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Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 03/2011; 25(6):1050-3. · 8.30 Impact Factor
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ABSTRACT: The thrombopoietin receptor gene (MPL) is expressed in megakaryocytes and exhibits the gain of function point mutation W515K/L in approximately 5% of patients with primary myelofibrosis/idiopathic myelofibrosis (PMF) representing one subtype of the chronic myeloproliferative disorders (myeloproliferative neoplasm). A series of primary and secondary acute myeloid leukaemias (AML) with megakaryoblastic phenotype and myelofibrosis unrelated to PMF (n=12) was analysed for the MPL(W515K/L) mutation by pyrosequencing. In three cases (25%), MPL(W515L) was found and in two of these a combination with trisomy 21 or the Philadelphia chromosome occurred. None of the secondary AML cases evolving from pre-existing PMF showed MPL(W515K/L) (n=4). We conclude that MPL(W515L) occurs in a considerable proportion of acute megakaryoblastic leukaemias with myelofibrosis unrelated to PMF.
Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 03/2009; 23(5):852-5. · 8.30 Impact Factor
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ABSTRACT: Better understanding of early onset of interstitial fibrosis and tubular atrophy (IF/TA), as the morphological surrogate of renal allograft deterioration might improve outcome after renal transplantation.
We quantified mRNA expression of 3 profibrotic (transforming growth factor-beta (TGF-beta), tissue transglutaminase (tTG), tissue inhibitor of matrix metalloproteases (TIMP-1)) and 1 antifibrotic (matrix metalloprotease-2 (MMP-2)) molecule in protocol biopsies from renal allografts. From 107 transplants, two sequential protocol biopsies (6 weeks and 6 months) were analyzed. We evaluated a control group showing no IF/TA in both biopsies (n = 65) and a IF/TA group developing IF/TA at 6 months (n = 42). Expression data were correlated with clinical and histological risk factors for IF/TA and allograft function.
The expression of the genes correlated strongly with each other, particularly the profibrotic genes and in patients who developed IF/TA. Analyzing protocol biopsies from stable grafts, not all patients in both groups showed increased gene expression. In patients with increased gene expression a significantly higher tTG expression (matrix stabilization) at 6 weeks and a significantly lower MMP-2 expression (failure in matrix degradation) at 6 months were observed in the IFTA group compared to controls. Multivariate logistic regression revealed donor age positively and TIMP-1 expression at 6 weeks inversely correlated with IF/TA at 6 months.
We conclude that a disturbance in the equilibrium of pro- and antifibrotic pathways is decisive for early onset of IF/TA in renal allografts: insufficient degradation of exaggerated matrix production apparently changes the balance in the direction of IF/TA.
Clinical nephrology 07/2008; 69(6):408-16. · 1.17 Impact Factor
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Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 06/2008; 22(5):1059-62. · 8.30 Impact Factor
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G Buesche,
H Teoman,
W Wilczak,
A Ganser,
H Hecker,
L Wilkens,
G Göhring,
B Schlegelberger, O Bock,
A Georgii,
H Kreipe
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ABSTRACT: Marrow fibrosis (MF) has rarely been studied in myelodysplastic syndromes (MDS). There are no data on occurrence and significance of MF in the context of the World Health Organization (WHO) classification of disease. In total, 349 bone marrow biopsies from 200 patients with primary MDS were examined for MF and its prognostic relevance. MF correlated with multilineage dysplasia, more severe thrombopenia, higher probability of a clonal karyotype abnormality, and higher percentages of blasts in the peripheral blood (P<0.002). Its frequency varied markedly between different MDS types ranging from 0 (RARS) to 16% (RCMD, RAEB, P<0.007). Two patients with MF showed a Janus kinase-2 mutation (V617F). Patients with MF suffered from marrow failure significantly earlier with shortening of the survival time down to 0.5 (RAEB-1/-2), and 1-2 (RCMD, RA) years in median (P<0.00005). The prognostic relevance of MF was independent of the International Prognostic Scoring System and the classification of disease. Conclusion: The risk of MF Differs markedly between various subtypes of MDS. MF indicates an aggressive course with a significantly faster progression to fatal marrow failure and should therefore be considered in diagnosis, prognosis and treatment of disease.
Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 02/2008; 22(2):313-22. · 8.30 Impact Factor
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Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 01/2008; 21(12):2566-8. · 8.30 Impact Factor
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ABSTRACT: In chronic myeloid leukemia (CML), imatinib may reverse bone marrow fibrosis (MF). Whether the unfavorable prognosis of MF is also reversed and whether imatinib guarantees against evolution of MF are unclear as yet. Fifty-nine patients with Ph+ CML treated with > or = 400 mg imatinib/day were examined for MF in 6- to 12-month intervals. Imatinib effectively reversed initial MF (P<0.0005). However, during a follow-up period of up to 4.8 years, small foci with abnormal fiber increase (FFI) emerged in 8 of 30 pretreated and 6 of 29 non-pretreated patients. Patients with FFI showed a significantly lower probability of achieving a complete cytogenetic or major molecular response (36 versus 81%; P<0.007). During the further follow up, 57% of patients with FFI but none of the other patients suffered from full-blown MF (P=0.00005). None of the patients with FFI or MF showed a Janus kinase-2 mutation (V617F). Evolutions of FFI and MF were independent significant predictors of imatinib failure (P=0.0031), accelerated phase and death of patients (P=0.0001; multivariate analyses). Imatinib effectively reverses initial MF in CML, but neither eliminates its unfavorable prognosis nor guarantees completely against new evolution of MF.
Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 12/2007; 21(12):2420-7. · 8.30 Impact Factor
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Leukemia 07/2007; 21(12):2566-2568. · 9.56 Impact Factor
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ABSTRACT: The discovery of the Janus kinase 2 gain-of-function V617F mutation (JAK2V617F) provided a major breakthrough in the understanding of Philadelphia chromosome-negative chronic myeloproliferative disorders (Ph– CMPD). Among haematologic neoplasm the mutation appears to be almost specific for Ph– CMPD but the different entities comprising polycythaemia vera (PV), essential thrombocythaemia and chronic idiopathic myelofibrosis (CIMF) are not discriminated by the mutation. It is unclear how the diversity with heterogeneous clinical and pathoanatomical presentations comes about. It has been suggested that differences in JAK2V617F gene dosage or different degrees to which the haematologic lineages are affected by the mutation could explain the heterogeneity of morphology and prognosis. Indeed the mutation mediates a PV-like phenotype but with regard to myelofibrosis JAK2V617F does not appear to be a causative factor. Megakaryocytes are homozygous in the majority of fibrotic CIMF and PV, whereas JAK2V617F heterozygosity is predominantly encountered in prefibrotic CIMF and essential thrombocythaemia but transition from hetero- to homozygosity with onset of fibrosis is rare. In conclusion, JAK2V617F provides a valuable adjunct to the diagnosis of Ph– CMPD, in particular with regard to discrimination from reactive proliferations, but the challenge of correct subtyping and hence prognostication persists for clinicians and bone marrow pathologists.
Pathobiology 02/2007; 74(2):72-80. · 1.18 Impact Factor
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O Bock
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ABSTRACT: Histomorphological evaluation of bone marrow trephines and smears represents the major approach to diagnose the chronic myeloproliferative diseases (CMPD) and the myelodysplastic syndromes (MDS). However, rising insights into molecular pathogenesis of human diseases strengthen the attempt of pathologists to define and to detect underlying defects beyond the microscope. Since discovery of the Philadelphia chromosome in chronic myeloid leukemia as the first specific molecular abnormality ever detected in a human neoplasia the gain of knowledge of molecular pathomechanisms in Philadelphia chromosome negative (Ph-) CMPD was rather sparse. A decisive breakthrough in Ph CMPD was the finding of JAK2 (V617F) derived from a somatic point mutation in the majority of patients with polycythemia vera (P.vera) and half of patients with essential thrombocythemia (ET) and primary myelofibrosis (PMF). It therefore can not be overestimated that detection of JAK2 (V617F) in a suspective myeloproliferation now enables a clearcut discrimination of a true Ph CMPD from a reactive state, e.g. P.vera from reactive erythrocytosis. Interestingly, a basic principle of molecular defects demonstrable in CMPD and related disorders seems to be the involvement of genes with kinase activities. Some of those genes will be discussed in more detail. In primary MDS, karyotyping via classical cytogenetics is the predominant molecular approach to estimate prognosis, e.g. -Y, del(5q) and del(20q) represent favourable anomalies. Indeed, in 5q- syndromes karyotyping enables definite subtyping and allows clinicians and patients to expect a good prognosis. Until now, dozens of molecular abnormalities such as mutations in AML1, FLT3 and Ras as well as epigenetic alterations of genes have been identified to various degrees in MDS subtypes. Some of them seem to be involved in disease initiation ("master event") and others might indicate disease progression. However, even though useful for further dissection of molecular pathomechanisms the majority of aberrations currently does not serve as potent markers in the daily routine. Nevertheless, in CMPD and MDS the importance of molecular analyses for diagnosis, estimation of prognosis, and disease monitoring will further increase in a foreseeable period of time.
Verhandlungen der Deutschen Gesellschaft für Pathologie 02/2007; 91:140-53.
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ABSTRACT: Various immunological and non-immunological pathomechanisms are responsible for the cellular damage in renal allografts. Since the kidney is an anatomically complex organ with functional and morphological heterogeneous compartments (interstitium, tubuli, vessels, glomeruli), the local response to injury maybe variable, therefore, the identification of local pathomechanisms is important.
To elucidate any discrepancies in quantitative mRNA expression profiles between a total specimen analysis and a cell-specific evaluation after laser microdissection.
Real-time RT-PCR was performed for complement component C3 and heme oxygenase-1 (HO-1) genes compared to the housekeeping gene beta-actin using whole section RNA extracted from formalin-fixed and paraffin-embedded archival material of 16 explanted, rejected renal allografts. Ten non-transplant nephrectomies served as controls. For five cases from each group, five different compartments of the organs (interstitium, proximal tubuli, distal tubuli, vessels, glomeruli) were microdissected and quantitative analysis for C3 and HO-1 was performed identically.
Whole section mRNA expression analysis: the data showed a constant expression of the housekeeping gene beta-actin, a 7-fold increased expression of C3 and a 3-fold decreased expression of HO-1 in the allograft group as compared to the control group. mRNA expression results from microdissected compartments: in the control group, C3 and HO-1 expression could only be detected in the proximal tubuli of all cases whereas all five compartments analyzed from the rejecting kidneys showed expression of the two genes. In the allografts, expression levels of the investigated genes varied considerably not only among the different compartments but between individual cases as well.
Laser microdissection combined with real-time RT-PCR is a feasible approach for retrospective quantitative gene expression analysis in formalin-fixed and paraffin-embedded renal allograft specimens. As shown for C3 and HO-1, cell-specific expression patterns ofpathogenetically relevant genes vary considerably between individual cases. A close correlation of morphology and cell-specific gene expression analysis will contribute to the elucidation of the complex pathogenesis of chronic renal allograft nephropathy.
Clinical nephrology 04/2005; 63(3):193-201. · 1.17 Impact Factor
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ABSTRACT: Assessment of clonality either by demonstrating light chain restriction or showing monoclonal immunoglobulin gene rearrangement is a valuable and indispensable adjunct to diagnosis in hematopathology. The study of light chain restriction by immunohistochemistry on archival material is hampered by a very low sensitivity especially regarding low grade lymphomas of B cell origin. DNA rearrangement studies of the immunoglobulin locus do improve sensitivity markedly but for lymphomas of follicle center origin they are prone to false negative results due to hypermutations. Therefore we developed a new clonality assay based on the quantification of immunoglobulin light chain transcripts using real-time polymerase chain reaction technology, which is also suitable for the analysis of archival bone marrow trephines. We tested the reproducibility and sensitivity of this approach by comparatively analyzing a series of bone marrow trephines with multiple myeloma (n = 26), reactive lymphoid hyperplasia (n = 37), and focal infiltration by low grade B cell lymphoma (n = 29). We could raise the detection rate of clonality from an average of 17% by immunohistochemistry and 66% as assessed by polymerase chain reaction rearrangement studies to 83% by this new technique. Despite false negative results due to light chain hypermutation in some cases, the detection rate of clonality could be improved even for B cell lymphomas of follicle center origin (follicular lymphoma or marginal zone lymphoma) thus making this novel approach a valuable additional tool for the hematopathologist.
American Journal Of Pathology 01/2002; 159(6):2023-9. · 4.89 Impact Factor
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Analytical Biochemistry 09/2001; 295(1):116-7. · 3.00 Impact Factor
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ABSTRACT: Laser microdissection enables the contamination-free isolation of morphologically defined pure cell populations from archival formalin-fixed paraffin-embedded tissue specimens. Cells isolated by this method have been characterized by a wide variety of qualitative molecular assays, e.g. loss of heterozygosity, point mutations, clonality and lineage origin. The recently introduced real-time PCR technology renders the reliable quantification of very small amounts of nucleic acids possible. Several groups including our own showed that this technique can be successfully applied for the quantification of DNA and RNA isolated from microdissected archival tissue sections, even after immunohistochemical staining. The exact analysis of quantitative changes of nucleic acids during the course of pathological alterations has thus become possible. In many situations these quantitative changes can be expected to be more important than qualitative changes. The new technology for the quantification of structural genomic alterations and changes in the gene expression pattern in conjunction with microdissection have equipped morphologists with a powerful tool to study reactive and neoplastic changes of tissues.
Pathobiology 02/2000; 68(4-5):202-8. · 1.18 Impact Factor
