[New diagnostic approaches in malignant B-cell lymphomas]. / Nouvelles approches diagnostiques des lymphomes malins B.

The new WHO classification of B-cell neoplasms published in 2001 gathers leukaemia and lymphoma. This classification highlights the stage of differentiation of the tumour cell. It groups B-cell neoplasms morphology, histology (nodular or diffuse pattern, cell morphology), immunophenotypes, cytogenetic, molecular abnormalities and clinical data. Recent advances of the normal B cell differentiation understanding and the development of new types of analysis have shown that mature B-cell neoplasms appear to recapitulate stages of normal B-cell differentiation, leading to such a new classification.

Morphometric and colorimetric analysis of peripheral blood smears lymphocytes in B-cell disorders: proposal for a scoring system.

Distinguishing leukemic phases of B-cell disorders in peripheral blood smears is well recognized to be difficult in some cases since it depends on subtle and subjective criteria. In order to quantify cytological features and to assess objective descriptions, a morphometric analysis was performed on 83 peripheral blood smears of B-cells disorders (n = 77) and healthy donors (n = 6). Using standardized May-Grunwald Giemsa staining, standardized image acquisition system and well defined microscopic fields, we have analyzed lymphoid cells, measuring morphometric and color parameters. By combining seven relevant morphometric criteria (the nuclear shape, the cellular shape and area, the nucleo-cytoplasmic ratio, the nuclear red/blue ratio, the cytoplasmic green/blue ratio and the proportion of cells with nucleolus), we have established a score that could range from a minimum of -3 (large B-CLL type) to a maximum of +8 (large MCL type): negative scores corresponds to different types of B-CLL (n = 30), including "atypical B-CLL" (n = 6), the score zero correspond to healthy donors (n = 6) used as baseline, the positive score values correspond to +1 for Follicular lymphoma (n = 2), +3 for Splenic Lymphoma with Villous Lymphocytes (n = 12), +4 for Hairy Cell Leukemia (n = 7), for Hairy Cell Leukemia-variant (n = 2), +6 for B-prolymphocytic leukemia (n = 6) and +7 and +8 for most Mantle Cell Lymphoma (n = 18). Testing T-cell disorders samples (n = 10) using the same protocol, the profile is different and cannot be confused with B-cell diseases. Our scoring system indicates that measurement of some common morphologic features in standardized conditions provides objective criteria to characterize those diseases and might be helpful for diagnosis.

A new morphologic classification system for acute promyelocytic leukemia distinguishes cases with underlying PLZF/RARA gene rearrangements.

Acute promyelocytic leukemia (APL) is typified by the t(15;17) translocation, which leads to the formation of the PML/RARA fusion gene and predicts a beneficial response to retinoids. However, approximately 10% of all APL cases lack the classic t(15;17). This group includes (1) cases with cryptic PML/RARA gene rearrangements and t(5;17) that leads to the NPM/RARA fusion gene, which are retinoid-responsive, and (2) cases with t(11;17)(q23;q21) that are associated with the PLZF/RARA fusion gene, which are retinoid-resistant. A key issue is how to rapidly distinguish subtypes of APL that demand distinct treatment approaches. To address this issue, a European workshop was held in Monza, Italy, during June 1997, and a morphologic, immunophenotypic, cytogenetic, and molecular review was undertaken in 60 cases of APL lacking t(15;17). This process led to the development of a novel morphologic classification system that takes into account the major nuclear and cytoplasmic features of APL. There were no major differences observed in morphology or immunophenotype between cases with the classic t(15;17) and those with the cryptic PML/RARA gene rearrangements. Auer rods were absent in the t(5;17) case expressing NPM/RARA. Interestingly, this classification system distinguished 9 cases with t(11;17)(q23;q21) and, in addition, successfully identified 2 cases lacking t(11;17), which were subsequently shown to have underlying PLZF/RARA fusions. The PLZF/RARA cases were characterized by a predominance of blasts with regular nuclei, an increased number of Pelger-like cells, and by expression of CD56 in 4 of 6 cases tested. Use of this classification system, combined with an analysis for CD56 expression, should allow early recognition of APL cases requiring tailored molecular investigations. (Blood. 2000;96:1287-1296)

Morphometry and quality control for a May-Grunwald Giemsa stained preparation. A 40 centers cooperative study.

A multicentric study including 40 medical centers has been done on May-Grunwald Giemsa (MGG) stain, to obtain precise information concerning the size, shape and color of cellular elements. A number of factors are taken into consideration including the zone of microscopic observation, the quality of staining, condition of grabbing images and the size of the sample. Morphometric and colorimetric measurements have been applied as objective tools for demonstrating the predominance of each Red/Green/Blue component, and more particularly their reciprocal ratio. The results obtained for each center may constitute a guideline for adjusting their procedure, in order to reach a consensus regarding similar staining by all centers.

Comparison of the classical manual pushed wedge films, with an improved automated method for making blood smears.

Comparison of apparatus for automatically spreading peripheral blood films (GENES) to manual wedge-pull technique has been performed, showing that the average size of optimal area for counting is twice larger in automated smears compared to manual ones. The variability of WBC repartition has been studied in doing differential count on manual and automated smears and both are compared to results obtain with an independent technique (Coulter STKS) for differential. The correlation coefficients for each WBC population show a closer relationship between automated smear preparations and reference instrument (STKS) than with manual spreading smears. Monocyte differential is particularly influenced by the variability of manual spreading procedures.