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Biomedical subjects

H Fujimori

Publications and source records attributed to H Fujimori.

At least 19 recordsLinked to original sources
PubMed2001

In situ demonstration of intraepithelial lymphocyte adhesion to villus microvessels of the small intestine.

The recirculation of lymphocytes through the intestinal mucosa is important for specific immune defense, but the origin and differentiation of intraepithelial lymphocytes (IEL) are not fully understood. The present study therefore used intravital microscopy to investigate the migration of IEL to the villus mucosa and Peyer's patches of the small intestine. IEL were separated from inverted murine small intestine and mesenteric lymph node (MLN) T cells were also isolated. The adhesion of fluorescence-labeled lymphocytes to postcapillary venules (PCV) of Peyer's patches and arcade microvessels of small intestinal villi was observed after injection. In some experiments, the effect of antibodies against adhesion molecules on cell kinetics were investigated. IEL time-dependently accumulated in villus microvessels of the small intestine, whereas few MLN cells did. Few IEL adhered to the PCV of Peyer's patches. IEL were shown to express alpha(E)beta(7)-integrin but not L-selectin. The accumulation of IEL in villus archade was significantly inhibited by antibody against beta(7)-integrin or mucosal addressin cell adhesion molecules (MAdCAM)-1, but not by alpha(E)-integrin. The combined blocking of beta(7)-integrin and MAdCAM-1 further attenuated the sticking of IEL in this area, although it did not entirely block the IEL adherence. The adherence of CD4(+) or TCRalphabeta IEL to villus microvessels was significantly greater than that of CD4(-) or TCRgammadelta IEL. It was demonstrated in situ for the first time that IEL adhered selectively to the villus microvessels of the small intestine partly via beta(7) and MAdCAM-1.

Animals↗
PubMed2001

Intravital demonstration of modulation of T lymphocyte migration by CINC/gro in rat Peyer's patches.

BACKGROUND/AIMS: Cytokine-induced neutrophil chemoattractant (CINC/gro), a member of interleukin-8 family, was found as a potent chemotactic factor for rat neutrophils. Although several chemokines have been shown to be potent regulators of T cell chemotaxis in vitro, the potential role of chemokines in T-cell migration in gut-associated lymphoid tissues has not been investigated in vivo. In the present study, the effects of CINC/gro on T-lymphocyte migration were examined in rat Peyer's patches. METHODS: T lymphocytes collected from intestinal lymph of rats were fluorescence-labeled and injected into the jugular vein. Peyer's patches of the recipient rats were observed with intravital fluorescence microscopy and the effects of CINC/gro infusion was investigated. Lymphocyte flux in mesenteric collecting lymphatics was also observed. RESULTS: In vivo infusion of CINC/gro significantly attenuated the initial lymphocyte interaction with postcapillary venules of Peyer's patches. However, once these lymphocytes adhered to venules, CINC/gro treatment significantly accelerated the transendothelial migration of T lymphocytes and they also significantly increased their subsequent flux in collecting lymphatics. CONCLUSION: There is a possibility that CINC/gro could modulate the characteristics of T lymphocyte homing in the inflammatory sites of gut.

Animals↗
PubMed2001

Intercellular cell adhesion molecule-1 regulates lymphocyte movement into intestinal microlymphatics of rat Peyer's patches.

The objective of this study was to determine whether specific adhesion molecules modulate lymphocyte movement from Peyer's patches into intestinal microlymphatics. The fluorochrome acridine orange was injected via a micropipette into Peyer's patches to fill lymphatics. The flux of labeled lymphocytes into intestinal microlymphatics was monitored with intravital fluorescence microscopy. The lymphatic microvessels in the perifollicular area of Peyer's patches were filled with lymphocytes, most of which remained within the lymphatics. Some lymphocytes became detached and were drained into intestinal lymph. Administration of antibodies directed against ICAM-1 significantly increased lymphocyte flux into interfollicular lymphatics. The immunohistochemical study showed intense ICAM-1 expression on the lymphocytes densely packed in the lymphatics surrounding follicles in Peyer's patches. A large number of lymphocytes are normally sequestered in the lymphatic network of Peyer's patches. This sequestration of lymphocytes is largely mediated by ICAM-1-dependent cell-cell interactions.

Animals↗
PubMed2000

The receptor for advanced glycation end products is induced by the glycation products themselves and tumor necrosis factor-alpha through nuclear factor-kappa B, and by 17beta-estradiol through Sp-1 in human vascular endothelial cells.

The binding of advanced glycation end products (AGE) to the receptor for AGE (RAGE) is known to deteriorate various cell functions and is implicated in the pathogenesis of diabetic vascular complications. Here we show that AGE, tumor necrosis factor-alpha (TNF-alpha), and 17beta-estradiol (E(2)) up-regulated RAGE mRNA and protein levels in human microvascular endothelial cells and ECV304 cells, with the mRNA stability being essentially invariant. Transient transfection experiments with human RAGE promoter-luciferase chimeras revealed that the region from nucleotide number -751 to -629 and the region from -239 to -89 in the RAGE 5'-flanking sequence exhibited the AGE/TNF-alpha and E(2) responsiveness, respectively. Site-directed mutation of an nuclear factor-kappaB (NF-kappaB) site at -671 or of Sp-1 sites at -189 and -172 residing in those regions resulted in an abrogation of the AGE/TNF-alpha- or E(2)-mediated transcriptional activation. Electrophoretic mobility shift assays revealed that ECV304 cell nuclear extracts contained factors which retarded the NF-kappaB and Sp-1 elements, and that the DNA-protein complexes were supershifted by anti-p65/p50 NF-kappaB and anti-Sp-1/estrogen receptor alpha antibodies, respectively. These results suggest that AGE, TNF-alpha, and E(2) can activate the RAGE gene through NF-kappaB and Sp-1, causing enhanced AGE-RAGE interactions, which would lead to an exacerbation of diabetic microvasculopathy.

Blotting, Western↗
PubMed1999

Advanced glycation endproducts accelerate calcification in microvascular pericytes.

Vascular calcification in advanced atherosclerosis is frequently associated with diabetes, and is a predictor of future cardiovascular events. To investigate the molecular mechanisms of vascular calcification, we examined whether advanced glycation endproducts (AGE) formed at an accelerated rate under diabetes induce the osteoblastic differentiation of pericytes, a mesenchymal progenitor. First, von Kossa staining demonstrated that AGE significantly increased the number of calcified nodules in a bovine pericyte culture. AGE were also found to induce calcium accumulation in the pericyte monolayer in time- and dose-dependent manners. Second, quantitative reverse transcription-polymerase chain reaction revealed that AGE increased the pericyte levels of mRNAs coding for alkaline phosphatase and osteopontin, the representative markers for early and late osteoblastic differentiation, respectively. Alkaline phosphatase activity was actually enhanced by AGE. The results suggest that AGE have the ability to induce the osteoblatic differentiation of pericytes, which would contribute to the development of vascular calcification in diabetes.

Alkaline Phosphatase↗
PubMed1999

Volumetric MRI measurements of the tree shrew hippocampus.

Protocols suitable for repeated magnetic resonance imaging (MRI) studies of the tree shrew's brain were established. This included the development of (i) a technique for prolonged inhalation anesthesia by endotracheal intubation; (ii) a reproducible fixation of the animal's head in a stereotaxic frame and finally (iii) the set-up of the hardware (rf coil) and software (MRI sequences) of the MRI system. The endotracheal intubation as well as the repeated and prolonged anesthesia showed no complications. The in vivo measurements of the tree shrew's hippocampal formation revealed a high reproducibility. Right and left hippocampal volume was determined as 85.2 mm3 +/- 8% and 87.4 mm3 +/- 10%, respectively. The utility of MRI in delineating alterations in brain anatomy was demonstrated in three animals receiving cortisol via the drinking water (5 mg/animal/day). After a 4-week treatment, in two of the three tree shrews a reduction in hippocampal volume was observed. Thus, the MRI protocols used here allow for repeated and non-invasive measurements of changes in hippocampal anatomy within the same animal and to monitor the temporal dynamics of structural alterations within this brain structure.

Animals↗
PubMed1999

Proton MRS of oral creatine supplementation in rats. Cerebral metabolite concentrations and ischemic challenge.

Proton magnetic resonance spectroscopy (MRS) was employed to determine the concentrations of N-acetylaspartate (NAA), total creatine (tCr), choline-containing compounds (Cho), myo-inositol (Ins), glucose (Glc), and lactate (Lac) in rat brain before and after 10 days of oral supplementation of 2.6 g Cr-monohydrate per kg body weight per day. Measurements were performed both in vitro (n = 16) and in vivo (n = 6). The neuroprotective potential of oral Cr was assessed by dynamically monitoring brain Glc and Lac in response to transient global ischemia (12 min). In comparison to controls the in vitro concentrations of Cr (13.1 +/- 9.3%) and Ins (12.7 +/- 14. 0%) were significantly increased in Cr-fed rats. Under in vivo conditions, the data revealed trends for elevated tCr (4.7%) and Ins (10.6%) which were enhanced in the concentration ratios of tCr:Cho (10.2%) and Ins:Cho (17.8%). Together with an increased Glc level (27.3%), the observation of a statistically significant decrease of brain Lac (-38.5 +/- 19.3%) in Cr-fed rats may reflect a shift of the energy metabolism from non-oxidative toward oxidative glycolysis. One hour after global ischemia most of the metabolic differences between Cr-fed rats and controls were retained. The increased Glc level (44.4 +/- 33.3%) reached statistical significance, but the accumulation of Lac and its time course during ischemia and early reperfusion showed no differences between Cr-fed rats and controls.

Administration, Oral↗
PubMed1999

Brain water diffusion in normal and creatine-supplemented rats during transient global ischemia.

Brain water diffusion in response to transient global ischemia (12 min), reperfusion (60 min), and cardiac arrest was monitored by localized proton magnetic resonance spectroscopy. The trace of the apparent diffusion coefficient tensor (ADC(Av)) was determined at high temporal resolution (10 sec) to assess the putative neuroprotective potential of oral creatine (Cr) in rats that received 2.2 g Cr-monohydrate per kg body weight per day for 10 days (n = 8) relative to controls (n = 9). Cr-fed rats revealed a statistically significant increase of the cerebral concentration ratio of Cr to choline-containing compounds (20%). The decrease of the ADC(Av) value during acute ischemia showed a three-phasic behavior in line with energy depletion, cytotoxic edema, and brain cooling. In Cr-fed rats, slightly less severe and mildly delayed diffusion changes during ischemia and similar beneficial trends during early reperfusion did not reach statistical significance. Magn Reson Med 42:798-802, 1999.

Animals↗
PubMed1999

Insulin stimulates the growth and tube formation of human microvascular endothelial cells through autocrine vascular endothelial growth factor.

Insulin treatment is known epidemiologically as an independent risk factor for the progression of diabetic retinopathy. However, how insulin exacerbates the retinopathy is not yet fully understood. In this study, we investigate the effects of insulin on the growth and tube formation of microvascular endothelial cells (EC). When human skin microvascular EC were grown under various concentrations of insulin, DNA synthesis as well as tube formation of EC was found to be significantly stimulated. We obtained evidence that it is mainly vascular endothelial growth factor (VEGF) that mediates the angiogenic activity of insulin as follows. (1) Insulin upregulates the level of mRNA coding for secretory forms of VEGF, while the expression of the two VEGF receptor genes, kinase insert domain-containing receptor (kdr) and fms-like tyrosine kinase1 (flt1), was essentially unchanged by exposure to insulin. (2) A monoclonal antibody against human VEGF can completely neutralize both the proliferation and the tube formation of EC induced by insulin. The angiogenic effects of insulin were additive with those of hypoxia, a principal factor that causes angiogenesis. Further, insulin significantly stimulated plasminogen activator inhibitor-1 activity in EC. The results thus suggest that insulin not only elicits angiogenesis through the induction of autocrine VEGF but also is a predisposing factor for thrombogenesis, which may give rise to focal ischemia that could superdrive angiogenesis, thereby leading to the exacerbation of diabetic retinopathy.

Autocrine Communication↗
PubMed1999

Dumbbell-shaped trigeminal neurinoma in a child.

We experienced a case of solitary trigeminal neurinoma in a 10-year-old boy without stigmata of neurofibromatosis. This boy presented with progressive cerebellar ataxia, right abducens palsy and right hemiparesis. Magnetic resonance images (MRI) demonstrated a tumour extending into both the middle and posterior fossas forming a dumbbell-shaped mass lesion with a size of 35 x 30 x 45 mm. The tumour was successfully removed free of complications by a two-stage operation composed of right subtemporal and suboccipital retromastoid approaches. It is extremely rare for a solitary neurinoma to occur at this site in a child without neurofibromatosis. Surgical approaches in this type of tumour are discussed with a review of the literature.

Child↗
PubMed1999

Altered migration of gut-derived T lymphocytes after activation with concanavalin A.

Although activation of lymphocytes is known to be associated with profound changes in homing behavior, it remains unclear how activation alters migration of gut-derived lymphocytes in lymphoid and nonlymphoid organs. The objectives of this study were 1) to compare migration of naive and concanavalin A (ConA)-activated T lymphocytes into the gut mucosa, spleen, and liver and 2) to define the role of specific adhesion molecules in this homing process. Fluorescently labeled T lymphocytes collected from rat intestinal lymph were injected into the jugular vein, and the kinetics of appearance of the infused lymphocytes were monitored in ileal Peyer's patches, spleen, and liver. The migration of naive and ConA-activated T lymphocytes into microvessels were compared using an intravital microscope. ConA stimulation significantly increased the rolling velocity of T lymphocytes in postcapillary venules of Peyer's patches, and ConA-stimulated lymphocytes exhibited a loss of the selective adherence properties in Peyer's patches that is normally observed with naive T cells. ConA activation also suppressed the accumulation of T cells in the spleen. On the other hand, the adherence of T cells to hepatic sinusoidal endothelium was significantly increased after ConA activation, especially in the periportal area, and this increase was attenuated by an anti-intercellular adhesion molecule (ICAM)-1 antibody. Flow cytometry analysis revealed a decline in L-selectin expression and an increase in CD11a expression and ICAM-1 on the surface of ConA-treated T cells. In conclusion, activation of gut-derived T lymphocytes with ConA significantly alters their migration path, with a diminished localization to Peyer's patches and spleen and a preferential accumulation in hepatic sinusoids. This altered migration pattern likely results from changes in the expression of leukocyte adhesion molecules such as L-selectin and CD11a.

Animals↗
PubMed1999

Vascular endothelial growth factor acts as a pericyte mitogen under hypoxic conditions.

Angiogenesis is the process by which new vascular networks are formed from preexisting capillaries. The small vessels are composed of two types of cells, namely endothelial cells (EC) and pericytes, with the former being encircled by the latter. We previously showed that hypoxia, the principal cause of angiogenesis, can induce the proliferation of pericytes as well as EC. In this report we present evidence that the hypoxic induction of pericyte growth can be ascribed at least in part to vascular endothelial growth factor (VEGF) produced by this very cell type. First, the finding that hypoxia can stimulate the proliferation of pericytes was confirmed by cultivating bovine retinal pericytes in a controlled-atmosphere culture chamber containing various concentrations of oxygen and then assaying pericyte synthesis of DNA. Second, Northern blot analysis revealed that pericyte levels of mRNA encoding VEGF increased as the atmospheric oxygen tension was decreased; this was accompanied by an increase in de novo synthesis of VEGF proteins. Third, pericytes were able to respond to exogenously added VEGF, resulting in a dose-dependent increase in viable cell numbers. Fourth, polyclonal antibodies against VEGF efficiently blocked the hypoxic induction of pericyte growth. Fifth, pericytes expressed the gene for fms-like tyrosine kinase 1 (flt1) as the predominant form of VEGF receptor, and tyrosine phosphorylation of this receptor protein was enhanced when pericytes were exposed to hypoxia, as it was when cells were exposed to VEGF. Sixth, the antisense DNA complement of flt1 mRNA abolished the hypoxia-induced stimulation of pericyte growth. Finally, exogenous VEGF stimulated the migration of pericytes in a dose-dependent manner. The results thus suggest that VEGF, which has been thought to be a specific mitogen for EC, also acts on neighboring pericytes, probably in both autocrine and paracrine manners, and that the hypoxia-induced overproduction of VEGF could promote not only EC sprouting but also the recruitment of pericytes, thereby contributing to the maturation of newly formed microvessels.

Animals↗
PubMed1998

Dual regulation of the skeletal muscle ryanodine receptor by triadin and calsequestrin.

Triadin, a calsequestrin-anchoring transmembrane protein of the sarcoplasmic reticulum (SR), was successfully purified from the heavy fraction of SR (HSR) of rabbit skeletal muscle with an anti-triadin immunoaffinity column. Since depletion of triadin from solubilized HSR with the column increased the [3H]ryanodine binding activity, we tested a possibility of triadin for a negative regulator of the ryanodine receptor/Ca2+ release channel (RyR). Purified triadin not only inhibited [3H]ryanodine binding to the solubilized HSR but also reduced openings of purified RyR incorporated into the planar lipid bilayers. On the other hand, calsequestrin, an endogenous activator of RyR [Kawasaki and Kasai (1994) Biochem. Biophys. Res. Commun. 199, 1120-1127; Ohkura et al. (1995) Can. J. Physiol. Pharmacol. 73, 1181-1185] potentiated [3H]ryanodine binding to the solubilized HSR. Ca2+ dependency of [3H]ryanodine binding to the solubilized HSR was reduced by triadin, whereas that was enhanced by calsequestrin. Interestingly, [3H]ryanodine binding to the solubilized HSR potentiated by calsequestrin was reduced by triadin. Immunostaining with anti-triadin antibody proved that calsequestrin inhibited the formation of oligomeric structure of triadin. These results suggest that triadin inhibits the RyR activity and that RyR is regulated by both triadin and calsequestrin, probably through an interaction between them. In this paper, triadin has been first demonstrated to have an inhibitory role in the regulatory mechanism of the RyR.

Animals↗
PubMed1998

Effects of glucocorticoids on apoptosis of infiltrated eosinophils and neutrophils in rats.

The effects of glucocorticoids on the survival of rat eosinophils and neutrophils infiltrated into the peritoneal cavity were examined. Glucocorticoids including dexamethasone, prednisolone and hydrocortisone inhibited the survival of rat peritoneal eosinophils at 10(-6) M, whereas they prolonged survival of rat peritoneal neutrophils at 10(-8) M. Sex steroids including estradiol and progesterone did not affect cell survival. Dexamethasone decreased the viability of eosinophils after 3 days of incubation and maintained the viability of neutrophils until 4 days after incubation concentration dependently. The EC50 of dexamethasone for inhibition of the survival of eosinophils was 1.5 x 10(-8) M, and that for the spontaneous death of neutrophils was 6.4 x 10(-10) M, suggesting that glucocorticoids at concentrations that inhibit eosinophil survival prolong neutrophil survival. Analysis of DNA fragmentation of cultured eosinophils and neutrophils revealed that glucocorticoids enhance eosinophil apoptosis but inhibit neutrophil apoptosis. The effects of dexamethasone on viability and DNA fragmentation were counteracted by the glucocorticoid receptor antagonist, mifepristone, concentration dependently. These findings indicate that glucocorticoids induce contradictory effects via the glucocorticoid receptor on rat eosinophils and neutrophils extravasated to an inflammatory locus such as the peritoneal cavity by modulating apoptosis.

Animals↗
PubMed1998

Proton T2 relaxation of cerebral metabolites during transient global ischemia in rat brain.

Putative changes of metabolite T2 relaxation times were investigated before and after a 20-min period of global ischemia in rat brain in vivo (n = 10) using localized proton MRS at different echo times (2.35 T). Neither absolute T2 relaxation times (TE = 20-270 ms) nor time courses of T2-weighted metabolite signals (TE = 135 ms) revealed statistically significant changes during the occlusion or early reperfusion relative to pre-ischemic baseline. These findings are in line with reports of relaxation changes at much later stages and further demonstrate that altered T2 relaxation is not a confounding factor in diffusion-weighted long-TE proton MRS during early ischemic events.

Animals↗
PubMed1998

Advanced glycation endproducts inhibit prostacyclin production and induce plasminogen activator inhibitor-1 in human microvascular endothelial cells.

Several thrombogenic abnormalities are associated with diabetes. To investigate the underlying molecular mechanisms, we examined the effects of advanced glycation endproducts (AGE), non-enzymatically glycated protein derivatives, on the production of prostacyclin (PGI2), an anti-thrombogenic prostanoid, and of plasminogen activator inhibitor-1 (PAI-1), a fast-acting serine protease inhibitor of fibrinolysis, in human microvascular endothelial cells (EC). Firstly, AGE-bovine serum albumin (BSA) but not non-glycated BSA, was found to considerably decrease the production of PGI2 to about two-thirds of the control value. Secondly, quantitative reverse transcription-polymerase chain reaction showed that AGE-BSA increased the EC levels of mRNA coding for PAI-1, this being associated with a concomitant increase in the immunoreactive PAI-1 contents and the anti-fibrinolytic activity. Thirdly, the effects of AGE on PGI2 and PAI-1 syntheses in EC were found to be mediated by a receptor for AGE (RAGE) because antisense DNA against RAGE mRNA could reverse the AGE effects. Further, it was found that AGE decreased the intracellular cyclic AMP concentrations in EC and that cyclic AMP agonists such as dibutyryl cyclic AMP, forskolin and PGI2 analogue reduced the AGE-stimulated PAI-1 production, suggesting the involvement of cyclic AMP in the AGE-signalling pathway. The results thus suggest that AGE have the ability to cause platelet aggregation and fibrin stabilization, resulting in a predisposition to thrombogenesis and thereby contributing to the development and progression of diabetic vascular complications.

Cyclic AMP↗