Murine Lens Research Articles

Morphological study of signal pathway between the somata and the axon terminals of L-type horizontal cells in carp retina

Ionising radiation interacts with lenses and retinae differently. In human lenses, posterior subcapsular cataracts are the predominant observation, whereas retinae of adults are comparably resistant to even relatively high doses. In this study, we demonstrate the effects of 2 Gy of low linear energy transfer ionising radiation on eyes of B6C3F1 mice aged postnatal day 2. Optical coherence tomography and Scheimpflug imaging were utilised for the first time to monitor murine lenses and retinae in vivo. The visual acuity of the mice was determined and histological analysis was conducted. Our results demonstrated that visual acuity was reduced by as much as 50 % approximately 9 months after irradiation in irradiated mice. Vision impairment was caused by retinal atrophy and inner cortical cataracts. These results help to further our understanding of the risk of ionising radiation for human foeti (∼ 8 mo), which follow the same eye development stages as neonatal mice.

The role of ascorbic acid in senile cataract.

The reductone ascorbic acid, present in the crystalline lens in concentrations higher than those of glucose, is capable of undergoing nonenzymatic "browning" in the presence of lenticular proteins. We studied the nonenzymatic browning with ascorbate in model systems employing bovine serum albumin and lens crystallins. When bovine serum albumin, alpha-crystallin, or gamma-crystallin was incubated with [14C]ascorbic acid, the formation of yellow and then brown condensation products appeared to correlate with increasing protein-associated radioactivity. The fluorescence spectrum of these products was similar to that of homogenates of human cataractous lenses. We suggest that the nonenzymatic reaction of lens crystallins with ascorbic acid may contribute, at least in part, to the color changes of aging lenses and to the physical lenticular deterioration leading to senile cataract. High dietary intake of ascorbic acid did not affect the fluorescence spectrum of murine lenses; thus, we assume that the speed and extent of the lenticular browning reactions must depend on a deterioration of other factors of the multicomponent antioxidant system of the eye.

Evidence against γ-crystallin DNA or RNA sequences in the chicken

A cloned cDNA (pM γ 1Crl) encoding about two-thirds of a γ-crystallin polypeptide from the murine lens was used as a hybridization probe to search for the presence of γ-crystallin-like RNA or DNA sequences in the chicken. The 15-day-old chicken lens did not contain any RNA sequences homologous to the murine γ-crystallin cDNA, as judged by Northern blot hybridization. An approximate 2·3 Kbp (kilobase pair) Bam HI fragment from the chicken genome hybridized to the murine γ-crystallin cDNA in Southern blots. Cloning and sequencing of this genomic fragment, however, did not reveal any homology with γ- or β-crystallin sequences. A stretch of 22 dG:dC nucleotides was present in the cloned DNA fragment and possibly this hybridized to the cloned γ-crystallin cDNA via its dG:dC nucleotide tails introduced during the cloning procedure. These data support those of McDevitt and Croft (1977) indicating that the chicken lens lacks γ-crystallin and provide evidence against γ-crystallin mRNA or a γ-crystallin gene or pseudogene in the chicken. The absence of γ-crystallin mRNA in the embryonic chicken lens cells makes them potentially useful recipients for investigating the expression of cloned γ-crystallin genes.

Comparative two-dimensional electrophoretic analysis ofwater soluble proteins from bovine and murine lenses

Total soluble proteins were prepared from the lenses of 40-day-old mice by standard methods andeluted from Sepharose 6B. The resulting elution profile consisted of five crystallin peaks the last two of which were incompletely resolved. The total water soluble proteins and proteins prepared from fraction pools corresponding to each of the five peaks were subjected to polyacrylamide gel electrophoresis in two dimensions. Twenty-two major subunits were identified, each of which could be assigned to one of the major crystallin classes. We had expected the five peaks to correspond to the α H -, α L -, β H -, β L -, and γ -crystallin classes, well documented in the bovine system. However, electrophoretic studies showed that the five murine peaks actually contained, in descending order of molecular weight, α H -, α L -, β L - and two forms of γ -crystallins. No β H component was found. For comparative purposes, we have also analysed by identical methods, the lens crystallins of 4-month-old calves. Elution of the bovine crystallins from Sepharose 6B yielded the five protein peaks of expected composition. The elution volumes of the α H -, α L -, and β L -murine crystallins were identical with those of the corresponding bovine crystallins and the two incompletely resolved murine peaks eluted in a region similar to that of the bovine γ -crystallins. However, no murine crystallins eluted at a position corresponding to that of the bovine β H -crystallins. Bovine soluble lens proteins examined by two-dimensional electrophoresis showed limited interspecific homology with similarly prepared proteins from murine lenses. Two-dimensional electrophoresis provides a powerful analytical tool for investigations on soluble lens proteins.

Identification of the cytoskeletal proteins in lens-forming cells, a special epithelioid cell type

Proteins of contractile and cytoskeletal elements have been studied in bovine lens-forming cells growing in culture as well as in bovine and murine lenses grown in situ by immunofluorescence microscopy using antibodies to the following proteins: actin, myosin, tropomyosin, α-actinin, tubulin, prekeratin, vimentin, and desmin. Lens-forming cells contain actin, myosin, tropomyosin, and α-actinin which in cells grown in culture are enriched in typical cable-like structures, i.e. microfilament bundles. Antibodies to tubulin stain normal, predominantly radial arrays of microtubules. In the epithelioid lens-forming cells of both monolayer cultures grown in vitro and lens tissue grown in situ intermediate-sized filaments of the vimentin type are abundant, whereas filaments containing prekeratin-like proteins (‘cytokeratins’) and desmin filaments have not been found. The absence of cytokeratin proteins observed by immunological methods is supported by gel electrophoretic analyses of cytoskeletal proteins, which show the prominence of vimentin and the absence of detectable amounts of cytokeratins and desmin. This also correlates with electron microscopic observations that typical desmosomes and tonofilament bundles are absent in lens-forming cells, as opposed to a high density of vimentin filaments. Our observations show that the epithelioid lens-forming cells have normal arrays of ( i) microfilament bundles containing proteins of contractile structures; ( ii) microtubules; and ( iii) vimentin filaments, but differ from most true epithelial cells by the absence of cytokeratins, tonofilaments and typical desmosomes. The question of their relationship to other epithelial tissues is discussed in relation to lens differentiation during embryogenesis. We conclude that the lens-forming cells either represent an example of cell differentiation of non-epithelial cells to epithelioid morphology, or represent a special pathway of epithelial differentiation characterized by the absence of cytokeratin filaments and desmosomes. Thus two classes of tissue with epithelia-like morphology can be distinguished: those epithelia which contain desmosomes and cytokeratin filaments and those epithelioid tissues which do not contain these structures but are rich in vimentin filaments (lens cells, germ epithelium of testis, endothelium).

Nonparametric Determination of Modifying Factors in Radiation Action

effectiveness of different radiation qualities are commonly derived from the comparison of dose-effect curves. In this approach, the statistical analysis is frequently made difficult or impossible by the uncertain influence of the scale of effect and of the method of curve fitting. To avoid this disadvantage, a method is used to establish the dose dependence of modifying factors and their confidence limits without the intermediate step of constructing dose-effect curves. The method is based on the comparison of effect levels in pairs of irradiated samples. The statistical tests suitable for this comparison of effect levels in typical experimental situations are considered, and the method is illustrated by its application to experimental studies of the opacification of the murine lens and to growth-reduction studies of Vicia faba.