[Genetics of dementia]. / Genetik der Demenzen.

Most psychiatric diseases in adulthood have a multifactorial origin. This also applies for most cases of dementia; however, rare familial forms of Alzheimer's disease and frontotemporal lobar degeneration follow an autosomal dominant (Mendelian) inheritance pattern. Alzheimer's disease that is caused by mutations in the genes for presenilin 1, presenilin 2 and amyloid precursor protein has an onset under the age of 65 years in most cases. Approximately 10 % of frontotemporal lobar degeneration cases display an autosomal dominant inheritance pattern. According to the current S3 guidelines on dementia of the German Association for Psychiatry, Psychotherapy and Psychosomatics and the German Society of Neurology, genetic counseling should be offered if an autosomal dominant disease pattern is suspected. Genetic counseling must conform to the German Genetic Diagnostics Act (Gendiagnostikgesetz).

Three-generational alkaptonuria in a non-consanguineous family.

OBJECTIVE: Alkaptonuria (AKU) is a rare inborn error of metabolism of aromatic amino acids and considered to be an autosomal recessive trait caused by mutations in the homogentisate 1,2-dioxygenase (HGD) gene. A dominant pattern of inheritance has been reported but was attributed to extended consanguinity in many cases. However, we have observed a non-consanguineous family segregating AKU in a dominant manner over three generations. RESULTS: All affected individuals presented with typical features of AKU including darkening of the urine, ochronosis, arthropathy, and elevated urinary excretion of homogentisic acid. Sequence analysis of the HGD gene from genomic DNA of two affected individuals, uncle and niece, revealed a heterozygous missense mutation (M368V) in the uncle that was not present in his niece. Microsatellite genotyping demonstrated that both were heterozygous at the HGD locus and shared one haplotype. This haplotype did not contain a detectable HGD mutation. The haplotype was also found in a healthy son of the niece, making a dominant HGD mutation unlikely. Moreover, sequencing of cDNA from lymphoblastoid cells of the niece did not reveal an HGD mRNA with a potentially dominant-negative effect. CONCLUSION: Rare causes of the uncommon AKU inheritance in this family have to be considered, ranging from the coincidence of undetectable HGD mutations to a dominant mutation of a second, hitherto unknown AKU gene.

Detection of a significant association between mutations in the ACVRL1 gene and hepatic involvement in German patients with hereditary haemorrhagic telangiectasia.

Hereditary haemorrhagic telangiectasia (HHT) is a heterogeneous multisystemic dysplasia of the vascular tissue. This autosomal dominant inherited disorder shows a wide variation in its phenotypic expression. Between 8 and 78% of the HHT patients show arteriovenous malformations of the liver. The molecular basis for hepatic manifestation is still unknown. Two genes are known to play a major role in the development of HHT: activin A receptor type II-like 1 gene (ACVRL1) and ENG. Previously, we and others showed that hepatic involvement is associated with mutations in the ACVRL1 gene, but rarely caused by ENG mutations. Here, we report about the sequencing analysis of a new cohort of 18 adult HHT patients. In these patients, we identified eight novel (four in ACVRL1 and four in ENG) and eight already known mutations. Statistical analysis of our entire data revealed significant differences in the distribution of ACVRL1 and ENG mutations among HHT patients with and without liver involvement (p = 0.0016). The positive predictive value for type 2 HHT (ACVRL1 positive) patients to develop liver disease until the age of 52 years is 68.4%. We conclude that molecular genetic testing of HHT patients is important for prognosis with respect to liver disease.

LDL receptor-related protein 5 (LRP5) affects bone accrual and eye development.

In humans, low peak bone mass is a significant risk factor for osteoporosis. We report that LRP5, encoding the low-density lipoprotein receptor-related protein 5, affects bone mass accrual during growth. Mutations in LRP5 cause the autosomal recessive disorder osteoporosis-pseudoglioma syndrome (OPPG). We find that OPPG carriers have reduced bone mass when compared to age- and gender-matched controls. We demonstrate LRP5 expression by osteoblasts in situ and show that LRP5 can transduce Wnt signaling in vitro via the canonical pathway. We further show that a mutant-secreted form of LRP5 can reduce bone thickness in mouse calvarial explant cultures. These data indicate that Wnt-mediated signaling via LRP5 affects bone accrual during growth and is important for the establishment of peak bone mass.

Cause of progression in Duchenne muscular dystrophy: impaired differentiation more probable than replicative aging.

Replicative aging of myogenic cells (satellite cells) owing to enhanced myofiber turnover is a common explanation of the progression of Duchenne muscular dystrophy (DMD). This hypothesis has been reexamined recently by telomere length measurements in dystrophic tissue. We evaluate the controversial results of these studies. We also review a large body of in vitro, animal (mdx), and patient data which indicate that impaired differentiation, but not replicative aging, is the leading cause of progression in DMD. We recommend in vivo investigations of cell kinetics in DMD muscle, as well as telomere length and telomerase analyses of DMD satellite cells in vitro for a definite judgement of the replicative aging hypothesis. Analogous investigations were helpful in AIDS research where replicative aging was embraced as a simple explanation of the paradigmatic CD4 lymphocyte decline but had to be rejected in favour of more complex models of disturbed lymphocyte homeostasis and regeneration. The question of replicative aging versus impaired differentiation is relevant for the understanding of therapeutic failures and the design of new strategies. Impaired differentiation is compatible with the failure of myoblast transfer in DMD and calls for further studies on the myofiber environment. Replicative aging, on the other hand, could possibly be treated by telomerase gene delivery.

PMP22 Thr118Met is not a clinically relevant CMT1 marker.

It is controversial if peripheral myelin protein 22 gene (PMP22) Thr118Met represents a functionally irrelevant polymorphism or, since hemizygosity for this variant has been found in two patients with Charcot-Marie-Tooth disease type 1 (CMT1 patients), it can act as a recessive CMT1 mutation. To shed further light on this variant and its diagnostic value we searched for carriers in 1018 individuals from the German general population, in 104 probands with hereditary neuropathy with liability to pressure palsies (HNPP) who were carriers of the 1.5-Mb deletion frequently associated with this disorder, in 187 patients with the 1.5-Mb duplication, and in 22 patients with a CMT1 phenotype who did not have any detectable anomaly in the PMP22 gene. Using allele-specific PCR we identified 14 [allele frequency (AF)=0.007] in the German general population, one (AF=0.01) in the HNPP group and six (AF=0.016) and two (AF=0.05) carriers of the PMP22 Thr118Met mutation in the CMT1 groups with and without gene defect. Carriers from all groups showed nerve conduction velocities which did not differ from typical values for these groups. We conclude that the hemizygous occurrence of the 118Met allele does not usually cause CMT1. Because of previous reports on its association with disease, and because its allele product shows abnormalities in in vitro expression systems, it seems possible that this mutation, together with yet unidentified factors, predisposes to CMT1. Alternatively, previously reported disease associations occurred by chance, and the 118Met allele causes biochemical abnormalities irrelevant for CMT1 formation. In either case this mutation is not a clinically relevant disease marker.

Mutations in the skeletal muscle alpha-actin gene in patients with actin myopathy and nemaline myopathy.

Muscle contraction results from the force generated between the thin filament protein actin and the thick filament protein myosin, which causes the thick and thin muscle filaments to slide past each other. There are skeletal muscle, cardiac muscle, smooth muscle and non-muscle isoforms of both actin and myosin. Inherited diseases in humans have been associated with defects in cardiac actin (dilated cardiomyopathy and hypertrophic cardiomyopathy), cardiac myosin (hypertrophic cardiomyopathy) and non-muscle myosin (deafness). Here we report that mutations in the human skeletal muscle alpha-actin gene (ACTA1) are associated with two different muscle diseases, 'congenital myopathy with excess of thin myofilaments' (actin myopathy) and nemaline myopathy. Both diseases are characterized by structural abnormalities of the muscle fibres and variable degrees of muscle weakness. We have identified 15 different missense mutations resulting in 14 different amino acid changes. The missense mutations in ACTA1 are distributed throughout all six coding exons, and some involve known functional domains of actin. Approximately half of the patients died within their first year, but two female patients have survived into their thirties and have children. We identified dominant mutations in all but 1 of 14 families, with the missense mutations being single and heterozygous. The only family showing dominant inheritance comprised a 33-year-old affected mother and her two affected and two unaffected children. In another family, the clinically unaffected father is a somatic mosaic for the mutation seen in both of his affected children. We identified recessive mutations in one family in which the two affected siblings had heterozygous mutations in two different exons, one paternally and the other maternally inherited. We also identified de novo mutations in seven sporadic probands for which it was possible to analyse parental DNA.

Telomere length distribution and Southern blot analysis.

Southern blot analysis of terminal restriction fragments (TRFs) is the standard method for quantitative examination of telomere length distributions. Since TRFs contain a subtelomeric component, central parameters of the TRF distribution n(L) such as the arithmetic mean (M) or the median (Me) cannot be derived directly from Southern blot data, i.e. from the optical density distribution OD(L). Several estimates have been applied instead; the seeming arithmetic mean A, the "center of mass" C, and the positions of maximal (P) and half-maximal optical density (P(1/2)). We show that C> A> M for any non-truncated distributions n(L), and P> M> P1/2 for any symmetrical unimodal n(L). Symmetric appearance on a Southern blot, however, suggests positive skewness of n(L). Thus, a lognormal form of n(L) may be considered. Then, C> A> M> P=Me> P(1/2). Alternatively, a Weibull distribution may be assumed. The latter is compatible with negative feedback-regulation of the telomere lengths. Using the maximum likelihood method we compare these distributions with FISH-data on telomere lengths in different cell types. The fit of the lognormal distribution is clearly superior. Lognormal genesis may relate to telomere breakage and recombination. Truncation of the upper end of the TRF distribution is possible due to Southern blot artifacts. Thereby, the order of the estimates may change to P> C> A. Having minimal sensitivity to truncation, P seems to be the optimal choice. however, the variability of P is high since peakedness of OD(L) and DNA length resolution are inversely related.

Examination of telomere lengths in muscle tissue casts doubt on replicative aging as cause of progression in Duchenne muscular dystrophy.

The mean telomere length (TL) of somatic cells indicates their replicative age. In comparison with normal leukocytes (-0.03 kbp/y, 6.2 kbp at 80 y), we found advanced TL shortening in premature aging due to ataxia-telangiectasia or the Nijmegen chromosomal breakage syndrome. Duchenne muscular dystrophy (DMD) has been related to replicative senescence of satellite cells (SCs) caused by increased fiber turnover. Therefore, we determined TLs in DMD muscle. Because the regenerated fiber nuclei are produced by SCs. telomeres of both fiber and SC nuclei should be shortened. In DMD the SC number is increased. We determined that up to the age of 7 y the sum of fiber and SC nuclei should be large enough (73%) for the detection of TL shortening. Normal muscle fibers have negligible turnover rates, and, as expected, we did not find age-related TL shortening (10-83 y, n = 24, 8.3 +/- 0.5 kbp). Surprisingly, there was only slight TL shortening in patient muscles (DMD, 0.3-4.8 y, n = 4, 8.3 +/- 0.7 kbp; 5-7 y, n = 7, 7.9 +/- 0.4 kbp; limb-girdle muscular dystrophy 2C, 13 y, 7.6 kbp; Becker muscular dystrophy, 7 y, 8.5 kbp). Similarly, the peak positions of the telomere blots varied only slightly (DMD, 10.0 +/- 0.9 kbp; normal: 10.7 +/- 0.9 kbp). In accordance with our TL findings we derived less than 4 annual doublings per SC from published histologic data on DMD.

Advanced telomere shortening in respiratory chain disorders.

Cell and tissue damage in respiratory chain disorders have been related to increased production of reactive oxygen species (ROS). We measured telomere lengths in such disorders since ROS have also been implicated with telomere shortening. We investigated whole blood cell DNA of 14 patients with MELAS-related mitochondriopathy and two patients with the LHON-associated G11778A mutation of the mitochondrial genome. The phenotypes were variable and included an unusual case of schizophrenia-like psychosis associated with the A3243G mutation. As compared to healthy controls telomere shortening in the patient group was advanced (P < or = 0.006). We compare this finding with the accelerated telomere shortening in Down's syndrome and in chromosomal breakage syndromes. We discuss possible relations between advanced telomere shortening and selective constraints that act on proliferating cells with respiratory chain dysfunction.

Congenital myopathy with excess of thin myofilaments.

Three unrelated young children are reported to have suffered since birth from muscle hypotonia and two of them from fatal respiratory insufficiency. Muscle tissues were found to contain large masses of thin myofilaments, immunologically identified as containing actin, but without further morphological features. These masses of thin filaments were found in different muscles at different occasions in the three children, suggesting a disease-specific morphological and possibly nosological feature all of them justifying classification as congenital myopathy with excess of actin or actin myopathy. The lesions were dissimilar to hyaline bodies in that the latter consist of granular material which is faintly positive for ATPase activity whereas the masses of thin filaments are devoid of ATPase activity. Two of our three patients also had intranuclear rods with virtually no sarcoplasmic rods suggesting the term of this congenital myopathy as actin myopathy with intranuclear rods.

Wheat kernel ingestion protects from progression of muscle weakness in mdx mice, an animal model of Duchenne muscular dystrophy.

A simple, reproducible test was used to quantify muscle weakness in mdx mice, an animal model of Duchenne muscular dystrophy. The effect of bedding on wheat kernels and of dietary supplementation of alpha-tocopherol on the progression of muscle weakness was investigated in mdx mice. When measured during the first 200 d of life, mdx mice developed muscle weakness, irrespective of bedding and diet. When kept on wood shavings and fed a conventional rodent diet, mdx mice showed progressive muscle weakness over the consecutive 200 d, and eventually showed a significant weight loss during the next 200-d observation period. Progression of muscle weakness and weight loss were almost completely prevented in mdx mice that were kept on wheat kernel bedding. In contrast, only incomplete maintenance of muscle strength and body weight was observed in mdx mice kept on wood shavings and fed the alpha-tocopherol-supplemented diet. It is concluded from these experiments that a component of wheat kernels other than alpha-tocopherol is essential to prevent the progression of muscle weakness in mdx mice.

A cysteine 3340 substitution in the dystroglycan-binding domain of dystrophin associated with Duchenne muscular dystrophy, mental retardation and absence of the ERG b-wave.

We report the first C-terminal missense mutation in a Duchenne muscular dystrophy patient. A G10227A transition of the dystrophin gene was found which resulted in the substitution of a highly conserved cysteine at position 3340 within the second half of the dystroglycan-binding domain. Residual amounts of 427 kDa dystrophin were detected in western blot analysis of the patient's muscle tissue, and immunohistological examination revealed weak traces of dystrophin on all fibers. Sarcolemmal staining intensity of 43 kDa beta-dystroglycan was also reduced. Mental retardation in our patient and absence of the b-wave in his electroretinogram indicate that central nervous functions of dystrophin isoforms also depend on the presence of cysteine 3340.

Neurosensory hearing loss in secondary adhalinopathy.

We report mild-to-moderate neurosensory hearing loss and severe childhood autosomal recessive muscular dystrophy with adhalin-deficiency in two siblings from a Bulgarian sibship of Turkish origin. Microsatellite analysis excluded linkage to the adhalin gene, mutations of which cause limb girdle muscular dystrophy (LGMD) 2D, but was compatible with linkage to the gene locus of LGMD 2C on chromosome 13q12. Compound heterozygosity of the affected siblings was detected in this chromosomal region. A severe autosomal recessive form of neurosensory deafness has been linked to the same region (locus NSRD1) which is now contained in a 7 Mb YAC contig. Using polymorphic markers and STS PCR primers mapping in this contig, we did not find evidence for major rearrangements in the suspected region. These preliminary findings are not in favor of, but do not completely exclude a contiguous gene syndrome in these cases. Therefore, we consider a potential role of the putative 13q12 gene product and/or adhalin in neurosensory hearing.

NADH in the pyramidal cell layer of hippocampal regions CA1 and CA3 upon selective inhibition and uncoupling of oxidative phosphorylation.

N2-laser-induced fluorescence in combination with the time and spectral resolution of fluorescent NADH molecules allows on-line measurement of relative NADH concentration with high spatial resolution (diameter of optical fibre 200 microns, lambda(exc) = 337 nm, lambda(det) = 460 nm). Energy metabolism was impaired in submerged rat hippocampal slices using the inhibitors amytal, 3-nitropropionate (3-np), sodium cyanide (1 mM each) and the uncoupling agent 2,4-DNP (200 microM). A microprocessor-controlled repeated positioning of the optical fibre in CA1 and CA3 pyramidal cell layers, and CA1 stratum radiatum (CA1SR). Time-dependently, NADH fluorescence increased reversibly upon perfusion with amytal and cyanide. It was unchanged by perfusion with 3-np for 40 min and rapidly decreased upon perfusion with 2,4-DNP. The CA1/CA3 ratio of NADH fluorescence mildly decreased to 0.92 +/- 0.04 (mean +/- S.D.) at 10 min (P < 0.05) and 0.89 +/- 0.05 at 20 min (P < 0.01) upon perfusion with amytal. The CA1/CA3 ratio increased to 1.56 +/- 0.28 at 10 min (P < 0.01) and 1.29 +/- 0.35 at 20 min (P < 0.05) upon application of 2,4-DNP. Fluorescence in CA1SR was similar to fluorescence in CA1 upon perfusion with 2,4-DNP and similar to CA3 upon perfusion with amytal. We conclude that NADH fluorescence can be measured with high regional selectivity and specificity in hippocampal slices. Selective inhibition of mitochondrial complex I and uncoupling of energy metabolism differentially impair NADH concentration in different hippocampal areas.

Islet cell antibodies in diabetes mellitus associated with a mitochondrial tRNA(Leu(UUR)) gene mutation.

An A3243G point mutation of the mitochondrial tRNA(Leu(UUR)) gene was detected in a Caucasian family with maternal diabetes mellitus and signs of mitochondrial dysfunction such as muscular hypotonia, encephalopathy, lactic acidosis, stroke-like episodes (MELAS), neurosensory hearing loss, cardial pre-excitation, and short stature. Low levels (10 JDF) of islet cell antibodies (ICA) in insulin-treated diabetes of the mother and impaired glucose tolerance with high levels of ICA (80 JDF) in her older son indicated that mitochondrial diabetes mellitus may involve beta cell damage. Furthermore, exocrine pancreas cell damage may also occur since the stroke-like episodes of this son were combined with pancreatitis. In all family members HLA types and plasma antioxidants were determined. Normal concentrations of hydro- and lipophilic antioxidants (including ubiquinol-10) were found.