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Biomedical subjects

Akira Hirata

Publications and source records attributed to Akira Hirata.

At least 55 records · Page 3Linked to original sources
PubMed2004

Sensitivity to gefitinib (Iressa, ZD1839) in non-small cell lung cancer cell lines correlates with dependence on the epidermal growth factor (EGF) receptor/extracellular signal-regulated kinase 1/2 and EGF receptor/Akt pathway for proliferation.

Gefitinib (Iressa, ZD1839), a quinazoline tyrosine kinase inhibitor that targets the epidermal growth factor receptor (EGFR), is approved for patients with advanced non-small cell lung cancer (NSCLC) in several countries including Japan. However, the mechanism of drug sensitivity to gefitinib is not fully understood. In this study, we examined the molecular basis of sensitivity to gefitinib using nine human lung cancer cell lines derived from NSCLC. PC9 was the most sensitive to gefitinib of the nine NSCLC cell lines when assayed either by colony formation or MTS assays. The various cell lines expressed different levels of EGFR, HER2, HER3, and HER4, but there was no correlation between levels of EGFR and/or HER2 expression and drug sensitivity. Phosphorylation of EGFR, protein kinase B/AKT (Akt), and extracellular signal-regulated kinase (ERK) 1/2 was inhibited by much lower concentration of gefitinib in PC9 cells than in the other eight cell lines under exponential growing conditions. About 80% of cell surface EGFR in PC-9 was internalized within 10 min, whereas only about 30-50% of the cell surface EGFR was internalized in more drug-resistant cell lines in 15-60 min. The present study is the first to demonstrate that sensitivity to growth inhibition by gefitinib in NSCLC cell lines under basal growth condition is associated with dependence on Akt and ERK1/2 activation in response to EGFR signaling for survival and proliferation and also that drug sensitivity may be related to the extent of EGF-induced down-regulation of cell surface EGFR.

Carcinoma, Non-Small-Cell Lung↗
PubMed2003

Structural and enzymatic analysis of soybean beta-amylase mutants with increased pH optimum.

Comparison of the architecture around the active site of soybean beta-amylase and Bacillus cereus beta-amylase showed that the hydrogen bond networks (Glu380-(Lys295-Met51) and Glu380-Asn340-Glu178) in soybean beta-amylase around the base catalytic residue, Glu380, seem to contribute to the lower pH optimum of soybean beta-amylase. To convert the pH optimum of soybean beta-amylase (pH 5.4) to that of the bacterial type enzyme (pH 6.7), three mutants of soybean beta-amylase, M51T, E178Y, and N340T, were constructed such that the hydrogen bond networks were removed by site-directed mutagenesis. The kinetic analysis showed that the pH optimum of all mutants shifted dramatically to a neutral pH (range, from 5.4 to 6.0-6.6). The Km values of the mutants were almost the same as that of soybean beta-amylase except in the case of M51T, while the Vmax values of all mutants were low compared with that of soybean beta-amylase. The crystal structure analysis of the wild type-maltose and mutant-maltose complexes showed that the direct hydrogen bond between Glu380 and Asn340 was completely disrupted in the mutants M51T, E178Y, and N340T. In the case of M51T, the hydrogen bond between Glu380 and Lys295 was also disrupted. These results indicated that the reduced pKa value of Glu380 is stabilized by the hydrogen bond network and is responsible for the lower pH optimum of soybean beta-amylase compared with that of the bacterial beta-amylase.

Bacillus cereus↗
PubMed2003

Lens epithelium-derived growth factor: neuroprotection on rat retinal damage induced by N-methyl-D-aspartate.

The purpose of this study is to investigate possible neuroprotective effects of lens epithelium-derived growth factor (LEDGF) against cell death induced by N-methyl-D-aspartate (NMDA) in the rat retina. LEDGF and/or NMDA were intravitreally injected into rat eyes. NMDA-induced retinal death and protective effects of LEDGF were evaluated by morphometric analysis, cell numbers in the ganglion cell layer (GCL) and the thickness of the inner plexiform layer (IPL). Retrograde labeling with a fluorescent tracer (Fluoro-Gold) was applied for counting retinal ganglion cells (RGCs) that survived after NMDA injection. Terminal deoxyribonucleotidyl transferase (TdT)-mediated fluroscein-16-dUTP nick end-labeling (TUNEL) staining was used to evaluate of retinal cell death. Morphometric analysis and retrograde labeling analysis showed that retinal damage induced by NMDA was protected significantly by LEDGF. TUNEL assay revealed that pretreatment with LEDGF prevents NMDA-induced apoptosis. Retinal damage (ganglion and amacrine cells) induced by NMDA was protected by an intravitreal injection of LEDGF.

Animals↗
PubMed2003

In situ PCR for visualizing distribution of a functional gene "amoA" in a biofilm regardless of activity.

In this study, ammonia-oxidizing bacteria present in biofilms resulting from a nitrifying reactor were detected by both a conventional FISH technique and an original in situ PCR technique. Both techniques showed that ammonia-oxidizing bacteria were found near the surface of the biofilms. However, after the biofilm had been exposed to 2 weeks of ammonia starvation, ammonia-oxidizing bacteria present in the biofilm could not be detected by fluorescence in situ hybridization (FISH) because they did not have sufficient copies of rRNA. In contrast, ammonia-oxidizing bacteria could be detected by in situ PCR with strong signal. It was thus demonstrated that a cell possessing a specific functional gene is detectable by in situ PCR regardless of its activity.

Ammonia↗
PubMed2003

Influence of growth phase on bacterial cell electrokinetic characteristics examined by soft particle electrophoresis theory.

The influence of incubation time on the electrokinetic properties of Escherichia coli, Pseudomonas putida, Alcaligenes faecalis, and Alcaligenes sp., was examined by electrophoretic mobility measurements and the results were discussed based on Ohshima's soft particle theory. The electrophoretic mobility of E. coli plotted against incubation time revealed that the mobility gradually increased from the outset of incubation to 7 h, which corresponded to the exponential growth and early stationary phase, and then decreased. For all strains, electrophoretic mobility leveled off to nonzero values in accordance with the increase in ionic concentration, which was a characteristic feature of soft particles. Soft particle analysis was carried out at the outset of incubation and the time when mobility reached a maximum in order to obtain the spatial charge density (ZN) and cell surface softness (1/lambda). ZN at the maximum electrophoretic mobility was more negative than that at the outset of incubation for E. coli, P. putida, and A. faecalis, while the ZN of Alcaligenes sp. almost remained unchanged. 1/lambda decreased for E. coli, but increased for Alcaligenes sp. and A. faecalis. These findings indicated that the cell growth phase affects both ZN and 1/lambda, although the dependencies of these parameters are unique to each bacterial strain.

Journal Article↗
PubMed2003

Extracellular polymeric substances responsible for bacterial adhesion onto solid surface.

The influence of extracellular polymeric substances (EPS) on bacterial cell adhesion onto solid surfaces was investigated using 27 heterotrophic bacterial strains isolated from a wastewater treatment reactor. Cell adhesion onto glass beads was carried out by the packed-bed method and the results were discussed in terms of the amount of each EPS component produced and cell surface characteristics such as zeta potential and hydrophobicity. Protein and polysaccharides accounted for 75-89% of the EPS composition, indicating that they are the major EPS components. Among the polysaccharides, the amounts of hexose, hexosamine and ketose were relatively high in EPS-rich strains. For EPS-poor strains, the efficiency of cell adhesion onto glass beads increased as the absolute values of zeta potential decreased, suggesting that electrostatic interaction suppresses cell adhesion efficiency. On the other hand, the amounts of hexose and pentose exhibited good correlations with cell adhesiveness for EPS-rich strains, indicating that polymeric interaction due to the EPS covering on the cell surface promoted cell adhesion. It was concluded that, if the EPS amount is relatively small, cell adhesion onto solid surfaces is inhibited by electrostatic interaction, and if it is relatively large, cell adhesion is enhanced by polymeric interaction.

Bacteria↗
PubMed2003

Long-term alteration in the air-infused rabbit retina.

PURPOSE: To investigate the long-term histological changes in rabbit retina damaged by infusion air during vitrectomy. METHODS: Fourteen eyes of 14 rabbits were used. A standard three-port vitrectomy with artificial posterior vitreous detachment followed by fluid-air exchange was performed. After the fluid-air exchange, one side port was kept open for 30 s to freely introduce the air into the vitreous cavity under a pressure of 40 mmHg. Four weeks after surgery, indocyanine green angiography and histological examination were performed. RESULTS: At the air-infused area, indocyanine green angiography revealed the filling delay of the choroidal circulation in the early phase. The retinal surface was smooth and the internal limiting membrane was revealed to be intact by scanning electron microscopy. Light microscopy documented the thinning of the photoreceptor layer in the air-infused area. Transmission electron microscopy showed the disarrangement of the retinal pigment epithelium and loss of choriocapillaries. CONCLUSION: Infusion air during vitrectomy causes long-term outer retinal damage after surgery. Thinning of the photoreceptor cells, disarrangement of the retinal pigment epithelium and loss of choriocapillaries reflect the findings that we have observed previously in clinical cases. Visual field defect is a complication of vitreous surgery using fluid-air exchange. Although the visual field defects were observed just after surgery, abnormal fundus lesions continued to appear over time after the surgical procedure.

Air↗
PubMed2003

Neuroprotective effects of interleukin-6 on NMDA-induced rat retinal damage.

This study shows that interleukin-6 (IL-6) combined with soluble interleukin-6 receptors (sIL-6R) modulates N-methyl-D-aspartate (NMDA)-induced retinal damage. Eyes pretreated with a combined injection of IL-6 and sIL-6R had NMDA administered into the vitreous cavity. Morphometric analysis and retrograde labeling analysis found that pretreatment with either IL-6 or sIL-6R alone did not bring about any neuroprotective effect. However, pretreatment with a combined administration of IL-6 and sIL-6R induced a significant neuroprotective effect against NMDA-induced retinal damage. Apoptotic changes in the retina were assessed by the TUNEL method. The results indicated that pretreatment with IL-6 combined with sIL-6R prevents NMDA-induced apoptosis. Western blotting studies demonstrated upregulation of gp130 expression in the NMDA-injected retina. Present studies suggest that IL-6 combined with sIL-6R provides a neuroprotective effect on NMDA-induced retinal damage.

Animals↗
PubMed2003

Simultaneous nitrification and denitrification by controlling vertical and horizontal microenvironment in a membrane-aerated biofilm reactor.

Nitrogen and carbon components in domestic modified wastewater were completely removed by simultaneous nitrification and denitrification using a membrane-aerated biofilm reactor where biofilm was fixed on a hollow-fiber membrane. To measure the spatial distribution of pH, ammonium and nitrate ions and to observe microbes inside the biofilm fixed on the membrane, microelectrodes and the fluorescence in situ hybridization (FISH) method were applied. Due to plug flow in the vertical direction (from the bottom to the top of the reactor), ammonium nitrogen was gradually removed and negligible nitrate nitrogen was detected throughout the reactor. FISH revealed that ammonia-oxidizing bacteria were mainly distributed inside the biofilm and other bacteria, which included denitrifying bacteria, were mainly distributed outside the biofilm and over the suspended sludge. In order to characterize bacterial activity in the vertical direction of the reactor, nitrification rates at lower, central and upper points were calculated using microelectrode data. The nitrification rate at the lower point was 7 and 125 times higher than those at the central and upper points, respectively. These results show that the removal of carbon and nitrogen compounds was accomplished efficiently by using various kinds of bacteria distributed vertically and horizontally in a single reactor.

Bacteria, Aerobic↗
PubMed2003

Secondary glaucoma in patients with familial amyloidotic polyneuropathy.

OBJECTIVE: To elucidate the clinical features and surgical outcomes of the treatment of secondary glaucoma associated with transthyretin (TTR)-related familial amyloidotic polyneuropathy (FAP). DESIGN: Retrospective case study. PARTICIPANTS: Forty-nine Japanese patients with FAP. METHODS: For all patients, measurement of best-corrected visual acuity, intraocular pressure, and visual fields as well as slitlamp and ocular fundus examinations were conducted and compared. In addition, the exact mutation of the amyloidogenic TTR variants was analyzed for all 49 patients with FAP. The TTR mutations included amyloidogenic TTR (ATTR) Val30Met in 41 patients, ATTR Tyr114Cys in 6, ATTR Ser50Ile in 1, and a compound heterozygous mutation of ATTR Val30Met + Arg104His in 1. RESULTS: The onset of secondary glaucoma was defined as elevation of intraocular pressure and glaucomatous changes in visual field defects. Secondary glaucoma was detected in 12 (24%) of the 49 patients. The incidence of secondary glaucoma in patients with the Val30Met mutation (17%) was lower than for the other FAP genotypes (P =.02 using the chi(2) test). Of 20 glaucomatous eyes, amyloid deposition on the pupil and anterior surface of the lens was found in 18 eyes. Amyloid deposition was found prior to glaucoma in 11 eyes and at the first visit to our clinic in another 7 eyes. In the 11 eyes in which the onset of glaucoma occurred following amyloid deposition along the pupil, the mean +/- SD period between the onsets of pupillary amyloid deposition and glaucoma was 2.55 +/- 1.43 years (range, 0.2-4.0 years). Further statistical analyses revealed significant relationships between the onset of secondary glaucoma and both amyloid deposition (P<.001) and vitreous opacity (P<.001). Surgical treatment was required in 15 (75%) of the 20 glaucomatous eyes. In 9 (81%) of the 11 eyes that underwent trabeculectomy, the intraocular pressure was well controlled at or lower than 20 mm Hg during the follow-up period. In the eyes that underwent combined trabeculotomy and sinusotomy (2 eyes), nonpenetrating trabeculectomy (1 eye), or a cyclodestructive procedure (1 eye), the intraocular pressure was poorly controlled. CONCLUSIONS: Glaucoma is not a rare condition in patients with FAP, especially because liver transplantation now enables patients with FAP to live longer. Careful observation of amyloid deposition along the pupil allows the prediction of glaucoma onset.

Adult↗
PubMed2003

Characterization of nitrifying granules produced in an aerobic upflow fluidized bed reactor.

Since nitrification is the rate-determining step in the biological nitrogen removal from wastewater, many research studies have been conducted on the immobilization of nitrifying bacteria. In this research, granulation of nitrifying bacteria in an aerobic upflow fluidized bed (AUFB) reactor in a nitrification process for inorganic wastewater containing 500 g/m(3) of NH(4)(+)-N was investigated. It was observed that spherical, pseudocubic and elliptical granules with a diameter of 346 microm were produced at the bottom of the reactor after 300 days. Denaturing gradient gel electrophoresis analysis revealed that Nitrosomonas-like bacteria were the dominant ammonia-oxidizing species in the granules. Many colonies of Nitrosomonas-like bacteria were found in the outer part of the granules based on the spatial distribution analysis by fluorescence in situ hybridization. By stepwise reduction of the hydraulic retention time, the ammonia removal rate of the AUFB reactor containing these nitrifying granules finally reached 1.5 kg-N/m(3)/day. Results suggested that the use of granules realizes the retention of a large amount of nitrifying bacteria in the reactor, which guarantees a highly efficient nitrification.

Ammonia↗
PubMed2003

Vitreous opacities and outcome of vitreous surgery in patients with familial amyloidotic polyneuropathy.

PURPOSE: To report the prevalence of vitreous opacities and the outcome of vitreous surgery in patients with familial amyloidotic polyneuropathy (FAP). DESIGN: Observational case series. METHODS: In 37 patients with FAP and the ATTR Val30 Met mutation, vitreous opacities were present in 14 eyes of 9 patients and vitrectomy combined with phacoemulsification and intraocular lens implantation was performed in five eyes of three patients. In six patients with the ATTR Tyr114Cys mutation, vitreous opacities were present in both eyes of all six patients and vitrectomy combined with phacoemulsification and intraocular lens implantation was performed in nine eyes of six patients. The mean follow-up period after vitreous surgery was 20.9 +/- 16.8 months (range, 3 to 52 months). RESULTS: The prevalence of vitreous opacities is much higher in patients with ATTR Tyr114Cys (100%) than in those with ATTR Val30 Met (24%). The mean age at the onset of vitreous opacities was significantly lower in the patients with ATTR Tyr114Cys (37.0 +/- 5.3 years) than in the nine patients with ATTR Val30 Met (52.8 +/- 9.1 years; P <.005). Visual acuity improved in all 14 eyes after vitreous surgery; however, final visual acuity decreased in one eye owing to the occurrence of a central retinal vein occlusion. Vitreous opacities mildly increased in two eyes. CONCLUSIONS: Our data suggest that the ATTR Val30 Met and ATTR Tyr114Cys mutations induce different clinical features of vitreous opacities. Vitreous surgery combined with phacoemulsification and implantation of an intraocular lens is a safe and useful treatment. Careful long-term follow-up should be performed.

Adult↗
PubMed2003

Persistent subretinal indocyanine green induces retinal pigment epithelium atrophy.

PURPOSE: To describe a case of a patient with macular hole with subretinal indocyanine green (ICG) during vitrectomy. DESIGN: Interventional case report. METHODS: A 66-year-old woman with macular hole underwent a vitrectomy with ICG. RESULTS: After application of ICG into the vitreous, ICG was introduced in the subretinal space. Indocyanine green was found to be present for more than 6 months. Retinal pigment epithelium atrophy appeared at the site of the lesion. CONCLUSIONS: Although ICG may be a useful tool for distinguishing the internal limiting membrane and other tissues careful application is required to prevent side effects.

Aged↗
PubMed2003

A case of primary B-cell type non-Hodgkin lymphoma originating in the iris.

PURPOSE: To report a case of primary malignant B-cell type non-Hodgkin lymphoma originating in the iris. DESIGN: Interventional case report. METHODS: An 83-year-old woman presented with anterior uveitis resulting from primary malignant lymphoma in the iris. Ultrasound biomicroscopy and indocyanine green angiography using a scanning laser ophthalmoscope showed abnormalities in the iris. Diagnostic biopsy of the iris revealed B-cell type non-Hodgkin lymphoma. RESULT: The patient was treated with radiotherapy, and the tumorous lesion resolved. CONCLUSION: Primary lymphoma localized in the iris only is rare. In this case, diagnostic biopsy and radiotherapy of the iris lymphoma provided good results.

Aged↗
PubMed2003

Expression profiling of cytokines and related genes in regenerating skeletal muscle after cardiotoxin injection: a role for osteopontin.

To examine the roles of cytokines in muscle regeneration, we injected cardiotoxin into mouse tibialis anterior muscle and examined the expression profiles of cytokines and related genes in the regeneration process. Expression of 40, 64, and 7 genes among 522 genes spotted on a cytokine expression array were increased more than fivefold at 48 hours, 96 hours, and 7 days after toxin injection, respectively, when compared with those of the control muscle. Especially the levels of mRNA for chemokines and chemokine receptors, many of which are potent regulators of macrophages, were highly elevated 48 hours after injury. The expression of osteopontin (OPN), a versatile regulator of inflammation and tissue repair, was up-regulated more than 118-fold in regenerating muscle at 48 hours after injury. Northern blotting confirmed that the expression of OPN was highest at 48 hours after cardiotoxin injection and declined sharply thereafter. Immunohistochemistry showed that OPN was detected both in the cytoplasm of macrophages and in necrotic muscle infiltrated with macrophages. Our studies suggest OPN may serve as an adhesion molecule that promotes macrophage binding to necrotic fibers and may be an important mediator in the early phase of muscle regeneration.

Animals↗
PubMed2003

Nitrogen removal characteristics and biofilm analysis of a membrane-aerated biofilm reactor applicable to high-strength nitrogenous wastewater treatment.

A membrane-aerated biofilm reactor (MABR) capable of simultaneous nitrification and denitrification in a single reactor vessel was developed to investigate the characteristics of nitrogen removal from high-strength nitrogenous wastewater, and biofilm analysis using microelectrodes and the fluorescence in situ hybridization (FISH) technique was performed. Mean removal percentages of total organic carbon (TOC) and nitrogen were 96% and 83% at removal rates of 5.76 g-C m(-2) d(-1) and 4.48 g-N m(-2) d(-1), respectively. For stable removal efficiency, constant washing of the biofilm was needed. Dissolved oxygen microelectrode measurement revealed that the biofilm thickness was about 1600 microm, and that oxygen penetrated about 300 to 700 microm, from the outer surface of the membrane. Furthermore, FISH analysis revealed that ammonia-oxidizing bacteria (AOB) were located near the outer surface of the membrane, whereas other bacteria were located from the inner to the outer part of the biofilm. Combining these results demonstrated that simultaneous nitrification and denitrification occurred in the biofilm of the MABR system. In addition, stoichiometric analysis revealed that after 130 d(-1), the free ammonia (FA) concentration ranged within the concentration causing inhibition of the growth of nitrite oxidizing bacteria (NOB) and that AOB consumed 86% of the oxygen supplied through the intra-membrane. These results indicate that nitrogen removal not via nitrate but via nitrite was mainly achieved in the MABR system.

Journal Article↗
PubMed2003

Expression of glycosaminoglycans during development of the rat retina.

PURPOSE: To investigate the spatiotemporal expression of glycosaminoglycans during development of the rat retina. METHODS: Hyaluronan and sulfated glycosaminoglycans, including chondroitin sulfate, heparan sulfate and keratan sulfate were detected using biotinylated hyaluronan binding protein, immunohistochemical analysis, respectively, in the rat retina at various stages of development. RESULTS: Hyaluronan was expressed in the nerve fiber layer, inner plexiform layer and outer plexiform layer during early postnatal stages (postnatal day 1-14; P1-P14) and was undetectable after P21. In contrast, hyaluronan was faintly observed in the photoreceptor layer on P7, and gradually increased up to P49. The spatiotemporal expression pattern of chondroitin sulfate was similar to that of hyaluronan. Heparan sulfate was also detected in the nerve fiber layer, inner plexiform layer and outer plexiform layer during early postnatal stages (P1-P14). In addition, heparan sulfate was expressed in the inner limiting membrane during all stages of development. Keratan sulfate was not detected in the retina at any stage of development. CONCLUSIONS: Hyaluronan, chondroitin sulfate and heparan sulfate are expressed in nerve fiber-rich layers during early postnatal stages and may regulate neurite outgrowth. In adulthood, both hyaluronan and chondroitin sulfate are expressed in the photoreceptor layer and may consist of the interphotoreceptor matrix. In addition, heparan sulfate is expressed in the inner limiting membrane throughout the various stages of development and may be associated with the structure of the inner limiting membrane.

Animals↗