Mammalian oocyte quality reduces with age. We show that prior to the occurrence of significant aneuploidy (9M in mouse), heterochromatin histone marks are lost, and oocyte maturation is impaired. This loss occurs in both constitutive and facultative heterochromatin marks but not in euchromatic active marks. We show that heterochromatin loss with age also occurs in human prophase I‐arrested oocytes. Moreover, heterochromatin loss is accompanied in mouse oocytes by an increase in RNA processing and associated with an elevation in L1 and IAP retrotransposon expression and in DNA damage and DNA repair proteins nuclear localization. Artificial inhibition of the heterochromatin machinery in young oocytes causes an elevation in retrotransposon expression and oocyte maturation defects. Inhibiting retrotransposon reverse‐transcriptase through azidothymidine (AZT) treatment in older oocytes partially rescues their maturation defects and activity of the DNA repair machinery. Moreover, activating the heterochromatin machinery via treatment with the SIRT1 activating molecule SRT‐1720, or overexpression of Sirt1 or Ezh2 via plasmid electroporation into older oocytes causes an upregulation in constitutive heterochromatin, downregulation of retrotransposon expression, and elevated maturation rates. Collectively, our work demonstrates a significant process in oocyte aging, characterized by the loss of heterochromatin‐associated chromatin marks and activation of specific retrotransposons, which cause DNA damage and impair oocyte maturation.

Summary The TEL/AML1 (ETV6/RUNX1) fusion gene is the most common genetic rearrangement in paediatric acute lymphoblastic leukaemia (ALL). Although considered to be a low‐risk leukaemia, it is associated with a relapse rate of 10–20%. The coexistence of different subclones at diagnosis, based on polymerase chain reaction (PCR) studies of IG/TCR gene rearrangement, with differential response to chemotherapy, was recently reported in this subtype of ALL. We wished to demonstrate such subclones at diagnosis by a recently developed technique of quantitative multiparametric fluorescence in situ hybridization (FISH). Bone marrow cells from 80 paediatric patients with ALL at diagnosis were analysed for the presence of the TEL/AML1 fusion gene by interphase FISH. Fourteen patients were positive for the translocation. Four of them had several subclones associated with various combinations of additional chromosomal abnormalities. The most striking was an atypical and unexpected hybridization pattern consistent with a submicroscopic deletion of the 5′ region of the AML1 breakpoint. Other abnormalities included TEL deletion, trisomy and tetrasomy 21 as well as double TEL‐AML1 fusion. The presence of numerous subclones in about 25% of patients with TEL/AML1+ ALL suggests extensive clonal evolution by the time of diagnosis.

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Summary:Successful stem cell transplantation for patients with severe combined immunodeficiency (SCID) from matched family donors without conditioning results in engraftment of T lymphocytes. B lymphocytes engraft in only 50% of the cases, while myelopoiesis and erythropoiesis remain of host origin. Full hematopoietic engraftment was reported in one case after bone marrow transplantation without conditioning for a SCID patient. We studied three SCID patients who were transplanted with unmodified mobilized peripheral blood from HLA-identical family sex-mismatched members. They received megadoses of stem cells (18-23 Â 10 6 CD34/kg). In contrast to the expected mixed chimerism that usually occurs in the absence of conditioning, we found in our patients 100% donor cell engraftment based on fluorescence in situ hybridization (FISH) and microsatellite techniques. Subset analysis of the engrafted cells using a multiparametric system enabling a combined analysis of morphology, immunophenotyping and FISH showed that both T and B lymphocytes and myeloid cells were of donor origin in two patients, while T lymphocytes and myeloid cells were of donor origin in the third. In the two cases with ABO incompatibility, erythroid engraftment was evidenced by blood group conversion from recipient to donor type. Multilineage donor engraftment is possible in SCID patients even without conditioning. Successful stem cell transplantation without conditioning in these patients results in T-lymphocyte engraftment and correction of the immune dysfunction. However, engraftment of B lymphocytes occurs in only 50% of the cases, NK cells are either donor or host in origin, whereas myelopoiesis and erythropoiesis remain of host origin. 1,2The preparative regimen ablates the host stem cells and enables the engraftment of donor stem cells which later differentiate to all cell lineages. In the absence of conditioning, the host stem cells are not ablated, and as donor stem cells do not have an advantage over host stem cells they usually do not engraft. Full donor hematopoietic engraftment after allogeneic bone marrow transplantation without conditioning has been reported in only one patient with adenosine deaminase (ADA)-deficient SCID. 3Fluorescence in situ hybridization (FISH) using sexchromosome-specific probes is being increasingly used for the assessment of chimerism in sex-mismatched transplants. 4 However, this technology sometimes lacks both specificity and sensitivity. Flow-sorting of cell subsets provides sensitive assessment of changes in the pattern of chimerism which had escaped detection in assays using whole blood cell samples.5 This technology, however, is laborious and requires high personnel skills. A simultaneous analysis of morphology, immunophenotyping and FISH of the same cell using a multiparametric cell-scanning system enables the detection of the cell origin of different cell subsets without the need of cell sorting. 6 We have followed the chimerism status of different cell subsets of three SCID patients after peripheral bl...