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Biomedical subjects

N Hashizume

Publications and source records attributed to N Hashizume.

At least 37 records · Page 2Linked to original sources
PubMed1997

Detection of Clostridium septicum hemolysin gene by polymerase chain reaction.

A polymerase chain reaction (PCR) was developed for the detection of the hemolysin (alpha toxin) gene of Clostridium septicum. The PCR primers were designed from the sequence of the hemolysin gene and synthesized. A DNA fragment of 270 bp was amplified from 10 strains of C. septicum, but was not from strains of C. chauvoei, C. perfringens, C. novyi, or C. haemolyticum. When the PCR product was digested with Sau3AI, two DNA fragments of the expected 148 bp and 122 bp were recognized. The lowest detectable threshold of PCR for the hemolysin gene was 3.8 x 10(3) cells/ml. The PCR technique may be useful for rapid detection or identification of C. septicum associated with malignant edema.

Animals↗
PubMed1997

The alginate reduce the postprandial glycaemic response by forming a gel with dietary calcium in the stomach of the rat.

Soluble dietary fibers include an alginate which reduces the postprandial glycaemic response. We speculated that the cause of the reduction of alginate on the glycaemic response is the gel formation of alginate with dietary calcium in the stomach. The aim of the present study was to confirm our hypothesis and to examine the effect of the sugar composition of alginates on the glycaemic response and the effect of continuous feeding of alginate-containing diet to rats. Diabetic rats were made by injecting streptozotocin (STZ) intravenously. These rats were fed 5 g of the experimental diet with or without sodium alginate and with or without calcium after overnight fasting. In the rats fed the diet containing both alginate and calcium, the postprandial glycaemic response was lower than in the rats fed other diets and gastric contents were retained as gel form. Three beta-d-mannuronic acid (M) to alpha-l-guluronic acid (G) (ratios (M/G 0.5, 1.3, 2.1) of alginates were not associated with the acute glycaemic response in STZ rats. Then, we fed normal rats with an alginate-free or one of three respective alginate-containing diets for 4 weeks. During the first 2 weeks of the feeding period, the total amount of food intake of rats fed the G-rich alginate diet was smaller than that of rats in the other dietary groups. Moreover, the weight of the dorsal abdominal adipose tissue of rats given the G-rich alginate diet was the lowest among the dietary groups. We conclude that the reductive effect of alginate feeding on the postprandial glycaemic response was not caused only by their inherent viscosity, but also by gel formation with dietary calcium in the stomach. Moreover, it is necessary to pay attention to the M/G ratio of alginates for estimating their physiological effect.

Adipose Tissue↗
PubMed1997

[Blood amino acid levels in sarin poisoning patients].

Since plasma is generally employed for amino acid analysis, we compared amino acid levels in plasma with those in serum for healthy individuals and examined the influence of separation and storage conditions on the stability of the samples. Then, we determined the amino acid levels of frozen serum samples obtained from sarin poisoned patients. A. Comparison of Amino Acid Levels in Plasma and Those in Serum Blood was collected from 5 healthy individuals. Then, heparinated plasma and serum were separated by centrifugation immediately after blood collection. Serum was also separated by centrifugation after standing whole blood at room temperature for 1 hour. Frozen plasma and serum were store at -40 degrees C for 5 months. All were subjected to analysis in an amino acid analyzer. It was found that the cystine (Cys) and 3-methyl-histidine (3-M-His) levels in serum and plasma were affected when stored in a frozen state, that the aspartate (Asp) level was changed according to the method of collecting serum, and that the taurine (Tau) and ornithine (Orn) levels were affected by standing blood. B. Analysis of Blood Taken from Sarin Poisoned Patients Twelve sarin poisoned patients were selected as subjects, and serum cholinesterase (Ch-E) and serum albumin (Alb) levels were determined. Amino acid analysis was conducted using an amino acid analyzer. Serum samples which had been obtained from the 6 patients and frozen and stored at -40 degrees C from 5 months were used for amino acid analysis. As a result, the serum Ch-E level decreased and the Alb level tended to rise. Since the Ch-E/Alb ratio was reduced in the sarin poisoned patients, it is considered useful for discrimination from liver cirrhosis in which both Ch-E and Alb levels decreased. Amino acid levels in the serum obtained from the sarin poisoned patients were compared with those of healthy individuals, both of which had been stored under the same conditions. There were significant differences in Asp, glutamate (Glu), phenylalanine (Phe), 3-M-His, glutamine (Gln), and Cys levels. The Glu, Phe, and Gln levels were not affected by storage of serum in a frozen state, while the Glu and Phe levels were elevated and the Gln level was reduced. Although Cys exhibited lower values in frozen serum samples, the Cys level was elevated with a rise in the serum Ch-E levels. Therefore, we deduced that Cys metabolism disorders also occur in sarin poisoning. As stated above, the Glu and Phe levels were elevated and the Gln and Cys levels were reduced, suggesting the presence of abnormal amino acid metabolism, in patients with sarin-poisoning.

Adolescent↗
PubMed1996

Expression of fos family and jun family proto-oncogenes during corneal epithelial wound healing.

PURPOSE: While transformation of epithelial cells to a motile form is the first step in wound healing of the corneal epithelium, the migratory mechanism in these cells is not fully understood. We studied the expression of proto-oncogene mRNAs: c-fos; c-jun; fos B; jun B; jun D in injured corneal epithelium using in situ hybridization. Moreover, we examined immunolocalization of c-Fos and c-Jun protein products to elucidate the transcriptional activation prior to the onset of migration in corneal epithelium. METHODS: An epithelial defect was made on one cornea of 60 Wistar rats. The affected eye was enucleated immediately (within 5 min) or was allowed to heal for 15, 30, 60, 90, 120 and 180 min. Frozen sections were processed for in situ hybridization with c-fos, c-jun, fos B, jun B and jun D mRNAs or were stained with anti-c-fos and anti-c-jun antibodies. RESULTS: Fifteen min after the epithelial ablation, weak signals for c-fos and c-jun mRNAs were detected in the corneal epithelium surrounding the wound. These signals reached a peak 30 to 60 min after ablation, but were no longer evident at 120 min. Immunoreactivities for these proteins were also detected in the same area at 60 to 120 min after the epithelial ablation. Fos B mRNA was detected in the same region at 30 min after the ablation, and reached its peak after 30 to 60 min, but was no longer evident at 120 min. Jun B mRNA was detected in the epithelium around the defect 60 min after the ablation, later than the other proto-oncogenes, and reached its peak after 90 min. The message for jun D was detected in normal epithelium, and was not affected by wounding. CONCLUSIONS: These findings indicate that transcriptional activation of epithelial cells is initiated in the early phase after epithelial ablation, before the cells start to migrate, and that these proto-oncogene products may play important roles in wound healing in corneal epithelium. The time lag of the peak of expression of these proto-oncogenes in this process.

Animals↗
PubMed1995

Effect of lysyl hydroxylase inhibitor, minoxidil, on ultrastructure and behavior of cultured rabbit subconjunctival fibroblasts.

BACKGROUND: Minoxidil is an inhibitor of lysyl hydroxylase, an enzyme involved in collagen production, and decreases collagen production in vitro. We investigated the in vitro effects of minoxidil on behavior such as proliferation and migration of rabbit subconjunctival fibroblasts (SCFs). The ultrastructural effect of the drug on SCFs was also examined. METHODS: Proliferation of SCFs and closure of the defect produced in monolayer cultures in the presence or absence of minoxidil was studied. The ultrastructure of SCFs treated with minoxidil was also examined. RESULTS: Minoxidil inhibited SCF proliferation and the closure of the defect produced in monolayer cell sheets. Ultrastructural observations revealed extensive areas of irregularly dilated endoplasmic reticulum in cells treated with minoxidil, indicating the accumulation of protein, probably underhydroxylated collagen precursors, in the cisternae of endoplasmic reticulum. CONCLUSIONS: The results indicated that minoxidil attenuated cellular activities of SCFs such as proliferation and migration in vitro. The exact mechanism of the inhibitory effects of minoxidil on these cellular activities is unknown. The findings suggest that the drug might help to prevent bleb scarring after glaucoma filtering surgery.

Animals↗
PubMed1995

Type IV collagen in the basement membrane of the corneal epithelium after alkali burns in guinea pigs.

To investigate the changes in the corneal epithelial basement membrane following an alkali burn, we examined the immunolocalization of type IV collagen and laminin in the eye of the guinea pig burned with alkali. The burn damaged the corneal, limbal and conjunctival epithelium. After regeneration, basement membrane was interrupted, as indicated by laminin immunoreactivity. Type IV collagen immunoreactivity was transiently expressed in the early healing phase in the epithelial derived from both the cornea and conjunctiva, but was not seen in the normal corneal epithelial basement membrane. Later in the healing process, following transdifferentiation of the conjunctival epithelium into a cornea-like epithelium, its type IV collagen immunoreactivity was weaker than that in the basement membrane of the nontransdifferentiated epithelium. Conjunctival transdifferentiation during healing may have led to transient development of type IV collagen immunoreactivity.

Animals↗
PubMed1995

Effect of a prolyl hydroxylase inhibitor on rabbit ocular fibroblasts.

We examined the effect of a prolyl hydroxylase inhibitor, ethyl-3,4-dihydroxybenzoate (E-DHB), on collagen production, proliferation, and spreading by cultured rabbit keratocytes and subconjunctival fibroblasts. Proliferating cells were incubated with or without E-DHB, and were examined by phase-contrast microscopy and proliferation assays. In addition, wounds were made in confluent cultures and were allowed to heal with or without exposure to E-DHB. Confluent cells were also observed by transmission electron microscopy after exposure to E-DHB (0.2-0.4 mM). Finally, confluent cells were incubated in the presence or absence of E-DHB, after which the concentration of soluble collagen in the culture medium was determined. E-DHB (0.2-0.4 mM) inhibited the growth of rabbit keratocytes and subconjunctival fibroblasts and decreased collagen production without inducing cytotoxicity.. Wounds in the control culture were initially healed by individual migrating cells and later by spreading monolayer sheets and proliferating cells, while E-DHB inhibited cell migration into the wound. Electron microscopy revealed large irregular cisternae of endoplasmic reticulum in E-DHB-treated cells, possibly due to protein accumulation. These findings suggest that collagen production may be closely related to various cellular activities.

Animals↗
PubMed1995

Changes in the concentration and distribution of lipoprotein (a) in plasma after fat intake.

To clarify the association between apo(a) and TG-rich lipoproteins, we studied changes in plasma Lp(a) concentration and apo(a) distribution in lipoprotein fractions after fat intake. The subjects were 15 hyperlipidemic patients and 3 healthy volunteers with fasting Lp(a) concentrations ranging from 2.5-52 mg/dl. They were given a fatty meal (50 g fat/m2 and 60 mg retinyl palmitate) after a 12-hour overnight fast and venous blood samples were taken at 0, 3, 4.5, 6, 7.5, 9, 12 and 24 hours. Fractions of sf > 400, sf 20-400 and d > 1.006 g/ml were isolated from plasma samples by ultracentrifugation. Plasma Lp(a) levels increased transiently at 4.5 hours, decreased between 4.5 and 12 hours, and recovered almost to initial levels by the next morning. Plasma Lp(a) peaked before the plasma RP peak appeared. Apo(a) associated with TG-rich lipoprotein fractions increased and apo(a) in the d > 1.006 fraction decreased after fat intake in most of the subjects, suggesting the transfer of apo(a) from the d > 1.006 fraction to the TG-rich lipoproteins. The increase in TG-rich lipoprotein apo(a) correlated with the RP area under the curve (r=0.79, p<0.05) and the decrease in d > 1.006 apo(a) (r = 0.80, p < 0.05). This distributional change of apo(a) after fat intake was confirmed by gel filtration and density gradient ultracentrifugation. Transfer of apo(a) from the main Lp(a) fraction of the plasma obtained from a subject with a high Lp(a)level to the TG-rich lipoprotein fraction of the plasma obtained 4.5 hours after fat intake from a subject with a low plasma Lp(a) level was also shown in vitro by density gradient ultracentrifugation. Our studies revealed a significant association between apo(a) and TG-rich lipoproteins in the postprandial. Further studies are necessary to clarify the pathophysiological role of Lp(a) in TG-rich lipoproteins.

Adult↗
PubMed1994

Prolyl hydroxylase inhibitor and lysyl hydroxylase inhibitor inhibit spreading of corneal epithelium.

BACKGROUND: The spreading of epithelium is critical in the healing of corneal wounds. Such epithelial spreading requires the continuous production of protein and glycoprotein. To determine whether collagen production is required for the spreading of corneal epithelium, we studied the effects of inhibitors for collagen production on spreading of corneal epithelium in vitro. METHODS: We examined the effect of two proline analogs, L-azetidine 2-carboxylic acid and cis-hydroxyproline, a prolyl hydroxylase inhibitor, ethyl-3,4-dihydroxybenzoate, and a lysyl hydroxylase inhibitor, minoxidil, on the spreading of epithelium of organ-cultured rabbit cornea. RESULTS: Both analogs and inhibitors inhibited epithelial spreading in a dose-dependent manner. CONCLUSION: These observations indicate that collagen production may be involved in the spreading of corneal epithelium.

Animals↗
PubMed1994

Effects of alcohol consumption on antioxidant content and susceptibility of low-density lipoprotein to oxidative modification.

OBJECTIVE: The effects of moderate alcohol intake on antioxidant content of low-density lipoprotein (LDL) and the susceptibility of LDL to oxidative modification were examined in 12 healthy adult males. DESIGN: Volunteers abstained from alcohol for 3 weeks and then 12 subjects (alcohol intake group) consumed alcohol (0.5 g/kg/day) as brandy for 4 weeks; 4 subjects (control abstinence group) did not consume any alcohol for the entire study. RESULTS: In the alcohol intake group, plasma total cholesterol (TC), triglyceride (TG), phospholipids (PL), free cholesterol (FC) and apoprotein B (apoB) levels in LDL decreased significantly after alcohol intake; however, since TC/apoB, TG/apoB, PL/apoB and FC/apoB ratios did not change significantly, it is clear that LDL particle numbers decreased. Vitamin E and vitamin A levels in plasma, and vitamin E content of LDL also did not change significantly. Beta-carotene levels in plasma and in LDL decreased significantly in the alcohol intake group. In the abstinence group, lipid levels and vitamin levels did not change. Lag time before the onset of LDL oxidation and propagation rate of LDL oxidation in the alcohol intake group did not change significantly. CONCLUSIONS: Moderate alcohol intake decreases particle numbers of LDL without any changes in chemical composition, vitamin E content and susceptibility of LDL to oxidative modification. However, beta-carotene content was decreased significantly by even moderate alcohol intake.

Adult↗
PubMed1994

Drug binding properties of glycosylated human serum albumin as measured by fluorescence and circular dichroism.

Binding properties of Sudlow's site-specific drugs to glycosylated human serum albumin (G-HSA) were investigated using fluorescence and circular dichroism (CD). Dansylamide, phenylbutazone and warfarin were used as site I-specific drugs, and dansylproline, ibuprofen and flufenamic acid were used as site II-specific ones. Similar changes in the fluorescence intensity of dansylamide occurred in the presence of both G-HSA and intact human serum albumin (HSA), while the fluorescence enhancement of dansylproline caused by G-HSA was extremely weakened in comparison with that by HSA. These results suggest that the glycosylation of HSA inhibits the binding of the site II-specific drug, dansylproline, to HSA, while it does not influence the binding of the site I specific drug, dansylamide. The induced ellipticities of the complexes of ibuprofen, flufenamic acid and phenyl butazone with G-HSA were diminished in comparison with those with HSA. With the complexes of warfarin, the induced ellipticity was enhanced. These CD results suggest that the glycosylation of HSA induces microenvironmental changes in the binding sites for the above site-specific drugs which influence the drug binding ability of HSA.

Binding Sites↗
PubMed1993

Epithelial basement membrane in alkali-burned corneas in rats. Immunohistochemical study.

We investigated the altered immunolocalization of the components of epithelial basement membrane (BM) in rats with alkali burns of the cornea. Type IV collagen, laminin, and fibronectin were immunohistochemically stained in the alkali-burned corneas after various intervals. Linear laminin immunoreactivity, which represented the localization of the BM, was interrupted, probably by epithelial enzymes, under the regenerating epithelium after 24 h. A normal configuration was restored on day 7, when the central corneal BM showed a type IV collagen immunoreactivity that was not detected in BM of normal cornea. The exact cause of the development of type IV collagen immunoreactivity in the healing BM was not determined. Laminin, type IV collagen, and fibronectin, which were not detected in normal stroma, were all detected in the healing corneal stroma with repopulated keratocytes. Moreover, the BM zone under the regenerating epithelium showed fibronectin immunoreactivity on days 5-14. Distribution of these components of BM seemed to be normal 2 months postburn. The alteration of BM components is thought to be an important marker of the healing process in corneas burned with alkali.

Animals↗
PubMed1993

Cis-hydroxyproline inhibits adhesion, migration and proliferation of cultured rabbit keratocytes.

Recruitment of keratocytes into injured corneal stroma, and secretion of proteins including collagen in the cells are essential for wound healing of the corneal stroma. We examined the effect of a proline analog, cis-hydroxyproline, on the adhesion, migration and growth of rabbit keratocytes in vitro. This agent decreased the plating efficiency, migration and growth of the keratocytes in a dose-dependent manner. Reduction in these cellular activities may reflect altered functions of pericellular proteins such as collagen. Further studies are needed to determine which specific protein is involved.

Animals↗
PubMed1993

[Vitamin deficiency and marginal vitamin deficiency].

Fat-soluble vitamin deficiency and marginal deficiency of this type of vitamin will be discussed. The trias of vitamin A deficiency is composed of nyctalopia, xerophthalmia and hyperkeratosis follicularis. Vitamin D deficiency causes rachitis and osteomalacia. In Japan, vitamin D deficiency is more often caused by impaired vitamin D activation due to renal failure, than by a lack of sunbathing or insufficient ingestion of vitamin D from diet. At present, it is quite rare that the physician encounters patients with deficiency of vitamin E or other vitamins. According to surveys of occult vitamin deficiency, the prevalence of marginal fat-soluble vitamin deficiency was lower than that of marginal water-soluble vitamin deficiency. Marginal vitamin deficiency seems to be absent or very rare among healthy individuals. In patients, however, marginal vitamin deficiency is sometimes observed. Since marginal vitamin deficiency can modify the underlying disease or trigger complications, the physician should take adequate measures to prevent the onset of marginal vitamin deficiency in managing patients with various diseases.

Humans↗