Biomedical subjects
Publications and source records attributed to M Poetsch.
A species-specific gene probe for Hyphomicrobium facilis was generated from a transposon Tn5-132 insertion mutant defective in methanol oxidation by the inverse PCR. With this probe, the abundance of H. facilis in a garden soil was determined as a subfraction of the total cultivable hyphomicrobia.
The French-American-British subtype acute myelomonocytic leukemia with abnormal eosinophils (FAB AML M4Eo) with pericentric inversion of chromosome 16 is cytomorphologically defined by a myelomonoblastic blast population and abnormal eosinophils. Until now, it remained an open question whether these abnormal eosinophils are part of the malignant clone or an epiphenomenon. We analyzed five cases of AML M4Eo with inv(16) and combined May-Grünwald-Giemsa staining with fluorescence in situ hybridization using yeast artificial chromosome clone 854E2, which spans the inv(16) breakpoint on 16p. In the case of inv(16), three instead of the normal two hybridization signals can be observed both on metaphase spreads and in interphase cells. With this approach, we were able to show inversion 16 in abnormal eosinophils and, therefore, identified them as a part of the leukemic cell population.
Three patients with secondary acute leukaemia after treatment with topoisomerase II inhibitor agents are described. Two patients had acute myeloid leukaemia (AML). FAB M5a, one had pro-B-acute lymphoblastic leukaemia (ALL). The interval between initiation of chemotherapy and the onset of secondary acute leukaemia was 19-20 months. 11q23 rearrangements were detected in all cases. They were due to translocations t(11;19) (q23;p13.3), t(11;16)(q23;p13) and t(4;11)(q21;q23), respectively. Fluorescence in situ hybridization (FISH) with Yeast Artificial Chromosome (YAC) probe 13HH4 spanning the ALL-1 gene on 11q23 confirmed that in each case the ALL-1 gene had been disrupted by the translocations. The study underlined the relationship between the development of secondary acute leukaemias with 11q23 rearrangement and previous chemotherapy with topisomerase II inhibitor agents. So far, however, only six adult patients with secondary ALL with t(4;11) after treatment with topoisomerase II inhibitor agents have been reported. All with t(4;11) mostly occurs in infants or young children. Our patient received epirubicin continuously for >19 months. This indicates that both myeloid and lymphoid leukaemias with involvement of the ALL-1 gene can be induced by exogenous agents, especially topoisomerase II inhibitors. Thus they may have a common biological background. This hypothesis was substantiated by means of combined immunophenotyping and FISH (FICTION). In the case of AML M5a with t(11;19), the tumour cells with ALL-1 rearrangement expressed CD34. Moreover, the pro-B-ALL with t(4;11) was CD34 positive. These findings suggest that the cell of origin of secondary AML and ALL with 11q23 rearrangement is an immature haemopoietic progenitor cell.
PURPOSE: The aim of this study was to establish a fluorescence in situ hybridization (FISH) technique for the detection of t(14;18)(q32;q21), characteristic for follicular lymphoma (Kiel classification: centroblastic centrocytic [cb-cc] lymphoma). MATERIALS AND METHODS: After the FISH system had been established, parallel studies of lymph node biopsy specimens from 30 patients with cb-cc lymphoma and from 32 patients with nonneoplastic lymphoproliferation were performed by means of chromosome analysis, polymerase chain reaction (PCR), and FISH analysis. Two differently labeled yeast-artificial-chromosome (YAC) probes that contained the entire bcl-2 gene and the C-region of the immunoglobulin H (IgH) gene, respectively, were used to detect t(14;18) by FISH. RESULTS: The presence of the translocation is indicated by a red (Cy3)/green (fluorescien isothiocyanate [FITC]) double signal, which corresponds to the IgH/bcl-2 fusion gene, whereas in normal cells the signals are separate. Control studies showed that the double signal is visible in less than 1% of normal cells. FISH analysis was able to identify the t(14;18) in all cases of cb-cc lymphoma we studied. All bcl-2 breakpoints can be detected. Combined immunophenotyping and interphase cytogenetics demonstrated that t(14;18) was restricted to CD22+ B lymphocytes and never occurred in CD3+ T lymphocytes. In four of 32 cases of nonneoplastic lymphoproliferation, t(14;18) was also detected. CONCLUSION: FISH turned out to be the most sensitive method to detect t(14;18). Our FISH results confirm PCR data from other groups that found evidence for the presence of t(14;18) in nonneoplastic lymphoproliferation. It needs to be determined whether, in morphologically nonneoplastic processes, t(14;18) is associated with an increased risk for the development of non-Hodgkin's lymphoma.
A number of neoplastic disorders are characterized by recurrent chromosome aberrations. One of these is the translocation t(2;5), which is found in a considerable percentage of large-cell anaplastic lymphomas. This translocation results in the fusion of two genes, alk and npm. The recent discovery of alk/npm mRNA in 11 of 13 cases of Hodgkin's disease has caused a controversial discussion concerning the question of whether t(2;5) is also present in Hodgkin and Reed-Sternberg cells. We tackled this problem on the molecular cytogenetic level by combined CD30 immunophenotyping and interphase cytogenetics. Using a pair of DNA probes flanking both sides of the npm gene breakpoint at 5q35 we were able to prove, at least in 12 of 13 cases of Hodgkin's disease, that all CD30-positive Hodgkin and Reed-Sternberg cells lacked the translocation t(2;5). Fifteen to forty-five Hodgkin/Reed-Sternberg cells were analyzed per case (mean, 27). Our findings indicate that this translocation is not a primary event in the development of Hodgkin's disease.
In Hodgkin's disease, cytogenetically aberrant clones have been demonstrated in a minority of cases studied. In the remaining cases, only normal metaphases have been found, but it is questionable whether normal karyotypes actually correspond to the pathognomonic Hodgkin and Reed-Sternberg (HRS) cells. Numerical aberrations could be studied by fluorescence in situ hybridization (FISH). However, in Hodgkin's disease, the percentage of tumor cells is mostly below the detection limit of FISH, which is near 1%. With the technique of simultaneous fluorescence immunophenotyping and interphase cytogenetic analysis (FICTION), this problem can be overcome. By FICTION, hybridization signals can selectively be evaluated within the CD30a+ cell population. We have studied 30 cytogenetically analyzed cases of Hodgkin's disease by means of FICTION. In all cases, we found numerical chromosome aberrations within the majority of CD30+ HRS cells. In cases with complex and hyperdiploid karyotypes, the cytogenetic results agreed with the FICTION data. There was considerable variability in the chromosome numbers, demonstrating that karyotype instability is an in vivo phenomenon of HRS cells. Lymphocytes never displayed numerical chromosome changes. Our results indicate that HRS cells regularly exhibit numerical chromosome aberrations and that the chromosome numbers are always in the hyperploid range.
Cytogenetic studies on Hodgkin's disease (HD) typically reveal very complex karyotypes with a variety of numerical and structural abnormalities. The confusing thing is that about 10% of cases contain relatively discrete chromosome aberrations, for example a simple trisomy or loss of one single chromosome. Whether these karyotypes really correspond to Hodgkin and Reed-Sternberg (HRS) cells is uncertain. They could, for example, represent early stages in the evolution of the karyotype of the pathognomonic HRS cells. On the other hand, they could be artificial events that occur during the cytogenetic procedure. In our experience, isolated loss of the Y chromosome is the most frequent finding of this type. This aberration is usually considered to be a preparation artifact. However, if one takes into account that in HD up to 50% of male cases with complex karyotypes also lack the Y chromosome, a possible relation to HRS cells must be considered. The technique of simultaneous fluorescence immunophenotyping and interphase cytogenetic analysis (referred to as FICTION) is a powerful tool for studying the nature of cytogenetically abnormal cells. With the FICTION technique we studied four cases of HD in which the chromosome analysis had shown only the loss of the Y chromosome. Our aim was to clarify whether these karyotypes corresponded to the CD30-positive HRS cells. In two cases we found that HRS cells actually lacked the Y chromosome. There was strong evidence, however, that the HRS cells additionally had other chromosome aberrations and thus could not correspond to the cytogenetically determined karyotypes.
Four new and two known peptide antibiotics containing amiclenomycin (Acm) have been isolated from a culture of Streptomyces venezuelae Tü 2460: L-MeIle-L-Acm (1), L-Ile-L-Acm (2), L-MeVal-L-Acm (3), L-MeIle-L-Acm-L-Gln (4), L-Ile-L-Acm-L-Gln (5) and L-Val-L-Acm-L-Gln (6). These di- and tripeptides exhibited antimicrobial activity on a minimal medium against Gram-negative bacteria, which could be reversed by biotin. It was shown that the antibiotics were decomposed by peptidases to provide amiclenomycin (7) after their uptake into cells of Escherichia coli via peptide-permeases. The antimicrobial activity of the amiclenomycin-peptides was the inhibition of DAPA-aminotransferase by the amiclenomycin-warhead, however, amiclenomycin itself was hardly transported into the cells. Since the amiclenomycin peptides misuse general transport systems, they are presented here as examples for the illicit transport concept.
The beta-lactamase TEM-1 was purified from an Escherichia coli ATCC 11 775 R+TEM culture by preparative isoelectric focussing. This preparation was used to establish a calibration curve between the concentration of the enzyme and the cleavage of nitrocefin, by which the quantitative production of beta-lactamase TEM-1 in the growing culture was determined. The effect of purine derivatives on the synthesis of the beta-lactamase during the growth of the bacterium was tested in order to determine the essential basic structure of the molecule. It is suggested that the keto-methylamino part of the pyrimidine nucleus is decisive for the potency of the structure. Inhibition studies on the purified enzyme demonstrated that the purine compounds act by inhibiting the synthesis of the beta-lactamase and not by inactivation of the enzyme. This mode of action manifests itself as a synergism with amoxycillin against the beta-lactamase producing strain.
We describe the first inverted duplication of the p21.3p26 region of chromosome 3 in a child with phenotypic features of the trisomy 3p syndrome. This uncommon type of aberration was verified by multicolour fluorescence in situ hybridisation (FISH) using yeast artificial chromosome (YAC) clones from chromosome 3 (CEPH library). With a newly constructed YAC clone from the 3p26 region an unexpected subtelomeric deletion was diagnosed in the aberrant chromosome 3. Using the primed in situ labelling (PRINS) method, telomeres were found to be present on the recombinant chromosome 3. The repeated appearance of concomitant distal deletions in inverted duplications suggests that an overall mechanism exists for the origin of such duplications/deficiencies.
The paper reports on molecular biological investigations in a case of poaching which resulted in considerable damage for property. Blood traces from poaching sites have been analysed and compared with blood from two knives and a pair of trousers of the suspects. In two of three poaching sites, genomic DNA could be amplified by PCR and assigned to fallow deer. The authors could demonstrate identical allelic combinations between one of the poaching sites and a part of the traces. Problems of the legal appreciation in this case are described.