Biomedical subjects

H M Müller

Publications and source records attributed to H M Müller.

At least 19 recordsLinked to original sources

PubMed2005

Gene methylation data--a new challenge for bioinformaticians?

OBJECTIVES: Changes in the status of DNA methylation, known as epigenetic alterations, are among the most common molecular alterations in human neoplasia. For the first time, we reported on the analysis of fecal DNA from patients with CRC to determine the feasibility, sensitivity and specificity of this approach. We want to present basic information about DNA methylation analysis in the context of bioinformatics, the study design and several statistical experiences with gene methylation data. Additionally we outline chances and new research questions in the field of DNA methylation. METHODS: We present current approaches to DNA methylation analysis based on one reference study. Its study design and the statistical analysis is reflected in the context of biomarker development. Finally we outline perspectives and research questions for statisticians and bioinformaticians. RESULTS: Identification of at least three genes as potential DNA methylation-based tumor marker genes (SFRP2, SFRP5, PGR). CONCLUSIONS: DNA methylation analysis is a rising topic in molecular genetics. Gene methylation will push the extension of biobanks to include new types of genetic data. Study design and statistical methods for the detection of methylation biomarkers must be improved. For the purpose of establishing methylation analysis as a new diagnostic/prognostic tool the adaptation of several approaches has become a challenging field of research activity.

Biomarkers, Tumor↗

PubMed2003

MeCP2 and MBD2 expression in human neoplastic and non-neoplastic breast tissue and its association with oestrogen receptor status.

This study analysed mRNA expression of two members of the methyl-CpG-binding protein family - MeCP2 and MBD2 - in human non-neoplastic (n=11) and neoplastic (n=57) breast tissue specimens using a quantitative real-time PCR method. We observed higher expression levels of MeCP2 mRNA in neoplastic tissues than in non-neoplastic tissues (P=0.001), whereas no significant differences for MBD2 were detected. When studying the relations between the most important clinicopathologic features of breast cancer and the mRNA expression level of both MBDs, we found that oestrogen receptor (OR)-positive breast cancer specimens contained higher levels of MeCP2 mRNA than did OR-negative cancers (P=0.005). Furthermore, we observed statistically significantly higher levels of MeCP2 in non-neoplastic tissues expressing high levels of OR as compared to those expressing low levels (P=0.017). Finally, using a linear regression model, we identified a statistically significant association between OR expression and MeCP2 mRNA expression in neoplastic and non-neoplastic breast tissue specimens (P=0.003). In conclusion, we were able to demonstrate for the first time that there exists a strong association between OR status and MeCP2 mRNA expression. Furthermore, we speculate that MeCP2, regulated by OR, plays a key role in the differentiation processes in human breast tissues.

Adult↗

PubMed2002

Fusarium toxins in wheat flour collected in an area in southwest Germany.

A total of 60 samples of wheat flour were collected during the first 6 months of 1999 from mills and food stores in an area in southwest Germany. Samples included whole-grain and two types of white flour with these three groups characterized by a high, medium and low ash content. The contents of deoxynivalenol (DON), nivalenol (NIV), 3- and 15-acetyldeoxynivalenol, HT-2 toxin (HT-2), T-2 toxin (T-2) and fusarenon-X (FUS-X) were determined by gas chromatography/mass spectrometry, and those of zearalenone (ZEA), alpha- and beta-zearalenol (alpha- and beta-ZOL) by high performance liquid chromatography with fluorescence detection. FUS-X, alpha- and beta-ZOL were not detected in any sample. Based on incidence and level, DON was the predominant toxin followed by NIV and ZEA for all three flour types. The overall degree of toxin contamination was lower with decreasing ash content. This suggests a localization of the toxins analyzed primarily in the outer parts of the original wheat kernels. The median DON content was significantly (P<0.05) higher for wheat flour originating from wheat of conventional than of organic production.

Chromatography, High Pressure Liquid↗

PubMed2001

Gambicin: a novel immune responsive antimicrobial peptide from the malaria vector Anopheles gambiae.

A novel mosquito antimicrobial peptide, gambicin, and the corresponding gene were isolated in parallel through differential display-PCR, an expressed sequence tag (EST) project, and characterization of an antimicrobial activity in a mosquito cell line by reverse-phase chromatography. The 616-bp gambicin ORF encodes an 81-residue protein that is processed and secreted as a 61-aa mature peptide containing eight cysteines engaged in four disulfide bridges. Gambicin lacks sequence homology with other known proteins. Like other Anopheles gambiae antimicrobial peptide genes, gambicin is induced by natural or experimental infection in the midgut, fatbody, and hemocyte-like cell lines. Within the midgut, gambicin is predominantly expressed in the anterior part. Both local and systemic gambicin expression is induced during early and late stages of natural malaria infection. In vitro experiments showed that the 6.8-kDa mature peptide can kill both Gram-positive and Gram-negative bacteria, has a morphogenic effect on a filamentous fungus, and is marginally lethal to Plasmodium berghei ookinetes. An oxidized form of gambicin isolated from the cell line medium was more active against bacteria than the nonoxidized form from the same medium.

Amino Acid Sequence↗

PubMed2001

Substitution of the thioredoxin system for glutathione reductase in Drosophila melanogaster.

The disulfide reducing enzymes glutathione reductase and thioredoxin reductase are highly conserved among bacteria, fungi, worms, and mammals. These proteins maintain intracellular redox homeostasis to protect the organism from oxidative damage. Here we demonstrate the absence of glutathione reductase in Drosophila melanogaster, identify a new type of thioredoxin reductase, and provide evidence that a thioredoxin system supports GSSG reduction. Our data suggest that antioxidant defense in Drosophila, and probably in related insects, differs fundamentally from that in other organisms.

Amino Acid Motifs↗

PubMed2001

Nuclear factors bind to a conserved DNA element that modulates transcription of Anopheles gambiae trypsin genes.

The Anopheles gambiae trypsin family consists of seven genes that are transcribed in the gut of female mosquitoes in a temporal coordinated and mutually exclusive manner, suggesting the involvement of a complex transcription regulatory mechanism. We identified a highly conserved 12-nucleotide motif present in all A. gambiae and Anopheles stephensi trypsin promoters. We investigated the role of this putative trypsin regulatory element (PTRE) in controlling the transcription of the trypsin genes. Gel shift experiments demonstrated that nuclear proteins of A. gambiae cell lines formed two distinct complexes with probes encompassing the PTRE sequence. Mapping of the binding sites revealed that one of the complex has the specificity of a GATA transcription factor. Promoter constructs containing mutations in the PTRE sequence that selectively abolished the binding of either one or both complexes exerted opposite effects on the transcriptional activity of trypsin promoters in A. gambiae and Aedes aegypti cell lines. In addition, the expression of a novel GATA gene was highly enriched in A. gambiae guts. Taken together our data prove that factors binding to the PTRE region are key regulatory elements possibly involved in the blood meal-induced repression and activation of transcription in early and late trypsin genes.

Amino Acid Sequence↗

PubMed2001

Innate immune defense against malaria infection in the mosquito.

Anopheles gambiae, the most important vector of malaria, employs its innate immune system in the fight against Plasmodium. This can affect the propagative capacity of Plasmodium in the vector and, in some cases, leads to total refractoriness to the parasite. The components operating in the mosquito's innate immune system and their potential relevance to antimalarial responses are being systematically dissected.

Animals↗

PubMed2001

Epigenetic downregulation of the retinoic acid receptor-beta2 gene in breast cancer.

A growing body of evidence supports the hypothesis that the retinoic acid receptor beta2 (RAR-beta2) gene is a tumor suppressor gene which induces apoptosis and that the chemopreventive and therapeutic effects of retinoids are due to induction of RAR-beta2. During breast cancer progression, RAR-beta2 is reduced or even lost. It is known from studies of other tumor-suppressor genes that methylation of the 5'-region is the cause of loss of expression. Several groups demonstrated that this is also true for the RAR-beta2 in breast cancer by treating breast cancer cell lines with a demethylating agent and examining expression of the RAR-beta2 gene in response to a challenge with retinoic acid. Studies using sodium bisulfite genomic sequencing as well as methylation specific PCR showed that a number of breast cancer cell lines as well as breast cancer tissue showed signs of methylation. The RAR-beta2 gene was unmethylated in non-neoplastic breast tissue as well as in other normal tissues. A combination of retinoic acid with demethylating agents as well as with histone deacetylase inhibitors acts synergistically to inhibit growth. This review presents data that suggest that treatment of cancer patients with demethylating agents followed by retinoic acid may offer a new therapeutic modality. Both the time of commencement of chemoprevention and the choice of substances that are able either to prevent de novo methylation or to reverse methylation-caused gene silencing may be important considerations.

Base Sequence↗

PubMed2001

Characteristics and clinical relevance of chronic anemia in adult heart transplant recipients.

BACKGROUND: Mild chronic anemia following heart transplantation (HTX), with hemoglobin (Hb) values of 10-14 g/dL in men and 10-12 g/dL in women, is frequent. It has continued to be of uncertain etiology yet clinical relevance. Nonetheless, therapeutic immunosuppression has been regarded as a major cause of chronic anemia in HTX patients. METHODS: Sixty outpatients were observed over a period of 5 yr after HTX. Laboratory values related to anemia such as Hb, erythropoietin (EPO), ferritin, transferrin, iron, and vitamin levels were obtained and analyzed monthly. Patients were divided into two groups retrospectively. Patients with persistent anemia for more than 1 yr were compared with non-anemic patients. RESULTS: Forty-three (72%) of the 60 patients were anemic. Anemia was normochromic, normocytic, and slightly anisocytic. Anemic and non-anemic patients showed EPO levels within the expected range as defined by Erslev (Erythropoietin. N Engl J Med 1991: 324: 1339). Reticulocyte counts were found to be normal in all patients. Iron deficiency and deficiency of vitamin B12 or folic acid were not observed. Patients with persistent anemia showed a significantly shorter survival period than non-anemic patients (p<0.02). CONCLUSIONS: Mild anemia following HTX shows the same characteristics as anemia in chronic diseases. Persisting mild anemia used to be associated with a shorter life expectancy. There is no evidence that standard immunosuppression causes anemia.

Anemia↗

PubMed2001

Blood digestion in the malaria mosquito Anopheles gambiae: molecular cloning and biochemical characterization of two inducible chymotrypsins.

The elucidation of digestive processes in the Anopheles gambiae gut leading to the utilization of the blood meal will result in a deeper understanding of the physiology of blood digestion and its impact on parasite-vector interactions. Accordingly, the identification of digestive serine proteases in A. gambiae has implications for the development of alternative strategies for the control of mosquito-borne diseases. We report here on the cDNA and genomic cloning and on the expression analysis of two closely related chymotrypsin genes, Anchym1 and Anchym2. Genomic cloning revealed that Anchym1 and Anchym2, which map on chromosomal division 25D, are clustered in tandem within 6 kb, both genes being interrupted by two short introns. After blood feeding, transcription of Anchym1 and Anchym2 is induced in the midgut epithelium, followed by secretion of the translated products into the midgut lumen where the Anchym1 and Anchym2 zymogens are activated by partial tryptic digestion. The amino-acid residues forming the substrate pocket of Anchym1 and Anchym2 suggested chymotryptic cleavage specificity. This was confirmed by mass spectrometry analysis and Edman degradation sequencing of proteolytic products generated by the recombinant, trypsin-activated Anchym1.

Amino Acid Sequence↗

PubMed2001

Further survey of the occurrence of Fusarium toxins in wheat grown in southwest Germany.

A total of 53, 54, 57, 52 and 60 wheat samples for feed use were collected randomly after the 1989, 1990, 1991, 1992 and 1993 crops, respectively, from farms in an area of southwest Germany. Deoxynivalenol (DON), 3- and 15-acetyldeoxynivalenol (3-, 15-ADON), nivalenol (NIV), HT-2 toxin (HT-2), T-2 toxin (T-2), diacetoxyscirpenol (DAS), and fusarenon-X (FUS-X) were determined by gas chromatography, combined with mass selective detection (GC-MS), zearalenone (ZEA), alpha- and beta-zearalenol (alpha-beta-ZOL) were determined by HPLC. DON was the major toxin, with incidences at 77 to 93% and mean contents at 167 to 735 micrograms/kg. In contrast, incidences of ZEA, 3-ADON, NIV, HT-2, and T-2 were at 13 to 37%, 10 to 44%, 15 to 67%, 0 to 11%, and 0 to 12%, respectively, with mean contents in positive samples between 2 and 73 micrograms/kg, except for 948 micrograms/kg 3-ADON in samples from 1993. 15-ADON and FUS-X were assayed in samples from 1991, 1992 and 1993. 15-ADON was found in 0 to 11% of samples at mean levels < or = 17 mu/kg, DAS, alpha- and beta-ZOL, and FUS-X were not detected in any sample. Over the years, incidences and levels of toxins remained constant, decreased or increased, with most differences between years being slight and insignificant. The risk for livestock due to DON, HT-2 and ZEA was estimated based on maximum tolerated levels recommended for these toxins in some countries.

Animal Feed↗

PubMed2001

Effect of feeding regime on the metabolism of ochratoxin A during the in vitro incubation in buffered rumen fluid from cows.

Pure ochratoxin A (OA) was added to buffered rumen fluid collected from two fistulated cows and incubated under anaerobic conditions. Both animals were fed six diets containing grass, grass silage or hay, and two different amounts of concentrate consisting of barley and soybean meal. Four incubations per animal and diet were carried out at consecutive days. The concentration of OA declined exponentially to a very low or non-detectable level under all conditions examined, with half-lives at 0.51 to 2.76 h. The disappearance of OA was accompanied by the appearance of ochratoxin alpha (O alpha) with an average amount of O alpha formed relative to the disappearance of OA near 100%, independent of diet and animal. Based on four incubations per animal and diet the rate of OA disappearance was affected (P < 0.1) by the origin of rumen fluid from two animals, as well as by the type of basic component and amount of concentrate in the diet, with interactions between these factors. The disappearance of OA mostly was accelerated (P < 0.1) by replacing grass silage or hay by fresh grass and by increasing the content of concentrate from 10 to 50% of dry matter. It is concluded that the capacity of the rumen to detoxify OA is not limited by the yield of Oa from OA but is strongly dependent on animal and diet.

Animal Feed↗

PubMed2000

Theta synchronization predicts efficient memory encoding of concrete and abstract nouns.

Functional and topographical differences between processing of spoken nouns which were remembered or which were forgotten were shown by means of EEG coherence analysis. Later recalled nouns were related with increased neuronal synchronization (= cooperation) between anterior and posterior brain regions regardless of presented word category (either concrete or abstract nouns). However, theta coherence exhibited topographical differences during encoding of concrete and abstract nouns whereby former were related with higher short-range (mainly intrahemispheric), later with higher long-range (mainly interhemispheric) coherence. Thus, theta synchronization possibly is a general phenomenon always occurring if task demand increases and more efficient information processing is required. Measurement of EEG coherence yields new information about the neuronal interaction of involved brain regions during memory encoding of different word classes.

Adult↗

PubMed2000

A modular chitin-binding protease associated with hemocytes and hemolymph in the mosquito Anopheles gambiae.

Sp22D, a modular serine protease encompassing chitin binding, low density lipoprotein receptor, and scavenger receptor cysteine-rich domains, was identified by molecular cloning in the malaria vector, Anopheles gambiae. It is expressed in multiple body parts and during much of development, most intensely in hemocytes. The protein appears to be posttranslationally modified. Its integral, putatively glycosylated form is secreted in the hemolymph, whereas a smaller form potentially generated by proteolytic processing is associated with the tissues. Bacterial challenge or wounding result in low-level RNA induction, but the protein does not bind to bacteria, nor is its processing affected by infection. However, Sp22D binds to chitin with high affinity and undergoes transient changes in processing during pupal to adult metamorphosis; it may respond to exposure to naked chitin during tissue remodeling or damage.

Animals↗

PubMed2000

Methylation and silencing of the retinoic acid receptor-beta2 gene in breast cancer.

BACKGROUND: A growing body of evidence supports the hypotheses that the retinoic acid receptor beta2 (RAR-beta2) gene is a tumor suppressor gene and that the chemopreventive effects of retinoids are due to induction of RAR-beta2. RAR-beta2 expression is reduced in many malignant tumors, and we examined whether methylation of RAR-beta2 could be responsible for this silencing. METHODS: RAR-beta2 expression was studied by reverse transcription-polymerase chain reaction (RT-PCR) analysis in eight breast cancer cell lines that were either treated with the demethylating agent 5-aza-2'-deoxycytidine and subsequently with all-trans-retinoic acid (ATRA) or left untreated. Sodium bisulfite genomic sequencing was used to determine the locations of 5-methylcytosines in the RAR-beta2 genes of three of these cell lines. In 16 breast cancer biopsy specimens and non-neoplastic breast tissue, methylation-specific PCR was used to determine the methylation status of RAR-beta2, and, in 13 of the specimens, RT-PCR analysis was used to detect RAR-beta2 expression. RESULTS: Cell lines SK-BR-3, T-47D, ZR-75-1, and MCF7 exhibited expression of RAR-beta2 only after demethylation and treatment with ATRA. The first exon expressed in the RAR-beta2 transcript was methylated in cell lines ZR-75-1 and SK-BR-3. Six breast cancer specimens showed methylation in the same region of the gene. No expression of RAR-beta2 was found in any grade III lesion. An inverse association between methylation and gene expression was found in all grade II lesions. The RAR-beta2 gene from non-neoplastic breast tissue was unmethylated and expressed. CONCLUSIONS: Methylation of the RAR-beta2 gene may be an initial step in breast carcinogenesis; treatment of cancer patients with demethylating agents followed by retinoic acid may offer a new therapeutic modality.

Base Sequence↗

PubMed2000

Detection of cattle-derived BSE prions using transgenic mice overexpressing bovine PrP(C).

In interspecies transmissions of transmissible spongiform encephalopathies, the agent has to overcome a species barrier that is largely influenced by the rate of homology between the prion proteins (PrP(C)) of the two involved species. Generating transgenic mice expressing PrP(C) of a foreign species is an approach to develop TSE models that are at least partly devoid of a species barrier. The availability of such animals would enable the detection of low doses of infectivity in tissues or bodily fluids derived from other species. We generated transgenic mice that overexpress bovine PrP(C) (Tgbov XV mice) for the development of an improved detection assay for cattle derived BSE prions. These mice succumbed to the disease 250 days after inoculation with a brain homogenate from BSE diseased cattle. Diagnosis of BSE in transgenic mice was confirmed using Western blot, as well as histological and immunohistochemical methods. In contrast, transgenic mice overexpressing murine PrP(C) (tga20 mice) did not display shorter incubation times than nontransgenic RIII mice infected with the same inoculum. The expression of a chimaeric PrP of murine and bovine sequences rendered such mice partly, if not entirely resistant to an infection with the BSE agent.

Animals↗

PubMed2000

Cloning and analysis of a cecropin gene from the malaria vector mosquito, Anopheles gambiae.

Parasites of the genus Plasmodium are transmitted to mammalian hosts by anopheline mosquitoes. Within the insect vector, parasite growth and development are potentially limited by antimicrobial defence molecules. Here, we describe the isolation of cDNA and genomic clones encoding a cecropin antibacterial peptide from the malaria vector mosquito Anopheles gambiae. The locus was mapped to polytene division 1C of the X chromosome. Cecropin RNA was induced by infection with bacteria and Plasmodium. RNA levels varied in different body parts of the adult mosquito. During development, cecropin expression was limited to the early pupal stage. The peptide was purified from both adult mosquitoes and cell culture supernatants. Anopheles gambiae synthetic cecropins displayed activity against Gram-negative and Gram-positive bacteria, filamentous fungi and yeasts.

Amino Acid Sequence↗

PubMed2000

Effects of lovastatin and pravastatin on the survival of hamsters with inherited cardiomyopathy.

Cardiomyopathic hamsters develop heart disease early in life, which leads to congestive heart failure and death as these hamsters age. Hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitors have been reported to reduce ubiquinone concentrations and to deteriorate myocardial function in humans and in experimental animals. HMG-CoA reductase inhibitors differ regarding their ability to penetrate extrahepatic tissues. As a consequence, lovastatin inhibits cholesterol biosynthesis at least 100-fold more effectively than pravastatin in extrahepatic cells. We examined the effect of lovastatin and pravastatin (approximately 10 mg per kilogram of body weight and per day mixed in the diet) compared with controls on the lifespan of cardiomyopathic hamsters (BIO 8262 strain) in the heart-failure period. In male hamsters, neither lovastatin nor pravastatin significantly affected survival. In female hamsters, lovastatin reduced median survival time from 89 days (control animals) to 30 days (P <.05); pravastatin (median survival, 115 days) had no statistically significant effect. We conclude that lovastatin, but not pravastatin, at a daily dose of 10 mg per kilogram of body weight significantly increases the mortality of cardiomyopathic hamsters. This effect may be the result of inhibition of myocardial ubiquinone supply.

Administration, Oral↗