Hermann Rink

Thyroid stimulating hormone — Receptor overexpression in brain of patients with Down Syndrome and Alzheimer's disease

Thyroid hormone abnormalities are strongly associated with Down Syndrome (DS) with elevated thyroid stimulating hormone (TSH) levels as the most consistent finding. Using subtractive hybridization for gene hunting we found significant overexpression of mRNA levels for the TSH-receptor (TSH-R) in brain of a fetus with DS. Based upon this observation we determined TSH-R protein levels in five brain regions of patients with DS (n=8), Alzheimer disease (AD, n=8) and controls (C, n=8). Western blots revealed significantly elevated immunoreactive TSH-R protein(s) 40 kD and 61 kD in temporal and frontal cortex of patients with DS and, unexpectedly, in AD. Levels for the 40 kD protein in temporal cortex were 1.00+/-0.036 (arbitrary units+/-SD) in C, 1.35+/-0.143 in DS, 1.52+/-0.128 in AD; in frontal cortex: 1.00+/-0.046 in C, 1.10+/-0.03 in DS, 1.10+/-0.038 in AD. Levels for the 61 kD protein in temporal cortex were 1.01+/-0.015 in C, 1.47+/-0.013 in DS, 1.623+/-0.026 in AD; in frontal cortex: 1.02+/-0.020 in C, 1.18 +/-0.123 in DS, 1.48+/-0.020 in AD. These results show that elevated brain immunoreactive TSH-R is not specific for DS and maybe reflecting apoptosis, a hallmark of both neurodegenerative disorders, as it is well-documented that the thyroid hormone system is involved in the control of programmed cell death.

Decreased transcription factor junD in brains of patients with Down syndrome.

JunD is a member of the Jun family of transcription factors (TF), recently shown to negatively regulate cell growth and antagonizes transformation by the protooncogene ras: c-jun decreases while junD is accumulating when fibroblasts become quiescent. Furthermore, overexpression of junD resulted in slower growth and an increase in cells in G0/G1. Performing gene hunting on fetal Down syndrome (DS) brain we found a sequence downregulated and homologous to junD. This observation made us examine junD protein levels in adult brain specimens. Western blot experiments were carried out in five brain regions of aged patients with DS (n = 9), controls (n = 9) and patients with Alzheimers disease (AD, n = 9). We found that junD in AD brains were comparable to controls, whereas junD levels were significantly and remarkably reduced in frontal, temporal lobe and cerebellum of patients with DS. These findings may indicate a specific finding in DS and were not linked to the AD-like-neuropathological changes of plaques and tangles, observed in DS from the fourth decade, which is also suggested by the findings of downregulated junD at the mRNA level revealed by the gene hunting technique (subtractive hybridization) in fetal DS brain. We propose that junD plays a role for the impaired development and wiring of DS brain, maybe already early in life.

Increased steady state mRNA levels of DNA-repair genes XRCC1, ERCC2 and ERCC3 in brain of patients with Down syndrome.

Although deficient DNA-repair was proposed for neurodegenerative disorders including Down Syndrome (DS), repair genes for nucleotide excision repair or X-ray repair have not been studied in brain yet. As one of the hypotheses for the pathogenesis of brain damage in DS is oxidative stress and cells of patients with DS are more susceptible to ionizing irradiation, we decided to study ERCC2, ERCC3 and XRCC1, representatives of repair genes known to be involved in the repair of oxidative DNA-damage. mRNA steady state levels of ERCC2, ERCC3, XRCC1, a transcription activator (TAF-DBP) and an elongation factor (EF1A) were determined and normalized versus the housekeeping gene beta-actin in five individual brain regions of nine controls and nine DS patients. Although different in the individual regions, DNA-repair genes were consistently higher in temporal, parietal and occipital lobes of patients with DS accompanied by comparable changes of TFA-DBP and EF1A. Our results are the first to describe DNA-repair gene patterns in human brain regions providing the basis for further studies in this area. We showed that DNA-repair genes ERCC2 and ERCC3 (excision-repair-cross-complementing-) for nucleotide excision repair and XRCC1 (X-ray-repair-cross-complementing-) for X-ray-repair, were increased at the transcriptional level with the possible biological meaning that this increase may be compatible with permanent (oxidative?) DNA damage.

Guidelines for the Classification of Lenses and the Characterization of Lens Proteins

In November 1980 a workshop for the classification and characterization of lenses and lens proteins was held in Louvain-La-Neuve, Belgium. Some essential results are summarized in this paper.

Comparison of DNA Strand Break Induction in CHO Cells Measured by Alkaline Elution and by Fluorometric Analysis of DNA Unwinding (FADU)

DNA damage in X-irradiated CHO cells was measured by alkaline filter elution and compared to fluorometric analysis of DNA unwinding (FADU). The FADU method proved to be as sensitive as the alkaline filter elution technique in detecting X-ray induced DNA breaks. Strand break induction was also measured after treatment with four radical generating chemicals (hydrogen peroxide, bleomycin, mitomycin C and methyl viologen) using the FADU technique.

Sodium, Potassium and Calcium Contents of Bovine Lenses in Dependence on Age

The electrolyte content in whole bovine lenses as well as in the cortical and nuclear regions was determined in a longitudinal study. K+, Na+ and calcium contents show distinct a

Gene expression in fetal Down syndrome brain as revealed by subtractive hybridization.

Information on gene expression in brain of patients with Down Syndrome (DS, trisomy 21) is limited and molecular biological research is focussing on mapping and sequencing chromosome 21. The information on gene expression in DS available follows the current concept of a gene dosage effect due to a third copy of chromosome 21 claiming overexpression of genes encoded on this chromosome. Based upon the availability of fetal brain and recent technology of gene hunting, we decided to use subtractive hybridization to evaluate differences in gene expression between DS and control brains. Subtractive hybridization was applied on two fetal brains with DS and two age and sex matched controls, 23rd week of gestation, and mRNA steady state levels were evaluated generating a subtractive library. Subtracted sequences were identified by gene bank and assigned by alignments to individual genes. We found a series of up- and downregulated sequences consisting of chromosomal transcripts, enzymes of intermediary metabolism, hormones, transporters/channels and transcription factors (TFs). We show that trisomy 21 or aneuploidy leads to the deterioration of gene expression and the derangement of transcripts described describes the involvement of chromosomes other than chromosome 21, explains impairment of transport, carriers, channels, signaling, known metabolic and hormones imbalances. The dys-coordinated expression of transcription factors including homeobox genes, POU-domain TFs, helix-loop-helix-motifs, LIM domain containing TFs, leucine zippers, forkhead genes, maybe of pathophysiological significance for abnormal brain development and wiring found in patients with DS. This is the first description of the concomitant expression of a large series of sequences indicating disruption of the concerted action of genes in that disorder.

Immunological Properties of Rat Lens Gamma-Crystallins

The rat lens contains three immunologically different γ-crystallins, γ1-, γ2-, and γ3-crystallins. With the aid of specific antisera, it could be established that part

Altered gene expression in fetal Down syndrome brain as revealed by the gene hunting technique of subtractive hybridization.

Information on gene expression in brain of patients with Down Syndrome (DS, trisomy 21) is limited and molecular biological research is focussing on mapping and sequencing chromosome 21. The information on gene expression in DS available follows the current concept of a gene dosage effect due to a third copy of chromosome 21 claiming overexpression of genes encoded on this chromosome. Based upon the availability of fetal brain and recent technology of gene hunting, we decided to use subtractive hybridization to evaluate differences in gene expression between DS and control brains. Subtractive hybridization was applied on two fetal brains with DS and two age and sex matched controls, 23rd week of gestation, and mRNA steady state levels were evaluated generating a subtractive library. Subtracted sequences were identified by gene bank and assigned by alignments to individual genes. We found a series of up- and downregulated sequences consisting of chromosomal transcripts, enzymes of intermediary metabolism, hormones, transporters/channels and transcription factors (TFs). We show that trisomy 21 or aneuploidy leads to the deterioration of gene expression and the derangement of transcripts describes the impairment of transport, carriers, channels, signaling, known metabolic and hormone imbalances. The dys-coordinated expression of transcription factors including homeobox genes, POU-domain TFs, helix-loop-helix-motifs, LIM domain containing TFs, leucine zippers, forkhead genes, maybe of pathophysiological significance for abnormal brain development and wiring found in patients with DS. This is the first description of the concomitant expression of a large series of sequences indicating disruption of the concerted action of genes in this disorder.

Crystallins of Lens Epithelial Cells during Aging and Differentiation

Lens epithelial cells from rats were serially subcultured. Five distinct stages (A–E) developed, which have been classified according to morphological and cellular parameters. The proteins of these cells were analyzed by isoelectric focusing. The differences in the protein patterns (β- and γ-region) in diploid as well as in aneuploid stages are discussed. Changes in the protein patterns of diploid stage B cells with different CPD numbers are considered as age-related effects. The occurrence of elongated cell forms and the synthesis of γ-crystallin (only in the diploid stage) favours the assumption that lens epithelial cells from the rat retain some features of their differentiation capacity in vitro.