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Biomedical subjects

T Koide

Publications and source records attributed to T Koide.

At least 109 records · Page 6Linked to original sources

Human histidine-rich glycoprotein expressed in SF9 insect cells inhibits apatite formation.

Histidine-rich glycoprotein (HRG) is structurally related to the alpha2-HS glycoprotein/fetuin family of mammalian plasma proteins; both belong to the cystatin superfamily of proteins. We expressed recombinant human HRG and alpha2-HS in Sf9 insect cells for functional analysis. Recombinant HRG bound heparin and fibrinogen while alpha2-HS did not. Both proteins inhibited the formation of apatite, recombinant HRG (IC50 approximately 1 microM) with 2-fold lower molar activity than alpha2-HS (IC50 approximately 0.5 microM). The inhibition in vitro of apatite formation suggests a new function for plasma HRG protein, inhibition of phase separation in blood vessels.

Animals↗

Intracellular degradation of secretion defect-type mutants of antithrombin is inhibited by proteasomal inhibitors.

To examine the cellular basis for secretion defect-type antithrombin deficiency, we expressed two mutants, P --> stop (Pro429 to stop codon) and deltaGlu (deletion of Glu313). Pulse-chase experiments using stably transfected BHK cells showed that little (< 5%) of P --> stop mutant as well as deltaGlu mutant was secreted and the total amount of radioactivity was significantly reduced, suggesting an intracellular degradation. The degradation was not inhibited by brefeldin A, indicating it occurring in a preGolgi apparatus. However, the degradation was strongly inhibited by proteasomal inhibitors, such as carbobenzoxy-L-leucyl-L-leucyl-L-leucinal (LLL), carbobenzoxy-L-leucyl-L-leucyl-L-norvalinal (LLnV) and lactacystin. By endoglycosidase H digestion and immunofluorescence staining, these mutants were shown to localize in the endoplasmic reticulum (ER). These results suggest that the secretion defect-type mutants of antithrombin are degraded by proteasome through the ER-associated quality control mechanism in the cells.

Acetylcysteine↗

Characterization of protein components of human urinary crystal surface binding substance.

We previously extracted crystal surface binding substance (CSBS) from human urine and showed that it appeared to constitute a substantial proportion of urinary macromolecular inhibitors of calcium oxalate crystallization. CSBS was isolated from human urine and fractionated by three consecutive chromatography procedures in order to characterize protein inhibitors of calcium oxalate crystallization. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and NH2-terminal amino acid sequencing revealed that inhibitory fractions eluted from a final, hydroxyapatite column contained prothrombin and osteopontin. Hydroxyapatite column fractions also contained other, unidentified protein inhibitors of calcium oxalate crystallization. CSBS contained also human serum albumin, alpha 1-acid glycoprotein, alpha 1-microglobulin, alpha 2-HS glycoprotein, retinol-binding protein, transferrin, and Tamm-Horsfall protein, but these proteins seemed to play no direct role in inhibitory activity.

Adult↗

Glycosaminoglycans in crystal-surface binding substances and their role in calcium oxalate crystal growth.

OBJECTIVE: To clarify the role of glycosaminoglycans (GAGs) in crystal-surface binding substances (CSBS) on the growth of calcium oxalate crystals in urine. MATERIALS AND METHODS: Urine samples (24 h) were collected from healthy men (aged 25-42 years) and CSBS were obtained from the pooled urine samples. The CSBS were digested with heparitinase or proteinase and the inhibition of crystal growth assessed before and after enzyme digestion. Anion-exchange chromatography and high-performance liquid chromatography (HPLC) were used to determine the types of GAGs contained in the CSBS. RESULTS: The inhibitory activity of CSBS on crystal growth decreased with concentration when digested with heparitinase or proteinase. HPLC showed that CSBS contained a small amount of dermatan sulphate and abundant heparan sulphate, both of which inhibited crystal growth. CONCLUSION: Both heparan sulphate and dermatan sulphate may inhibit calcium oxalate crystallization, the former being the predominant GAG in CSBS.

Adult↗

Influence of flavonoids on cell cycle phase as analyzed by flow-cytometry.

Flavonoids were previously reported to have cytostatic activity in vitro and in vivo. In the present study each phase of the cell cycle was observed by flow-cytometry after HCT-15 cells had been cultured with flavonoids for 2 days. In the control group, two peaks were observed on the histogram, the first peak representing double strands of DNA, and the second, tetra strands. Chalcone, which is known to be a precursor of all flavonoids, seemed to block the passage from S to G2 and M phase. Flavonone, which is freely transformed into chalcone in vivo, showed a similar histogram as chalcone. Among the anthocyanins most effective in suppressing the tumor cell growth, cyanin and pelargonidin made a mild rise between the 1st and 2nd peak suggesting block between G1 and S phase. Delphinidin gave a high percent of cells in G1 phase, suggesting prolongation of G1 phase. When quercetin or kaempherol, both of which are flavonols were added to the culture, the second peak rose significantly, indicating a block between G2 and M phases. Thus, the site of these cytostatic agents seems to differ depending on the kind of flavonoid.

Cell Cycle↗

Effect of allomelanin on tumor growth suppression in vivo and on the cell cycle phase.

Allomelanins are nitrogen-free macromolecular polymers of simple phenols produced by higher plants and fungi. In an earlier work we found that allomelanins were effective in suppressing the growth of cultured tumor cells. In the present study we examined the effect of these polymers on the survival curve of Balb/C mice inoculated i.p. with Meth/A cells (3 x 10(4) cells/mouse), which originated from a malignant lymphoma. Allomelanins were extracted from black sesame seeds and black soybeans, and given p.o. via the drinking water. The dose of allomelanins was approximately 3 mg/mouse/day. The percent of survivals was significantly higher in the experimental groups than in the control. Also, when HCT-15 cells were cultured for 2 days in medium containing 400 micrograms/ml of protein-free allomelanins, histograms obtained from flow-cytometry data showed a significant increase in the fraction between the diploid and tetraploid peaks, indicating blockage of the S phase of the cell cycle. These results suggest that allomelanins suppressed the tumor cell growth in vitro and in vivo, and that the effect was cytostatic, mostly by blockage of the S phase.

Animals↗

Antitumor effect of anthocyanin fractions extracted from red soybeans and red beans in vitro and in vivo.

Many bioflavonoids extracted from petals of higher plants and from fruit rinds, as well as purified flavonoids, have been reported to have antitumor effects in vitro and in vivo. Bioflavonoids extracted from red soybeans are mostly cyanin conjugated with glucose and rhamnose, whereas bioflavonoids of red beans are cyanin conjugated with rhamnose as revealed by thin-layer chromatogram. Flavonoids extracted from red soybeans were effective in inhibiting the growth of HCT-15 cells in vitro. On the other hand, flavonoids from red beans were not effective, although their hydrolyzed sugar-free forms were growth inhibitory. Sugar-bonded bioflavonoids extracted from both red soybeans and red beans were effective in prolonging the survival of Balb/C mice bearing syngeneic tumor-Meth/A cells, when they were dissolved in drinking water and given at a dose of approximately 500 micrograms/mouse/day.

Animals↗

Direct tumor growth suppressive effect of melanoidin extracted from immunomodulator-PSK.

Melanoidin, which belongs to the melanin group of molecules, was extracted from the polysaccharide biological response modifier PSK. Melanoidin was cultured together with HCT-15 cells derived from human colon cancer and with AGS cells derived from human gastric carcinoma. After four days of culture, cell count was compared with that of the control cells. Significant suppression was observed, that is, 50% suppression was shown at concentrations of melanoidin between 200 and 100 micrograms/ml. A histogram generated by flow cytometry showed elevation of the tetraploid peak and of that between diploid and tetraploid peaks, suggesting blockage of S phase and G2 to M phase of the cell cycle. Thus, melanoidins contained in the immunomodulator PSK revealed to have a direct tumor cell growth inhibitory effect.

Adjuvants, Immunologic↗

Tumor cell growth-inhibiting effect of melanoidins extracted from miso and soy sauce

Melanoidins, a group of plant-produced melanins, were extracted from miso and shoyu (soy sauce), which are products of soybeans and widely consumed as food in oriental countries. The molecular weight of these melanoidins was approximately 5600 as revealed by Sepharose CL-4B column chromatography. The melanoidins were cultured together with HCT-15 cells derived from human colon carcinoma and AGS cells derived from human gastric carcinoma. Significant cell growth suppression was observed by incubating the cells with these melanoidins. The 50% growth suppression dose ranged between 100 micrograms/ml and 25 micrograms/ml, that is, between 2 x 10(-5) mM/ml and 0.5 x 10(-5) mM/ml. Flow cytometric analysis suggested that these melanoidins blocked the S phase and G2 to M phase of the cell cycle.

Journal Article↗

Suppression of growth of cultured malignant cells by allomelanins, plant-produced melanins.

Allomelanins, which belong to the melanins produced by higher plants and fungi, were studied with respect to their growth suppressive effects on cultured HCT-15 cells derived from human intestinal carcinoma and Meth/A cells derived from a Balb/C mouse lymphoma. Allomelanins were extracted from black soy beans and black sesame seeds with 0.2% NaOH in water. Proteins bonded to the allomelanins were removed by boiling the extracts for 20 hours in 30% NaOH. Although native allomelanins conjugated with protein did not suppress the growth of the cells, protein-free allomelanins did suppress it. That is, 50% suppression of HCT- 15 cell growth was found at a concentration between 100 and 200 micrograms/ml of the melanin prepared from the black soy beans, and between 25 and 100 micrograms/ml of that from the sesame seeds. For Meth/A cells, 50% suppression by allomelanin from the black beans was approximately 100 micrograms/ml; and that by the allomelanin preparation from sesame seeds, between 25 and 100 micrograms/ml. Thus, protein-free allomelanins seemingly suppress cultured mammalian and animal tumor cells.

Animals↗

Tumor cell growth suppression by chalcone (1,3-diphenyl-2-propen-1-one).

Chalcone or 1,3-diphenyl-2-propen-1-one which is widely distributed in higher plants and is considered to be a precursor of all flavonoids, was studies in terms of its anti-tumor growth activity in vitro and in vivo. Mice given the chalcone, which was suspended in the drinking water, showed a statistically significant rise in percent survival: 66% at the 35th day for the chalcone-treated animals vs. 30% for the control ones. The chalcone also gave significant suppression of growth of HCT-15 and Meth/A cells in vitro. The dose of 50% reduction in growth was between 2.5 micrograms/ml and 0.6 microgram/ml for the HCT-15 cells and less than 10 micrograms/ml for the Meth/A cells. Histograms obtained by flow cytometric examination showed an elevated region between the diploid and the tetra-ploid nucleoidal peaks. A third peak heavier than the tetra-ploid peak was also observed, which peak was not observed in the control. The percentage of cells in the S phase was significantly raised. Microscopically, large nucleated cells were observed following addition of the chalcone. These results suggest that the chalcone induced abnormal DNA synthesis and mitosis in the cultured cells.

Animals↗

Imprinting of Igf2 and H19 from a 130 kb YAC transgene.

A stringent test for imprint control elements is to examine their function at ectopic loci in transgenic experiments. Igf2 and H19 are part of a larger imprinting region and as a first step, we examined these reciprocally imprinted genes in transgenic experiments using a 130 kb YAC clone. After paternal inheritance, H19 was appropriately repressed and Igf2 was expressed, irrespective of copy number or genetic background. After maternal inheritance H19 was consistently expressed, albeit with some variability. The levels of H19 expression per copy of the transgene inversely correlated with Igf2 (-lacZ) expression in cis. The consistent imprinting of H19 from this YAC contrasts with the previously described imprinting of mini-H19 transgenes, which only occurs at multi-copy loci, is inconsistent, and is prone to genetic background effects. We propose a novel model in which silencing of the H19 gene is the default state and its activation after maternal inheritance is the key mechanistic event for imprinting in this region. In addition, in situ analysis of the Igf2-lacZ reporter indicates that additional mesoderm-specific enhancers are present within the YAC clone. No obvious phenotype was detected from the excess gene dosage of H19.

Animals↗

Effects of various collagen crosslinking techniques on mechanical properties of collagen film.

We examined the relationship between collagen crosslinking techniques and dentinum reinforcement. In the present study, the tensile tests, imbibition test and antienzyme test were performed on collagen films mainly composed of type I collagen after ultraviolet irradiation and immersion in tannic acid and glutaraldehyde. The elastic modulus increased with the reaction time, but reduction of elongation at breaking point and increased stiffness were observed. This tendency was enhanced by ultraviolet irradiation. Tannic acid treatment resulted in slower crosslinking than the other two techniques. Antienzyme activity was markedly increased by treatment with tannic acid and glutaraldehyde.

Animals↗

[Postprostatectomy dysuria: restored voiding function by perineal hot water spray in an old male with a contractile bladder].

The hot water spray on perineum restored voiding function in a 77-year-old male patient with the poorly contractile bladder. He noticed the gradual decrease of bladder sensation and voiding stream, and needed clean intermittent catheterization to empty bladder even after TUR-P. Neurological examination revealed only weakened anal and bulbocavernosus reflex and impaired skin sensation of foot plate and perineum. Simple cystometry showed impaired perception of bladder filling and underactive bladder. Electromyography of the pelvic floor displayed decreased responses to bladder filling and voluntary sphincter contraction. Spine-X-ray examination and computed tomography of brain and spinal cord did not reveal any neurological lesions except for the silent small infarction of subcortex. Trial of drug therapy with Distigmine bromide, Cernilton and Prazosin did not restore voiding function. By accident, however, patient himself found spontaneous voiding could be induced by hot water spray on perineum and/or scrotum during bidet using. Since then, he has been continuing this stimulation daily for 4 years, with which almost complete voiding is possible without catheterization, though bladder sensation remains impaired. The pathophysiology and affecting factors on micturition reflex in this case were discussed.

Aerosols↗

[A case of Guillain-Barré syndrome after the travel in southeast Asia--isolation of Campylobacter jejuni PEN 5 serotype].

A 57-year-old man, while on travel in Malaysia, suffered from diarrhea after he ate fruits. He developed limbs weakness without sensory disturbance after his return to Japan. Serum from the patient had high IgG anti-GM1 antibody titer. Campylobacter jejuni was isolated from his stool. The serotype belonged to PEN 5. The patient received double-filtration plasmapheresis 7 times during from days 6 to 17. Muscle strength began to recover gradually on day 10, and returned to normal 5 months after the onset of neurologic symptoms. Repeated neurophysiologic studies indicated that the axonal degeneration of motor nerves was predominant process. This case suggests that Guillain-Barré syndrome is a complication of traveler's diarrhea.

Campylobacter Infections↗

[Renal cell carcinoma in acquired cystic disease of kidney (ACDK) manifested by spontaneous renal rupture: a case report].

We report a case of renal cell carcinoma in acquired cystic disease of the kidney (ACDK) presenting with a spontaneous renal rupture. A 41-year-old man on chronic hemodialysis for 16 years was referred to our hospital with sudden left back pain. On arrival, the patient was in a state of hemorrhagic shock but his general condition improved by conservative therapy. Computed tomography demonstrated a left renal rupture as well as bilateral ACDK. Although there was no definite evidence of a renal tumor, a left radical nephrectomy was performed one week later. The resected kidney contained a yellowish-brown tumor of 5.5 x 5.0 x 4.5 cm in size. Pathological examination revealed renal cell carcinoma, cystic type, clear cell subtype, pT2pN0M0. He has been free of recurrence for 3 months.

Adult↗

[Inhibitory effect of oral administration of Takusya on calcium oxalate crystallization in human whole urine].

We previously reported that Takusya had the inhibitory effect on in vitro calcium oxalate crystallization and in vivo stone formation in an animal model and it could be a prophylactic agent against calcium oxalate stone formation. We studied the effect of Takusya on calcium oxalate crystallization in human urine. Takusya (500 mg/day and 1,000 mg/day) was administered to 16 healthy men for 3 days and then 24-hour urine samples were collected to measure the urinary excretion of calcium, phosphate, magnesium, uric acid, creatinine, citric acid and oxalic acid. The urine samples before the administration of Takusya was used as a control. The size and distribution of crystals, which were formed in the urine samples by adding calcium chloride and sodium oxalate, were measured using the Coulter counter technique. Urinary magnesium excretion was significantly reduced by 1,000 mg/day of Takusya compared with the control (p < 0.05). The growth of crystals was significantly inhibited by 500 mg/day of Takusya in the large crystal formers whose urine created crystals more than 3.5 microns before the administration of Takusya (p < 0.05). These findings suggested that Takusya inhibited the growth of crystals formed in human urine.

Administration, Oral↗