Cell sources, regulatory factors and gene expression in the regeneration of the crystalline lens and retina in vertebrate animals

Over the past century extensive experimental materials have been accumulated concerning cell sources of lens and retina regeneration, successive transformations of the cells, regulatory factors, and gene expression during restitution of these eye structures. The use of nuclear and cytoplasmic markers provided convincing evidence that the removed lens is restituted from the dorsal iris cells in vivo or from embryonic cells of the pigment epithelium and retina in vitro. The removed or destroyed retina is restituted as a result of transdifferentiation of the pigment epithelium cells in amphibians, fish, birds, and mammals during embryogenesis, in larvae of some anuran amphibians, and in adult newts. Cell precursors of rods are a cell source of retina regeneration in adult fish. A subpopulation of randomly distributed cells, which are a cell source of rod formation during the normal development of the eye was found in the external nuclear layer with the use of electron microscopy and nuclear and cytoplasmic markers. These cells are not only a source of regeneration of rods, but also of cones and cells of the internal nuclear layer after destruction of the corresponding retina layers. There is a peripheral growth area in the retina of vertebrates, where multi- and unipolar cells are localized, which provide for the retina growth during ontogenesis. A paradox of retina regeneration consists in that these little differentiated cells are not a source of complete restitution of the removed or destroyed retina. They make only a small contribution to its regeneration corresponding to the growth potential of cells of this eye region, while restitution of the retina proceeds only at the expense of cells of another type of differentiation. A factor controlling the differentiated state of the cell was found in the dorsal iris during studies of lens regeneration. Removal of this factor in the early stages of cell transformations leads to the initiation of lens regeneration. The factor is not specific and was identified in many cells of vertebrates, including the pigment epithelium and limb tissues, which, as is known, may be fully restituted. Studies of gene expression during lens and retina regeneration are now at the initial stage. The greatest advances were achieved on the model of transdifferentiation of the pigment epithelium cells of chick embryos into lentoids. Expression of genes MMP115 and pP344 was established in the pigment epithelium cells, which characterize the pigmented phenotype of the initial cells. Expression of the alpha-, beta-, and delta-crystallin genes was found in the lentoids, which characterize the phenotype of regenerating structures. The gene activity appears to be switched at an intermediate stage during cell dedifferentiation. Expression of the gamma-crystallin genes during lens regeneration in adult newts is initiated after completion of dedifferentiation and cell proliferation in the dorsal iris. The genes specifically expressed in the dorsal and ventral iris and in the retina rudiment have been identified by the method of gene subtraction. Expression of homeobox-containing genes from the family of PAX genes was found during lens regeneration in adult newts and retina regeneration in adult fish. The role of growth factors (FGF) as morphogenetic factors was proved, which are involved in a yet unknown way of altering the differentiation pathway of the initial cells during formation of the neuroepithelium rudiment in chick embryos, adult newts, and fish.     

Partial properties of four glycosidases in normal human lens and variations in their enzyme activities during aging and with the advance of lens coloration

This paper reports the active glycosidases in normal human lenses and their partial properties. In addition, variations in their enzyme activities during aging and with the advance of lens coloration were also examined. Five glycosidases, alpha-glucosidase, beta-D-glucuronidase, N-acetyl-beta-D-glucosaminidase, beta-D-cellobiosidase, and alpha-L-fucosidase, were detected as active glycosidases in the normal human lens. However, the activity of beta-D-cellobiosidase was considerably low as compared to the other four glycosidases. Thus, this enzyme was omitted from this study. The four glycosidases, alpha-D-glucosidase, beta-D-glucuronidase, N-acetyl-beta-D-glucosaminidase, and alpha-L-fucosidase, showed that their enzyme activities fell abruptly between the ages of 40 and 50. Furthermore, the Km values of their enzymes exhibited some variability during aging. Namely, the Km values of their enzymes indicated the lowest value between the 40 age group and 50 age group, suggesting that the substrate affinity became the strongest at these age groups. Then, variations in enzyme activity with the advance of lens coloration were examined. In each cases, the specific activity of detectable glycosidases in color lenses, white to brown, decreased. In particular, the specific activity of enzymes in the brown lens was very low, indicating that glycosidases in the brown lens may scarcely display their enzyme activities.     

Effects of theophylline and cyclohexyladenosine on brain injury following normo- and hyperglycemic ischemia: a histopathologic study in the rat

The present study was designed to determine the effects of theophylline, an adenosine receptor antagonist, and cyclohexyladenosine (CHA), an adenosine receptor agonist, on ischemic brain injury following normo- and hyperglycemic ischemia and reperfusion in fasted male Wistar rats. Moderate hyperglycemia was achieved by administering 17% D-glucose (3 g/kg i.p.), whereas normoglycemic animals received an equal volume of saline. The animals were further divided into two groups: One group was pretreated with either theophylline (0.20 mumol/g i.p.) or an equal volume of saline; the second group received either intraventricular CHA (6.25 nmol) or mock CSF prior to the onset of ischemia. During ischemia, pericranial temperature was maintained at 36 degrees C and EEG was monitored. Cerebral ischemia was induced for 15 min, after which flow was restored and the animals were allowed to recover completely. There were no significant differences in physiologic parameters among the groups studied. Five days following the ischemic episode, the rats were perfused with formalin and the brains subserially sectioned (8 microns) in the coronal plane and stained with celestine blue/acid fuchsin. Histopathologic analysis was performed in a blinded fashion to determine percentage of dead neurons. Hyperglycemic animals had significantly greater ischemic injury in CA1, cortex, and caudate than the normoglycemic group (p < 0.01). Moreover, rats pretreated with theophylline had a significantly (p < 0.01) higher percentage of dead neurons in CA1, cortex, and caudate than corresponding controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Body composition in normal subjects: relation to lipid and glucose variables

OBJECTIVE: To describe sex- and age-dependent values of total and regional body composition as determined by dual-energy X-ray absorptiometry (DXA) in normal subjects, and furthermore to relate body composition measurements to blood lipids, glucose and insulin concentrations. DESIGN: A cross-sectional study. SUBJECTS: 173 (84 male and 89 female) healthy subjects, BMI < 30 kg/m2. MEASUREMENTS: Body composition parameters including data on total bone mineral content (TBMC), total bone mineral density (TBMD), lean body soft tissue mass (LTM), total and regional fat mass (FM) were estimated in all subjects. In 87 of the subjects fasting blood glucose, S-insulin and lipid profile were measured. RESULTS: The study population was for each sex divided into five decades for which results on body composition and blood lipids are presented. Body weight increased 2 kg per age decade, representing a significant increase in both total FM and relative FM (FM%BW) with age, and in males a central accumulation of FM. LTM decreased significantly in males but not in females, whereas TBMC and TBMD remained constant in males, but decreased in females. A significant correlation between relative FM and S-cholesterol, S-triglyceride, and in males S-insulin was found. CONCLUSION: The present study gives coherent data on bone mineral content, lean body soft tissue mass total and regional fat mass for 173 healthy subjects with a BMI below 30 kg/m2. Total body fat mass increases, and lean mass decreases with age. In males a simultaneous central accumulation of fat mass is observed. The well-known relationship between central obesity and lipids is confirmed even in non-obese subjects.     

Excitotoxic neurodegeneration induced by deprivation of oxygen and glucose in isolated retina

PURPOSE: Ischemic neurodegeneration contributes to many retinal diseases. An isolated retina model has been used to examine the neuronal cell death induced by deprivation of oxygen and glucose (simulated ischemia) as a model for ischemic disease. METHODS: Neurodegeneration in the isolated chick embryo retina was induced by simulated ischemia and assessed using biochemical (lactate dehydrogenase release) and morphologic (light microscopy) techniques. RESULTS: Simulated ischemia led to lactate dehydrogenase release gradually in a period of 6 to 24 hours. Light microscopic observations demonstrated morphologic cell degeneration well before lactate dehydrogenase release occurred. N-Methyl-D-aspartate (NMDA) and non-NMDA receptor blockers individually provided partial protection, and the combination was fully protective. No protection was provided if the antagonists were added after simulated ischemia. When NMDA receptors were blocked by MK-801, cyclothiazide, an inhibitor of desensitization at non-NMDA receptors, enhanced lactate dehydrogenase released after 1 or 2 hours of simulated ischemia. Low concentrations of glucose effectively prevented lactate dehydrogenase release, despite anoxic conditions. CONCLUSIONS: The isolated retina provided a convenient system to characterize quantitatively ischemic cell death. Retinal ischemic neurodegeneration is an excitotoxic process that involves overactivation of NMDA and non-NMDA glutamate receptors. Blockade of both of these receptor subtypes was necessary for complete neuroprotection. Receptor desensitization played a protective role. If even low concentrations of glucose were delivered to an ischemic retina in vitro, substantial neuroprotection could be achieved. This may have implications for the management of acute retinal ischemic episodes.     

The Müller (glial) cell in normal and diseased retina: a case for single-cell electrophysiology

In the retina of most vertebrates there exists only one type of macroglia, the Müller cell. Müller cells express voltage-gated ion channels, neurotransmitter receptors and various uptake carrier systems. These properties enable the Müller cells to control the activity of retinal neurons by regulating the extracellular concentration of neuroactive substances such as K+, GABA and glutamate. We show here how electrophysiological recordings from enzymatically dissociated mammalian Müller cells can be used to study these mechanisms. Müller cells from various species have Na(+)-dependent GABA uptake carriers, but only cells from primates have additional GABA receptors that activate Cl- channels. Application of glutamate analogues causes enhanced membrane currents recorded from Müller cells in situ but not from isolated cells. We show that mammalian Müller cells have no ionotropic glutamate receptors but respond to increased K+ release from glutamate-stimulated retinal neurons. This response is involved in extracellular K+ clearance and is mediated by voltage-gated (inwardly rectifying) K+ channels which are abundantly expressed by healthy Müller cells. In various cases of human retinal pathology, currents through these channels are strongly reduced or even extinguished. Another type of voltage-gated ion channels, observed in Müller cells from many mammalian species, are Na+ channels. In Müller cells from diseased human retinae, voltage-dependent Na+ currents were significantly increased in comparison to cells from control donors. Thus, the expression of glial ion channels seems to be controlled by neuronal signals. This interaction may be involved in the pathogenesis of retinal gliosis which inevitably accompanies any degeneration of retinal neurons. In particular, Müller cell proliferation may be triggered by mechanisms requiring the activation of Ca(2+)-dependent K+ channels. Ca(2+)-dependent K+ currents are easily elicitable in Müller cells from degenerating retinae and can be blocked by 1 mM TEA (tetraethylammonium). In purified Müller cell cultures, the application of 1 mM TEA greatly reduces the proliferative activity of the cells. These data clearly show that Müller cells are altered in cases of neuronal degeneration and may be crucially involved in pathogenetic mechanisms of the retina.     

Electrosynthesized non-conducting polymers as permselective membranes in amperometric enzyme electrodes: a glucose biosensor based on a co-crosslinked glucose oxidase/overoxidized polypyrrole bilayer

The sperm plasma membrane is segregated into functionally, biochemically, and structurally distinct domains yet the protein sorting pathways and assembly mechanisms that assemble these domains during spermiogenesis are incompletely understood. We previously characterized two structurally related size-variant, integral membrane proteins of 52 kDa (PM52) and 35 kDa localized to the periacrosomal plasma membrane of guinea pig cauda epididymal spermatozoa (Westbrook-Case et al., 1994). In this study we used light and electron microscopic immunocytochemistry to define the expression pattern and sorting pathway that establishes the domain-specific distribution of PM52 during spermiogenesis. The PM52 is first expressed in acrosome-phase spermatids and it localizes exclusively to the cytoplasmic lobe. Immunoelectron microscopy revealed that both cytoplasmic vesicles and the plasma membrane of the cytoplasmic lobe labeled with anti-PM52. During early stages of expression, PM52 appeared to be absent from the head region, but significant PM52 accumulation over the spermatid head was noted in late acrosomal phase spermatids. Throughout spermiogenesis PM52 extended posteriorly to the annulus, which represents a barrier preventing PM52 diffusion into the posterior tail. Following the migration of the annulus to the midpiece-principal piece junction, PM52 began to disappear from the flagellar region and at the completion of spermiogenesis most of the PM52 was restricted to the acrosomal segment. Spermatids and epididymal sperm PM52 exhibited identical sizes by SDS-PAGE and immunoblotting, indicating that they are not proteolytically modified during epididymal maturation. The PM52 antibodies were also used to screen a guinea pig testis cDNA library, and sequence determination of full-length PM52 clones demonstrated identity of a sperm membrane protein recently termed "sperad" (Quill and Garbers, 1996). Membrane barriers and potential mechanisms establishing the domain-specific residence of PM52 are discussed.     

NADP-malic enzyme from maize leaves: a fluorescence study

NADP-malic enzyme from maize leaves is covalently labeled with a fluorescent-SH reactive probe eosin-5-maleimide (EMA), which reacts with groups that are totally protected by NADP against inactivation. The comparison of the emission fluorescence spectra of the native and the modified enzyme suggests the proximity of the fluorescent groups of the native enzyme (probably tryptophanyl groups) and the EMA modified residues. Intrinsic fluorescence quenching studies shows that NADP is the only substrate capable to interact with the fluorescent excited groups of the enzyme, while Mg2+ is able to increase this interaction. Quenching studies of EMA-bound fluorescence shows that the NADP-binding site was modified and thus uncapable of further interaction with the nucleotide. When the results of protection studies are combined with those of extrinsic quenching experiments, we must conclude that EMA reacts with sulfhydryl groups that are involved in the NADP-binding site of the enzyme.     

Feline immunoglobulin E: induction of antigen-specific antibody in normal cats and levels in spontaneously allergic cats

A number of lines of evidence suggest that immunotherapy with the cytokine interleukin-2 (IL-2) may boost the immune system to fight tumors. CD4+ T cells, the cells that orchestrate the immune response, use these cytokines as signaling mechanisms for immune-response stimulation as well as lymphocyte stimulation, growth, and differentiation. Because tumor cells begin as 'self', the immune system may not respond in an effective way to eradicate them. Adoptive cellular immunotherapy can potentially restore or enhance these effects. We illustrate through mathematical modeling the dynamics between tumor cells, immune-effector cells, and IL-2. These efforts are able to explain both short tumor oscillations in tumor sizes as well as long-term tumor relapse. We then explore the effects of adoptive cellular immunotherapy on the model and describe under what circumstances the tumor can be eliminated.     

Intraocular lens performance in air, in water, and in situ: a computer study

Computer analysis is used to predict performance of four intraocular lenses with assigned values of aberration. Cylinder error and asphericity error are used as examples of possible manufacturing errors. Three measures of performance are calculated: maximum optical path difference, root mean square optical path difference, and modulation transfer function. For evaluation in air these standard test conditions are assumed: collimated green light incident on the convex surface of a plano-convex lens, with a 3 mm aperture. All four lenses show substantially improved performance in water compared to air, and a further improvement in the simulated eye (i.e., in situ). However, an aberrated 30 diopter (D) lens with performance in air comparable to an aberrated 20 D lens, performs worse in situ than does the 20 D lens. This suggests that a performance test in air that is suitable for a 20 D lens (e.g., 100 line pairs per millimeter resolution) may not be adequate for a 30 D lens. A test in air at 30% resolution efficiency may be more suitable.     

Influence of glucagon replacement on the hyperglycemic and hyperketonemic response to prolonged somatostatin infusion in normal man

Somatostatin was infused for 6 h into seven normal subjects with and without a replacement dose of glucagon. The addition of glucagon to somatostatin resulted in a 30-40% rise in plasma glucagon, whereas plasma insulin declined by 40-50% in both treatment groups. Plasma glucose and glucose production initially increased 2-fold with glucagon replacement, and subsequently declined by 2-3 h to levels comparable to those observed with somatostatin alone. After 6 h plasma glucose and glucose kinetics were no different whether or not glucagon was present. The rise in blood ketones after somatostatin was not exaggerated by glucagon replacement. We conclude that glucagon lack is not a modifying factor in the late hyperglycemic and hyperketonemic response to prolonged infusions of somatostatin.     

Effects of caffeine on glucose tolerance: a placebo-controlled study

BACKGROUND: Leishmaniasis recidiva cutis (LRC) is rare in New World leishmaniasis. Only seven cases have been reported so far. MATERIALS AND METHODS: Four cases are reported here. Parasite diagnosis was performed by classical methods of touch preparations, histopathologic sections, and cultures. In addition, the detection of parasite DNA by the polymerase chain reaction (PCR) was performed in all cases. RESULTS: Parasites were detected by at least one of the classical methods in all primary lesions; however, only the PCR was positive in the recidivant lesions. DISCUSSION: LRC cases most likely represent a reactivation of an initial infection, probably due to the persistence of parasites in scarred tissue. Although lupoid leishmaniasis (LL) has been used as a synonym of LRC, a clear difference between LRC and LL can be defined as LL is the initial clinical presentation while LRC is a recurrent lesion. CONCLUSIONS: The results indicate that it is not appropriate to use these two denominations as synonyms. The designation of LRC should be maintained in order to define recidives occurring at the border of an old scar of cutaneous leishmaniasis, avoiding the confusion with the lupoid form of the disease.     

The study of compensation systems through the lens of self-determination theory: Reconciling 35 years of debate.

Although compensation specialists generally argue for incentive systems that link rewards to performance, self-determination theory argues that such contingent rewards can have detrimental effects on autonomous motivation. The authors present a model of the motivational effects of compensation systems that attempts to reconcile the self-determination theory view and the literature on compensation. This model evaluates how compensation system characteristics, such as the amount and variability of pay, can influence the satisfaction of the needs for autonomy, competence, and relatedness, which in turn influence autonomous work motivation. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Diabetes-induced changes in lens antioxidant status, glucose utilization and energy metabolism: effect of DL-alpha-lipoic acid

The study was aimed at evaluating changes in lens antioxidant status, glucose utilization, redox state of free cytosolic NAD(P)-couples and adenine nucleotides in rats with 6-week streptozotocin-induced diabetes, and to assess a possibility of preventing them by DL-alpha-lipoic acid. Rats were divided into control and diabetic groups treated with and without DL-alpha-lipoic acid (100 mg x kg body weight(-1) x day(-1), i.p.). The concentrations of glucose, sorbitol, fructose, myo-inositol, oxidized glutathione, glycolytic intermediates, malate, alpha-glycerophosphate, and adenine nucleotides were assayed in individual lenses spectrofluorometrically by enzymatic methods, reduced glutathione and ascorbate--colorimetrically, and taurine by HPLC. Free cytosolic NAD+:NADH and NADP+:NADPH ratios were calculated from the lactate dehydrogenase and malic enzyme systems. Sorbitol pathway metabolites were found to increase, and antioxidant concentrations were reduced in diabetic rats compared with controls. The profile of glycolytic intermediates (increase in glucose 6-phosphate and fructose 6-phosphate, decrease in fructosel,6-diphosphate, increase in dihydroxyacetone phosphate, 3-phosphoglycerate, phosphoenolpyruvate, pyruvate, and no change in lactate), and 5.9-fold increase in alpha-glycerophosphate suggest diabetes-induced inhibition of glycolysis. Free cytosolic NAD+:NADH ratios, ATP levels, ATP/ADP x inorganic phosphate (Pi), and adenylate charge were reduced in diabetic rats while free cytosolic NADP+:NADPH ratios were elevated. Diabetes-induced changes in the concentrations of antioxidants, key glycolytic intermediates, free cytosolic NAD+:NADH ratios, and energy status were partially prevented by DL-alpha-lipoic acid, while sorbitol pathway metabolites and free cytosolic NADP+:NADPH ratios remained unaffected. In conclusion, diabetes-induced impairment of lens antioxidative defense, glucose intermediary metabolism via glycolysis, energy status and redox changes are partially prevented by DL-alpha-lipoic acid. The findings support the important role of oxidative stress in lens metabolic imbalances in diabetes.     

Confocal microscopy of human lens membranes in aged normal and nuclear cataracts

PURPOSE: To visualize the structure and determine the continuity of lipid membranes in lens fiber cells (LFCs) from human aged normal and cataractous lenses. METHODS: Thick sections from human nuclear cataracts and aged normal lenses were stained with the lipophilic probe DiI, and then analyzed by confocal microscopy. Staining patterns of membranes were observed in individual optical sections or three-dimensional projections of z-series taken in longitudinal section and cross-section of LFCs from different regions within the lens nucleus. RESULTS: DiI bound to and delineated the plasma membrane of LFCs from all regions of the lens nucleus. Three-dimensional projections of z-series from aged normal and cataractous lenses suggested that some of the stained lipid membranes were not continuous with LFC plasma membrane of cataractous lenses. CONCLUSIONS: The results obtained using these methods demonstrated that lipid membranes, discontinuous with the plasma membrane of LFCs, were indicative of a novel process occurring predominately in cataractous human lenses.