We report two cases of aplastic anaemia following exposure to 'Ecstasy' (MDMA, 3,4-methylenedioxymethamphetamine). In both cases the aplastic anaemia resolved spontaneously 7-9 weeks after presentation. Long-term bone marrow culture study of one patient demonstrated complete normalization of haemopoiesis at time of haematological recovery, suggesting either that damage to the haemopoietic stem cell had been only transient, or that a more mature, committed progenitor cell was the target. Because MDMA may have been a factor in the aetiology of the bone marrow suppression in these two cases, we recommend close haematological monitoring of young adults presenting with toxicity from MDMA, and a detailed history of exposure to recreational drugs in all new patients presenting with aplastic anaemia.

We investigated the effects of granulocyte-macrophage colony- stimulating factor, interleukin-3, stem cell factor, interleukin-6, and granulocyte colony-stimulating factor (G-CSF) alone, and in combination, on the clonogenic potential of normal and aplastic anemia (AA) bone marrow mononuclear cells (BMMC and CD34+ cells. AA BMMC consistently produced a significantly lower absolute number of colonies than normal, but, when account was taken of the reduced proportion of CD34+ cells in AA BM, there was no significant difference in terms of cloning efficiency (CE). However, when removed from the influence of accessory cells, the CE of AA CD34+ cells decreased significantly more than normal, indicating a defect in their function, either in terms of dependence on accessory cell-derived factors or susceptibility to cell damage when sorted. Of the factors studied, G-CSF had the most significant effect on the response of CD34+ cells from both groups when removed from their accessory cells. This was particularly true for AA CD34+ cells, whose response to cytokine stimuli containing G-CSF enabled them to match the response of normal CD34+ cells.

Defective marrow stroma, or microenvironment, have been proposed as one of several mechanisms to account for bone marrow failure in aplastic anaemia (AA). This could involve defects in positive- or negative-acting haemopoietic regulator expression by AA stroma, or alteration of normal stroma-stem cell interactions. We have used a sensitive bioassay to investigate production of granulocyte-colony stimulating factor (G-CSF), granulocyte-macrophage-colony stimulating factor (GM-CSF), interleukin (IL)-3, IL-6 and stem cell growth factor (SCF), by normal and AA stroma in long-term bone marrow culture (LTBMC). LTBMC were grown to confluence, irradiated and harvested to yield a single cell suspension. These cells were cocultured with normal target bone marrow mononuclear cells (BMMC), or CD34+ cells, in clonogenic assays, in the absence of exogenous cytokines. Cytokines responsible for the colony-stimulating activity (CSA) and burst-promoting activity (BPA) produced by stromal cells were identified by neutralizing antibodies to specific cytokines. All normal stroma populations produced G-CSF and GM-CSF, 93% produced IL-3, 80% produced IL-6, and 70% produced SCF. Similarly, all AA stroma produced G-CSF and GM-CSF, and 71% produced SCF. In contrast, only 71% of AA stroma produced IL-3 and 36% produced IL-6. Target cell stimulation was not dependent on direct stroma-target cell contact, suggesting production of soluble cytokines. However, although both IL-6 and G-CSF were detected in LTBMC supernatants by enzyme-linked immunoassay (ELISA), IL-3 and GM-CSF were undetectable, perhaps indicating low-level local production of these factors.

Interleukin-3 (IL-3) has been shown to have significant effects on haemopoiesis in vitro, but early investigations of normal human long-term bone marrow cultures (LTBMC) have failed to demonstrate IL-3 production by stromal cells, either by Northern blotting for mRNA, or assaying for bioactivity in culture supernatants. One recent report, using reverse transcription-polymerase chain reaction (RT-PCR), demonstrated IL-3 expression in only one of eight cultures. We have developed a sensitive bioassay for the detection of IL-3 production from normal stroma in LTBMC. LTBMC were grown until confluent, irradiated, and stroma harvested by trypsinization to yield single-cell suspensions. These cells were then cocultured with target bone marrow mononuclear cells (BMMC), or CD34+ cells in clonogenic assays, either in the presence or absence of anti-IL-3 neutralizing antibodies. We have demonstrated IL-3 production in 32/34 cases. In addition, by separating stroma from target cells using cell culture inserts, we have shown that direct stroma:stem cell contact is not necessary for colony growth, suggesting that IL-3 diffuses into the supernatant. However, when supernatants from LTBMC were assayed by enzyme-linked immunoassay (ELISA), no IL-3 was detected. This suggests that IL-3 is probably produced at low levels and has a short-range interaction. Stroma production of IL-3 was confirmed by the detection by RT-PCR of IL-3 mRNA in 3/3 cases. The simultaneous detection of CD2 mRNA demonstrated that T cells are part of the bone marrow stroma. It is therefore possible and probably likely that these cells are the source of IL-3.