Clonal patterns of X‐chromosome inactivation in female patients withaplastic anaemia studied using a novel reverse transcription polymerase chain reaction method
Abstract: Conflicting results have been published on the frequency of clonal patterns of X‐chromosome inactivation in female patients with aplastic anaemia. Previous studies have used DNA methylation to measure X‐inactivation, but aberrant methylation is known to occur in some situations. We have developed a non‐radioactive reversetranscription polymerase chain reaction (RT‐PCR) method to study expression of the polymorphism at nt.1311 of the G6PD gene at the RNA level. Using this, and a similar method for the iduronate‐2‐sulfatase (IDS) gene, we have re‐evaluated X‐inactivation in AA patients. 32/35 normal individuals showed polyclonal haemopoiesis. Patients with presumed clonal diseases showed both monoclonal and polyclonal patterns, consistent with previous reports. Overall, clonal patterns were observed in granulocytes of 10/26 AA patients (38%), a significantly higher proportion than in controls (p<0.01). Two cases showed discordance between lymphocytes and granulocytes, indicating clonality arising within the myeloid lineage. Eight cases showed clonal patterns in both myeloid and lymphoid cells, indicating the involvement of a pluripotent stem cell. Clonal patterns did not correlate with age, but there appeared to be an association with duration of disease. In PNH patients, CD59‐negative cells showed clonal patterns of X‐inactivation. In two cases, however, clonal patterns were also detected in CD59‐positive cells.
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