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Biomedical subjects

K Horie

Publications and source records attributed to K Horie.

At least 73 records · Page 4Linked to original sources
PubMed1996

Immunohistochemical study of advanced glycation end products in aging and Alzheimer's disease brain.

Advanced glycation end products (AGEs) in the brain were immunohistochemically examined in Alzheimer's disease (AD) and aging using anti-AGE antibody recognizing mainly carboxymethyllysine. AGE positive staining diffusely located in the neuronal perikarya of hippocampus and parahippocampus in AD and aged brains without dementia, but not in young brains less than 17 years of age. Extra-neuroperikaryal AGE deposits were also detected in the neuropil of AD and aged brains. The extra-neuroperikaryal AGE deposits markedly increased in AD brains as compared to aged brains. These AGE-positive deposits in the neuropil were not related to the senile plaque identified by anti-beta amyloid protein antibody. These findings suggest a potential link of AGE accumulation in the central nervous system to the aging process of neurons and the degenerating process of AD neurons.

Adolescent↗
PubMed1996

Chemonucleolytic effects of chondroitinase ABC on normal rabbit intervertebral discs. Course of action up to 10 days postinjection and minimum effective dose.

STUDY DESIGN: This study demonstrated the chemonucleolytic effects of chondroitinase ABC and its histologic and biochemical background. OBJECTIVES: To determine the course of chondroitinase ABC action on normal rabbit discs, and to find its minimum effective dosage. SUMMARY OF BACKGROUND DATA: No previous study has assessed the chemonucleolytic action of chondroitinase ABC in a time- and dose-dependent manner. This study also investigated the biochemical causes of radiologic and histologic changes in the discs. METHODS: Rabbits were injected with 4 U of pharmaceutical-grade chondroitinase ABC intradiscally. They were radiologically and histologically observed, and biochemical analyses of the discs were conducted on days 1, 3, 5, 7, and 10 postinjection in the time course study. Different doses of chondroitinase ABC were injected, and radiologic observations and water content of the discs were measured in the dose-finding study. RESULTS: The time course study revealed that the chondroitin sulfate content of discs significantly decreased from day 1 postinjection until the end of the experimental period. The weight and water content of the nucleus pulposus decreased on day 3, and disc space narrowing was observed from the day after injection. The dose-finding study showed that a dose of 0.0002 U/disc still induced disc space narrowing and a decrease in water content. CONCLUSIONS: Chondroitinase ABC is estimated to have a chemonucleolytic effect at least by day 3 postinjection at a dose level of 0.0002 U/disc or higher in rabbits.

Animals↗
PubMed1996

Alpha 1b-adrenoceptor-mediated calcium oscillation is specific for the S phase in cell cycle and dependent on the extracellular calcium.

Using Chinese hamster ovary cells stably expressing alpha 1b-adrenoceptor (alpha 1bAR) as a model, we examined the effect of the cell cycle on the agonist-promoted intracellular [Ca2+]i oscillation. In cells synchronized into either the G1 state or the M phase, no oscillatory behavior was observed under any extracellular Ca2+ concentrations ([Ca2+]o < or = 3 mM), whereas in cells synchronized into the S phase, norepinephrine caused [Ca2+]i oscillation in a [Ca2+]o-dependent manner, indicating that alpha 1AR-mediated [Ca2+]i oscillation is specific for the S phase in cell cycle and dependent on [Ca2+]o. The S phase-specific occurrence of alpha 1AR-mediated [Ca2+]i oscillation is not associated with changes in alpha 1AR density. As the cells consistently developed [Ca2+]i oscillation in the S phase, the cells would provide a valuable system to study further the biochemical mechanism for agonist-induced [Ca2+]i oscillation phenomenon.

Animals↗
PubMed1996

Selected salivary-gland cell culture and the effects of isoproterenol, vasoactive intestinal polypeptide and substance P.

To establish a selected salivary-gland cell culture and determine the effect of neuropeptides, monolayers were cultured using 3T3 cells as a feeder layer. To confirm the origin of these cultured cells, amylase production was examined by electron microscopy and periodic acid-Schiff staining, together with immunocytochemical analysis of myosin, anti-cytokeratin (CK-1, CK 10/13, CK MNF116, CK LMW, CK HMW and CK-19) and amylase antibody. The cultured cells demonstrated secretion granules containing amylase and presented features characteristic of acinar cells, which they retained until passage two. By using a feeder layer in conjunction with a newly formulated culture medium, the selectability of these cells was improved. Changes in proliferation of cultured salivary-gland cells in the presence of selected neurotransmitters were also examined. Isoproterenol enhanced cellular proliferation. On the other hand, vasoactive intestinal polypeptide and substance P, which increase the weight of salivary glands in vivo, showed no significant enhancement of proliferation.

3T3 Cells↗
PubMed1996

Effect of 'bakumondo-to', a Chinese-Japanese herbal medicine, on cultured and dispersed salivary gland cells.

'Bakumondo-to', a Chinese-Japanese herbal medicine, has been used for patients with xerostomia in Japan. Although the efficacy of this medicine for xerostomia has been reported, the pharmacological basis was only partially understood. The aim of this study was to clarify the direct effect of Bakumondo-to on salivary gland cells using isolated and cultured cells. In the physiological experiment using the fluorescent dye fura-2, Bakumondo-to showed no direct effect on isolated parotid gland cells. On the other hand, Bakumondo-to, when applied to cultured salivary gland cells, showed enhancement effects on cell proliferation. After inspection by transmission electron microscopy, we concluded that Bakumondo-to did not show an increase in the number of secretion granules but did increase the mean size of secretion granules in parotid gland cells. These mechanisms, together with other in vivo effecters, may contribute to clinical efficacy.

Bromodeoxyuridine↗
PubMed1996

Structural changes and cell viability of cultured epithelium after freezing storage.

Numerous clinical reports have shown the utility of cultured epithelial grafting in the field of plastic and reconstruction surgery. Recently, freezing storage of the cultured epithelium has been tried and has successfully grafted after thawing. It is clinically convenient if it is possible for cultured epithelium to keep its normal structure and viability. However, few papers have described the structural changes in cultured epithelium after freezing storage. In the present study, the morphological changes and cell viability of cultured mucosal epithelial sheets after freezing were studied in comparison with cultured epidermal sheets. Furthermore, we discuss the effect of storage temperature and cryoprotectants. As a result, there were some structural changes such as vacuolar degeneration in the cultured mucosal sheets using dimethyl sulphoxide (DMSO) as a cryoprotectant. Such changes were more clearly observed at -80 degrees C than at -196 degrees C with DMSO. However, little morphological change was observed in both epithelial sheets cultured with glycerin. The cell viability analysed by flow cytometry showed that more than 62% of the cells kept their viability after freezing storage. These results suggest that the optimum conditions of freezing for cultured epithelium were -196 degrees C storage by slow cooling methods with glycerin as a cryoprotectant.

Cell Survival↗
PubMed1996

Protective efficacy of sodium hyaluronate on the corneal endothelium against the damage induced by sonication.

The protective efficacy of sodium hyaluronate (Na-HA) on the corneal endothelium against the damage induced by sonication was investigated using enucleated rabbit eyes and Na-HA fluorescence labeled with 5-aminofluorescein (FA-HA). The anterior chamber was reformed by injecting a 1% solution of FA-HA, and then sonication, irrigation and aspiration were performed in the anterior chamber using a phaco-needle attached to phacoemulsification and aspiration (PEA) equipment. The protective efficacy was evaluated by the area of damaged corneal endothelium. When the anterior chamber was reformed by the solution of FA-HA with a molecular weight of 2010 x 10(3) (FA-HA (2010 x 10(3) group), most of the FA-HA was eliminated within 20 seconds after starting the irrigation and aspiration, and the area of damaged endothelium was the same with the case when the FA-HA was not pre-injected (Control group). On the other hand, when the anterior chamber was reformed by the solution of FA-HA with a molecular weight of 1130 x 10(3) (FA-HA (1130 x 10(3) group), the FA-HA was gradually eliminated after mixing with an irrigating solution, and the area of damaged endothelium was significantly smaller than those of the Control and FA-HA (2010 x 10(3)) groups. These results suggest that the corneal endothelial damage induced by PEA can be avoided by pre-injecting a viscoelastic material which can remain in the anterior chamber during PEA.

Animals↗
PubMed1996

The effect of sodium hyaluronate on the growth of rabbit corneal epithelial cells in vitro.

The effect of sodium hyaluronate (Na-HA) on the growth of rabbit corneal epithelial cells was investigated in vitro by using MTT assay and [3H]-thymidine incorporation. Na-HA, with a molecular weight of 86 x 10(4), enhanced the growth of the corneal epithelial cells cultured on a fibronectin (FN) coated dish, but when cells were cultured on a noncoated dish, Na-HA did not enhance the growth of the cells. Na-HA, labeled with 5-aminofluorescein (FA-HA), bound to the corneal epithelial cells, and the CD44-like molecule was identified on the cells by using a monoclonal antibody, OS/37. Furthermore, the binding of FA-HA to the cells was significantly inhibited by the monoclonal antibody. These results suggest that Na-HA binds to the CD44-like molecule associated with FN and then enhances the growth of the corneal epithelial cells.

Animals↗
PubMed1996

Pathophysiology of advanced glycation end-products in renal failure.

beta 2-Microglobulin is a major constituent of amyloid fibrils in dialysis-related amyloidosis, a serious complication leading to bone and joint destruction in long-term haemodialysis patients. However, the molecular pathogenesis of this complication remains unknown. Intact beta 2-microglobulin per se seems an unlikely contributor to the pathogenesis, because no difference in the plasma levels of intact beta 2-microglobulin has yet been found between haemodialysis patients with and without this complication. Some investigators have therefore focused on the modification of this molecule. Recent studies have revealed a new modification of beta 2-microglobulin in amyloid fibrils-the advanced glycation end-products (AGEs) formed by a non-enzymatic reaction between sugar aldose and protein. Further studies have suggested that the interaction of AGE-modified beta 2-microglobulin with monocyte/macrophage and osteoclast/osteoblast gives a plausible albeit partial explanation for the mechanism of bone and joint destruction in dialysis-related amyloidosis. This article discusses the pathophysiology of AGEs in renal failure and the modification of beta 2-microglobulin with AGEs, especially focusing on their structure and pathological role in dialysis-related amyloidosis.

Amyloid↗
PubMed1996

Advanced glycation end-products in diabetic nephropathy.

Advanced glycation end-products (AGEs) are the pigmented and fluorescent adduct formed by a non-enzymatic reaction between sugar and protein. Since AGEs are generated in high glucose milieu, then induce the structural and functional alteration of matnx proteins, and have biological effects on various kinds of cells including mesangial cells, AGEs have been implicated in tissue damage of diabetic nephropathy. To elucidate the factor(s) that determine the AGEs level in diabetic nephropathy, we quantitated the plasma pentosidine level of different status of diabetic nephropathy by HPLC assay. The plasma pentosidine level in diabetic nephropathy was found to be determined by factors such as renal function control of glucose and the patient's age; of these, renal function was the most critical factor. For a better understanding of the pathological role of AGEs in diabetic nephropathy, we then examined renal tissues of diabetic nephropathy immunohistochemically using antibodies specific for AGE proteins. Immunohistochemistry revealed the positive immunostaining for AGEs in the expanded mesangial area of diffuse diabetic glomerulosclerosis. The degree of staining was stronger than that in patients of IgA nephropathy with a similar degree of mesangial expansion. The nodular lesions, characteristic of diabetic nephropathy, were also stained positive for AGEs. These findings suggests a potential link of AGEs accumulation, which may be determined by renal function, control of glucose and age, to renal tissue damage in diabetic nephropathy.

Animals↗
PubMed1996

Oxytocin receptors expressed and coupled to Ca2+ signalling in a human vascular smooth muscle cell line.

1. In a human vascular smooth muscle cell line (HVSMC), binding experiments with [3H]-arginine8-vasopressin (AVP) have shown the existence of a homogeneous population of binding sites with affinity (Kd value) of 0.65 nM and a maximum number of binding sites (Bmax) of 122 fmol mg-1 protein. 2. Nonlabelled compounds compete for [3H]-AVP binding in the HVSMC membrane with an order of potency of oxytocin > lyspressin > or = AVP > Thr4, Gly7-oxytocin > (beta-mercapto-beta-beta-cyclopentamethylenepropionyl-O-Me Tyr2, Arg8) vasopressin > desmopressin > OPC21268 > OPC31260. This order was markedly different from that observed in rat vascular smooth muscle cells (A10), a well-established V1A receptor system. 3. In HVSMC both oxytocin and AVP increased inositol 1,4,5-trisphosphate (IP3) production and [Ca2+]i response, but the efficacy of the responses was greater for oxytocin than AVP. 4. Reverse transcription-polymerase chain reaction (RT-PCR) assay detected only oxytocin receptor but not V1A or V2 receptors in HVSMC, whereas only V1A receptors were found in A10 cells. 5. In conclusion, in HVSMC only oxytocin receptors are expressed among the vasopressin receptor family, and they coupled to phosphatidyl inositol (PI) turnover/Ca2+ signalling. This unexpected observation should provide new insight into the functional role of the oxytocin receptor in a human vascular smooth muscle cell line.

Animals↗
PubMed1996

Hyaluronan and chondroitin sulfate in rabbit tears.

The concentrations of hyaluronan (HA) and chondroitin sulfate (CS) in rabbit tears were determined using high performance liquid chromatography. The tears were taken from both normal eyes and eyes with a corneal epithelial defect. The concentration of CS was 7.61 times higher than that of HA in the tears of normal eyes. When the concentration of CS isomers was compared, the highest concentration was observed in chondroitin 6-sulfate (C6S), followed by chondroitin 4-sulfate (C4S) and chondroitin (C0S), in that order. When the corneal epithelium was removed from the left eye using a trephine and microscissors, a significant increase was detected in the concentrations of HA and C6S in tears of the left eye one day after removal, whereas no change was observed in those of the right eye. In addition, a significant positive correlation was observed between the increased amount of HA concentration and the healing rate of the corneal epithelium. These results suggest that the increase of HA concentration in tears is associated with spontaneous corneal epithelial healing.

Animals↗
PubMed1996

Hyaluronan, CD44 and fibronectin in rabbit corneal epithelial wound healing.

Changes in hyaluronan (HA) concentration and stainability were investigated in the rabbit cornea after wounds made by exposure to n-heptanol. The HA concentration in the cornea increased gradually until day 14 after wounding, and then decreased. The HA concentration returned to the normal level 56 days after wounding. In the normal control cornea, HA staining was observed in the epithelium and stroma. The intensity of HA staining in the epithelium and stroma increased until day 3 after wounding, when the epithelium had completely covered the defect. At day 28, when the thickness of the corneal epithelium returned to the normal level, the intensity of HA staining in the epithelium also decreased. However, staining in the stroma was still strong. HA staining in the stroma decreased by day 56 after wounding. In parallel experiments, the immunostaining for CD44, an HA receptor, and fibronectin (FN) was carried out in the same model. The immunostaining in the epithelium of both CD44 and FN was synchronistic with the HA staining during the early stages after wounding. These events suggest that HA, CD44 and FN cooperatively play important roles in corneal epithelial wound healing.

Alcohols↗
PubMed1996

GpIIb/IIIa+ subpopulation of rat megakaryocyte progenitor cells exhibits high responsiveness to human thrombopoietin.

The recently cloned factor thrombopoietin (TPO) has been shown to exhibit megakaryocyte colony-stimulating activity in vitro. In this investigation, to further evaluate the action of TPO on megakaryocyte progenitor cells (colony-forming units-megakaryocyte [CFU-MK]), GpIIb/IIIa+ and GpIIb/IIIa- populations of CFU-MK were prepared from rat bone marrow cells based on their reactivity with P55 antibody, a monoclonal antibody against rat GpIIb/IIIa, and their responsiveness to recombinant human TPO (rhTPO) and recombinant rat interleukin-3 (rrIL-3) was examined using a megakaryocyte colony-forming assay (Meg-CSA). rhTPO supported only megakaryocyte colony growth from both fractions in a dose-dependent fashion. The mean colony size observed with the GpIIb/IIIa+ population was smaller than that seen with the GpIIb/IIIa- population. With the optimal concentration of either rhTPO or rrIL-3, similar numbers of megakaryocyte colonies were formed from the GpIIb/IIIa+ population previously shown to be highly enriched for CFU-MK. In contrast, the maximum number of megakaryocyte colonies from the GpIIb/IIIa- population stimulated by rhTPO was only 24.2% of that achieved with rrIL-3. Morphologic analysis of rhTPO-promoted megakaryocyte colonies from the GpIIb/IIIa+ population showed that the average colony size was smaller but that the mean diameter of individual megakaryocytes was larger than in megakaryocyte colonies promoted with rrIL-3. rhTPO plus rrIL-3, each at suboptimal concentrations, had an additive effect on proliferation of CFU-MK in the GpIIb/IIIa+ fraction, whereas rhTPO plus murine IL-6 or murine granulocyte-macrophage colony-stimulating factor (mG-M-CSF) modestly but significantly reduced megakaryocyte colony growth. These results indicate that TPO preferentially acts on GpIIb/IIIa+ late CFU-MK with lower proliferative capacity and interacts with some other cytokines in CFU-MK development.

Animals↗
PubMed1995

A replacement vector used to introduce subtle mutations into mouse genes.

A replacement vector convenient for introducing subtle mutations into various mouse genes has been developed using, a model system, the mouse transthyretin-encoding gene (ttr) and mouse embryonal carcinoma F9 cells. The vector consists of part of ttr carrying a subtle mutation in its second exon, and a cassette of the neomycin-resistance (neo)- and herpes simplex virus thymidine kinase (HSV-tk)-encoding genes flanked with a 3-kb duplication of mostly the second intron of ttr. In the first step ('replacement'), part of the endogenous ttr was replaced by vector DNA via homologous recombination, and two such clones, #33 and #77, were isolated from 185 G418-resistant clones by allele-specific PCR. In the second step ('excision'), gancyclovir-resistant colonies were screened, and 7 and 84% of those isolated from clones #33 and #77, respectively, were demonstrated to carry the subtle mutation in ttr, without the cassette of selection markers. In five independently isolated random integrants of the same vector DNA, the cassette of selection markers was excised efficiently by recombination within the duplication.

Animals↗
PubMed1995

NS-49, a novel alpha 1a-adrenoceptor-selective agonist characterization using recombinant human alpha 1-adrenoceptors.

alpha 1-Adrenoreceptors comprise a heterogeneous family and subtype-selective ligands are valuable in studying the functional role of each receptor subtype. Using the Chinese hamster ovary (CHO) cells stably expressing the cloned human alpha 1-adrenoceptor subtypes (alpha 1a, alpha 1b, and alpha 1d)1, we have compared a newly synthesized phenethylamine class agonist (R)-(-)-3'-(2-amino-1-hydroxyethyl)-4'-fluoromethanesulfonanilide hydrochloride (NS-49) with imidazoline class agonist oxymetazoline in their binding affinities and intrinsic activities in causing transient increases of cytosolic Ca2+ concentrations ([Ca2+¿i response). Radioligand binding studies with 2-[beta-(4-hydroxyl-3-[125I]iodophenyl)ethylamino-methyl]tetralone ([125I]HEAT) showed NS-49 and oxymetazoline had higher affinities at alpha 1a-than at alpha 1b- and alpha 1d-subtypes (-log Ki values at alpha 1a-, alpha 1b-and alpha 1d-subtype: 6.18, 5.13, and 5.38 for NS-49: 8.19, 6.50, and 6.44 for oxymetazoline, respectively). In functional studies, both oxymetazoline and NS-49 worked as a selective and partial agonist at alpha 1a-subtype: however, NS-49 is more efficacious than oxymetazoline. NS-49 is the phenethylamine class of alpha 1-adrenoceptor partial agonist relatively selective and efficacious for the human alpha 1a-adrenoceptor subtype, NS-49 would be potentially useful for studying the physiological role of alpha 1-adrenoceptor subtype.

Adrenergic alpha-Agonists↗
PubMed1995

Cloning, functional expression and tissue distribution of human alpha 1c-adrenoceptor splice variants.

We report the cloning and characterization of two isoforms of human alpha 1c-adrenoceptor cDNA (alpha 1c-2, alpha 1c-3). These isoforms are generated by alternative splicing and differ from the clone we previously isolated (alpha 1c-1) in their length and sequences of the C-terminal domain. Tissue distribution of mRNAs showed that these variants co-express with alpha 1c-1 in the human heart, liver, cerebellum and cerebrum. Despite the structural differences, functional experiments in transfected CHO cells showed that the three isoforms have similar ligand binding properties, and all couple with phospholipase C/Ca2+ signaling pathway.

Alternative Splicing↗