The role of mitochondria-derived reactive oxygen species in the pathogenesis of non-steroidal anti-inflammatory drug-induced small intestinal injury.

Non-steroidal anti-inflammatory drugs (NSAIDs) have been implemented in clinical settings for a long time for their anti-inflammatory effects. With the number of NSAID users increasing, gastroenterological physicians and researchers have worked hard to prevent and treat NSAID-induced gastric mucosal injury, an effort that has for the large part being successful. However, the struggle against NSAID-induced mucosal damage has taken on a new urgency due to the discovery of NSAID-induced small intestinal mucosal injury. Although the main mechanism by which NSAIDs induce small intestinal mucosal injury has been thought to depend on the inhibitory effect of NSAIDs on cyclooxygenase (COX) activity, recent studies have revealed the importance of mitochondria-derived reactive oxygen species (ROS) production, which occurs independently of COX-inhibition. ROS production is an especially important factor in the increase of small intestinal epithelial cell permeability, an early stage in the process of small intestinal mucosal injury. By clarifying the precise mechanism, together with its clinical features using novel endoscopy, effective strategies for preventing NSAID-induced small intestinal damage, especially targeting mitochondria-derived ROS production, may be developed.

[Histochemical studies of glycosaminoglycans in the developing eyelids of experimental microphthalmic (Cts) mice].

PURPOSE: The Cts mouse is a mutant strain which induces congenital cataract and small eyes in homozygotes. The process of eyelid development in Cts mice was examined histochemically. MATERIALS AND METHODS: Prenatal (from day 14 of gestation) and postnatal (to day 14) animals of unaffected and homozygous mice were examined. Sensitized high iron diamine staining was done after chondroitinase B/testicular hyaluronidase double digestion, and sensitized low iron diamine staining was done after Streptomyces hyaluronidase digestion. RESULTS: The main chondroitin sulfate isomer in the dermis of the eyelid changed from chondroitin sulfate A/C to chondroitin sulfate B at birth in the unaffected mice. In the homozygotes, the same change took place 4 days after birth. In the dermis of the eyelid, hyaluronic acid was first detected on day 14 of gestation and gradually increased until birth in both unaffected and homozygous mice. There was slightly more hyaluronic acid in homozygous mice until 4 days after birth. CONCLUSION: The present study showed that the maturation of glycosaminoglycan molecular species in the eyelid was retarded in Cts homozygotes.

Expression of tenascin-C and the integrin alpha 9 subunit in regeneration of rat nasal mucosa after chemical injury: involvement in migration and proliferation of epithelial cells.

Nasal mucosa covered by pseudostratified ciliated epithelia can be injured by microbial infection and physical and chemical agents. To elucidate mechanisms of regeneration, erosion of rat nasal mucosa was produced by intranasal instillation of trichloroacetic acid, and tissue specimens were then sequentially obtained after 1-14 days. Since tenascin-C (TN-C) and its receptor, alpha 9 beta 1 integrin, are assumed to play important roles in regeneration of stratified squamous epithelia, their expression was evaluated by immunohistochemistry and in situ hybridization. Three to five days after the injury, TN-C mRNA was found in epithelial cells of migrating fronts and in epithelial sheets recovering ulcerated surfaces between the fronts and normal regions. TN-C deposition was increased under such sheets. Enhanced alpha 9 staining was also evident in the involved epithelium. 5-Bromo-2'-deoxyuridine incorporation assays revealed significant increase in proliferating cells in cell sheets over TN-C deposits at 3-7 days. Therefore, we conclude that regenerating epithelial cells produce and secrete TN-C, associated with an increase in alpha 9 expression, and that interactions between these molecules could regulate migration and proliferation of the epithelial cells in an autocrine manner.

[Histological studies in the developing eyelids of congenital microphthalmic (Cts) mice].

The Cts mouse is a mutant of the Jcl:ICR strain with congenital cataract and small eyes in homozygotes. In the present study, attempts were made to measure postnatally the palpebral fissure and to examine the process of eyelid development by light microscopy in both normal (+/+) and homozygous (Cts/Cts) mice. The width of the palpebral fissure in homozygotes was significantly smaller than in unaffected mice. To investigate the process of eyelid development, tissue specimens from normal and Cts homozygous mice were prepared with hematoxylin and eosin, and observed by light microscopy on days 14, 16, and 18 of gestation and on days 0, 7, and 14 after birth. In both groups, the eyelid fused on day 16 of gestation and opened completely on day 14 after birth. The primordium of the orbicularis oculi muscle was defined on day 16 of gestation, and the primordia of hair follicles and Meibomian glands were detected on day 18 of gestation. In the present study, there were no significant differences in the process of eyelid development between normal and homozygous mice. Eyelid development has thus far been thought to be self-determining and not affected by tissue interaction, and the present findings support this line of thought.

[Four cases of persistent hyperplastic primary vitreous].

We evaluated four cases of persistent hyperplastic primary vitreous (PHPV) encountered at Nagoya City University Hospital in 1995. PHPV was seen unilaterally in three cases and bilaterally in one. The series comprised two males and two females, ranging in age from three to eight months, with an average of 4.8 months. Case 1 had a white strand running from the optic disc to the posterior surface of the lens in the left eye. Case 2 showed leukocoria in the right eye and central corneal opacity in the left. Magnetic resonance imaging (MRI) revealed total retinal detachment in both eyes. Case 3 exhibited retinal folds running from the optic disc to the posterior surface of the lens in the left eye. Case 4 showed elongation of the ciliary processes and leukocoria in the right eye. Ipsilateral total retinal detachment was seen in MRI. Three eyes of two cases were microphthalmic. Associated ocular anomalies included, posterior embryotoxon, sclerocornea, hypoplasia of the iris stroma and peripapillary staphyloma. There were accompanying systemic anomalies such as arachnoidal cyst, syndactyly, microcephalus, heart anomalies, pulmonary atresia and asplenia. Patients with PHPV should be carefully examined for the possible presence of other ocular and systemic anomalies caused by neural crest disorders.

[Developmental eye abnormalities in mouse fetuses induced by retinoic acid].

To clarify the relationship between neural crest cells and ocular anomalies, pregnant mice were treated intraperitoneally with 12.5 mg/kg retinoic acid suspended in corn oil on day 7 of pregnancy (RA group). Control mice were given an equal volume of corn oil (control group). Each group consisted of 5 mother mice, and the offsprings were removed on day 18 of gestation. The fetal mortality was 46.3% in the RA group and 2.2% in the control group. Twenty-two live fetuses of the RA group and 45 of the control group were grossly observed, and the eyes were examined histologically. In the RA group, gross malformations such as microphthalmos (95.5%), cleft lip and palate (36.4%), and central nervous system anomalies (31.8%) were observed, and in the control group, malformations such as microphthalmos (6.7%), central nervous system anomalies (2.2%), and low set ears (2.2%) were seen. Histological examination revealed microphthalmos (47.7%), anophthalmos (38.6%), faulty closure of the embryonic fissure (36.4%), developmental abnormalities of the vitreous (34.1%), aphakia (22.7%), goniodysgenesis (18.2%), and faulty separation of the lens vesicle (15.9%) in the RA group. These anomalies arose from abnormal neural crest cell migration induced by retinoic acid. They were detected in only 3.3, 1.1, 3.3, 8.9, 1.1, 2.2 and 2.2%, respectively of the control group.

[Histochemical studies on the separation of the lens vesicle].

We studied histologically the changes and distribution patterns of glycosaminoglycan molecular species during the separation of the lens vesicle in the mouse. Embryos were obtained by sacrificing pregnant mice of the Jcl: ICR strain on day 10.5 and 11 of pregnancy. Serial frontal sections were stained with hematoxylin-eosin and a sensitized high iron diamine method. To identify glycosaminoglycan molecular species in tissues, enzyme digestion (double digestions with chondroitinase B and testicular hyaluronidase) and chemical modification (nitrous acid treatment) were performed in combination with the sensitized high iron diamine method. Before separation of the lens vesicle, the glycosaminoglycan molecular species, identified in the basement membrane of the presumed corneal epithelium and intercellular matrices between the presumed corneal epithelium and lens vesicle, were chondroitin sulfate A/C and B, and those in the lens capsule were chondroitin sulfate A/C. After separation of the lens vesicle, heparan sulfate emerged in the basement membrane of the presumed corneal epithelium and intercellular matrices between the presumed corneal epithelium and lens vesicle. These results are thus taken to indicate that the changes and distribution patterns of glycosaminoglycan molecular species play an important role during separation of the lens vesicle.

Local hypothermia protects the retina from ischaemic injury in vitrectomy.

AIMS: Hypothermic irrigating solutions were used during vitrectomy in pressure induced ischaemic eyes so that their effects on retinal function and histological changes could be investigated. METHODS: After anaesthetised albino rabbits underwent closed vitrectomy, their vitreous cavities were continuously irrigated for 30 minutes at a perfusion pressure of 140 mm Hg. The rabbits were divided into three groups according to their intraocular perfusion temperatures--8 degrees C, 22 degrees C, and 38 degrees C. Electroretinograms were taken before and after irrigation. Glutamate levels in the vitreous were examined after irrigation. Eyes were enucleated on the seventh postoperative day and examined histologically. RESULTS: On the seventh postoperative day, the recovery rate of a-wave amplitudes was significantly lower in the 38 degrees C group than in the 8 degrees C group, and that of b-wave amplitudes was significantly lower in the 38 degrees C group than in either the 8 degrees C or 22 degrees C group. Retinal damage in the 38 degrees C group revealed more severe histological impairment than in either the 8 degrees C or 22 degrees C group. Oedema of the inner retinal layer was significant in both the 22 degrees C and 38 degrees C groups. Glutamates reached peak values 30 minutes after the end of ischaemia in the 38 degrees C group. However, no significant glutamate increases were detected 15 to 60 minutes after ischaemia in either the 8 degrees C or 22 degrees C group. CONCLUSION: Local hypothermia during vitrectomy in acute ischaemic eyes appears to decrease retinal damage.

Protective effects of local hypothermia in vitrectomy under fluctuating intraocular pressure.

The effects of local hypothermia on the blood-aqueous barrier and retina were investigated in experimental vitrectomy under wide-ranging fluctuations of intraocular pressure. After anesthetized albino rabbits had undergone closed vitrectomy, their vitreous cavities were continuously irrigated for 60 minutes. Perfusion pressures were fluctuated alternately between 10 and 80 mmHg. The rabbits were divided into two groups in accordance with their intraocular perfusion temperatures: 8 degrees C and 22 degrees C. Electroretinograms were taken both before and after the irrigation. Protein and prostaglandin E2 (PGE2) concentrations in the aqueous humor were measured on the 1st and 7th postoperative days. Histologic examination was performed on the eyes enucleated on the same days. There were no significant differences in either a- or b-wave amplitudes between the 8 degrees C and 22 degrees C groups. Both aqueous protein and PGE2 concentrations were significantly higher in the 22 degrees C group than in the 8 degrees C group on the 1st postoperative day. Histologically, the ciliary body showed remarkably more stromal edema in the 22 degrees C group than in the 8 degrees C group. The severity of the retinal damage did not seem to differ between the two groups. Local hypothermia during vitrectomy under fluctuating intraocular pressure inhibits the breakdown of the blood-aqueous barrier in the early postoperative stages.

Clinical evaluation of posterior embryotoxon in one institution.

To elucidate the pathogenesis of posterior embryotoxon, we estimated its incidence in our clinic and evaluated its associated ocular and systemic anomalies. Slit-lamp and gonioscopic examinations were performed on 440 randomly selected patients at Nagoya City University Hospital over a 10-month period. Posterior embryotoxon was detected in 107, 50 bilateral and 57 unilateral, cases (24.3%). Twelve (11.2%) of the 107 cases had open-angle glaucoma. Accompanying ocular anomalies included six cases of sclerocornea, two each of persistent pupillary membrane and familial exudative vitreoretinopathy, and 1 each of melanocytoma of the optic nervehead, choroidal nevus and subconjunctival dermoid cyst. Associated systemic anomalies included three cases of Alagille syndrome, two of congenital biliary atresia, and one each of congenital facial palsy with microtia, congenital adrenal hyperplasia, empty sella syndrome, Hirschsprung disease and Wilson disease. Many of these ocular and systemic anomalies were caused by the maldevelopment of neural crest cells. Patients with posterior embryotoxon should be examined for the possible presence of open-angle glucoma and for ocular and systemic anomalies related to maldevelopment of neural crest cells.

[A case of nonrhegmatogenous retinal detachment in Dandy-Walker Syndrome].

A 2-month-old female presented with nonrhegmatogenous retinal detachment in Dandy-Walker syndrome. At the fist examination, coloboma involving the optic disc in both eyes was detected. The left eye showed microphthalmos with sclerocornea, persistent pupillary membrane, hypoplasia of the iris stroma, and bullous retinal detachment near the optic disc. Chromosomal analysis revealed a mosaic pattern: 46, XX/47, XXX. Increased intracranial pressure associated with Dandy-Walker syndrome was detected by a neurosurgeon at the age of 3 months. The patient was followed for several weeks, and then nonrhegmatogenous retinal detachment appeared in the right eye. Subretinal fluid alternately increased and decreased in both eyes. A ventriculo-peritoneal shunt was performed at the age of 6 months, and the retinal detachment was remarkably reduced in both eyes after lowering of intracranial pressure. Coloboma involving the optic disc, sclerocornea, persistent pupillary membrane, hypoplasia of iris stroma, and Dandy-Walker syndrome were thought to be caused by the abnormal development of neural crest cells. We surmised that the retinal detachment in this case might have resulted from a communicating pathway between the subarachnoid space and the subretinal space. We concluded that the etiology of retinal detachment associated with optic disc anomaly should be investigated to determine adequate treatment.

[Histochemical studies on two cases of Peters' anomaly].

We examined glycosaminoglycan molecular species in the corneal stroma and the sclera histochemically in two cases of Peters' anomaly. Paraffin-embedded sections were stained with either hematoxylin-eosin or sensitized high iron diamine combined with enzyme digestion and then examined with a light microscope. In the center of the cornea, the histological specimens revealed defects of the corneal endothelium and Descemet's membrane, together with a local defect of the corneal stroma. Disorganized lens materials were detected in the corneal stroma. In the peripheral cornea, the undifferentiated iris adhered to the corneal posterior surface. Based upon the findings mentioned above, these two cases were diagnosed as having Peters' anomaly caused by faulty separation of the lens vesicle. The sclera showed normal structures in the specimen stained with hematoxylin-eosin. In both cases, the corneal stroma involved nearly equal amounts of chondroitin sulfate A/C and B. The sclera, on the other hand, contained a large amount of chondroitin sulfate A/C and a relatively small amount of chondroitin sulfate B. Keratan sulfate was never detected in either the cornea or the sclera. Thus, disorders of glycosaminoglycan molecular species were detected in the sclera as well as in the corneal stroma.

Histology of fetal eyes with oculocutaneous albinism.

The diagnosis of tyrosinase-negative oculocutaneous albinism (OCA) was made in a 19-week-old fetus by skin biopsy. Because the parents had an 11-year-old son with tyrosinase-negative OCA, they requested that the fetus be aborted at the 20th week of gestation. A histological analysis of the eyes was performed. Throughout the retina, the ganglion cell layer was separated from the inner neuroblastic layer by the inner plexiform layer. However, the number of ganglion cells was decreased and the nerve fiber layer was immature. Bipolar and horizontal cells had begun to segregate into the inner nuclear layer. Rods and cones were identifiable in the posterior, but not peripheral, retina. Cones were more numerous in the center of the retina, and no rod-free area was identifiable. In addition, the ciliary body (epithelial folds, blood vessels in the mesodermal connective tissue core, and ciliary muscle) was less developed than in a normal fetus. Melanosomes in the retinal pigment epithelium only contained filaments without melanization and were therefore classified as stage I or II melanosomes. However, the ciliary epithelium also contained some stage III melanosomes with melanin adherent to the filaments.

[Histochemical studies on glycosaminoglycans in Bruch's membrane of experimental microphthalmic mice].

The Cts mouse is a mutant mouse of the Jcl:ICR strain which has congenital cataracts and small eyes in homozygotes. Therefore, this mouse could be a model for autosomal dominant cataract with microphthalmia in humans. In the present study, attempts were made to demonstrate light-microscopically the molecular species of glycosaminoglycans (GAG) in Bruch's membrane of Cts mice on day 14, 16, and 18 of gestation and at 0, 4, and 7 days after birth by means of combined sensitized high iron diamine staining and testicular hyarulonidase digestion procedures. In Bruch's membrane of the unaffected mice at all stages examined, no substantial amount of chondroitin sulfate A/C could be detected. In homozygous mice, however, a large amount of chondroitin sulfate A/C was contained in Bruch's membrane. These results indicate that the abnormal change in GAG molecular species induced the functional disorder of Bruch's membrane and could play a certain role in the etiology of microphthalmia in homozygotes.

Implications of the severity of retinopathy of prematurity on childhood development.

This study investigated the severity of retinopathy of prematurity (ROP) and its relationship to childhood development. The subjects were 21 children with a birthweight of 2500 g or less who were tested for flash visual evoked potential (FVEP) at conceptual ages ranging from 1 to 3 years. We also examined 73 children with a birthweight of 1500 g or less, using the Developmental Quotient (DQ) test, at 2 1/2 years of age. Twenty-five of this group were given an Intelligence Quotient (IQ) test at 6 to 7 years of age. The P100 latency of FVEP in the severe ROP group was significantly longer than that of the mild ROP group (P < 0.05). The main DQ in the severe ROP group was 99.1 +/- 17.9, significantly lower than the 108.7 +/- 15.5 of the mild ROP group (P < 0.05). No significant IQ differences were found between the two groups. The proportion of children with verbal-performance IQ discrepancies was, however, significantly greater in the severe ROP group than in the mild ROP group (P < 0.05). Since infants with severe ROP are more likely to suffer developmental disorders, long-term follow-up is absolutely necessary and must be done in cooperation with pediatricians.

Histological and histochemical studies of the normal and faulty closure of the embryonic fissure in the eye of ICR mouse.

Histological and histochemical studies were carried out in Jcl:ICR mice to determine the changes in microscopic structures and glycosaminoglycan molecular species in the tissues involved in normal or faulty closure of the embryonic fissure. The purpose of this study was to elucidate the mechanism underlying the faulty closure of the embryonic fissure and to identify the key substances involved in normal and faulty closure. To obtain mice with an appropriate faulty closure of the embryonic fissure, ochratoxin A was employed as a teratogenic agent. Serial sections from tissues undergoing normal and faulty closure of the embryonic fissure were cut at a right angle to the fissure. As the staining procedures, a hematoxylin-eosin procedure and a sensitized high iron diamine method were used. A chemical modification (nitrous acid treatment) or an enzyme digestion procedure (chondroitinase ABC digestion procedure) was employed in combination with the sensitized high iron diamine method to identify the glycosaminoglycan molecular species in the tissues. The results obtained in the present study have substantiated the histophysiological importance of glycosaminoglycan molecular species during the course of histogenesis in the normal and the faulty closure of the embryonic fissure of developing murine eyes.

Effects of local hypothermia on uveal blood flow and postoperative inflammation in vitrectomy.

Effects of local hypothermia on uveal blood flow and postoperative inflammation were evaluated in experimental vitrectomy in albino rabbits. Solutions used for intraocular perfusion were maintained at 9 degrees C, 22 degrees C or 37 degrees C. Following closed vitrectomy, the vitreous cavity was irrigated for 60 minutes. Temperatures at various sites and uveal blood flow were measured before and during the procedure. Aqueous protein concentrations were checked on postoperative days 1, 7 and 14. There was a larger decrease in temperature at the retina than at the choroid or the ciliary body. Blood flow at the ciliary body was reduced to 76.0% and that at the choroid to 77.0% of the preoperative level after 60 minutes of irrigation at 9 degrees C. The decrease was 91.0% and 88.3%, respectively, after 60 minutes of irrigation at 22 degrees C. Aqueous protein concentrations in the 9 degrees C and 22 degrees C groups were significantly lower than those in the 37 degrees C group on the first postoperative day in the eyes irrigated for 60 minutes. In the eyes irrigated for 30 minutes, however, no significant differences were seen. Local hypothermia during prolonged vitrectomy seems to decrease inflammation in the early postoperative stage.

[Microphthalmos and its pathogenic classification].

Congenital microphthalmos is a common malformation encountered clinically. Microphthalmos in adults is here defined as eyes whose axial length is below 20.4 mm in males and 20.1 mm in females; in under 14-year-old children, it is eyes at least 3 square root of 2/3 below the mean for age-similar controls. Experimental animals with hereditary microphthalmos have been widely investigated, and many environmental factors given to pregnant animals frequently induce microphthalmos. In both clinical and experimental microphthalmos, there are conspicuous variations in size, and various kinds of ocular and systemic complications. Recently, fetal alcohol syndrome produced by alcohol intake during pregnancy has been reported. In this syndrome, microphthalmos is one of the important symptoms. Experimentally, microphthalmos also developed at a high incidence among mouse fetuses whose mothers were given ethanol during pregnancy. The present investigator established a preliminary etiological classification of microphthalmos in 1984. In this paper summing up newly obtained results, the relationship to neural crest cells and histochemical changes of glycosaminoglycan molecular species, the author presents a final pathogenic classification of microphthalmos, which consists of developmental disturbance of the optic vesicle, malformation of the optic cup, mesenchymal dysgenesis of the anterior ocular segment, maldevelopment of the lens, maldevelopment of the vitreous, faulty closure of the embryonic fissure and developmental disturbance of the wall of eyeball.

[Hypothermic effects on uveal blood flow and postoperative inflammation in vitrectomy].

Hypothermic effects on uveal blood flow and postoperative inflammation were evaluated in experimental vitrectomy on albino rabbits. Solutions used for intraocular perfusion were maintained at 9 degrees C and 22 degrees C. Following the vitrectomy, the rabbits' intraocular spaces were irrigated for 60 minutes. The temperature at various sites and the uveal blood flow were measured before and during the procedure. Aqueous protein concentrations were checked on postoperative days 1, 7, and 14. Another solution maintained at 37 degrees C was used as a control. There was a larger decrease in temperature at the retina than there was at the choroid and the ciliary body. Blood flow at the ciliary body decreased to 76.0% and 77.0% at the choroid after 60 minutes of irrigation at 9 degrees C. The decrease at 22 degrees C were 91.0% and 88.3%, respectively. Aqueous protein concentrations at both 9 degrees C and 22 degrees C were significantly lower than at 37 degrees C on the first postoperative day in the eyes irrigated for 60 minutes. In the eyes irrigated for 30 minutes, however, no significant differences were seen. Hypothermia during a prolonged vitrectomy operation seems to decrease inflammation in early postoperative stages.

[Multicenter prospective study of retinopathy of prematurity--II. Optimum timing of the first examination].

We reviewed the records of 360 cases of retinopathy of prematurity (ROP) in 12 institutions in Japan, and investigated the optimum timing of the first examination. On the basis of the onset of ROP and the timing of the first treatment, the first ophthalmological examination should be performed 3 weeks after birth or 29 weeks of postconceptional age at the latest. Regarding the visibility of the fundus, there was little hazy media after 29 weeks of postconceptional age. Under the basic policy that the severe progressive "plus" disease should be properly treated, we concluded that the first ophthalmological examination should be performed 3 weeks after birth at the latest. When the gestational age at birth is less than 26 weeks, funduscopy at the post conceptional age of 29 weeks is advisable.