Red cells I: inherited anaemias.

Examination of the genetic mechanisms underlying the thalassaemias has led to a clearer understanding of the control of eukaryotic genes in general. Inherited disorders of haemoglobin synthesis are an important cause worldwide of morbidity and mortality, and place a large burden on patients, families, and ultimately communities. The haemoglobin disorders can be controlled, by counselling and prenatal diagnosis. Treatment is usually symptomatic, though bone-marrow transplantation for beta-thalassaemia may be successful in suitable patients.

The effect of rh-cytokines on the sensitivity of normal human CFU-GM progenitors to Ara-C and on the S-phase activity of light density human bone marrow cells.

This study examines the effect of pre-incubation in liquid phase of normal human CFU-GM progenitors with recombinant human (rh) cytokines prior to exposure to cytosine arabinoside (Ara-C) in clonogenic culture. Light density marrow cells (LMDCs) were preincubated in Biorich (chemically defined serum-free complete medium) with G + GM-CSF or IL-3 prior to semi-solid culture with Ara-C (10(-4) M-10(-12) M). Cell cycle analysis was performed by flow cytometry pre- and post-rh-cytokine treatment. Data from 26 normal marrows studied clonogenically, demonstrated a 60% reduction in CFU-GM with 10(-4) M Ara-C. Preincubation in Biorich medium with the addition of G + GM-CSF or IL-3 caused a significant enhancement of sensitivity to Ara-C across the dose curve. Biorich medium alone had an apparent protective effect resulting in amelioration of the cytotoxic effect of Ara-C. Cell cycle analysis of eight subjects showed significant recruitment into S-phase following pre-treatment with cytokines. A reduction in S-phase activity was noted in cells pre-incubated in Biorich alone. In summary, it would appear that cytokines can enhance the sensitivity of normal CFU-GM to Ara-C in vitro possibly due to an increase in S-phase activity. Pre-incubation in nutrient medium without cytokines resulted in cell quiescence.

Use of paraffin wax embedded bone marrow trephine biopsy specimens as a source of archival DNA.

AIMS: To evaluate the use of DNA extracted from paraffin wax embedded trephine biopsy specimens as a source of archival nucleic acid for Southern hybridisation studies and polymerase chain reaction (PCR) amplification. METHODS: DNA was extracted simultaneously from paraffin wax embedded bone marrow trephine and lymph node biopsy specimens after incubation of tissue sections for one to five days in lysis mix and proteinase K with periodic sampling. DNA from 10 trephine biopsy specimens was subjected to PCR amplification using HLA-DPB primers to determine whether the extracted nucleic acid was of sufficient quality to permit amplification. RESULTS: For most specimens the greatest yield of high molecular weight DNA was seen after five days' incubation. Unlike lymph node material the quality of extracted nucleic acid and the quantity obtained from trephines was insufficient for Southern blot analysis. PCR amplification using HLA-DPB primers yielded positive results in six out of 10 trephine biopsy specimens. CONCLUSIONS: DNA extracted from paraffin wax embedded trephine biopsy specimens is largely degraded and unsuitable for Southern analysis but serves as a useful source of archival nucleic acid for PCR amplification.

Chronic myelomonocytic leukaemia associated with T cell receptor delta gene rearrangement.

Morphological, immunophenotypic, and genetic analyses were carried out on peripheral blood, bone marrow, and pharyngeal biopsy material from a patient with chronic myelomonocytic leukaemia (CMML). Morphological analysis of bone marrow was diagnostic of CMML; immunophenotypic analysis of peripheral blood and bone marrow were negative for B and T cell antigens, and immunochemistry performed on the pharyngeal extramedullary infiltrate showed the presence of large monocytoid cells which stained positively for muramidase. Genotypic analysis, however, showed clonal rearrangement of the T cell receptor (TCR) delta chain gene, a marker of T cell or, less commonly, B cell lymphoid neoplasms. Other TCR genes, beta and gamma, were germline in all tissues examined. TCR delta is rearranged in precursor B cell and most T lymphoid neoplasms. A small proportion of cases (10%) of acute myeloid leukaemia (AML) also show rearrangement of the TCR delta gene. To date TCR delta rearrangement has not been described in CMML. The aberrant TCR delta rearrangement shown in this patient with CMML provides further evidence of the clonal nature of this disorder.