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Biomedical subjects

M Kado

Publications and source records attributed to M Kado.

At least 19 recordsLinked to original sources

Transient corneal edema induced by nitric oxide synthase inhibition.

The aim of the study is to identify nitric oxide synthase (NOS) in the rabbit cornea and further investigate the physiological role of nitric oxide in the rabbit cornea. For histological identification, an immunohistochemical technique using anti-NOS monoclonal antibodies was employed. For the physiological study, we measured the corneal thickness in vivo as an indicator of corneal edema by ultrasonic pachymetry. The measurements were repeated before and after ipsilateral injections of N(G)-nitro-L-arginine methyl ester (L-NAME) or N(G)-nitro-D-arginine methyl ester (D-NAME) or 6-anilino-5,8-quinolinedione (LY-83583) with contralateral injection of vehicle (balanced salt solution) into the anterior chamber of the rabbit. We also monitored intraocular pressure (IOP) by pneumatonometry. Endothelial NOS (eNOS) immunoreactivity was demonstrated both in the corneal epithelium and the endothelium. The corneal thickness significantly increased after L-NAME or LY-83583 without significant rise of IOP, whereas no change was detected after vehicle or D-NAME. These results suggest that NO is spontaneously produced in the corneal endothelium and the NO/cyclic GMP pathway is involved in maintainance of corneal thickness.

Aminoquinolines

X-ray micrography and imaging of Escherichia coli cell shape using laser plasma pulsed point x-ray sources.

High-resolution x-ray microscopy is a relatively new technique and is performed mostly at a few large synchrotron x-ray sources that use exposure times of seconds. We utilized a bench-top source of single-shot laser (ns) plasma to generate x-rays similar to synchrotron facilities. A 5 microlitres suspension of Escherichia coli ATCC 25922 in 0.9% phosphate buffered saline was placed on polymethylmethyacrylate coated photoresist, covered with a thin (100 nm) SiN window and positioned in a vacuum chamber close to the x-ray source. The emission spectrum was tuned for optimal absorption by carbon-rich material. Atomic force microscope scans provided a surface and topographical image of differential x-ray absorption corresponding to specimen properties. By using this technique we observed a distinct layer around whole cells, possibly representing the Gram-negative envelope, darker stained areas inside the cell corresponding to chromosomal DNA as seen by thin section electron microscopy, and dent(s) midway through one cell, and 1/3- and 2/3-lengths in another cell, possibly representing one or more division septa. This quick and high resolution with depth-of-field microscopy technique is unmatched to image live hydrated ultrastructure, and has much potential for application in the study of fragile biological specimens.

Escherichia coli

X-ray microscopy and imaging of Escherichia coli, LPS and DNA.

Ultrastructural examination by transmission and scanning electron microscopy involves a series of specialized preparation steps which may introduce artefacts in the micrographs. X-ray microscopy can take instant images of specimens but is mostly restricted to a few synchrotron X-ray sources. We have utilized a bench-top nanosecond laser-plasma to produce a single-shot source of nanosecond X-rays tuned for maximum contrast with carbon-rich material. To examine the ultrastructure by absorption profiles, we utilized a laser-produced plasma generated by a single-shot laser (1.06 microns wavelength, 5 x 10(12) W cm-2 intensity) focused on to a silicon target as an X-ray source for high-resolution X-ray microscopy. This approach eliminates the specimen preparation steps. Whole hydrated cells of Escherichia coli and purified preparations of lipopolysaccharide (LPS) and chromosomal DNA (cDNA) were streaked onto poly(methyl methacrylate) (PMMA)-coated grids (resist). This resist was exposed to X-rays under vacuum at a distance of 2.5 cm from the target disc. The silicon plasma produced by a 10-ns burst of laser energy (at 20J) radiates strong emission lines in the region of 300 eV. The X-rays penetrate the sample and their absorption profile is transferred on to the resist where PMMA acts as a negative to generate an image. By atomic force microscopy imaging of this photoresist we have visualized layers around cells of E.coli, darker areas inside the cell probably corresponding to cDNA, and preliminary images of LPS and DNA molecules. This technique has resolution at the 100 A level, produces images similar to the space-filling models of macromolecules and may be of great value in the study of the ultrastructure of hydrated live biological specimens.

DNA, Bacterial

Variations of posterior vitreous detachment.

AIMS: To identify variations in posterior vitreous detachment (PVD) and establish a clinical classification system for PVD. METHODS: 400 consecutive eyes were examined using biomicroscopy and vitreous photography and classified the PVD variations-complete PVD with collapse, complete PVD without collapse, partial PVD with thickened posterior vitreous cortex (TPVC), or partial PVD without TPVC. RESULTS: In each PVD type, the most frequently seen ocular pathologies were as follows: in complete PVD with collapse (186 eyes), age related changes without vitreoretinal diseases (77 eyes, 41.4%) and high myopia (55 eyes, 29.6%); in complete PVD without collapse (39 eyes), uveitis (23 eyes, 59.0%) and central retinal vein occlusions (8 eyes, 20.5%); in partial PVD with TPVC (64 eyes), proliferative diabetic retinopathy (30 eyes, 46.9%); and inpartial PVD without TPVC (111 eyes), age related changes without vitreoretinal diseases (62 eyes, 55.9%). This PVD categorisation was significantly associated with the prevalence of each vitreoretinal disease (p < 0.0001, chi 2 test on contingency table). CONCLUSIONS: PVD variations can be classified into four types, which is clinically useful because each type corresponds well to specific vitreoretinal changes.

Adult

Copper-ion-catalyzed vitreous liquefaction in vivo.

To investigate copper-ion (Cu2+)-catalyzed vitreous liquefaction in vivo, Cu2+ solution (10 mumol) was injected into the vitreous cavity of rabbits. At 24 h after the injection, the gel and liquid vitreous were weighed, and the percent of vitreous liquefaction was calculated. Cu2+ injection resulted in liquefaction of 58% of the vitreous, although control eyes had 12% liquefaction (p < 0.1). The vitreous liquefaction was more pronounced in the presence of exogenous ascorbic acid. However, the addition of mannitol, a hydroxyl-radical-specific scavenger, significantly suppressed the Cu(2+)-catalyzed vitreous liquefaction. The free radicals generated by the Cu(2+)-catalyzed oxidation system may cause vitreous liquefaction in vivo.

Animals

Lenticular autofluorescence in normal tree shrews.

Lenticular autofluorescence was measured in vivo in 25 normal tree shrews using a fluorophotometer with a specially designed small animal adapter to assess the usefulness of this model of lens pathology. The average variability in the lenticular autofluorescence measurements was 8.7%. The average ratio between lenticular autofluorescence of the left eye and that of the right eye was 0.99 +/- 0.13 SD. There was a strong correlation between age and mean lenticular autofluorescence in both eyes (correlation coefficient, 0.988; p < 0.001). The calculated annual increase in lenticular autofluorescence was 23.7 ngEq/ml. This method of measurement is reliable, and lenticular autofluorescence in tree shrews varies little among animals other than a variation caused by aging. Moreover, the tree shrew, which is believed to be a primate, is small and easy to breed. This could be a useful animal model to study various lens pathologies in humans.

Aging

[Fluorophotometry of the animal eye].

We assessed the usefulness of the Fluorotron Master fitted with a small animal adapter. We also discuss the measurement conditions for the tupai, which is a promising experimental mammal. A good concentration for measurements with this instrument ranged from 0.5 x 10(-6) to 1.0 x 10(-6) g/ml. A suitable time for fluorophotometry of the tupai was 30 min. after injection of fluorescein-Na, and a suitable dose of the fluorescein-Na was 2 mg/kg. We could do ultrafiltration for measurement of the protein unbound fluorescein concentration from a minimal sample of the blood using a hematocrit tube, and thus could reduce the deleterious effects of blood sampling on the animal. This instrument is useful for the estimation of the blood-ocular barrier permeability of animal models.

Animals

Fluorophotometry with a small animal adapter.

The usefulness of the Fluorotron Master fluorophotometer fitted with a small animal adapter was evaluated with tree shrews. We also determined that the optimal measurement parameters for these animals, which are regarded as promising for experimental use in ophthalmic research, are: fluorescein-Na concentration, 0.5 x 10(-9) to 1.0 x 10(-6) g/mL; fluorescein-Na dosage, 2 mg/kg; measurement time, 30 minutes after injection of fluorescein-Na into the ocular compartments. Results indicated that ultrafiltration for the measurement of protein-unbound fluorescein can be done with a hematocrit tube and a minimal blood sample, thereby reducing the impact of the sampling on the animal's general condition. The Fluorotron Master with the small animal adapter can offer advantages in estimating the blood-ocular barrier permeability using disease models in tree shrews.

Animals

Clinical characteristics of traumatic macular holes.

The clinical characteristics of 20 eyes of 20 consecutive patients with traumatic macular hole were studied retrospectively. The macular holes were elliptical with irregular edges in 19 eyes (95%) and ranged in size from 0.2 to 0.5 DD (disc diameter). Posterior vitreous detachment (PVD) was found in only 3 eyes (15%); the vitreous was detached from the macula in only one of these 3. Other conditions found in the 20 patients included commotio retinae, vitreous hemorrhage, hyphema, and choroidal rupture. These findings strongly suggest that most traumatic macular holes develop in the absence of PVD and that the pathogenesis is independent of the occurrence of PVD. We believe that the macular rupture (hole) is caused mechanically by the force of the impact on the posterior pole and the ocular deformity which result from blunt trauma.

Accidents

The successful repair of annuloaortic ectasia using Cabrol's operation in a 5-year-old child with Marfan's syndrome of the forme fruste type.

This report describes our experience of treating a 5-year-old boy with annuloaortic ectasia and a presumptive diagnosis of Marfan's syndrome. He had elongation of the distal aortic arch and dilatation of the abdominal aorta. Surgical repair of annuloaortic ectasia was successfully carried out using Cabrol's operation, following which no significant perioperative complications developed. To our knowledge, this is the first reported case of Cabrol's operation being successfully performed on a child aged 5 years or less.

Aortic Diseases

Biomicroscopic vitreous videography.

PURPOSE: The authors report on the technique of biomicroscopic vitreous videography using the recently developed charge coupled device (CCD) video camera. METHODS: The authors performed biomicroscopic vitreous videography using the CCD video camera connected to a slit-lamp biomicroscope to examine 50 eyes with age- or disease-related vitreous changes in 50 patients. The cases were categorized as follows: 20 normal senile eyes; 10 eyes with retinal breaks; 10 eyes with diabetic retinopathy; 5 eyes with uveitis; and 5 eyes with high myopia. Noncontact positive preset lenses were used for observing the posterior vitreous; a wide-angle funduscopic contact lens and a three-mirror contact lens were used to observe the peripheral vitreoretinal changes. RESULTS: Using the CCD video camera, dynamic vitreous changes were documented clearly using real-time television monitoring. The video system also allowed rapid review of the vitreous changes. CONCLUSION: This new biomicroscopic vitreous videography system will contribute to the study of vitreoretinal diseases and be valuable as an educational tool.

Adult

[Traumatic macular hole and posterior vitreous].

We studied retrospectively the clinical features of 18 eyes of 18 patients with traumatic macular hole. The macular hole was elliptical with irregular edges in 17 eyes (94%) and its size ranged from 1/5 to 1/2 of the disc diameter. Posterior vitreous detachment was found in 3 eyes (17%); the vitreous was detached from the macula in only one of these 3 eyes. Commotio retinae, vitreous hemorrhage, hyphema, or choroidal rupture was also found in several eyes. These findings suggest that traumatic macular hole develops usually in the absence of posterior vitreous detachment and its pathogenesis is independent of the occurrence of posterior vitreous detachment. We speculate that blunt trauma-induced deformity of the eyeball or impact on the posterior pole provides a mechanical cause for the macular rupture.

Adolescent

[Vitreous videography with a scanning laser ophthalmoscope].

We used a scanning laser ophthalmoscope (SLO) to attempt better visualization of the posterior vitreous than conventional biomicroscopy. We observed the posterior vitreous using the SLO in patients with age- or disease-related vitreous changes. We compared the results using lasers of several wavelengths. The detailed structure of the posterior hyaloid membrane, its collagen mesh structure with premacular defect, and the prepapillary glial ring were clearly visualized by direct laser illumination. Persistent attachment of the vitreous gel to the macula through the oval defect in the posterior hyaloid membrane was seen even with repeated eye rotations. Short wavelength laser was preferable to a long wavelength one for observing the posterior hyaloid membrane. Vitreous opacities were clearly visualized with all lasers as dark spots with fundus monitoring by retro illumination. Thus in vivo visualization of the posterior hyaloid membrane and vitreous gel was achieved using this new technique, which is more advantageous than the conventional biomicroscopy because of its monochromatic laser delivery system with confocal light detection.

Adolescent

[Biomicroscopic findings of premacular posterior vitreous].

The presence of vitreous gel in front of the macular area is still controversial. In order to understand the anatomy of the premacular vitreous, the posterior vitreous was observed biomicroscopically and slit-lamp photographs were taken in 100 eyes without posterior vitreous detachment. We defined the premacular vitreous as the lacuna seen when optically empty space, a demarcated oval shaped dark area without Tyndall phenomenon, was observed in front of the macula. Although the premacular vitreous showed liquefaction, the Tyndall-phenomenon, indicating the presence of formed vitreous gel, was observed in most cases. Premacular lacuna was observed in 3 cases with high myopia, and in one case with vitreoretinal degeneration syndrome. The bursa premacularis or premacular precortical vitreous pocket observed in autopsy eyes could barely be observed in living eyes.

Adolescent

Vitreous videography using the scanning laser ophthalmoscope.

To establish the method of vitreous videography using the scanning laser ophthalmoscope, we observed the vitreous in patients with age- or disease-related vitreous changes. We compared the results using four laser wavelengths. The detailed structures of the posterior vitreous cortex, a premacular defect, and a prepapillary glial ring were clearly shown under direct laser illumination. A short wavelength laser was found to be superior to a long wavelength laser for observing the posterior vitreous cortex. Vitreous opacities were clearly shown as dark spots under retroillumination using any of the four laser wavelengths. In vivo visualization of the vitreous was achieved using this new technique, which is superior to conventional biomicroscopy because of its monochromatic laser delivery system and confocal light detection capacity.

Adolescent

Determination of trace amounts of albumin in human bronchoalveolar lavage fluids by fluorometry with chromazurol S.

Fluorometry using chromazurole S (CAS) was applied to determine trace amounts of albumin in human bronchoalveolar lavage fluids (BALF). The calibration curve was linear in the range of 5-60 micrograms/ml of albumin. The CAS method was proven to be much more selective for albumin than for IgG. Freezing of BALF samples did not affect albumin analysis by the CAS method after storage at -20 degrees C for 80 days. This finding suggests that albumin in the BALF samples is stable under these conditions. The correlation was highly linear (r = 0.966) between the albumin levels in concentrated BALF samples (n = 47) determined by the CAS method and by radial immunodiffusion. The CAS method is sensitive enough to determine albumin levels in unconcentrated BALF samples, whereas radial immunodiffusion often requires concentration. The former method is more suitable for measuring albumin in BALF samples than the latter, because concentration by ultrafiltration results in poor reproducibility. The concentration of albumin in BALF samples of healthy volunteers (n = 5) and patients with sarcoidosis (n = 32) was determined by the CAS method. There was a statistically significant difference (P < 0.01) in the albumin levels in BALF samples between healthy subjects and patients with sarcoidosis at a clinically active state (n = 15). This finding shows that the determination of albumin levels in BALF samples is useful for investigating lung diseases and that the CAS method is promising in the determination of trace albumin in BALF samples, because it is simple, sensitive and precise.

Bronchoalveolar Lavage Fluid

[Property and clinical availability of the noncontact autofocus specular microscope].

Reproducibility of results and possible clinical use of a newly developed noncontact autofocus specular microscope were studied. Morphological parameters of corneal endothelium were analyzed on images taken with the noncontact autofocus specular microscope and the ordinary contact specular microscope simultaneously in 10 eyes of 10 normal subjects. Intraphotographic and interphotographic study by autoanalysis of 5 images each of 10 eyes taken by the noncontact instrument showed small coefficient variation of mean cell area and cell density (less than 5%, individually). These morphological parameters showed about 7% difference between autoanalysis and manual analysis of the images taken by the noncontact instrument. In comparison with the images taken by the contact instrument, the images taken by the noncontact instrument showed about 7% difference in mean cell area and cell density (p < 0.001, individually). In respect of reproducibility, this newly developed noncontact autofocus specular microscope is clinically useful. The differences of morphological parameters between the new and ordinary instruments might be caused the differences in corneal shapes.

Adult