Hematogones are identified by 4-color flow cytometry in most bone marrow specimens. They are more commonly found and are generally present in higher numbers in children. There is a general decline in hematogones with increasing age but a broad range exists at all ages and marrow from some adults contains relatively high numbers. They are often increased (> 5%) in regenerating marrow and in some clinical conditions, particularly various types of cytopenias and neoplastic diseases. Hematogones may morphologically resemble the neoplastic lymphoblasts of precursor B ALL and their immunophenotype also has features in common with neoplastic lymphoblasts. Distinguishing hematogones from neoplastic lymphoblasts may be problematic in post-chemotherapy and post-bone marrow transplant regenerating marrow. With 4-color flow cytometry using optimal antibody combinations the distinction can nearly always be made. Hematogone populations always exhibit a continuous and complete maturation spectrum of antigen expression typical of the normal evolution of B-lineage precursors; they lack aberrant or asynchronous antigen expression. The neoplastic lymphoblasts in precursor B ALL deviate from the normal B-lineage maturation spectrum and exhibit maturation arrest and over-, under-, and asynchronous expression of antigens observed on normal B-cell precursors and they often aberrantly express myeloid-associated antigens.

The simian polyoma virus SV40 has been detected in specific human tumors including non-Hodgkin's lymphomas, although a causative role for the virus has not been convincingly demonstrated. Aberrant methylation of CpG islands in promoter regions is a frequent method of silencing tumor suppressor genes (TSGs) in cancers and may be induced by oncogenic viruses. We investigated the relationship between the presence of SV40 or EBV DNA sequences and the methylation profiles for 10 TSGs in 90 cases of non-Hodgkin's lymphomas/leukemias and 56 control tissues. SV40 sequences were present in 33/90 (37%) non-Hodgkin's lymphomas/leukemias, and EBV was present in 11/42 (26%) of non-Hodgkin's lymphomas. We found a highly significant correlation between the presence of SV40 and methylation of seven genes (P values, 0.006 to <0.0001). In lymphomas, there was no relationship between EBV and methylation. Oncogenic viruses and methylation were rarely present in control tissues. We investigated methylation of the same 10 TSGs in peripheral blood mononuclear cells (PBMC) from a healthy volunteer infected with EBV or EBV and SV40. Promoter methylation of CDH1 and CDH13 were noted in dual SV40-and EBV-infected PBMC, and these two genes were also highly significantly correlated to the presence of SV40 sequences in tumors. SV40 infection also resulted in appearance of the lymphoma/leukemia-specific marker, methylated SHP1. Methylation was completely absent in uninfected and EBV-infected PBMC. Our results demonstrate that the presence of SV40 in hematological malignancies is associated with promoter methylation of TSGs and that in all probability, the virus plays a role in tumor pathogenesis.

The presence of prominent proliferation centers (PCs) in lymph nodes (LNs) involved with chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) has been associated with atypical blood smear morphology. Atypical CLL has in turn been associated with variant immunophenotypes and poor outcome. However, the significance of abundant PCs remains controversial. We have analyzed the flow cytometric immunophenotypic features of 54 CLL/SLL LNs and correlated these findings with the morphologic and clinical features. The LN histology was assigned to one of two groups based on the prominence of PCs: Group I LNs contained scattered small, sometimes ill-defined PCs in a background of monotonous small round lymphocytes. Group II LNs had increased numbers and sizes of PCs resulting in an obviously nodular appearance at low magnification. Flow cytometry was performed using broad three- or four-color antibody panels that included anti-CD5, CD19, CD20, CD23, CD38, FMC7, and surface immunoglobulin (sIg). The intensity of expression of all markers was scored semi-quantitatively using isotypic controls and internal positive and negative populations as standards. There were 32 group I and 22 group II LNs that, by definition, expressed CD19, CD5, and CD23. Little variability was seen in the intensity of expression of CD19, and the majority of cases expressed CD23 brightly. CD5 varied from very dim to an intensity similar to that of normal T cells; the majority had an intermediate level of CD5 expression. FMC7 was expressed to a significant extent in 11 cases (21%). CD20 was relatively bright in 17 cases (32%). sIg was dim in 29 cases (55%) and moderate or bright in 24 cases (45%). CD38 was expressed significantly in 25 cases (47%). There was no correlation between histologic group and intensity of expression of any individual marker or with an immunophenotypic atypia score based on FMC7, CD20, and sIg. There was also no correlation between morphology or immunophenotype and clinical features. These findings do not support the interpretation that the prominence of proliferation centers in CLL/SLL LNs defines biologically distinct subtypes.

exclusively to the most mature maturational stage. These findings have implications for PB analysis of minimal residual disease in acute lymphoblastic leukaemia and follicular lymphoma.

Four patients presented with acute leukemia of ambiguous or myeloid lineage in association with Langerhans cell histiocytosis and provide evidence suggesting a common origin of the two neoplasms. One patient had a non-constitutional trisomy 21 in both the leukemic blasts and the Langerhans cells indicative of a clonal relationship. A second case expressed CD2, CD13, and CD117 on both the Langerhans cells and the blasts suggesting a possible clonal relationship. All four cases exhibited geographic intermingling of the Langerhans cell histiocytosis and acute leukemia and shared unique features including extramedullary leukemia involving lymph nodes in all cases with Langerhans cell histiocytosis only present in sites involved by acute leukemia. T-cell antigen expression was present in all cases with one meeting criteria for mixed phenotype acute leukemia, T/myeloid, not otherwise specified. These findings support the concept that coexistent Langerhans cell histiocytosis and acute leukemia is clonally related in some cases. Furthermore, these cases of acute myeloid or acute leukemia of ambiguous lineage with Langerhans cell histiocytosis share some unique features suggesting a common underlying neoplastic hematopoietic stem cell.