CLONALITY ANALYSIS OF HEMATOPOIESIS IN ESSENTIAL THROMBOCYTHEMIA - ADVANTAGES OF STUDYING T-LYMPHOCYTES AND PLATELETS

Essential thrombocythemia (ET) is a myeloproliferative disorder charac terized by a sustained elevation of the platelet count in the absence of other causes of thrombocytosis, Ef is difficult to diagnose, and th e demonstration of clonal hematopoiesis may be of value. However, clon ality analysis of hematopoietic cells based on the study of the X-chro mosome inactivation pattern is complicated by the observation that som e normal females present skewed lyonization. Moreover, DNA methylation of X-linked genes in hematopoietic cells may differ from that in othe r tissues. Appropriate controls for skewed lyonization are therefore c ritical for the study of clonality. We developed two techniques based on X-chromosome inactivation and polymerase chain reaction (PCR) analy sis of polymorphisms, to study clonality in ET patients. Reverse trans criptase-PCR analysis of IDS, P55, and G6PD mRNAs was used to examine the different hematopoietic cell lineages including platelets in patie nts heterozygous for these polymorphisms and analysis of the HUMARA ge ne methylation pattern permitted us to study clonality in all nucleate d cell fractions of the other patients. Using both types of assay and T lymphocytes as a control tissue for lyonization, clonal hematopoiesi s was demonstrated in 28 patients. In 14 patients, the granulocytes we re polyclonal; among these patients, platelets were monoclonal in 3 ca ses, polyclonal in 7 cases, and in the remaining 4 cases this fraction could not be studied because the patients were homozygotes for all RN A markers, Mo conclusion about clonality could be drawn in 6 cases, Po lyclonal hematopoiesis was found in all the cases of reactive thromboc ytosis. These findings confirm the high frequency of monoclonal hemato poiesis in ET, the utility of studying platelets, and the possibility of using T lymphocytes as a control tissue for X-chromosome inactivati on patterns. (C) 1997 by The American Society of Hematology.