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PubMed2005

Mitochondrial tRNA gene mutations in patients having mitochondrial disease with lactic acidosis.

Lactic acidosis has been associated with a variety of clinical conditions and can be due to mutation in nuclear or mitochondrial genes. We performed mutations screening of all mitochondrial tRNA genes in 44 patients who referred as hyperlactic acidosis. Patients showed heterogeneous phenotypes including Leigh disease in four, MELAS in six, unclassified mitochondrial myopathy in 10, cardiomyopathy in five, MERRF in one, pure lactic acidosis in six, and others in 12 including facio-scaplo-femoral muscular dystrophy (FSFD), familial cerebellar ataxia, recurrent Reye syndrome, cerebral palsy with mental retardation. We measured enzymatic activities of pyruvate dehydrogenase complex, and respiratory chain enzymes. All mitochondrial tRNA genes and known mutation of ATPase 6 were studied by single strand conformation polymorphism (SSCP), automated DNA sequence and PCR-RFLP methods. We have found one patient with PDHC deficiency and six patients with Complex I+IV deficiency, though the most of the patients showed subnormal to deficient state of respiratory chain enzyme activities. We have identified one of the nucleotide changes in 29 patients. Single nucleotide changes in mitochondrial tRNA genes are found in 27 patients and one in ATPase 6 gene in two patients. One of four pathogenic point mutations (A3243G, C3303T, A8348G, and T8993G) was identified in 12 patients who showed the phenotype of Leigh syndrome, MELAS, cardimyopathy and cerebral palsy with epilepsy. Seventeen patients have one of the normal polymorphisms in the mitochondrial tRNA gene reported before. SSCP and PCR-RFLP could detect the heteroplasmic condition when the percentage of mutant up to 5, however, it cannot be observed by direct sequencing method. It is important to screen the mtDNA mutation not only by direct sequence but also by PCR-RFLP and the other sensitive methods to detect the heroplasmy when lactic acidosis has been documented in the patients who are not fulfilled the criteria of mitochondrial disorders.

Acidosis, Lactic↗
PubMed2004

Autosomal-dominant Alport syndrome: natural history of a disease due to COL4A3 or COL4A4 gene.

BACKGROUND: Alport syndrome is a clinically and genetically heterogeneous nephropathy. The majority of cases are transmitted as an X-linked semidominant condition due to COL4A5 mutations. In this form males are more severely affected than females. Less than 10% of cases are autosomal recessive due to mutation in either COL4A3 or COL4A4. In this rarer form, both males and females are severely affected. Only two cases of autosomal-dominant Alport syndrome have been reported, one due to a COL4A3 mutation and the other due to a COL4A4 mutation. Because of the paucity of the reported families, the natural history of autosomal-dominant Alport syndrome is mostly unknown. METHODS: Four families with likely autosomal-dominant Alport syndrome were investigated. COL4A3 and COL4A4 genes were analyzed by denaturing high-performance liquid chromatography (HPLC). Automated sequencing was performed to identify the underlying mutation. RESULTS: Two families had a mutation in the COL4A4 gene and two in the COL4A3. Accurate clinical evaluation of family members showed interesting results. Affected individuals (22 persons) had a wide range of phenotypes from end-stage renal disease (ESRD) in the fifth decade to a nonprogressive isolated microhematuria. Finally, three heterozygous individuals (90, 22 and 11 years old, respectively) were completely asymptomatic. CONCLUSION: This paper demonstrated that patients affected by autosomal-dominant Alport syndrome have a high clinical variability. Moreover, a reduced penetrance of about 90% (3 of 25) may be considered for the assessment of recurrence risk during genetic counseling of these families.

Adult↗
PubMed2002

Use of the Cell-Dyn 3500 to predict leukemic cell lineage in peripheral blood of dogs and cats.

BACKGROUND: Morphology and cytochemistry are the foundation for classification of leukemias in dogs and cats. Advances in automated hematology instrumentation, immunophenotyping, cytogenetics, and molecular biology are significantly improving our ability to recognize and classify spontaneous myeloproliferative and lymphoproliferative disorders. OBJECTIVE: The purpose of this study was to assess the utility of flow cytometry-based light scatter patterns provided by the Cell-Dyn 3500 (CD3500) automated hematology analyzer to predict the lineage of leukemic cells in peripheral blood of dogs and cats. METHODS: Leukemic cells from 15 dogs and 6 cats were provisionally classified using an algorithm based on the CD3500 CBC output data and were subsequently phenotyped by enzyme cytochemistry, immunocytochemistry, indirect flow cytometry, and analysis of antigen receptor gene rearrangement. RESULTS: The algorithm led to correct predictions regarding the ontogeny of the leukemic cells (erythroid/megakaryocytic potential, myeloid leukemia, monocytic leukemia, chronic granulocytic leukemia, lymphoid leukemia) in 19/21 animals. Mismatches in the WBC impedance count and the WBC optical count in conjunction with microscopic assessment of blasts in the blood were useful for predicting myeloproliferative disorders with erythroid or megakaryocytic potential. The leukocyte light scatter patterns enabled distinction among myeloid leukemias (represented by acute myelomonocytic leukemia, acute monocytic leukemia, chronic granulocytic leukemia) and lymphocytic leukemias (including acute and chronic lymphocytic leukemias). One case of acute lymphocytic leukemia was misidentified as chronic lymphocytic leukemia. CONCLUSIONS: Algorithmic analyses can be applied to data generated by the CD3500 to predict the ontogeny of leukemic cells in the peripheral blood of dogs and cats. This rapid and quantitative technique may be used to improve diagnostic decisions, expand therapeutic choices, and increase prognostic accuracy.

Algorithms↗
PubMed2001

Mutations in the ELA2 gene encoding neutrophil elastase are present in most patients with sporadic severe congenital neutropenia but only in some patients with the familial form of the disease.

Severe congenital neutropenia (SCN) was originally described as an autosomal recessive disorder. Subsequently, autosomal dominant and sporadic forms of the disease have been recognized. All forms are manifest by persistent severe neutropenia and recurrent bacterial infection. In contrast, cyclical hematopoiesis is characterized by periodic neutropenia inter-spaced with (near) normal neutrophil counts. Recently, linkage analysis on 13 affected pedigrees identified chromosome 19p13.3 as the likely position for mutations in cyclical hematopoiesis. Heterozygous mutations in the ELA2 gene encoding neutrophil elastase were detected in all families studied. Further work also demonstrated mutations in ELA2 in sporadic and autosomal dominant SCN. However, all mutations described to date are heterozygous and thus appear to act in a dominant fashion, which is inconsistent with an autosomal recessive disease. Therefore, the current study investigated whether mutations in ELA2 could account for the disease phenotype in classical autosomal recessive SCN and in the sporadic and autosomal dominant types. All 5 exons of ELA2 and their flanking introns were studied in 18 patients (3 autosomal recessive, 5 autosomal dominant [from 3 kindreds], and 10 sporadic) using direct automated sequencing. No mutations were found in the autosomal recessive families. A point mutation was identified in 1 of 3 autosomal dominant families, and a base substitution was identified in 8 of 10 patients with the sporadic form, though 1 was subsequently shown to be a low-frequency polymorphism. These results suggest that mutations in ELA2 are not responsible for classical autosomal recessive Kostmann syndrome but provide further evidence for the role of ELA2 in SCN.

Adult↗
PubMed2002

Fabry disease: twenty novel alpha-galactosidase A mutations and genotype-phenotype correlations in classical and variant phenotypes.

BACKGROUND: Fabry disease (OMIM 301500) is an X-linked inborn error of glycosphingolipid metabolism resulting from mutations in the alpha-galactosidase A (alpha-Gal A) gene. The disease is phenotypically heterogeneous with classic and variant phenotypes. To assess the molecular heterogeneity, define genotype/phenotype correlations, and for precise carrier identification, the nature of the molecular lesions in the alpha-Gal A gene was determined in 40 unrelated families with Fabry disease. MATERIALS AND METHODS: Genomic DNA was isolated from affected males or obligate carrier females and the entire alpha-Gal A coding region and flanking sequences were amplified by PCR and analyzed by automated sequencing. Haplotype analyses were performed with polymorphisms within and flanking the alpha-Gal A gene. RESULTS: Twenty new mutations were identified (G43R, R49G, M72I, G138E, W236X, L243F, W245X, S247C, D266E, W287C, S297C, N355K, E358G, P409S, g1237del15, g10274insG, g10679insG, g10702delA, g11018insA, g11185-delT), each in a single family. In the remaining 20 Fabry families, 18 previously reported mutations were detected (R49P, D92N, C94Y, R112C [two families], F113S, W162X, G183D, R220X, R227X, R227Q, Q250X, R301X, R301Q, G328R, R342Q, E358K, P409A, g10208delAA [two families]). Haplotype analyses indicated that the families with the R112C or g10208delAA mutations were not related. The proband with the D266E lesion had a severe classic phenotype, having developed renal failure at 15 years. In contrast, the patient with the S247C mutation had a variant phenotype, lacking the classic manifestations and having mild renal involvement at 64 years. CONCLUSIONS: These results further define the heterogeneity of alpha-Gal A mutations causing Fabry disease, permit precise heterozygote detection and prenatal diagnosis in these families, and provide additional genotype/phenotype correlations in this lysosomal storage disease.

Fabry Disease↗
PubMed2004

Pharmacogenomic genotyping methodologies.

"Personalized medicine" based on an individual's genetic makeup is slowly becoming a reality as pharmacogenomics moves from the research setting to the clinical laboratory. Concordance studies between genotype and phenotype have shown that inherited mutations in several key drug-metabolizing enzymes, such as cytochrome P450 ( CYP ) 2D6 , 2C9 , and 2C19 , result in several distinct phenotypes that lead to different individual responses following drug administration. One of the major driving forces behind pharmacogenomics and its ability to be used effectively are the technologies that are available. A beneficial genotyping test must identify most or all of the mutations that have a significant impact on the expression or function of drug-metabolizing enzymes, transporter proteins, and/or drug receptors. Selection of the appropriate technology will be based on several issues, including prior knowledge of the mutation/polymorphism, sensitivity/specificity, sample requirements, and cost. Since the future volume of pharmacogenomic testing is anticipated to be large, automation of pharmacogenomics will also become increasingly important. This paper provides an overview of current technologies available for assessing polymorphisms on a small- to large-scale basis.

Alleles↗
PubMed1998

A novel mutation in the GLC1A gene causes juvenile open-angle glaucoma in 4 families from the Italian region of Puglia.

BACKGROUND: Primary open-angle glaucoma encompasses a complex of potentially blinding ocular diseases characterized by a normal-appearing angle of the anterior chamber, a characteristic degeneration of the optic nerve with resultant typical visual field defects, and usually, an elevated intraocular pressure. It can be subdivided into 2 groups according to the age at onset: the more prevalent chronic open-angle glaucoma diagnosed after 40 years of age, and the less common juvenile form, which occurs between 3 years of age and early adulthood. A locus for primary open-angle glaucoma (GLC1A) has been mapped to a 3-centimorgan region of the long arm of chromosome 1 (1q23-25). Recently, the myocilin (MYOC) gene, located in this chromosomal interval, has been found mutated in several patients affected by primary open-angle glaucoma. OBJECTIVE: To describe the clinical and molecular genetic features of 4 pedigrees affected by autosomal dominant juvenile open-angle glaucoma, all from the Italian region of Puglia. METHODS: Clinical study, gonioscopy, automated perimetry, and DNA analysis were performed on several members of the 4 families. RESULTS: We identified a new molecular defect (1177GACA-->T) in the third exon of the GLC1A gene. This mutation is present in all affected persons and in 2 still phenotypically normal persons. CONCLUSION: Our results are important for diagnostic purposes because it is now possible to identify asymptomatic carriers, for whom clinical surveillance for the early detection and treatment of glaucoma may be suggested.

Adolescent↗
PubMed2002

Candida dubliniensis, a new fungal pathogen.

There is a high interest in Candida species other than Candida albicans because of the rise and the epidemiological shifts in candidiasis. These emerging Candida species are favored by the increase of immunocompromised patients and the use of new medical practices, and m. Most oropharyngeal candidiasis can be foundare observed in those HIV-infected patients infected with human immunodeficiency virus (HIV). Candida dubliniensis is a recently described opportunistic pathogen that is closely related to C. albicans but differs from it with respect to epidemiology, certain virulence characteristics, and the ability to develop fluconazole resistance in vitro. C. dubliniensis has been linked to oral candidiasis in AIDS patients, although it has recently been associated to invasive disease. C. dubliniensis shares diagnostic characteristics with C. albicans, as germ tube- and chlamydospore-production, and it is generally misclassified as C. albicans by standard diagnostic procedures. Several recent studies have attempted to elucidate useful phenotypic and genotypic characteristics for separating both species. A large variety of methods have been developed with the aim of facilitating rapid and, accurate identification of this species. These have included differential chromogenic isolation platesculture media, direct immunological tests, and enhanced manual and automated biochemical and enzymatic panels. Chromogenic isolation media, as CHROMagar Candida, demonstrate better detection rates than traditional media, and allow the presumptive identification of C. dubliniensis by means of colony color (dark-green colonies). API 20 C AUX system is considered a reference method, but ID 32 C strip, the VITEK Yeast Biochemical Card and the VITEK 2 ID-YST system correctly identify most C. dubliniensis isolates, being the latter the most accurate. Spectroscopic methods, such as Fourier transformed-infrared spectroscopy, offer potential advantages. However, many authors consider that standard methods for differentiation of Candida species are time-consuming, often insensitive and can fail to distinguish C. dubliniensis. To overcome these low sensitivity, poor specificity and intolerable delay,drawbacks, molecular tools have been developed to discriminate C. dubliniensis, and particularly those based on the polymerase chain reaction. But, molecular tools prove difficult and too complex for routine use in the clinical laboratory setting and new developments are necessary. Moreover, an increased resistance to antifungal drugs has been described. Although preliminary studies indicate that most strains of C. dubliniensis are susceptible to antifungal agents, fluconazole-resistant strains have been detected. Furthermore, fluconazole-resistant strains are easily derived in vitro, showing an increased expression of multidrug resistance transporters, as MDR1.

Antifungal Agents↗
PubMed1994

DNA typing of grapevines: a universal methodology and database for describing cultivars and evaluating genetic relatedness.

With established ampelographic techniques for grapevine identification it is often difficult to achieve a satisfactory, objective result. We have developed a DNA typing system using sequence-tagged microsatellite site markers as a means of differentiating cultivars of grapevine. A semi-automated analysis procedure was linked to an electronic database and found to be an objective and reliable system for cultivar identification using this simple marker type. The accumulated DNA typing data from over eighty cultivars demonstrated that cultivars that are difficult to differentiate phenotypically using ampelographic techniques can be distinguished by DNA typing. Parentage analysis uncovered errors in parent assignment or cultivar identification in specific cases. The electronic database has a conservative format to take into account the occurrence of null alleles and the possibility of missed alleles. Computer-assisted comparisons of cultivars in the database can be performed and various approaches for estimating the match probability that two unrelated cultivars have the same genotype simply due to chance are discussed. We suggest that further development of the database through international co-operation using standardised sequence-tagged site markers offers the possibility of achieving a universal grapevine identification system.

DNA, Plant↗
PubMed2006

In vitro activity of beta-lactam and non-beta-lactam antibiotics in extended-spectrum beta-lactamase-producing clinical isolates of Escherichia coli.

The activity of different beta-lactam and non-beta-lactam antibiotics was assessed against extended-spectrum beta-lactamase (ESBL)-producing and non-ESBL-producing clinical isolates of Escherichia coli. A phenotypic study to discover the presence of ESBLs in 399 clinical isolates of E. coli was made by the disk diffusion method following the Clinical and Laboratory Standards Institute (formely NCCLS, 2004) guidelines. The activity of different antibiotics was subsequently studied using the automated VITEK 2 system (bioMérieux, Marcy l'Etoile, France). One hundred fifteen isolates proved to be ESBL-producing and 284 non-ESBL-producing. Among the former, percentage susceptibilities to the antibiotics assayed were meropenem and amikacin, 100%; piperacillin/tazobactam, 97.4%; cefepime, 94.8%; amoxicillin/clavulanic acid, 84.3%; tobramycin, 84.3%; gentamicin, 83.5%; cefoxitin, 83.5%; nitrofurantoin, 71.3%; cotrimoxazole, 46.1%; norfloxacin, 29.6%; ciprofloxacin, 27%; and ofloxacin, 26.1%.

Anti-Bacterial Agents↗
PubMed2001

Newborn screening for congenital adrenal hyperplasia.

Classic CAH (salt-wasting and simple virilizing) meets all of the recommended criteria for newborn screening. There are reliable and efficient newborn screening tests, the disorder results in high morbidity and mortality if left undetected, there is effective treatment that reduces negative outcomes, and there is a relatively high incidence. When compared with the case findings without the benefit of screening, the data from screening programs show reduced adrenal crises, reduced incorrect sex assignments, and reduced deaths. Racial/ethnic prevalence differences are present in newborn screening program data. The Texas data indicate a lower disease frequency in African-Americans when compared with Caucasians, and international data indicate higher frequencies in native Yupik Eskimos, Brazilians, residents of La Reunion, and Filipinos. When worldwide clinical ascertainment data are compared with newborn screenng data, it is clear that newborns with CAH (especially males) die when screening is not done. To be effective in reducing mortality, newborn screening must be performed soon after birth, and the results must be available quickly so that early salt-wasting crises can be averted. It is preferable that newborn screening laboratiories be operational 7 days a week, and that sample delivery from the collection site to testing laboratory be as efficient as possible, including weekends and holidays, so that undue testing delays are not encountered. These two requirements pose major challenges for most programs, but they are critical to optimal screening outcome. Based on the studies in Texas, with second screening samples collected at approximately 2 weeks of age, some newborns with simple virilizing CAH are missed on initial screening using current testing protocols. There is need to set a screening cut-off such that the false-positive rate does not oversaturate the follow-up system, in part owing to the insensitivity of current kit methodologies and the biochemical manifestations of CAH. With advances in genetic testing procedures and improved automation techniques, it may soon be possible for CAH screening programs to include genotyping as a second-tier confirmation as a part of the newborn screening protocol. Despite the fact that CAH is a continuum of disorders, the correlation between genotype and phenotype is fairly consistent in most cases. For the purpose of screening, genotyping will likely be useful only for differential diagnoses of non-salt wasters, given the necessary time constraints and expense of obtaining genotypes and the need for immediate diagnosis/treatment of salt wasters. It is hoped that newborn screening programs will begin to provide answers to some of these question in addition to their primary function of reducing the morbidity and mortality resulting from CAH.

17-alpha-Hydroxyprogesterone↗
PubMed2002

Development of a closed-system process for clinical-scale generation of DCs: evaluation of two monocyte-enrichment methods and two culture containers.

BACKGROUND: Clinical immunotherapy trials using DCs depend on large-scale methods for DC generation that fulfil current good manufacturing practice requirements. Our goal was to develop data on two variables, monocyte-enrichment method and culture container, which could be used to design a closed-system process for ex vivo generation of immature DCs. METHODS: Mononuclear cells were collected by leukapheresis and enriched for monocytes by either counterflow centrifugal elutriation, or immunomagnetic selection using Isolex, an automated closed-system device. Monocytes were cultured for 7 days in serum-free medium with GM-CSF and IL-4, using either plastic flasks or gas-permeable Stericell bags. Monocytes and cultured DCs were evaluated for yield, flow cytometric phenotype, and in vitro function in MLR, and autologous recall responses to tetanus toxoid and influenza virus. RESULTS: Enriched monocyte products from elutriation and immunomagnetic selection were equivalent in yield and purity, and were capable of generating immature DCs in either flasks or bags. DCs from all four culture conditions were equivalent in yield, phenotype, and in vitro function. Mean DC yield was 67-80% per seeding monocyte, and 11-13% per starting mononuclear cell (MNC). A leukapheresis product containing 5 x 10(9) MNCs processed by this method could therefore yield approximately 5 x 10(8) immature DCs. DISCUSSION: In this manufacturing process, the Isolex system was equivalent to elutriation, and Stericell bags were equivalent to flasks. Together, the Isolex system and Stericell bags can be incorporated into a closed-system process to generate immature DCs.

Adoptive Transfer↗
PubMed2004

Screening for presenilin inhibitors using the free-living nematode, Caenorhabditis elegans.

Caenorhabditis elegans contains 3 homologs of presenilin genes that are associated with Alzheimer s disease. Loss-of-function mutations in C. elegans genes cause a defect in egg laying. In humans, loss of presenilin-1 (PS1) function reduces amyloid-beta peptide processing from the amyloid protein precursor. Worms were screened for compounds that block egg laying, phenocopying presenilin loss of function. To accommodate even relatively high throughput screening, a semi-automated method to quantify egg laying was devised by measuring the chitinase released into the culture medium. Chitinase is released by hatching eggs, but little is shed into the medium from the body cavity of a hermaphrodite with an egg laying deficient (egl) phenotype. Assay validation involved measuring chitinase release from wild-type C. elegans (N2 strain), sel-12 presenilin loss-of-function mutants, and 2 strains of C. elegans with mutations in the egl-36 K(+) channel gene. Failure to find specific presenilin inhibitors in this collection likely reflects the small number of compounds tested, rather than a flaw in screening strategy. Absent defined biochemical pathways for presenilin, this screening method, which takes advantage of the genetic system available in C. elegans and its historical use for anthelminthic screening, permits an entry into mechanism-based discovery of drugs for Alzheimer's disease.

Animals↗
PubMed2006

[MECP2 gene mutations in twenty-six cases with atypical Rett syndrome].

OBJECTIVE: Rett syndrome (RTT) is an X-linked progressive neurodeveopmental disorder that almost exclusively affects girls, and is one of the most common causes of mental retardation in females, with an estimated prevalence of approximately 1 in 10,000 - 15,000 female individuals. Mutations in X-linked methyl-CpG-binding protein 2 (MECP2) gene, located on chromosome Xq28, have been found to be a cause of RS. A lot of mutations have been reported to be related to RS recently. Mutations are found in 70% - 85% of patients with classical RTT and in less than 50% of patients with atypical RS. Up to now, RTT is diagnosed based on a consistent counseling for clinical features and the established diagnostic criteria. The present study aimed to investigate frequency and type of mutation of MECP2 gene and if hot spot of mutation exits in patients with atypical RTT and find out the relationship between genotype and phenotype. METHODS: A systematic analysis of the entire coding region of MECP2 in 26 unrelated patients with atypical RTT was performed by polymerase chain reaction (PCR) and direct sequencing. Genomic DNA was extracted using standard procedures from the peripheral blood leukocytes of each patient. PCR amplification products were checked by 2% agarose gel electrophoresis and were subsequently sequenced with ABI 3730 Automated DNA Sequencer with both the forward and reverse primers. Mutational analyses were performed using normal human genomic MECP2 sequence as a reference (GenBank accession NO.AF030876). RESULTS: Seven mutations were identified in 12 of 26 patients. Most of the mutations were missense mutation; c.397C > T (R133C) was found in 3 of 26 patients; c.473C > T (T158M) and c.916C > T (R306C) were found in 2 of 26 patients, respectively; c.397A > G (R133H) and c.1005G > A (R335C) were found in 1 of 26 patients, respectively. One base pair deletion mutation (806delG) resulting in frameshift was found in 2 of 26 patients, and 1 base pair transversion at splice accept-site (IVS3-2A > T). CONCLUSION: The results of this study indicated that c.397C > T (R133C), c.473C > T (T158M) and c.916C > T (R306C) were hot spot mutations in MECP2 gene of patients with atypical RTT. There was some relationship between genotype and phenotype.

Base Sequence↗
PubMed1999

Testing for tumor drug resistance in the age of molecular medicine. A contribution to the Debate Round-Table on Phenotypic and Genotypic Analyses of Multidrug Resistance (MDR) in Clinical Hospital Practice.

The detection of multidrug resistance (MDR) in clinical hospital practice represents an important strategy to combat clinical tumor drug resistance. Predicting the response of tumors to cytostatic drugs is of prognostic value. The 'Debate Round-Table on Phenotypic and Genotypic Analyses of Multidrug Resistance (MDR) in Clinical Hospital Practice' was launched in 1997 to address specific questions on this topic. The results published thus far are a rich source to learn about the promises and pitfalls of methods, eg surrogate and functional tests and protein or mRNA expression assays as well. In the present paper, some requirements are discussed for applications of drug resistance testing in clinical routine diagnostics. To improve the detection of low-level resistance, established methodologies may be strengthened with respect to: (1) standardization of sample handling, antibodies, PCR primers, and detection reagents; (2) standardization of protocols and far reaching automation in performance and evaluation of results to ensure high quality control criteria. Sophisticated new techniques will feature: (1) high-throughput analyses for the 'horizontal screening' of single drug resistance genes in large numbers of patient samples at economically fair costs; (2) 'vertical screening' of a large number of resistance mechanisms operating upstream or downstream of the actual drug-target interaction sites, in order to detect more complex and multifaceted genotypes and phenotypes of multidrug resistance.

Drug Resistance, Multiple↗
PubMed2006

Mass-spectrometrical analysis of proteins encoded on chromosome 21 in human fetal brain.

Overexpression of chromosome 21 genes is directly or indirectly responsible for the Down syndrome phenotype. In order to analyse chromosome 21 gene products (Chr21Ps), we extracted proteins from fetal human brain cortex and applied an ultracentrifugal and chromatographic prefractionation principle followed by two-dimensional gel electrophoresis (2-DE) and mass-spectrometrical analysis using high-throughput automated MALDI-TOF/TOF. Nine Chr21Ps were identified: pyridoxal kinase; superoxide dismutase [Cu/Zn] 1; carbonyl reductase 1; ES1 protein homolog, mitochondrial [Precursor]; cystathionine-beta-synthetase; T-complex protein 1, theta subunit; cystatin B; 6-phosphofructokinase; glycinamide ribonucleotide synthetase. Mass-spectrometric characterisation of Chr21Ps following separation in 2-DE gels is a useful tool for the analysis of these structures in brain, independent of antibody availability and specificity.

Brain↗
PubMed2005

Genetic disposition to addictive disorders--current knowledge and future perspectives.

It is well established that addictive disorders have a strong genetic background. Multiple, and in part interacting, genes are likely to be responsible for the disease phenotype, making the search for underlying alleles a challenging and complicated task. Linkage analyses and association studies have failed to unequivocally identify underlying genes. Conversely, genome sequencing and the systematic search for polymorphic marker loci have yielded dense chromosome maps so that, along with automated genotyping, the identification of individual genes will soon become possible. Initial results provide hints that regulators of gene expression might play an important role in addiction.

Animals↗