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Biomedical subjects

S Horiuchi

Publications and source records attributed to S Horiuchi.

At least 181 records · Page 10Linked to original sources
PubMed1996

Accumulation of advanced glycation end products of the Maillard reaction with age in human hippocampal neurons.

The recent immunological demonstration of advanced glycation end products (AGE) of the Maillard reaction in several human tissues suggests a possible involvement of AGE in the aging process. We previously prepared a monoclonal anti-AGE antibody (6D12) which recognized N epsilon-(carboxymethyl)lysine. We examined, immunohistochemically, the effect of aging on AGE-immunoreactivity in hippocampal pyramidal neurons in ten brain tissue samples obtained at autopsy from subjects aged 20-85 years old. Using 6D12 antibody, our results demonstrated a positive correlation between AGE-immunoreactivity in hippocampal pyramidal neurons and age. A more intense immunoreaction was observed in the CA3-4 pyramidal neurons compared with that of the CA1 neurons, known to be vulnerable to the formation of neurofibrillary tangles. Our results suggest that AGE are probably involved in the aging process affecting the human central nervous system, and that AGE do not mainly contribute to the formation of neurofibrillary tangles, at least in the CA1 neurons.

Adult↗
PubMed1996

Identification of pentosidine as a native structure for advanced glycation end products in beta-2-microglobulin-containing amyloid fibrils in patients with dialysis-related amyloidosis.

beta-2-Microglobulin (beta-2m) is a major constituent of amyloid fibrils in patients with dialysis-related amyloidosis (DRA). Recently, we found that the pigmented and fluorescent adducts formed nonenzymatically between sugar and protein, known as advanced glycation end products (AGEs), were present in beta-2m-containing amyloid fibrils, suggesting the possible involvement of AGE-modified beta-2m in bone and joint destruction in DRA. As an extension of our search for the native structure of AGEs in beta-2m of patients with DRA, the present study focused on pentosidine, a fluorescent cross-linked glycoxidation product. Determination by both HPLC assay and competitive ELISA demonstrated a significant amount of pentosidine in amyloid-fibril beta-2m from long-term hemodialysis patients with DRA, and the acidic isoform of beta-2m in the serum and urine of hemodialysis patients. A further immunohistochemical study revealed the positive immunostaining for pentosidine and immunoreactive AGEs and beta-2m in macrophage-infiltrated amyloid deposits of long-term hemodialysis patients with DRA. These findings implicate a potential link of glycoxidation products in long-lived beta-2m-containing amyloid fibrils to the pathogenesis of DRA.

Amyloid↗
PubMed1996

Reconstituted high density lipoprotein reduces the capacity of oxidatively modified low density lipoprotein to accumulate cholesteryl esters in mouse peritoneal macrophages.

Oxidized low density lipoprotein (ox-LDL) was incubated with discoidal complexes of apolipoprotein A-I (apo A-I) and dimyristoylphosphatidylcholine (DMPC) (DMPC/apo A-I) in a cell-free system and re-isolated on Sephacryl S-400 gel filtration chromatography. Analyses of re-isolated ox-LDL showed that apo A-I was transferred from DMPC/apo A-I to ox-LDL, which accounted for 10% of the total protein of ox-LDL. Re-isolated ox-LDL also showed a 2.2-fold increase in phospholipid and a 14% decrease in cholesterol content on an apo B basis. The electrophoretic mobility of re-isolated ox-LDL was markedly reduced almost to that of native LDL. Moreover, the amounts of re-isolated ox-LDL to be degraded by mouse peritoneal macrophages as well as the capacity of re-isolated ox-LDL to accumulate cholesteryl esters (CE) in these cells were markedly reduced (60% and 80% reduction, respectively), suggesting that the ligand activity of ox-LDL for the scavenger receptor was significantly reduced upon treatment with DMPC/apo A-I. Parallel incubation of ox-LDL with free apo A-I led to a similar incorporation of apo A-I into ox-LDL. However, it had no effects on the ligand activity of ox-LDL. Thus, it is likely that the reduction in the ligand activity of ox-LDL by DMPC/apo A-I is explained by the change in the lipid moiety (mainly phospholipid) of ox-LDL. Since discoidal high density lipoprotein (HDL) is known to occur in vivo, this phenomenon might explain one of the anti-atherogenic functions of HDL.

Animals↗
PubMed1996

Purification of a marine bacterial glucose dehydrogenase from Cytophaga marinoflava and its application for measurement of 1,5-anhydro-D-glucitol.

A novel glucose dehydrogenase (GDH) from a marine bacterium Cytophaga marinoflava IFO 14170 was isolated from its membrane fraction. This GDH catalyzes the oxidation of a hydroxy group of glucose, but does not react in its C-1 position. This enzyme is composed of a single peptide with a mol wt of 67,000. The GDH can react under high salinity. The optimum pH is around 8.0, showing typical property of marine bacterial enzymes. Using this novel enzyme, and enzymatic determination of 1,5-anhydro-D-glucitol (1,5AG) utilizing 2,6-dichrolophenolindophenol (DCIP) and phenazine methosulfate (PMS) as electron mediators was carried out. A good linear correlation was observed from 0.5 mM to 4 mM of 1,5AG.

Chromatography↗
PubMed1996

The heparin-bound fraction of human lipoprotein-deficient serum inhibits endocytic uptake of oxidized low density lipoprotein by macrophages.

We recently demonstrated that bovine lactoferrin, a cationic whey protein from bovine milk, interacts with the negative charges of modified low density lipoproteins (modified LDL) such as acetylated LDL (acLDL) and oxidized LDL (oxLDL), which markedly interferes with their endocytic uptake by rat peritoneal macrophages (Kajikawa M, Ohta T, Takase M, Kawase K, Shimamura S, Matsuda I. Biochim Biophys Acta 1994;1213:82-90). In the present study, we examined whether human lipoprotein-deficient serum (LPDS) might contain protein(s) that could inhibit the endocytic uptake of oxLDL by mouse macrophages. We fractionated LPDS by heparin affinity chromatography and found that the cellular binding of oxLDL to mouse macrophages and subsequent endocytic uptake were inhibited by 50%-60% with the heparin-bound fraction, whereas the heparin-unbound fraction had no effect. Similar results were obtained in the experiments with acetylated LDL. Sephacryl S-300 gel-filtration chromatography of a mixture of oxLDL and the heparin-bound fraction revealed that a 150-kDa protein was associated with oxLDL. These results indicate that the electrostatic interaction of oxLDL with some component(s) of the heparin-bound fraction might interfere with the endocytic uptake of oxLDL by the macrophage scavenger receptor.

Animals↗
PubMed1996

Roles of advanced glycation end-products in the progression of diabetic nephropathy.

Available data indicate that the development of diabetic nephropathy is linked to hyperglycaemia. Glucose reacts nonenzymatically with proteins to form Schiff base and Amadori products. Further incubation of these early products leads to the formation of advanced glycation end-products (AGEs). AGEs seem to play a central role in the progression of diabetic nephropathy. Immunohistochemically, AGEs were also detected in an expanded mesangial matrix, especially in nodular lesions from patients with diabetic nephropathy. AGEs staining was noted in the Bowman's capsule, periglomerular fibrosis in sclerosing glomeruli. In our ultrastructural study of mesangial matrix from patients with diabetic nephropathy by high-resolution scanning electron microscopy after cellular removal, the meshwork structure was evident at higher magnification. In nodular lesions, the loose meshwork structure appeared to be composed of various sized strands, ranging from 6 to 24 nm (mean +/- SD: 11.4 +/- 3.8 nm). The pore sizes were variable, ranging from 4 to 70 nm (mean +/- SD: 23.6 +/- 12.3 nm), and were statistically larger than those of normal controls. As the AGEs are localized most notably in nodular lesions, advanced glycations play a role in the progression of diabetic nephropathy through impairment of the assembly of matrix proteins in vivo. Because type V and type VI collagens are the major components of nodular lesions, increases in these interstitial and fibril or microfibril collagens may contribute to the formation of wider strands in the mesangial matrix of a nodular lesion. As no metalloprotease that is specific for type VI collagen has been identified thus far, AGEs formation might occur preferentially in type VI collagen-rich nodular lesions, which are sites of slow turnover.

Diabetic Nephropathies↗
PubMed1996

Role of polyol pathway in nonenzymatic glycation.

In order to confirm the link between nonenzymatic glycation and the polyol pathway, we observed the effect of treatment with epalrestat (Ep), an aldose reductase inhibitor, on the concentration of advanced glycation end-products (AGEs) in erythrocytes from diabetic patients. We also examined the effect of the drug on erythrocyte fructose 3-phosphate (F3P), a novel metabolite that has been reported to relate to the polyol pathway, and ascertained the glycation capability of F3P and its possible breakdown product, 3-deoxyglucosone (3DG), by incubating the metabolites with bovine serum albumin (BSA). Incubation of BSA with F3P or 3DG resulted in a greater production of AGEs in comparison with the incubation with glucose or fructose. F3P was significantly increased in erythrocytes from diabetic patients compared with those from nondiabetic individuals and was lower in patients who had been treated with Ep than in those who were free from the compound. A treatment of patients with Ep for 1 month resulted in a significant decrease in F3P. Erythrocyte AGEs were significantly elevated in diabetic patients compared with nondiabetic individuals and tended to be lower in patients taking Ep than in those without Ep. Administration of Ep for 2 months decreased AGEs. These results show that the polyol pathway is likely to play a substantial role in the nonenzymatic glycation of proteins and the suppression of E3P as well as AGEs by an aldose reductase inhibitor may explain in part the preventive effect of the drug on diabetic complications.

Aldehyde Reductase↗
PubMed1996

Immunohistochemical study with anti-advanced glycation end-products antibody in murine amyloidosis.

Advanced glycation end-products (AGE) are formed in the late phase of the non-enzymatic glycosylation reaction in conditions such as diabetes mellitus and aging. In amyloidosis, AGE have been found in the A beta 2M amyloid associated with long-term hemodialysis and in the beta-protein in Alzheimer's disease. Murine AApoAII and AA amyloidosis were examined immunohistochemically using anti-AGE monoclonal antibody, 6D12. AApoAII amyloid deposits studied in one senescence-accelerated mouse P1 (SAMP1), congenic mice that have the amyloidogenic apolipoprotein A-II of SAMP1 mice, and AKR mice all reacted with biotinylated 6D12 by formic acid pretreatment, whereas AA amyloid deposits did not react with the antibody. The immunoreaction with anti-apolipoprotein A-II for amyloid deposits in senile mice was approximately homogeneous in intensity; on the other hand the reaction with biotinylated 6D12 was irregular in distribution and intensity over the amyloid deposits. These findings suggest that amyloid precursor proteins are not associated uniformly with AGE modification before deposition as amyloid; it is more likely that the AGE modification progresses gradually and unevenly after amyloid deposition. Murine amyloidosis may be a useful model to elucidate the role of AGE in amyloidosis.

Amyloidosis↗
PubMed1996

Fluoridation of hydroxyapatite powder by ammonium hexafluorosilicate.

Diamine silver fluoride [AgF: (NH3)2AgF] is a unique fluoride solution used clinically in Japan for primary teeth. AgF has been shown to reduce dental caries and dentine hypersensitivity. However, AgF also stains teeth black due to sulfide precipitation and thus is unacceptable for permanent teeth. In the present study, the potential value of ammonium hexafluorosilicate [SiF: (NH4)2SiF6], which has a formula similar to that of AgF but contains no Ag, was studied with respect to its ability to fluoridate apatite. Hydroxyapatite [HAP: Ca10(PO4)6(OH)2] powder was treated with several topical fluoride solutions, i.e. neutral sodium fluoride, acidulated phosphate fluoride (APF) along with AgF and SiF. Following treatment, the HAP powders were analyzed for their fluorine content, crystallinity changes, and solubility in acidic solution. The highest level of fluoridated HAP [FAP: Ca10(PO4)6(OH)xF2-x] was observed in HAP powder treated with SiF, even though more total fluorine in the form of FAP and CaF2 was observed in HAP powder samples treated with APF. Presumably as a consequence of this higher level of FAP formation after SiF treatment, the highest level of crystallinity along with the lowest solubility in acidic solution were observed in HAP powder treated with SiF solution. We concluded, therefore, that SiF may have potential value for use as a topical fluoride solution.

Acidulated Phosphate Fluoride↗
PubMed1996

Lysophosphatidylcholine potentiates the mitogenic activity of modified LDL for human monocyte-derived macrophages.

The growth of murine peritoneal macrophages is induced by oxidized LDL (Ox-LDL), and lysophosphatidylcholine (lysoPC) plays an important role in its mitogenic activity. In the present++ study, Ox-LDL-induced macrophage growth was examined with human monocyte-derived macrophages. The cell growth of human macrophages was significantly induced by Ox-LDL but not by acetylated LDL (acetyl-LDL). The treatment of acetyl-LDL with phospholipase A2, however, led to a marked increase in its mitogenic activity, with a concomitant conversion of 75% of its phospholipids to lysoPC. The growth-stimulating activity became positive only when both acetyl-LDL and lysoPC were coincubated, although neither of them exhibited cell growth-promoting activity. These results suggest that Ox-LDL could stimulate the growth of human monocyte-derived macrophages, and lysoPC may play an essential role in the mitogenic activity of Ox-LDL.

Cell Division↗
PubMed1996

Stimulation of macrophage DNA synthesis by polyanionic substances through binding to the macrophage scavenger receptor.

We previously demonstrated that ligands of macrophage scavenger receptors such as acetylated low density lipoprotein (LDL), oxidized LDL and advanced glycation-end products (AGE) of the Maillard reaction induce the growth of peritoneal exudate macrophages, and that the activity of AGE is inhibited by the presence of an antibody for granulocyte/macrophage colony-stimulating factor (GM-CSF). To evaluate the suggested role of the scavenger receptor in the induction of macrophage growth, we compared the effect of various polyanionic compounds which were reported to either have or not to have competent activity for the binding of acetylated LDL to scavenger receptors on macrophage DNA synthesis. Among the polyanions exhibiting such activity, polyguanilic acid (poly G) and dextran sulfate strongly augmented macrophage DNA synthesis, although they did not increase macrophage cell number. On the other hand, polyanions which are not ligands for the scavenger receptors did not show a significant augmenting effect, suggesting that the binding of polyanions to the scavenger receptor is important but not, by itself, sufficient. The augmentation of DNA synthesis in macrophages cultured with dextran sulfate or poly G was inhibited by the co-presence of anti-GM-CSF antibody, suggesting that the reaction is mediated by GM-CSF. However, dextran sulfate did not augment the production of GM-CSF in macrophages. Therefore, GM-CSF spontaneously present in macrophages might be a prerequisite for the induction of DNA synthesis.

Animals↗
PubMed1996

Quantitative analyses of lesion areas of coronary atherosclerosis in cholesterol-fed rabbits.

A simple method to quantitatively evaluate atherosclerosis in the rabbit coronary arteries by measuring macroscopic lesion areas (%) was attempted in the present study. Sixteen rabbits were fed a 0.5% cholesterol diet for 15 weeks and then 9 rabbits were sacrificed whereas the remaining 7 rabbits were maintained for further 9 weeks on a normal chow (at week 24). The left circumflex coronary arteries (LCX) were excised from the rabbit hearts under stereoscopic observation. The prepared arterial strips of LCX were 38.7 +/- 7.1 mm long and all of them reached the cardiac apex from the orifice. At week 15, the lesion area in LCX was negligible (3.2 +/- 0.4%) whereas the aortic lesions significantly developed (50.0 +/- 7.6%). At week 24, atherosclerotic lesions in both LCX and aortas increased to 32.8 +/- 9.2% and 85.9 +/- 5.6%, respectively. This is the first report that determined the luminal surface areas of atherosclerotic lesions in rabbit coronary arteries. This method may be more practical and useful for quantitative evaluation of coronary atherosclerosis in a large number of rabbits than histological observations of serial sections of rabbit hearts.

Animals↗
PubMed1996

Advanced glycation end products and their recognition by macrophage and macrophage-derived cells.

Modification of proteins by long-term incubation with glucose leads to the formation of advanced glycation end products (AGEs). AGE proteins are taken up by macrophages via the AGE receptor, which is similar to the macrophage scavenger receptor (MSR). In the present study, we compared the ligand specificity of the AGE receptor with that of MSR by three different experiments. The endocytic uptake of 125I-acetyl-LDL by RAW cells was effectively inhibited by unlabeled AGE-bovine serum albumin (BSA), whereas the inhibitory effect of acetyl-LDL on 125I-AGE-BSA was partial. Polyanions showing an effective inhibition for endocytic uptake of AGE-BSA were not always inhibitory for endocytic degradation of acetyl-LDL. These data, together with those obtained by three-dimensional fluorescence-activated cell sorter analysis, indicate that AGE proteins are recognized by more than two receptors, of which MSR is at least one. Finally, we examined whether MSR could mediate the endocytic uptake of AGE proteins by Chinese hamster ovary cells overexpressing bovine type II MSR (CHO-SRII cells). 125I-AGE-BSA underwent endocytic degradation by CHO-SRII cells, and this was effectively inhibited by unlabeled acetyl-LDL. These results clearly show that MSR mediates the endocytic uptake of AGE proteins, suggesting a new role of MSR in biological recognition of AGE in vivo.

Animals↗
PubMed1996

Advanced glycation end products of the Maillard reaction in aortic pepsin-insoluble and pepsin-soluble collagen from diabetic rats.

Recent immunohistological studies using antibodies against advanced glycation end products (AGEs) have demonstrated the presence of AGEs in several tissues. By an enzyme-linked immunosorbent assay using the monoclonal anti-AGE antibody, the present study aimed to determine AGEs in pepsin-insoluble collagen (PIC) as well as in pepsin-soluble collagen (PSC) from the aortas of streptozotocin (STZ)-induced diabetic rats (at 4, 16, and 28 weeks after STZ injection) and those of age-matched control rats. Addition of EDTA to the immunoassay buffer has led us to successful determination of AGEs in the aortic PIC samples with following results: 1) in diabetic rats, there was a time-related increase in the AGE contents at 28 weeks (n = 9, 226.4 +/- 13.5 ng/mg collagen [mean +/- SE]), compared with that at 4 and 16 weeks (n = 6, 79.6 +/- 9.5 ng/mg collagen, and n = 8, 149.4 +/- 30.9 ng/mg collagen at 4 and 16 weeks, respectively; both P < 0.05, between 4 and 16 weeks and 28 weeks); 2) after 28 weeks of diabetes, the AGE contents in PIC of aortas were significantly higher in diabetic rats than in controls (n = 9, 226.4 +/- 13.5 ng/mg collagen vs. n = 8, 129.6 +/- 14.9 ng/mg collagen, P < 0.01, diabetic vs. control); and 3) the level of the AGE content was strongly correlated with the PIC/total collagen (TC) ratio (n = 45, r = 0.698, P = 0.0001). By treating the samples of PSC with alkaline solution, the AGE content of PSC was also determined. In the PSC fraction, the AGE levels in the diabetic rats tended to increase with time and to be higher than those of control rats at 28 weeks although these changes were not statistically significant (diabetic: n = 4, 19.4 +/- 9.7; n = 6, 22.3 +/- 6.2; n = 6, 39.6 +/- 10.8; control: n = 4, 19.7 +/- 9.8; n = 6, 22.9 +/- 7.3; n = 7, 30.7 +/- 7.2; at 4, 16, and 28 weeks, respectively). Compared with the AGE levels of PSC, those of PIC were about four to seven times and four to five times higher in diabetic and control rats, respectively (PIC versus PSC in diabetic or control rats, all P < 0.001, at 4, 16, and 28 weeks, respectively). These findings provide the first immunochemical evidence that AGE adducts are present in the materials extracted sequentially by pepsin and collagenase and that these adducts in PIC accumulated as a function of the increase in the aortic PIC/TC ratio.

Animals↗
PubMed1996

Modification of vitronectin by advanced glycation alters functional properties in vitro and in the diabetic retina.

Early diabetic retinopathy is characterized by areas of acellular capillaries and the accumulation of advanced glycation end products (AGE). Endothelial cell maintenance, growth, and migration is regulated by a complex system involving the partitioning of growth factors between extracellular matrix-bound proteoglycans and cells, as well as by the activation and control of pericellular proteolysis. Using specific antibodies, we compared the immunolocalization of the vascular heparan sulfate proteoglycan (HSPG)-binding protein vitronectin (VN), HSPG, basic fibroblast growth factor, and collagen type 1 in retinae prepared from 11-month-old diabetic and nondiabetic Wistar rats. In normal rats, VN immunostaining was prominent in vascularized parts of the inner retina and colocalized with HSPG, which itself showed costaining with basic fibroblast growth factor in the extracellular matrix, vascular walls, and the inner limiting membrane. In contrast, the recognition of VN, HSPG, and basic fibroblast growth factor in diabetic retinae was greatly reduced in the inner limiting membrane and the extracellular matrix, where increased immunoreactive AGE colocalizing with VN were detectable. Capillary labeling of retinal vessel walls for plasminogen activator inhibitor-1 was moderate in both normal and diabetic retinae. AGE-VN was demonstrated in tissue extracts from retina by immunoprecipitation and Western blotting and presented a similar increase of AGE-related immunoreactivity compared with AGE-VN generated in vitro. AGE-VN in vitro had lost its native conformation, yielded high Mr SDS-resistant products, and was resistant to proteolysis because of modification of about 30% of lysines. Because binding of glycosaminoglycans, as well as interaction of type I collagen and the morphoregulatory proteins osteonectin and tenascin with AGE-VN, were reduced to at least 50% of control, alteration of basic residues in the heparin-binding domain of VN is plausible. In comparison to nonmodified VN, AGE-VN exhibited reduced cell attachment-promoting activity. Together, these in vitro results suggest that AGE-VN found in vivo is related to morphologic and functional changes in the diabetic retina and may contribute to the genesis of acellular capillaries in early diabetic retinopathy.

Animals↗