Conventional cytogenetics and FISH evaluation of chimerism after sex-mismatched bone marrow transplantation (BMT) and donor leukocyte infusion (DLI).
OBJECTIVE
Sensitive and quantitative cytogenetic methods to better assess the biological significance of post-BMT chimerism have been recently developed. In this study, we compared the results of chimerism analysis and evolution employing conventional cytogenetics and fluorescence in situ hybridization (FISH) in 16 patients after sex-mismatched BMT, and in 5 patients after donor lymphocyte infusion (DLI) to treat post-BMT relapse.


METHODS
FISH studies were performed using separate digoxigenin labeled centromeric DNA probes for the X (pDMX1) and Y (DYZ1/DYZ3) chromosomes. To this purpose, different types of samples were used: bone marrow (BM) and peripheral blood (PB) slides processed for conventional cytogenetics, and routine BM and PB smears.


RESULTS
Results of chimerism studies performed on different types of samples showed no significant differences. No significant differences in the ability to identify the sex of each cell with both pDMX1 and DYZ1/DYZ3 probes were found and the results obtained from independent experiments showed a high linear correlation. Chimerism analysis by FISH showed initial mixed chimerism after BMT in 10 patients. Seven of these patients were also studied by conventional cytogenetics and 2 of these showed mixed chimerism. Seven of the former 10 patients evolved to complete donor chimera. 6 patients showed cytogenetic or hematologic bone marrow relapse, 3 of which were preceded by mixed chimaerism as revealed by FISH studies. FISH studies permitted an easy and accurate monitorization of the response to DLI in 5 relapsed patients, showing an increase in the proportion of donor cells in 4 patients as they reached a new complete remission.


CONCLUSIONS
Both FISH and conventional cytogenetics are quantitative methods to assess chimerism. However, FISH is more sensitive, accurate and can even be applied on routine BM and PB smears. Furthermore, its combination with immunophenotyping approaches to quantify chimerism on cell subpopulations, will help to clarify post-BMT chimerism significance.
Díez-Martín
J L
JL
Department of Hematology, Clínica Puerta de Hierro, Madrid, Spain. jldiez@radonc.hggm.es
Llamas
P
P
Gosálvez
J
J
López-Fernández
C
C
Polo
N
N
de la Fuente
M S
MS
Buño
I
I
D000293:Adolescent; D000328:Adult; D001782:Blood Donors; D016026:Bone Marrow Transplantation; D002648:Child; D002675:Child, Preschool; D005260:Female; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007621:Karyotyping; D017708:Leukocyte Transfusion; D008297:Male; D018183:Transplantation Chimera