[Whole-body MRI in children with langerhans cell histiocytosis for the evaluation of the skeletal system]. / Ganzkörper-MRT bei Langerhans-Zell-Histiozytose im Kindes- und Jugendalter zur Beurteilung des Skelettsystems.

PURPOSE: The usefulness of whole-body MRI (WB-MRI) for the detection of skeletal lesions in patients with Langerhans cell histiocytosis should be documented on the basis of case presentations. MATERIALS AND METHODS: In six patients with histologically proven Langerhans cell histiocytosis, 14 WB-MRI examinations were performed to evaluate the skeletal system within disease staging (6 primary, 8 follow-up examinations). The examinations were performed on a 1.5 Tesla, 32-channel whole-body scanner. The examination protocol consisted of T 1-weighted and STIR sequences in coronal and sagittal orientation. For comparison, radiographs of the initial skeletal lesions and those that were additionally detected on WB-MRI were available. RESULTS: In 4 patients no additional skeletal lesions were found on WB-MRI besides the initial lesion leading to the diagnosis of unifocal single system disease. In 2 patients WB-MRI was able to identify additional skeletal lesions. In a 5 S year-old boy with the primary lesion located in the cervical spine, a second lesion was detected in the lumbar spine on the initial scan and in the skull and proximal femur during follow-up examination. In a 12 year-old girl with a primary lesion of the thoracic spine, WB-MRI diagnosed additional lesions in the pelvic bone and the tibia. In both patients the diagnosis of multifocal skeletal involvement led to chemotherapy. During follow-up examination, the healing response under therapy could be demonstrated. Comparison with conventional imaging showed that especially lesions located in the spine or the pelvis were not detectable on radiographs even when knowing the MR results. CONCLUSION: The extent of skeletal involvement in Langerhans cell histiocytosis has crucial impact on therapy and prognosis. Whole-body MRI has been reported to be an established method for the evaluation of disseminated skeletal disease with distinct advantages over conventional radiography and bone scintigraphy. Our results suggest that WB-MRI should also be the imaging modality of choice for the assessment of skeletal involvement in children with Langerhans cell histiocytosis.

Key importance of the Helicobacter pylori adherence factor blood group antigen binding adhesin during chronic gastric inflammation.

Helicobacter pylori has been assigned as a class I carcinogen because of its relation to gastric adenocarcinoma. Chronic H. pylori infection may lead to severe gastritis, glandular atrophy (AT), and intestinal metaplasia (IM). Strains secreting the vacuolating toxin VacA and producing the cytotoxin-associated antigen CagA (type 1 strains), as well as the blood group antigen binding adhesin (BabA) targeting Lewis(b) antigens, have been associated previously with distal gastric adenocarcinoma (M. Gerhard et al., Proc. Natl. Acad. Sci. USA, 96: 12778-12783, 1999) and may therefore also be related to lesions preceding gastric cancer. Antral and corpus biopsies were collected from 451 patients; 151 were H. pylori positive, as determined by PCR. Gastric biopsies were histologically evaluated for activity of gastritis (G0-G3, granulocyte infiltration), chronicity of gastritis (L1-L3, lymphocyte infiltration), and the presence of IM and/or AT according to the Sydney classification. Simultaneously, the presence of bacterial genes encoding virulence and adherence factors (racAs1/s2, cagA, and babA2) was determined by PCR. The presence of cagA+ and vacAs1 (alone or combined) both correlated with activity and chronicity of gastritis (P < 0.05); however, the overall prevalence of these genes was 60 or 72%, respectively, and was thus relatively frequent. The babA2 gene, encoding the adhesin BabA, was detected in 38% of infected patients and was correlated with the activity of gastritis in antrum and corpus (P < 0.005). cagA+/vacAs1+ strains (suggesting the presence of type 1 strains) that were also babA2 positive were detected more frequently in patients with severe histological alterations (such as G3, IM, or AT) compared with subjects without these changes (P < 0.01). cagA+/vacAs1+ strains that were babA2 negative, however, lacked a significant correlation with severe histological changes, activity, or chronicity of gastritis in antrum and corpus. Adherence of H. pylori via BabA appears to be of importance for efficient delivery of VacA and CagA and may play a special role in the pathogenesis of severe histological changes.

Evolution of DNA or amino acid sequences with dependent sites.

A framework is outlined to study the evolution of DNA or amino acid sequences, if sequence sites do not evolve independently. The units of evolution are nonoverlapping subsequences of length l. Each subsequence evolves independently of the others, but within a subsequence the sequences show a Markov order one dependency. We describe an algorithm to mimic the evolution of such sequences. The influence of dependencies between sites on distance estimates and the reliability of tree reconstruction methods is investigated. We show that an inappropriate model of sequence evolution in the tree reconstruction process will lead to a nonempty Felsenstein zone. Finally, we describe a method to infer l from sequence data. Examples from the evolution of DNA sequences as well as from amino acids are given.

[Telemetry during swimming in risk evaluation of heart patients in rehabilitation]. / Die Schwimmtelemetrie in der Risikobeurteilung von Herzpatienten während der Rehabilitation.

Swimming differs from other forms of exercise due to its additional hydrostatic and thermal burden. It was investigated whether additional pathologic findings in comparison to history and standard exercise tests can be obtained by holter monitoring during swimming. Symptoms and exercise electrocardiogram were compared with the holter ECG during swimming in 125 patients divided into 3 groups with different diagnoses and severity of cardiac diseases. In a considerable percentage of patients ischemic changes and severe rhythm disturbances were found only during swimming with further diagnostic and therapeutic consequences, though patients with moderate and severe angina and with significant ischemic signs in the exercise test were excluded and mainly patients with slight or absent symptoms were evaluated predominantly. Thus, since swimming is a favorite leisure-time occupation also in patients with diseases of heart and circulation, holter monitoring during swimming is of diagnostic importance in the rehabilitation of these patients.

[Allergic transfusion reactions in a patient with multiple food allergies]. / Allergische Transfusionsreaktionen bei einer Patientin mit multiplen Nahrungsmittelallergien.

HISTORY AND CLINICAL PRESENTATION: A 13-year-old girl with an osteosarcoma was treated by surgery and chemotherapy. During three transfusions of apheresis platelet concentrates allergic reactions occurred, partly in spite of premedication with an antihistamine and a corticoid. INVESTIGATIONS: As the patient declared to be allergic to some foods, in-vitro tests for allergen-specific IgE antibodies were performed and showed markedly positive results for specific IgE to carrot and celery, less so to hazelnut, peanut and a lot of other food antigens. The donor of one of the unsuitable platelet concentrates remembered when questioned, that he had eaten carrots and chocolate with hazelnuts during the evening before platelet donation. TREATMENT AND COURSE: Two washed platelet concentrates were transfused without any problem. Furthermore, transfusions of nine red blood cell concentrates and one unit of virus-inactivated frozen pooled plasma were well tolerated. CONCLUSION: Patients should be asked for allergies previous to transfusions to be alert to allergic reactions in patients with a positive history of food or drug allergies. If premedication with antihistamines does not prevent severe allergic transfusion reactions, transfusion of washed platelet concentrates and of virus-inactivated frozen pooled plasma can be considered.

Total body MRI-governed involved compartment irradiation combined with high-dose chemotherapy and stem cell rescue improves long-term survival in Ewing tumor patients with multiple primary bone metastases.

We examined the role of total body magnetic resonance imaging (TB-MRI)-governed involved compartment irradiation (ICI) and high-dose chemotherapy (HDC), followed by stem cell rescue (SCR) in patients with high-risk Ewing tumors (ETs) with multiple primary bone metastases (high-risk ET-MBM). Eleven patients with high-risk ET-MBM receiving initial assessment of involved bones by TB-MRI were registered from 1995 to 2000 (group A). In all, 6 patients out of 11 had additional lung disease at initial diagnosis; all had multifocal bone disease with more than three bones involved. After systemic induction with etoposide, vincristine, adriamycin (doxorubicin), ifosfamide, and actinomycin D (EVAIA) or VAIA chemotherapy, ICI of all sites positive by TB-MRI was administered, followed by HDC and SCR. A second group matched for observation period and consisting of 26 patients with more than three involved bones at diagnosis was treated with the European Intergroup Cooperative Ewing Sarcoma Study-92 (EICESS-92) protocol (group B). These patients did not receive TB-MRI and consequently did not receive TB-MRI-governed ICI, or HDC and SCR. Survival in group A vs group B was 45 vs 8% at 5 years and 27 vs 8% at 10 years after diagnosis (log rank and Breslow: P<0.005). We conclude that TB-MRI-governed ICI followed by HDC and SCR in ET-MBM is feasible and warrants further evaluation in prospective studies.

[False-positive hepatitis serology after administration of immunoglobulins]. / Falsch-positive Hepatitis-Serologie nach Immunglobulingabe.

HISTORY: A 17-year-old boy with chronic myelogenous leukemia received a bone marrow transplantation (BMT) from an unrelated donor. 22 days before BMT he had been HBs antigen and anti-HBc negative. 68 days after BMT he was tested again and showed a "seroconversion" for hepatitis B (anti-HBc positive, anti-HBs 59 IU/l), raising the suspicion of a posttransfusion hepatitis. The patient had not received any blood transfusion in the 6 months before BMT. From day 0 to day + 68 he received four red blood cell concentrates and 18 platelet concentrates. The bone marrow donor had been HBs ag and anti-HBc negative. INVESTIGATIONS: A stored serum sample of the recipient obtained on day -8 was available and proved to be negative for HBsAg and Anti-HBc IgM, but positive for anti-HBc and anti-HBs. A serum sample from day -22 was negative for all these parameters. DIAGNOSIS: These results can be explained by the administration of 17.5 g of a polyvalent immunoglobulin (Ig) concentrate for CMV prophylaxis on day -9: anti-HBc and anti-HBs (1814 IU/l) were found in the lot that the patient had received. Nine further doses of immunoglobulin concentrates were given up to day + 68. COURSE: Four months after the last administration of Ig concentrates, the patient was negative for HBs ag, anti-HBc and anti-HBs. CONCLUSION: Ig concentrates contain not only those antibodies, which are given to a patient for treatment, but also all other antibodies contained in the donor plasma pool. Thus administration of Ig concentrates can cause a "false-positive" hepatitis B serology by passive transfer of these antibodies. Such an artificial seroconversion may also lead to a false suspicion of a transfusion transmitted hepatitis B infection.

Efficient and fast targeted production of murine models based on ENU mutagenesis.

Mice with targeted genetic alterations are the most effective tools for deciphering organismal gene function. We generated an ENU-based parallel C3HeB/FeJ sperm and DNA archive characterized by a high probability to identify allelic variants of target genes as well as high efficiencies in allele retrieval and model revitalization. Our archive size of over 17,000 samples contains approximately 340,000 independent alleles (20 functional mutations per individual sample). Based on an estimated number of approximately 30,000 mouse genes, the parallel sperm/DNA archive should permit the identification and recovery of ten or more alleles per average target gene which translates to a calculated 99% success rate in the discovery of five allelic variants for any given average gene. The low rate of unrelated ENU-induced passenger mutations has no practical impact on the analysis of the allele-specific phenotype at the G3 generation because of dilution and free segregation of such unrelated passenger mutations. To date 39 mouse models representing 33 different genes have been recovered from our archive using in vitro fertilization techniques. The generation time for a murine model heterozygous for a mutation in a gene of interest is less than 2 months, i.e., three to four times faster compared with current embryonic stem-cell-based technologies. We conclude that ENU-based targeted mutagenesis is a powerful tool for the fast and high-throughput production of murine gene-specific models for biomedical research.

DISTREE: a tool for estimating genetic distances between aligned DNA sequences.

MOTIVATION: Substitution rates estimated from aligned DNA data can be used as genetic distances to investigate the phylogenetic relationship of those sequences. For this purpose, a Markov model of nucleotide substitution has to be assumed that describes this process most adequately. RESULTS: A program is presented that estimates substitution rates and their standard errors for a variety of Markov models. The model introduced by Hasegawa et al. (J. Mol. Evol., 22, 160-174, 1985) is the only one for which distances and standard deviations need to be calculated numerically, since analytical formulae cannot be derived. Each model is implemented in two different variants: (i) assuming rate homogeneity or (ii) starting from Gamma-distributed substitution rates across sequence sites. The estimation of heterogeneous substitution rates is based on a method suggested by Tamura and Nei (Mol. Biol. Evol., 10, 512-526, 1993). All required parameters are estimated from sequence data, hence the user is not asked to supply any additional input. One goal of the program is to support the user when choosing a particular model that describes most adequately the evolution of the given data set. For this purpose, a more detailed analysis of this model fit is provided. Phylogenetic trees reconstructed from the inferred distances using the neighbor-joining algorithm are also available.

Simulating efficiently the evolution of DNA sequences.

Two menu-driven FORTRAN programs are described that simulate the evolution of DNA sequences in accordance with a user-specified model. This general stochastic model allows for an arbitrary stationary nucleotide composition and any transition-transversion bias during the process of base substitution. In addition, the user may define any hypothetical model tree according to which a family of sequences evolves. The programs suggest the computationally most inexpensive approach to generate nucleotide substitutions. Either reproducible or non-repeatable simulations, depending on the method of initializing the pseudo-random number generator, can be performed. The corresponding options are offered by the interface menu.

A local algorithm for DNA sequence alignment with inversions.

A dynamic programming algorithm to find all optimal alignments of DNA subsequences is described. The alignments use not only substitutions, insertions and deletions of nucleotides but also inversions (reversed complements) of substrings of the sequences. The inversion alignments themselves contain substitutions, insertions and deletions of nucleotides. We study the problem of alignment with non-intersecting inversions. To provide a computationally efficient algorithm we restrict candidate inversions to the K highest scoring inversions. An algorithm to find the J best non-intersecting alignments with inversions is also described. The new algorithm is applied to the regions of mitochondrial DNA of Drosophila yakuba and mouse coding for URF6 and cytochrome b and the inversion of the URF6 gene is found. The open problem of intersecting inversions is discussed.

A stochastic model for the evolution of autocorrelated DNA sequences.

Currently used stochastic models of DNA sequence evolution assume independent and identically distributed nucleotide sites. They are too simple to account for dependence structures obviously present in molecular data. Up to now more realistic stochastic models for nucleotide substitutions have been considered intractable. In this paper a procedure that accounts for non-overlapping correlations among pairs of sites of a DNA sequence is developed. We show that currently used models that ignore correlated sites underestimate distances inferred from observed sequence dissimilarities. For the analyzed mitochondrial sequence data this underestimation is not drastic in contrast to paired regions (stems) of bacterial 23S rRNA sequences.

Toward assigning helical regions in alignments of ribosomal RNA and testing the appropriateness of evolutionary models.

We suggest a nucleotide substitution model that takes correlation between base-paired nucleotides into account. The model includes the estimation of the transition-transversion ratio and allows inference of the shape parameter of a discrete gamma distribution to include rate heterogeneity. A Cox-test statistic, applied to a diatom ribosomal RNA alignment, shows that the suggested correlation model explains evolution of the stem region better than usual independence models. Moreover, the Cox-test procedure is extended to shed some light upon the problem of assigning helical regions in a secondary structure based alignment. This approach provides an estimate of the percentage of stem positions that do not appear to be correlated.

Neutral adaptation of the genetic code to double-strand coding.

We lay new foundations to the hypothesis that the genetic code is adapted to evolutionary retention of information in the antisense strands of natural DNA/RNA sequences. In particular, we show that the genetic code exhibits, beyond the neutral replacement patterns of amino acid substitutions, optimal properties by favoring simultaneous evolution of proteins encoded in DNA/RNA sense-antisense strands. This is borne out in the sense-antisense transformations of the codons of every amino acid which target amino acids physicochemically similar to each other. Moreover, silent mutations in the sense strand generate conservative ones in its antisense counterpart and vice versa. Coevolution of proteins coded by complementary strands is shown to be a definite possibility, a result which does not depend on any physical interaction between the coevolving proteins. Likewise, the degree to which the present genetic code is dedicated to evolutionary sense-antisense tolerance is demonstrated by comparison with many randomized codes. Double-strand coding is quantified from an information-theoretical point of view.

Complementary coding conforms to the primeval comma-less code.

The hypothesis that the universal genetic code is adapted to double-strand coding is supported by its remarkable compatibility with the RNY comma-less hypothesis. Coding by a triplet code on a polynucleotide double-strand allows for enciphering of five additional messages with reference to a chosen primary reading frame. Assuming the acceptance of coupled mutations on both strands, the best codon register for two overlapping messages can be inferred. The idea of evolutionarily compatible coding of two proteins by one nucleotide double-strand is extended to complementary coding for one protein in folded, single-stranded RNA.

Stochastic traits of molecular evolution--acceptance of point mutations in native actin genes.

A stochastic matrix of nucleotide mutation probabilities is derived by counting differences and identities in alignments of native actin genes, with the aim of obtaining a more reliable data base for regular modes of molecular evolution. The evolution of DNA sequences is thereby considered as a Markov process consisting of events (point mutations) characterized by a stochastic matrix for codon-codon interchanges. The genetic distance is set to 1 PAM (percentage of accepted point mutations). The results can be reproduced by Monte Carlo simulations which are subjected to selective constraints. The latter are observed as nonrandom codon usage and ratios of silent to recognizable point mutations. Specific patterns within the matrix of mutation probabilities attest to preferences of natural selection in the evolution of a specific protein.

Concurrent neutral evolution of mRNA secondary structures and encoded proteins.

Messenger RNA sequences often have to preserve functional secondary structure elements in addition to coding for proteins. We present a statistical analysis of retroviral mRNA which supports the hypothesis that the natural genetic code is adapted to such complementary coding. These sequences are still able to explore efficiently the space of possible proteins by point mutations. This is borne out by the observation that, in stem regions of retroviral mRNA foldings, silent mutations on one strand are preferentially accompanied by conservative mutations on the other. Distances between amino acids based on physicochemical properties are used to quantify the conservation of protein function under the constraint of maintained RNA secondary structure. We find that preservation of RNA secondary structure by compensatory mutations is evolutionary compatible with the efficient search for new variants on the protein level.