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C M Pusch

Publications and source records attributed to C M Pusch.

At least 19 recordsLinked to original sources
PubMed2005

A genetic perspective on myopia.

Myopia is a refractive error of the eye that has a significant socioeconomic impact due to its increasing prevalence and the fact that it causes visual impairment. Its aetiology is complex and is likely to involve the interaction of environmental and genetic influences. Tight environmental influence is exemplified by defocus-induced myopia produced in animal models, while genetic factors predominate in familial occurrence of myopia with a Mendelian inheritance pattern. The involvement of numerous mediators, such as cytokines, neurotransmitters and transcription factors, in myopia development has been indicated through various lines of investigation, particular interest focussing on scleral extracellular matrix proteins and developmental genes of the eye. As high-throughput technology for large-scale genotyping and RNA expression analysis enters the field of myopia research, a productive avenue will open up for deciphering the aetiological heterogeneity of myopia and the biological pathways underlying its development.

Animals↗
PubMed2004

PCR-induced sequence alterations hamper the typing of prehistoric bone samples for diagnostic achondroplasia mutations.

Achondroplasia (ACH) is a skeletal disorder (MIM100800) with an autosomal dominant Mendelian inheritance and complete penetrance. Here we report the screening of ancient bone samples for diagnostic ACH mutations. The diagnostic G-->A transition in the FGFR3 gene at cDNA position 1138 was detected in cloned polymerase chain reaction (PCR) products obtained from the dry mummy of the Semerchet tomb, Egypt (first dynasty, approximately 4,890-5,050 BP [before present]), and from an individual from Kirchheim, Germany (Merovingian period, approximately 1,300-1,500 BP), both of which had short stature. However, these mutations were also reproducibly observed in four ancient control samples from phenotypically healthy individuals (false-positives), rendering the reliable molecular typing of ancient bones for ACH impossible. The treatment of a false-positive DNA extract with uracil N-glycosylase (UNG) to minimize type 2 transitions (G-->A/C-->T) did not reduce the frequency of the false-positive diagnostic ACH mutations. Recently, it was suggested that ancient DNA extracts may induce mutations under PCR. Contemporary human template DNA from a phenotypically healthy individual was therefore spiked with an ancient DNA extract from a cave bear. Again, sequences with the diagnostic G-->A transition in the FGFR3 gene were observed, and it is likely that the false-positive G-->A transitions result from errors introduced during the PCR reaction. Amplifications in the presence of MnCl(2) indicate that position 1138 of the FGFR3 gene is particularly sensitive for mutations. Our data are in line with previously published results on the occurrence of nonrandom mutations in PCR products of contemporary human mitochondrial HVRI template DNA spiked with ancient DNA extracts.

Achondroplasia↗
PubMed2004

Refinement of the DFNA4 locus to a 1.44 Mb region in 19q13.33.

Many forms of autosomal dominant non-syndromic hearing impairment are known. While the underlying gene defects and causative mutations have been discovered for some forms, the gene responsible for DFNA4 has remained elusive to date. Examination of a German four-generation kindred led to the identification of a 1.44 Mb map segment in contig NT_011109 as being the most likely DFNA4 candidate region in 19q13.33. The recombination breakpoints in this family and the intervals of two previously reported DFNA4 families allowed us to delineate a minimum consensus region between the markers D19S879 and D19S246. In our family, a maximum two-point LOD score of 4.5 was obtained at theta = 0 for the marker D19S867. Within the refined DFNA4 interval the public databases list more than 50 genes, from which several appear to be promising DFNA4 candidates due to similarities with animal models and with other causative genes involved in hearing disability.

Adult↗
PubMed2004

[Paraganglioma in the area of the head and neck. A review of molecular genetic research].

Paragangliomas of the head and neck region are usually benign tumors that develop from chemoreceptors of paraganglionic origin in the majority of patients. These receptors play an important role in sensing and regulation of the blood CO(2) level. Genetic alterations in the mitochondrial enzyme complex II (SDH), which is involved in respiratory chain and citric acid cycle reactions, have been shown to lead to sporadic as well as familial cases of these tumors. The gene encoding the subunit SDHD shows mutations in up to 50% of these cases. In addition, loss of heterozygosity (LOH) was demonstrated in these tumor samples and has been shown to be connected with oncogenesis of paragangliomas. Thus, SDHD is the first known tumor suppressor gene encoding a mitochondrial protein. In this article we summarize the current state of knowledge concerning the development of paragangliomas.

Cell Transformation, Neoplastic↗
PubMed2003

Palaeopathological and variant conditions of the Homo heidelbergensis type specimen (Mauer, Germany).

Although early Homo specimens are now known from a number of African, Asian and European Middle Pleistocene sites, the taxon Homo heidelbergensis was initially introduced for the Mauer jaw recovered in 1907. Fossil hominids from the earlier Middle Pleistocene of Europe are very rare and the Mauer mandible is generally accepted as one of the most ancient, with an age of approximately 700 kyr. A new preparation of the mandible was conducted in 1996 and gave rise to the detailed palaeopathological examination which is presented here. Based on comparative analyses, the extreme breadth of the mandibular ramus and its flat intercondylar incision, in conjunction with the flattening and broadening of the coronoid process tip, results either from an idiosyncratic pattern of the course and insertion of the temporalis muscle on the coronoid process or from the temporalis possessing an accessory head. The incidence of periodontal pocketing, together with a vertical reduction of the alveolar margin to approximately 3.00 mm, and a slight protuberance formed in vicinity of the right M(2)can safely be interpreted as pathognomonic indications of periodontal disease. The short distance between the enamel-dentine junction of the teeth and the horizontal alveolar margins could either be an inherited variant or may result from incipient osteoporosis. In addition, an arthrotic condition with slight osteophytic peripheral exostoses and an arthrolit (i.e. an articular calculus or "joint mouse") on the left condylus articularisand a depression in the medial part of the left mandibular condyle extending into the inferior part of the ramus are present. These features are indicative of a trauma-induced osteochondrosis dissecans. The diagnosis therefore suggests that the observed depression results from a well-healed fracture. This traumatic event illustrates the demanding living conditions endured by humans during the European Middle Pleistocene. The variations and pathological conditions observed in Mauer do not question the mandible's role as type specimen for the taxon Homo heidelbergensis.

Alveolar Process↗
PubMed2002

Substitutions in the conserved C2C domain of otoferlin cause DFNB9, a form of nonsyndromic autosomal recessive deafness.

DFNB, the nonsyndromic hearing loss with an autosomal recessive mode of inheritance constitutes the majority of severe to profound prelingual forms of hearing impairment, usually leading to inability of speech acquisition. We analyzed a consanguineous family with autosomal recessive deafness which has been shown to segregate within chromosomal region 2p23.1 (DFNB9; MIM 601071). By SSCP analysis and DNA sequencing of the 48 exons of the DFNB9 gene, coding for otoferlin, previously reported mutations in OTOF were excluded. Next to a frequent T > C single nucleotide polymorphism in exon 8, two novel mutations linked in exon 15 of the OTOF long splice form were identified comprising substitutions at positions 490 (Pro > Gln) and 515 (Ile > Thr), both located in the conserved Ca(2+) binding C2C domain of this peptide. Comparisons of homology using human and mice otoferlins and closely related peptides and computer simulation analyses suggest that changes in the mutated segment's secondary structure affect the Ca(2+) binding capacity of the C2C domain in otoferlin.

Alternative Splicing↗
PubMed2001

Quantitation of heteroplasmy in mitochondrial DNA mutations by primer extension using Vent(R)(exo-) DNA polymerase and RFLP analysis.

In this report we describe a simple and rapid protocol for reliable quantitation of mitochondrial DNA (mtDNA) mutations, which is basically a modification of the traditional polymerase chain reaction (PCR)/restriction fragment length polymorphism (RFLP) analysis technique. Up to now, the PCR/RFLP method has been of limited use for the accurate determination of ratios of mutant and wild type molecules, largely owing to the formation of heteroduplex molecules by PCR and incompleteness of restriction digestion. In order to overcome this problem, we have introduced a single-step primer extension reaction using Vent(R)(exo-) DNA polymerase and a fluorescence-labeled primer to the standard assay. The labeled homoduplex molecules are then digested with a restriction endonuclease, and the nucleic acids fractionated on an automated DNA sequencer equipped with GENESCAN analysis software. The amount of mutant mtDNA is readily estimated from fluorescence intensities of the wild-type and mutant mtDNA fragments corrected for incomplete digestion as monitored by a homologous control fragment. The accuracy of the improved protocol was determined by constructing standard curves obtained from defined mixtures of genomic DNA containing homoplasmic wild-type and mutant mtDNA. The expected values were obtained, with an observed correlation coefficient of 0.997 and a typical variability of +/-5% between repeated measurements. Further validation of the protocol is provided by the screening of five patients and unaffected subjects carrying the guanine to adenine transition at the nucleotide 3460 of the mitochondrial genome responsible for the mitochondrial disorder of Leber's hereditary optic neuropathy.

DNA Mutational Analysis↗
PubMed2001

Hominid skull fragments from Late Pleistocene layers in Leine Valley (Sarstedt, District of Hildesheim, Germany).

Three cranial fragments were recovered from coarse-grained deposits dug up by a suction dredge from gravel pits on the Leine river flats in the vicinity of Sarstedt (northwestern Germany). Also recovered were a number of artefacts which, upon careful inspection, could be assigned to the Middle Paleolithic. The geological pattern of the Leine Valley in this region suggests that these fragments were deposited in the lower terrace during a yet undetermined warm period-possibly Brörup or Odderade-during the Weichsel glaciation. However, attribution to the Eemian period or a Saale interstadial cannot be ruled out. The features of the Sarstedt (Sst) I infant temporal are known from Neanderthals (e.g., Weimar-Ehringsdorf, Engis, Krapina 1) and can be seen in specimens from the European late- Homo erectus group as well. Subadult individuals do not always exhibit full development of features characteristic for adults and-to some extent-anticipate the succeeding developmental stage (i.e., neoteny). The Neanderthal autapomorphies characterizing the fragments of the occipital and the parietal are certainly consistent with assigning both unequivocally to the species H. neanderthalensis. The presence of Middle Paleolithic artefacts recovered from the same deposits are commensurate with the presence of Neanderthals. However, there is no clear contextual association of any archaeological and fossil human material. Future DNA research will hopefully add up to the established morphological picture.

Animals↗
PubMed2001

Complete form of X-linked congenital stationary night blindness: refined mapping and evidence of genetic homogeneity.

A number of distinct, partly non-overlapping genetic loci have been reported for the complete type of X-linked congenital stationary night blindness (CSNB1), suggesting genetic heterogeneity. In order to refine the localization of the CSNB1 gene and to demonstrate genetic homogeneity, linkage analysis was performed in two large CSNB1 families. Clinical features consistent with the diagnosis of CSNB1 were documented in five patients from a German seven-generation kindred by full ophthalmological examination including psychophysical and electroretinographical testing. Haplotype analysis in 30 members of the large German family was performed with 38 polymorphic markers predominantly covering the critical region. Linkage analyses defined a locus for CSNB1 with flanking markers DXS8042 and DXS228, refining the interval to 2.5 cM in Xp11.4. In addition, two-point linkage analysis was carried out using the MLINK computer program. In agreement with meiotic breakpoints, lod scores of 3.0 and greater were obtained for markers located to the proximal site of the former 5 cM CSNB consensus interval. A large Dutch CSNB1 family was re-evaluated with markers from the Xp11.4 region, and supports the CSNB1 minimal interval found in the German family. Together with previous results from three unrelated families from Sweden, Sardinia and Great Britain, our results provide evidence of genetic homogeneity in the disorder. Subsequent mutation analyses in CSNB1 patients revealed no pathogenic sequence alterations in DFFRX and CASK genes, but retain candidates for other diseases mapping to that region.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed2001

Case populations must match the respective disease model: Genotype diversity causes linkage disequilibrium mapping failure in monogenic disorders.

Traditional linkage analysis in large families is the most promising approach for mapping disease genes of monogenic heritable disorders when the number of informative meioses is sufficient. With rare diseases, however, the low availability of informative pedigrees poses a significant limitation. As an adjunct to family linkage methods, association studies based on the investigation of individual haplotypes from a number of unrelated patients (i.e. linkage disequilibrium analysis) have recently been employed in mapping hereditary disease loci. However, such haplotype analysis is hampered by a number of effects that influence statistical evaluation, e.g. i) population history and size, ii) allele and haplotype frequencies in the respective population(s), iii) heterogeneous mutation and natural selection processes, and iv) small sample sizes of patient groups. The purpose of the present study was to determine the utility and limitations of haplotype-based genetic mapping in estimating the location of the NYX gene, which has recently been identified as the causative gene for a rare inherited retinal disorder known as the complete type of X-linked congenital stationary night blindness (CSNB1). For this purpose we recapitulated haplotypes and tested for linkage disequilibrium in 20 unrelated male CSNB1 patients from three European populations and 44 healthy individuals. All subjects were genotyped for 17 polymorphic microsatellite loci covering the Xp11.4 region with an average marker density of approximately 0.29 cM. We found that a precise model to describe mutations at loci that erroneously break up linkage is highly required, and that the case population must match the respective disease model.

Adolescent↗
PubMed2001

Segregation patterns and heteroplasmy prevalence in Leber's hereditary optic neuropathy.

PURPOSE: To investigate the segregation pattern of the mitochondrial DNA mutation at nucleotide position 3460 responsible for Leber's hereditary optic neuropathy (LHON) and to determine the prevalence of heteroplasmy for the three primary LHON mutations at positions 11778, 3460, and 14484. METHODS: Segregation analysis was performed in a cross-sectional study by determining the level of heteroplasmy in blood leukocytes of 23 LHON patients and unaffected carriers from four unrelated families. One family comprising two affected and three unaffected carriers was followed over 5.5 years for a longitudinal segregation analysis of heteroplasmy. The percentage of mutant mtDNA was determined using a novel procedure of fluorescence-based primer extension and restriction fragment length polymorphism analysis. The prevalence of heteroplasmy was assessed by determining the number of genealogically unrelated LHON pedigrees with heteroplasmic maternal family members from the LHON patient records of the Department of Ophthalmology, University of Tübingen, Germany. RESULTS: The authors observed a marked variability in the degree of heteroplasmy levels within each pedigree and a tendency toward a higher mutant allele frequency in offspring generations. Disease expression was correlated with higher levels of mutant mtDNA molecules. Longitudinal analysis revealed no statistically significant decrease in the heteroplasmy level in the family studied but a reduction of 11% and 12% in one affected and one unaffected individual, respectively. In 167 genealogically unrelated LHON families the prevalence of heteroplasmy was 5.6%, 40%, and 36.4% for the 11778, 3460, and 14484 LHON mutations, respectively. CONCLUSIONS: Cross-sectional studies of heteroplasmy for the 3460 LHON mutation suggest that the genotype shifts toward a higher mutational load in offspring generations. Long-term decrease in the blood mutant load in single cases indicates negative selection of the mutant allele in the hematopoietic cell system. The prevalence of heteroplasmy varies significantly between the different primary LHON mutations, suggesting genotypical differences in disease expression.

Chromosome Segregation↗
PubMed2001

Intrapopulational relationships in ancient societies: a multidisciplinary study.

Kinship determination is one of the major challenges for the anthropologist studying graveyard populations. Traditional techniques based on morphological comparisons of bone remains are limited. However, recent methods which generate and characterise DNA sequences derived from bones bear the possibility for a more accurate analysis. Extraction and characterisation of authentic nucleic acids was performed on a number of individuals from the early Medieval graveyard of Neresheim, South Germany. From this cemetery a total of 38 skeletal remains of individuals buried between 450 and 700 AD were examined using PCR-based methods. Comparisons were made using four human-specific short tandem repeat loci and the X/Y-specific amelogenin sex test. Twenty-eight of the approximately 1,500-year-old individuals yielded alleles in at least one of the polymorphic nuclear loci HumCD4, HumFES, HumTH01, HumVWA, and the sex test. These along with a 96 bp DNA variant previously unknown in recent CD4 contexts, and supporting evidence from anthropology and archaeology were used for defining one parental and one filial generation in each of three multiple burials (Ne 2, Ne 9 and Ne 78) in the cemetery.

Anthropology↗
PubMed2001

Slow and fast rod ERG pathways in patients with X-linked complete stationary night blindness carrying mutations in the NYX gene.

PURPOSE: To study the slow and fast rod signals of the scotopic 15-Hz flicker ERG in patients carrying mutations in the NYX gene, which has been recently identified as the cause of the complete form of congenital stationary night blindness, CSNB1. METHODS: Twenty eyes of 11 patients with CSNB1 who had nondetectable standard ERG rod b-waves were involved in the study. Scotopic ERG response amplitudes and phases to flicker intensities ranging from -3.37 to -0.57 log scotopic trolands. sec (scot td. sec) were measured at a flicker frequency of 15 Hz. ERG signals to flicker intensities between -3.37 and -1.97 and between -1.17 and -0.57 log scot td. sec were considered to represent primarily the slow and fast rod ERG pathway, respectively. Additionally, standard ERGs were performed. Twenty-two normal volunteers served as control subjects. RESULTS: For the slow rod ERG pathway, all patients exhibited ERG signals that were indistinguishable from noise. Accordingly, there was no systematic phase behavior for the slow rod signals. For the fast rod ERG pathway, the signals were significantly above noise, but they were significantly reduced in amplitude and advanced in phase. CONCLUSIONS: There is evidence that the slow and the fast rod ERG signals can be attributed to the rod bipolar-AII cell pathway and the rod-cone-coupling pathway, respectively. The current study provides evidence to suggest that a defective NYX gene product (nyctalopin) prevents detectable signal transmission through ON rod bipolar cells, but there is a residual transmission through rod-cone gap junctions in CSNB1, possibly through the OFF cone pathway.

Adolescent↗
PubMed2001

[Classification of a 300,000-year-old dental crown of the upper loamy deposit of the Bad Canstatter travertine zone].

Hominid dental remains were recovered in association with fossil bones and artifacts during systematic excavations in a loamy deposit located between the two travertine zones T4 and T5 at Stuttgart-Bad Cannstatt, Southwest Germany. Direct dating of a hominid tooth crown with thermoluminescence resulted in a date of 300 kya, which is in agreement with the Holstein Interglacial floral and faunal composition of this layer. The specimen is a lower left canine with hypoplastic morphology. This interpretation is supported by thorough assessment of its overall morphology, comparative metric evaluation, and by scanning electron microscopy analyses of the enamel prisms. Additional microstructural comparison of these dental remains with a tooth from the same site, but derived from a Cervidae specimen supported the distinct differences between both teeth. Here we discuss both the classification and significance of the specimen's evolutionary position as well as compare this specimen with stomatologic results from previous palaeopathological research.

Animals↗
PubMed2001

Large-scale analysis of sequence tags in Xp11.4-11.3 and evaluation of candidate genes for X-linked ocular diseases.

The gene-rich region of Xp11.4-Xp11.3 was characterized by increasing the physical marker density. Sequence tags (STSs) were generated by IRS- and DOP-PCR techniques, subsequent cloning, sequencing, and creation of primer pairs for single-copy sites. A total of 224 novel STSs were collected, providing an average marker density of 18 kb in the Xp11.4-Xp11.3 region which is assumed to be approximately 4 Mb in size. Sequence analysis of generated and established STSs via data base searches identified a novel gene highly homologous with the protein phosphatase 1 inhibitor 2 (IPP-2) and two pseudogenes; all of which map to the approximately 1.5 Mb proximal region of the critical region for X-linked congenital stationary night blindness type I (CSNB1) between markers DXS993 and DXS228. Using well-defined DNA panels, 69 STSs were fine-mapped to this approximately 1.5 Mb region, providing a marker coverage of one marker per 22 kb. No allelic loss was observed when the total STS content was applied to patient DNAs by PCR-mediated amplification. However, given the association of this region with a number of inherited ocular diseases, the data presented here provide valuable tools for genetic linkage and large-scale association studies.

Journal Article↗
PubMed2000

Genomic differentiation of Neanderthals and anatomically modern man allows a fossil-DNA-based classification of morphologically indistinguishable hominid bones.

Southern blot hybridizations of genomic DNA were introduced as a relatively simple fossil-DNA-based approach to classify remains of Neanderthals. When hybridized with genomic DNA of either human or Neanderthal origin, DNA extracted from two Neanderthal finds-the Os parietale, from Warendorf-Neuwarendorf, Germany, and a clavicula, from Krapina, Croatia-was shown to yield hybridization signals that differ by at least a factor of two compared to the signals obtained with the use of fossil DNA of an early Homo sapiens from the Vogelherd cave (Stetten I), Germany. When labeled chimpanzee DNA was used as a probe, Neanderthal and human DNA, however, revealed hybridization signals of similar intensity. Thus, the genome of Neanderthals is expected to differ significantly from the genome of anatomically modern man, because of the contrasting composition of repetitive DNA. These data support the hypothesis that Neanderthals were not ancestors of anatomically modern man.

Animals↗