Transplantation of cultured human retinal pigment epithelium into rabbit subretina

Transplantation of normal retinal pigment epithelium (RPE) into a diseased eye holds promise for treatment of several blinding disorders. Previous studies have involved immunosuppression and implantation of freshly isolated cells. We report here the successful transplantation of cultured human RPE cells into rabbits that were not immunosuppressed. A modified pars plana transvitreal technique was used for RPE transplantation. The cultured RPE cells, loaded with carbon as a marker, were transplanted into the denuded Bruch's membrane of albino rabbits. The animals were followed for from 1 week to 3 months. On histologic examination at 2 months, no infiltrating lymphocytes were found in the vitreous cavity or choroid, even though Bruch's membrane was damaged. At about 3 months there were some macrophages in the subretina of transplanted eyes, indicating that an immunoreaction does occur eventually. Electron microscopy of the transplanted RPE showed apical-basal polarity and gap junctions. Restored function was attested to by the presence of phagosomes and phagocytosed outer segments in the transplanted cells. Our findings suggest that there is a weak, delayed immunoreaction to human RPE cells transplanted beneath the retina of the rabbit; however, functional recovery of the transplanted cells occurs before this immune response develops.     

Ultrastructural studies of spermiogenesis in rabbit exposed to chronic fluoride toxicity

OBJECTIVE: To compare skinfold thickness measurements with bioelectrical impedance analysis (BIA) as a measure of body fat for use in a survey of children (the National Study of Health and Growth). DESIGN: Part cross-sectional, part repeated measurement study. SETTING: A junior school in Bath. SUBJECTS: 42 boys and 33 girls aged from 9 to 11 years. INTERVENTIONS: Measurements of BIA, height, weight, and triceps, biceps, subscapular and suprailiac skinfolds. RESULTS: All measurements were highly repeatable with intraclass correlation coefficients > 0.90. The level of agreement between estimates of percentage of body fat derived from prediction equations based on impedance or skinfold measurements respectively was poor: the mean difference (impedance estimate minus skinfold estimate) was 4.67% (95% range -3.47 to 12.82) for boys and 7.81% (95% range 1.27 to 14.34) for girls. The two estimates were found to correlate highly (r = 0.83 for boys and r = 0.81 for girls) because weight, used to convert estimates of fat-free mass derived from impedance to fat mass, was highly correlated with impedance and moderately highly correlated with skinfold thicknesses. The correlations of resistance (R) and (H)2/R with skinfold thicknesses were very low. There was a moderate correlation of R and H2/R with log(weight-for-height index), but lower than that of log(weight-for-height index) with each of the skinfolds. CONCLUSIONS: As currently available equations for converting impedance-based estimates of total body water to fat mass are not fully developed for use in children of varying ages, estimates of body fat calculated from skinfold thickness measurements remain preferable in epidemiological studies of children's health and growth.     

Medical surveillance studies of workers exposed to low level benzene

The paper presents th results of an investigation of haematotoxicity in workers exposed to low benzene concentrations. Forty-seven female workers in the shoemaking industry, exposed to solvent mixture and twenty-seven non-exposed controls were examined. Benzene concentrations in the working atmosphere ranged from 1.9 to 14.8 ppm. Significant differences in the levels of benzene in blood and phenols in pre- and post-shift urine between the exposed and control groups confirmed benzene exposure. Haemoglobin level and mean corpuscular haemoglobin concentration were significantly lower, and mean corpuscular volume was higher in the shoemaking workers than in controls. In the subgroup of shoemaking workers exposed to benzene concentrations of 5 ppm or lower, no differences in haematological parameters were found. In conclusion, exposure to a benzene concentration lower than 5 ppm does not appear to produce an increased level of abnormal haematological outcomes detectable in routine medical surveillance. The results of the study corroborate the present maximum permissible concentrations (5 ppm) as a protective limit preventing the onset of haematotoxic non-leukemogenic effects of chronic benzene exposure.     

Influence of hyperoxia on in vitro growth of rabbit middle ear epithelium and auditory meatal epithelium

As stated in the conclusion, "life is a thing of macromolecular cohesion in salty water." This brief historical overview shows that "compensatory" organic osmolytes take an essential place in this cohesion. It reviews the major steps of the study of these compounds over more than 100 years, from the early beginnings of 1885 until now, showing some of its fascinating developments and ending on the idea that the most fascinating is still to come. This study can be taken as an example of the richness of the comparative approach.     

Toxicity of Xylocaine to rabbit corneal endothelium

PURPOSE: To assess the toxicity of lidocaine hydrochloride (Xylocaine) to the corneal endothelium. SETTING: Department of Ophthalmology, H?tel-Dieu Hospital, Paris, France. METHODS: Rabbit corneas were excised and the endothelium was exposed to balanced salt solution (BSS), Xylocaine 1%, or Xylocaine 5% (5 corneas/group) for 20 minutes. The endothelium was then stained with trypan blue and alizarin red, and 5 photomicrographs were taken of each cornea at a standard magnification and analyzed with a digital imaging system (Biocom 200). RESULTS: Xylocaine solutions produced changes in endothelial cell morphology, but there was no cell staining with trypan blue. Corneas exposed to Xylocaine 5% had more marked cell alterations. Small areas of cells were lost from all 15 corneas, mainly at the periphery, but the differences among the 3 groups of corneas were not significant. CONCLUSION: Exposure of rabbit corneal endothelium to Xylocaine solutions in vitro was not associated with trypan blue staining of endothelial cells.     

Experimental studies of sensitization to beryllium, zirconium, and aluminum compounds in the rabbit

In a study designed to assess the potential sensitizing and granulomagenic capacities of selected metallic salts, rabbits were inoculated intradermally with zirconium aluminum glycinate (ZAG), sodium zirconium lactate (NZL), aluminum chlorhydrate (ACH), BeSO 4, and ovalbumin (OVA) by single and multiple injections. Animals immunized with BeSO4 and with OVA developed delayed skin reactivity as well as antigen-specific alveolar macrophage migration inhibition. Neither single nor multiple injections of ZAG or ACH resulted in clear-cut positive skin reactivity, macrophage migration inhibitory factor (MIF) production, or lymphocyte stimulation. Rabbits inoculated with multiple injections of NZL (500 microng) showed some marginally positive macrophage migration inhibition and skin reactivity. Histologically, ZAG and ACH were found to induce well-organized foreign-body granulomas after intradermal injection in both normal and inoculated rabbits. NZL and BeSO4 also induced skin granulomas, but these were less organized and distinct. Cell viability and ultrastructural studies indicated that BeSO4 was highly toxic for isolated alveolar macrophages in vitro at concentrations above 10 microng/ml, but NZL and ZAG did not exert such an effect at these dose levels. BeSO4 also depressed lymphocyte stimulation in sensitized animals which demonstrated delayed skin reactivity and macrophage migration inhibition.     

Enzyme-synthetic approach to demonstration of phosphorylase activity in the living rabbit cornea

An enzyme-synthetic method of demonstrating phosphorylase was applied to the living rabbit cornea, and polyglucose particles synthesized from glucose-1-phosphate in vivo were studied electron microscopically. In the corneas in which the medium for phosphorylase was applied from the anterior chamber or the bulbar subconjunctiva, synthesized polyglucose particles were found in the cytoplasmic matrices of the epithelium. When the medium was deposited in the conjunctival sac, a few synthesized polyglucose particles were found in the cytoplasmic matrices of only the superficial layer of the corneal epithelium. These findings suggest that metabolites for glycogen metabolism come mainly from the aqueous humor in the anterior chamber. The polyglucose particles synthesized by the enzyme-synthetic method in vivo resemble native glycogen particles. In addition, these particles were not overproduced because the synthesis of polyglucose is probably regulated in vivo.     

Cytokine-induced sequential migration of neutrophils through endothelium and epithelium

RATIONALE AND OBJECTIVES: To evaluate different vein graft-coated stent systems in the endovascular treatment of experimental arteriovenous fistulae (AVF) in a canine model. METHODS: Bilateral carotid-to-external jugular vein AVF were created. Two balloon-expandable tantalum stents (Strecker stent), two self-expanding nitinol stents (Strecker stent), and one stainless-steel stent (Wallstent) were coated with autologous vein grafts and placed using a transfemoral approach. Angiography was performed immediately after stent placement and at week 1 and 3, as well as at months 3, 6, and 9. All stents were removed and underwent histologic examination. RESULTS: Occlusion of the AVF succeeded with the Wallstent and both tantalum stents. The nitinol stents were misplaced, maintaining the AVF. One undersized tantalum stent and the Wallstent were occluded after 3 weeks. One nitinol stent was occluded at 3 months, whereas the two remaining stents were patent during the whole observation period. No inflammatory tissue response was seen, and no host-versus-graft reaction was present. CONCLUSIONS: Preparation and implantation of vein graft-coated stents, especially in the case of self-expanding stent systems, is cumbersome. This restricts the common use of such a coating, which shows an excellent biocompatibility. Vein graft-coated stents might be of use in infected endangered vessels.     

Experimental studies of polymorphonuclear leukocyte on mandibular bone infection model in rabbit

Polymorphonuclear leukocytes (PMN) play important roles in the prevention of infection at an early stage. We studied the function of these leukocytes using rabbit models of mandibular bone infection to evaluate the conditions which could not be reproduced in human beings Streptococcus milleri NCTC7331 and Bacteroides fragillis NCTC9343 were inoculated into the mandibular bone of rabbits using the Satoh-Heimdahl method, to produce supposed multiple infection models. Rabbits inoculated with these bacteria were used as a test group, and animals with surgically induced inflammation were used as a control group. We compared the number of leukocytes, the function of PMN, and histopathologic findings. 1) The number of leukocytes increased after treatment, reached a perk on day 3, gradually diminished later, but remained slightly higher than the baseline level on day 7, with persistence of inflammation in both groups. 2) Adhesiveness, ability to migrate and NBT reduction, were accelerated in both groups. 3) These functions of PMN accelerated more in the test group because the bacteria inoculated induced stronger inflammatory reactions and activated a series of infection defense mechanisms in the hosts. 4) Histopathologic examination after treatment showed invasion of inflammatory cells, predominantly leukocytes, in both groups, but heavier and more extensive infiltration in the group treated with the bacteria. All measurements were higher in the test group than the control group. These results showed that in the test group, causative or accompanied microorganisms activated the host's infection defense mechanisms and accelerated the functioning of PMN at an acute stage of infection.     

Immunohistochemical localization of cytosolic sialidase in the epithelium of rat cornea and conjunctiva

OBJECTIVE: To compare the hemodynamic change, course of recovery and adverse reaction in desflurane, sevoflurane and enflurane inhalation under low flow for patients undergoing selective abdominal surgery. METHODS: Following thiopental induction, 42 patients were divided into three groups: the first group received desflurane, the second sevoflurane and the third enflurane. During surgery, one of the agents around 1 minimum alveolar concentration (MAC) was used for maintenance, with fresh gas flow of 0.3-0.5 L/min for either desflurane or enflurane, and (0.8-1.0) L/min for sevoflurane. Heart rate (HR), blood pressure and end-tidal anesthetic concentration were monitored continuously. Time intervals from cutting off anesthetic to patient opening eyes, following commands, stating the time and location and recalling date of birth were all recorded. In addition, postoperative nausea or vomiting was traced. RESULTS: Desflurane caused the least cardiovascular depression. with mean arterial pressure (MAP) maintained significantly better at 10, 30 and 60 minutes of surgery and with HR stabilized right after incision as well. Its emergence was 2 times faster than sevoflurane, and 5-6 times quicker than enflurane. However, nausea or vomiting was found the lowest in patients receiving sevoflurane, though no distinct difference was shown between desflurane and enflurane. Nevertheless, patients under desflurane suffered less. CONCLUSIONS: Desflurane offers significant advantages for clinical anesthesia maintenance over sevoflurane and enflurane. It provides minimal cardiovascular depression, much quicker recovery, yet still causes some nausea during emergence.     

Oxidant injury to the alveolar epithelium: biochemical and pharmacologic studies

This multifaceted study involved a combined biochemical and cellular analysis of oxidant metabolism by a lung cell at risk from injury by endogenous and environmental oxidants, the pulmonary alveolar type II epithelial cell. Within the framework of this study, a method was developed for effectively delivering antioxidant enzymes and alpha-tocopherol to the intracellular compartment of alveolar epithelial cells. Alveolar type II cells are key sources of pulmonary surfactant phospholipids and apoproteins and serve as progenitors of type I alveolar epithelium, thus playing an important role in the re-epithelialization of the lung alveolus after exposure to pulmonary oxidants. The type I and II pulmonary epithelium also play an essential collaborative role in maintaining the integrity of the air-blood barrier of the lung. Because of these critical properties of the alveolar epithelium and their recognized sensitivity to oxidant stress derived from diverse sources, such as activated inflammatory cells, hyperoxia, the environmental oxidants and nitrogen dioxide, and surgical procedures, such as cardiopulmonary bypass and lung transplantation, we endeavored to understand more about the oxidant metabolism and antioxidant pharmacology of these cells. In our experiments, we made the observation that loss of differentiated oxidant generation and antioxidant properties of type II cells occurs very rapidly in vitro. For example, we observed a 50% to 75% reduction in the specific activities of type II cell superoxide dismutase, catalase, and glutathione peroxidase, all critical scavengers of cell superoxide and hydrogen peroxide and key enzymes in the attenuation of hydroxyl radical formation. Although the differentiated characteristics of the type II cell antioxidant defenses changed in vitro, they may have become more reflective of type I alveolar epithelial cells. The type I cell is the most vulnerable for oxidant damage in the alveolus because of its large surface area and the possibility of a reduced antioxidant capacity compared to type II alveolar epithelium. In spite of this limitation, we were able to culture type II cells and study their adaptive and toxic responses to exogenously administered oxidant stress. We also observed that a significant source of self-generated oxidants in type II cells was the enzyme xanthine oxidase. Normal rates of oxidant production by this enzyme had an inhibitory effect on incorporation of biosynthetic precursors into surfactant phospholipids; these effects were eliminated by the xanthine oxidase inhibitor, allopurinol.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of aflatoxin B and rubratoxin B on bacteriophage and rabbit cornea cells

Rabbit cornea cells exhibited a sensitivity to 1 mug aflatoxin B1 and 5 mug rubratoxin B per ml of growth medium. No changes were observed in the bacteriophages tested in the presence of 25 mug aflatoxin B1 or 100 mug rubratoxin B per ml of medium by the plaque-forming unit method or single-step growth curves.     

Effects of epidermal growth factor and insulin on migration and proliferation of primary cultured rabbit gastric epithelial cells

We assessed the influence of epidermal growth factor (EGF) and insulin on gastric epithelial restoration in vitro. Rabbit gastric epithelial cells were cultured and formed a complete monolayer cell sheet in 2 days. We created a wound (1.8 +/- 0.05 mm2) by denuding an area of cells, and EGF (0.1-30 ng/ml) and/or insulin (1 nM-1 microM) was added. The restoration process, which included cell migration and proliferation, was monitored by measuring the cell-free area every 12 h for 2 days. Proliferating cells were detected by sequential staining with bromodeoxyuridine (BrdU). Control cells showed complete repair in 36-48 h and restoration was accelerated dose-dependently by EGF or insulin. EGF plus insulin further accelerated restoration, which was then completed in 12-24 h. EGF and/or insulin increased the number of BrdU- positive cells. The results indicated that EGF and insulin additively accelerated gastric epithelial wound repair by stimulating both the migration and the proliferation of gastric epithelial cells (particularly the former).     

Hyperbaric oxygen inhibits the growth of cultured rabbit lens epithelial cells without affecting glutathione level

The influence of electron beam irradiation on migrational behaviour of additives present in food packaging material was studied. The migration experiments were carried out on irradiated and non-irradiated polypropylene pouches containing aqueous food simulating liquids (FSL) for 10 days at 40 degrees C. The controls were irradiated and non-irradiated pouches without FSL contact. After the contact period, the polypropylene and the FSL were analysed. A comparison between the results obtained by the two analyses showed the migration of three products of antioxidant degradation from the polypropylene into the FSL, and a partial decomposition of these migrants in the FSL.     

Lysosomal alterations in cultured macrophages exposed to anorexigenic and psychotropic drugs

Intravenous urography was performed in fourteen children with Ullrich-Turners syndrome. Renal abnormalities have been noted in twelve cases (85,7%). The most frequent kidney anomalies were malrotations (28,5%), horseshoe kidneys (21,4%) and double kidneys (21,4%). Malformations of kidney are thus a very frequent feature in Ullrich-Turners syndrome. It is therefore recommendable to perform in any case of Ullrich-Turners syndrome an intravenous urography, since these abnormalities are clinically latent.     

Specular microscopic observations of the corneal endothelium in the normal rabbit

Endothelial specular microscopy and pachometry were performed on both eyes of 14 young adult New Zealand white rabbits with clinically normal eyes. Endothelial cells of the central corneas formed a mosaic-like pattern of homogenous hexagonal cells with a mean diameter of 20.6 +/- 1.0 micron sd. The mean number of cells per mm was 2998 +/- 326 sd and the mean corneal thickness was 0.38 +/- 0.02 mm sd.     

Systematic growth studies, cocultivation, and cell hybridization studies of Werner syndrome cultured skin fibroblasts

The growth of 20 independently derived skin fibroblastlike (FL) cell strains from three individuals with Werner syndrome (adult progeria) was compared with the growth of ten FL cell strains from normal individuals. Population growth rates and total replicative life spans of Werner syndrome strains averaged 53% and 27%, respectively, of the growth rates and life spans of non-Werner strains. In the first few passages, four Werner syndrome strains demonstrated population growth rates in the low normal range, but the longest-lived Werner syndrome strain had only 75% of the total replicative potential of the shortest-lived normal strain. Exponential growth rates, cloning efficiencies, and saturation densities of Werner strains were also reduced, whereas cell attachment was normal. Viable cells (identified by dye exclusion) were maintained in post-replicative Werner syndrome and control cultures for periods of at least 10 months; there was no evidence of accelerated post-replicative senescence of cell death of Werner syndrome FL cells. Cocultivation of Werner syndrome and normal strains did not influence population growth rates of either strain. Two proliferating hybrid clones were obtained from fusions of normal and Werner syndrome FL cell strains and these hybrids displayed the reduced growth potential typical of Werner syndrome FL cells. These studies confirm that low growth rate and sharply reduced replicative life span are characteristic of cultured skin FL cells from patients with Werner syndrome, and they suggest that these characteristics are not affected by complementation with non-Werner FL cells.     

Transformation of tracheal epithelium exposed in vitro to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)

Using an organ culture/cell culture system, we transformed rat tracheal epithelial cells in vitro by exposure to MNNG. Ten tracheal organ cultures per group were exposed twice (at days 3 and 6) to 0,0.001, 1.0 or 10.0 microgram MNNG/ml of medium. Following this exposure, the explants were placed on the bottom of culture dishes to initiate epithelial cell outgrowths and establish primary cultures. Each explant was replanted 8-10 times to produce multiple outgrowths. The number of primary cultures and the number of subsequently established cell lines obtained was carcinogen-dose-dependent. The average number of primary epithelial cell cultures per explant after exposure to 0, 0.001 1.0 and 10.0 microgram MNNG/ml was 1.3, 1.5, 3.3, and 4.6, respectively. The average yield of cell lines per explant in these groups was 0, 0.8, 1.3, and 2.0, respectively. It is apparent that cell lines could only be established from carcinogen-exposed epithelium. These cell lines are currently being tested for tumorigenicity in vivo. To date, of 35 cell lines tested between the 5th and 15th passages, 5 cell lines from the 10 microgram MNNG/ml group and 2 from the 1.0 microgram MNNG/ml group have produced palpable tumors upon injection into immunosuppressed recipients.