferredoxins in the photosynthetic tissue of a single plant species is reported here for the first time. Each individual ferredoxin component was isolated, and its primary structure was analyzed for comparison with the two leaf ferredoxins, L-Fds A and B, isolated by Wada et al. (J. Biochem. 105, 619-625 (1989)). The primary structures of components a1, a2, and b were all found in 12 possible structures of L-Fd A deducible from its microheterogeneities. This led us to conclude that L-Fd A is a mixture of three ferredoxins corresponding to components a1, a2, and b. The complete primary structure of component c was established by sequencing of the whole protein and of peptides generated by protease digestion. The N-terminal structure of component c was identical with L-Fd B up to the 11th amino acid residue, suggesting that this component is the same ferredoxin as L-Fd B.

Amino Acid Sequence↗
PubMed1995

Multilayer peroxidase-labeled antibody method: comparison with labeled streptavidin-biotin method, avidin-biotin-peroxidase complex method, and peroxidase-antiperoxidase method.

A new, extremely sensitive, "multilayer peroxidase-labeled antibody method" is introduced. This method utilizes peroxidase-labeled antibodies that immuno-react with their antigen, but also can be an antigen to other peroxidase-labeled antibodies. An example of this method is: use rabbit IgG as the first antibody, peroxidase-labeled goat antirabbit IgG as the second antibody, peroxidase-labeled rabbit antigoat IgG as the third antibody, peroxidase-labeled goat antirabbit IgG as the fourth antibody, and peroxidase-labeled rabbit antigoat IgG as the fifth antibody in succession. We compared the sensitivities of this multilayer peroxidase-labeled antibody method with that of commercially available immuno-peroxidase staining kits, i.e., avidin-biotin-complex method and peroxidase-antiperoxidase method, both by immunostaining and enzyme-linked immunosorbent assay. The generated signal and background in immunostained sections were determined by a computer-assisted image analyzer and by enzyme-linked immunosorbent assay spectrophotometrically. It was found that a maximum signal/background ratio was obtained with those sections reacted with the first four antibodies in succession by the multilayer peroxidase-labeled antibody method, and those reacted with the five antibodies in succession by the multilayer peroxidase-labeled antibody method in the enzyme-linked immunosorbent assay. Thus, the multilayer peroxidase-labeled antibody method is more sensitive than the avidin-biotin-complex method and the peroxidase-antiperoxidase method.

Animals↗
PubMed1995

Effect of feeding clofibrate-containing diet on the hepatic NAD+ level in rats.

Feeding rats with a diet containing 0.25% clofibrate for 2 weeks elevated the hepatic NAD+ and total nicotinate levels significantly. Other peroxisome proliferators, such as 2-(4-chlorophenoxy)propionic acid and di(2-ethylhexyl)phthalate, had similar effects. When rats were fed the control diet without clofibrate for 1 week after 2 weeks of the clofibrate diet, the hepatic NAD+ level returned to the control value. Muscular NAD+ content was not affected by the peroxisome proliferators. The results were discussed in relation to induction of peroxisomal beta-oxidation enzymes by the peroxisome proliferators.

Animals↗