Molecular mass distribution of water-soluble crystallins from the human foetal lens during development.

The water-soluble crystallins of twenty human foetal lenses with gestational ages of 112-231 days were analysed by size-exclusion chromatography. The crystallin distribution showed similar patterns for all foetal lenses, but clear changes in the proportions of different crystallins were evident. The distribution showed that the water-soluble part of all the lenses already contained high-molecular-mass material. Also beta-crystallins of high molecular mass (beta H), formed by post-translational changes, were detected in all stages. During gestation, the percentage of high-molecular-mass crystallins and of alpha-crystallins of low molecular mass (alpha L) decreased significantly. The total beta-crystallins (beta T) and the total gamma-crystallins (gamma T) increased significantly. The low Mr crystallins were resolved into three peaks, designated beta s-, gamma H- and gamma L-crystallins. They increased significantly during development. These significant increases of the low Mr crystallins took place exclusively in the developing lens. The rate of protein synthesis of the low Mr crystallins was 23% of the total water-soluble crystallin synthesis rate.

Biochemical analysis of bovine lens sections obtained by a new sectioning device.

One mm thick layers were reproducibly sliced from the punched-out inner cylinder of bovine lenses. The layers were obtained by a special designed freeze-sectioning device. Biochemical analysis of these layers revealed a detailed picture of the lenticular distribution of the amount of water-insoluble and urea-insoluble fractions. Furthermore, the distribution of the content of glucose, fructose and sorbitol was established. The results clearly show the usefulness of a microsectioning device to obtain layers for subsequent biochemical analysis which may lead to a comprehensive understanding of age- and/or cataract-related alterations within one lens.

Non-tryptophan fluorescence of crystallins from normal and cataractous human lenses.

The soluble proteins of a newborn human lens exhibit almost no non-tryptophan fluorescence. On aging, an increase in the fluorescence of all native crystallins is found except for gamma 2-crystallin. With formation of nuclear cataract, a further increase is seen for gamma 1-crystallin. The fluorophore, excitation 355 nm/emission 420 nm, is mainly associated with one species of the gamma 1-crystallin population. It is also present as such in the soluble fraction and increases significantly with nuclear cataract formation. At least one of the gamma 1-crystallins seems to play an important role in the cataractogenic process.

Distribution of aromatic and fluorescent compounds within single human lenses.

The concentration of the free aromatic amino acids tryptophan and tyrosine, the antioxidant uric acid and some unidentified compounds have been determined in the water-soluble extracts of parts from clear and nuclear-cataractous human lenses. With age (29-90 years) and cataract formation, associated with increasing nuclear pigmentation, no significant changes were observed in content of tryptophan, tyrosine and uric acid. Furthermore, these compounds did not show variation in distribution within the lens. An unidentified fluorophore, excitation and emission maxima of 345 and 425 nm, respectively, increases significantly in content with nuclear cataract formation, especially in the nucleus. Another unidentified, presumably aromatic compound shows a striking age-related decrease.

Lanthionine, a protein cross-link in cataractous human lenses.

Oxidative processes are suggested to be a main cause of the covalent cross-linking of lenticular proteins. For a major part these cross-links are disulfides (cystine). With progression of nuclear cataract, cystine may be oxidized further to cysteic acid. Another oxidative degradation product of cystine is the symmetric thioether lanthionine. In this study, we clearly demonstrate that lanthionine is a protein cross-link of cataractous human lenses. Possible mechanisms of its formation are discussed.

Age-related variations in the distribution of crystallins within the bovine lens.

The native water-soluble proteins of equator, anterior cortex, posterior cortex and nucleus from bovine lenses in the age range 0.3-33.7 years were analyzed by high-pressure gel-permeation chromatography and high-pressure ion-exchange chromatography. Unlike the equator and cortices, the nucleus shows a gradual decrease in alpha-crystallin proportion with age which is not compensated for by an increase in HM-crystallin. The beta 6H-crystallin species, almost the only beta H-component in the youngest lens, is largely replaced by at least four fractions with higher and lower molecular weights in the older lenses. In the nucleus a beta L-component (39,000 MW) increasingly seems to replace the major beta L-crystallin (beta 2L, 50,000 MW). Moreover, a switch in the synthesis of monomeric crystallins is demonstrated. This study clearly reveals an age-related increase in the size heterogeneity of the native soluble crystallins with age.

Variation in proportion and molecular weight of native crystallins from single human lenses upon aging and formation of nuclear cataract.

Extracts of water-soluble proteins from cortex and nucleus of single human lenses have been analyzed by high-pressure gel permeation chromatography in combination with a size- and a concentration-sensitive detector. Using this rapid and accurate method, changes in molecular weight and proportion of the native crystallins upon aging and formation of nuclear cataract are observed. In the range from 30 to about 40 years the gamma-crystallin content of normal lenses increases and that of alpha-crystallin decreases; over 40 years the level of gamma-crystallin decreases and that of alpha-crystallin remains constant in the cortex (11%) as well as in the nucleus (1-2%). These changes are accompanied by an increase in the water-insoluble fraction. With progressing nuclear cataract an increase in beta 3- and a drastic decrease in gamma-crystallin content, especially in the nucleus, are accompanied by a steep increase in the water-insoluble fraction. The molecular weights of the crystallins do not change significantly, except that of alpha- and beta 1-crystallin which show an increase with age.

A comparison of two methods for the determination of thallium in urine. Differential pulse anodic stripping voltammetry and flameless atomic absorption spectrophotometry.

Two analytical procedures for the determination of thallium in urine are evaluated and compared. The two methods consist of differential pulse anodic stripping voltammetry and flameless atomic absorption spectrophotometry. Both procedures correlated well and were found to be acceptable in terms of precision and analytical recovery. However, a solvent extraction step proved to be necessary for the AAS procedure, and the linearity with respect to the concentration range was rather limited. The method based on DPASV is simple, accurate, precise, and sensitive and does not require any pretreatment of the sample. We therefore recommend the voltammetric procedure for the determination of thallium in urine, as described in this paper.

Comparative and age-dependent aspects of crystallin size and distribution in human, rabbit, bovine, rat, chicken, duck, frog and dogfish lenses.

Lens extracts obtained from eight vertebrate species were analyzed using high-performance gel permeation chromatography with differential refractive index and low-angle laser light scattering detection. The elution patterns enabled the direct determination of the proportions of the crystallin classes as well as the molecular weights of the numerous size fractions in a more rapid and accurate way than previously. Concerning the mammalian species, also age-dependent aspects could be assessed by comparing the data obtained from the extracts of differently aged lenses and/or lens parts.