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Biomedical subjects

H J Hoenders

Publications and source records attributed to H J Hoenders.

At least 19 recordsLinked to original sources
PubMed1992

Glycation of crystallins in lenses from aging and diabetic individuals.

Water-soluble crystallins were obtained from clear human lenses of different age (4-81-year-olds) and lenses of individuals showing senile or diabetic cataracts. Levels of early glycation products were high in the high molecular weight material (HM) and the alpha-crystallin fractions, compared with beta- and gamma-crystallins. This difference becomes more prominent upon aging. The content of total early glycation products in HM and alpha-crystallin increases clearly with age, whereas levels remain relatively constant in the beta- and gamma-crystallins. There is an elevation of early products in cataractous lenses from diabetic individuals compared with those suffering from senile cataract. Specific non-tryptophan fluorescence (excitation/emission wavelengths 370/440 nm), used as an indicator for late glycation products, increased dramatically with age and was 2-fold higher in the diabetic subjects. Levels of fluorescence decreased in the order HM > alpha- > beta- > gamma-crystallins. The results suggest an increase in glycation rate in alpha-crystallin as a result of aging and diabetes, while the rate of glycation of beta- and gamma-crystallins remains almost constant.

Adolescent↗
PubMed1992

In vivo glycation of bovine lens crystallins.

To investigate the possible role of glycation in the aging of lens proteins, we used bovine lenses as a model. We studied crystallins isolated from prenatal bovine lenses, calf lenses and lenses from mature animals (up to 20 years old). The experiments show an increase in glycation levels with age in all crystallin fractions. Regarding the lysine content of the different crystallins, gamma-crystallin showed relatively high levels of early glycation products. The results also revealed high levels of early glycation products for the HM material (containing mainly alpha-crystallin). In alpha-crystallin, alpha A-subunits were glycated to a higher extent compared with the alpha B-subunits. There is an age-related increase in advanced glycation products, measured as specific fluorescence (excitation/emission wavelengths 370/440 nm), mainly present in the HM and alpha-crystallin fraction.

Animals↗
PubMed1992

Glycation of human serum albumin: inhibition by Diclofenac.

Glycation is a non-enzymatic modification of proteins by sugars, probably responsible for the initiation of complications in diabetes patients and aging individuals. Our in vitro experiments show an inhibition of sugar attachment in the presence of Diclofenac. The levels of advanced glycation products, measured as specific fluorescent groups, were also lowered due to Diclofenac. These results were compared with inhibition by Aspirin (acetylsalicylic acid), a known inhibitor of the glycation process. The protection by Diclofenac is based on a non-covalent interaction of the drug with serum albumin. There is evidence that Diclofenac specifically blocks at least one of the major glycation sites of human serum albumin.

Aspirin↗
PubMed1991

Glycation-induced crosslinking of calf lens crystallins.

To investigate the possible role of glycation in the onset of diabetic cataract we used calf lens crystallins as a model. After incubation with reducing sugars, the proteins were investigated by high-pressure gel permeation chromatography, SDS-PAGE and analytical ultracentrifugation. Glucose-6-phosphate incubation resulted in an increase in mean molecular weight of all crystallin fractions and the occurrence of high-molecular weight material, partly formed by disulphide bonds. The glycated crystallins showed a decrease of tryptophan fluorescence and an increase of a specific non-tryptophan fluorescence. This fluorescence was, however, not exclusively associated with the high molecular weight protein, but was present in all protein fractions.

Animals↗
PubMed1990

Separation of serum proteins by high-performance gel-permeation column systems.

Combined TSK-Gel G4000SW-G3000SW and Zorbax GF450-GF250 columns and a Superose 6 column were evaluated for the analytical gel permeation chromatographic separation of serum proteins. Serum elution profiles showed four distinct peaks, which were attributed to immunoglobulin M (IgM), alpha 2-macroglobulin, IgG and albumin. A reproducible shoulder on the IgG peak could be attributed to IgA. These purified serum proteins and other commercially obtained proteins with relative molecular masses between 1000 kDa and 10 kDa were chromatographed on the combined systems and the separate columns. The frictional coefficient-based hydrodynamic radii of these proteins showed a linear relationship with the inverse error function complement of their partition coefficients. Using this relationship, theoretical summation plots of the data obtained from the separate columns of a system correspond to plots that were calculated by treating the combined systems as one column. The best resolution was obtained with the TSK colums, but all three column systems were suitable for the separation of clinically important immunoglobulins in serum. The retention of positively charged proteins by the TSK columns becomes noticeable after relatively short usage and is a first sign of progressive loss of resolution.

Animals↗
PubMed1990

A dynamic quaternary structure of bovine alpha-crystallin as indicated from intermolecular exchange of subunits.

The structural bovine eye lens protein alpha-crystallin was dissociated in 7 M urea and its four subunits, A1, A2, B1, and B2, were separated by means of ion-exchange chromatography. Homopolymeric reaggregates of these subunits were prepared by removal of the denaturant via dialysis. It was found that subunits were exchanged upon incubation of mixtures of two homopolymers under native conditions. New hybrid species were formed within 24 h as demonstrated by isoelectric focusing. Moreover, native alpha-crystallin molecules also exchanged subunits when incubated with homopolymeric aggregates of B2 subunits. Subunit exchange between native alpha-crystallin molecules is postulated, and a "dynamic quaternary structure" is presented that allows the polydisperse protein to adapt to changes in cytoplasmic conditions upon aging of the lens tissue.

Animals↗
PubMed1989

Protein folding and aggregation studied by isoelectric focusing across a urea gradient and isoelectric focusing in two dimensions.

Isoelectric focusing across a concentration gradient of urea was used to study the folding-unfolding and association-dissociation processes of proteins. Myoglobulin, albumin, RNase, papain, beta L- and alpha-crystallin were analyzed with this technique, and examples are given of visualized dissociation steps and of equilibrium-unfolding intermediates. Furthermore, a two-dimensional isoelectric focusing technique is presented that is useful to deduce whether a transition of a protein aggregate observed upon urea-gradient isoelectric focusing must be attributed to a change in the protein's tertiary or quaternary structure.

Animals↗
PubMed1989

Folding-unfolding and aggregation-dissociation of bovine alpha-crystallin subunits; evidence for unfolding intermediates of the alpha A subunits.

The aggregation and dissociation behavior of bovine alpha-crystallin as well as the folding and unfolding of its subunits were investigated by equilibrium studies using tryptophan fluorescence measurements and two isoelectric focusing techniques, viz. isoelectric focusing across a urea gradient and isoelectric focusing in two dimensions with different concentrations of urea. It was found that the alpha B chains lose their ability to aggregate and start unfolding at a lower concentration of urea than the alpha A chains. Equilibrium intermediates were found upon unfolding or refolding of alpha A subunits, which can be explained by a two-domain organization of these molecules.

Animals↗
PubMed1989

Racemization of aspartyl residues in proteins from normal and cataractous human lenses: an aging process without involvement in cataract formation.

A highly sensitive method was used to determine D- and L-aspartyl residues in protein fractions from normal and cataractous human lenses. A linear relationship with age was found in all fractions from normal lenses. However, no correlation with cataract could be established. It is concluded that racemization of aspartyl residues is a continuous process in eye-lens proteins but plays no role in cataract formation.

Adult↗
PubMed1987

Analysis of aspartic acid racemization. Evaluation of a chiral capillary gas chromatographic and a diastereomeric high-performance liquid chromatographic method.

A recently developed chiral gas chromatographic method and a diastereomeric high-performance liquid chromatographic method for the analysis of aspartic acid enantiomers in protein hydrolyzates have been evaluated. Although both techniques are fast and convenient, the latter is preferred because of its higher reproducibility and shorter analysis time. Furthermore, this method offers the possibility of on-line derivatization and analysis.

Aspartic Acid↗
PubMed1987

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.

Adult↗
PubMed1987

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.

Alanine↗
PubMed1987

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.

Adult↗
PubMed1986

Detection of aspartic acid enantiomers by chiral capillary gas chromatography. Determination of in vivo racemization and reduction of metal-induced background.

Racemization of aspartyl residues in proteins is a post-translational process, related to ageing. A method is presented for the detection of aspartic acid enantiomers in protein hydrolysates, based on chiral capillary gas chromatography. It is fast, easy and preferable to the usual diastereomeric dipeptide technique. We present evidence that traces of metals that are extracted from the glassware during acidic hydrolysis are the main cause for high background racemization, which often troubles accurate measurements. Effective ways to reduce this background and its standard deviation to acceptable levels are discussed, and a mathematical approach to correct for background racemization is given. Hydrolysates of aged human eye lens proteins were used to demonstrate the enantiomeric separation.

Amino Acids↗
PubMed1986

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.

Aging↗