Heterogeneous expression of B cell‐associated markers in follicular lymphoma

Sir: The conventional view of follicular lymphoma is that it comprises a relatively uniform proliferation of germinal centre cells with little heterogeneity in phenotype. Immunostaining for B cell-associated markers such as CD20 and CD79a shows a very similar labelling pattern and follicles are usually stained homogeneously. An exception is the expression of BCL2 protein, where in a minority of cases it has been found to be expressed heterogeneously. We studied the immunohistochemical expression of eight B cell-associated molecules in 10 t(14;18)positive follicular lymphomas. They included four intracellular signal transduction molecules (Syk, BLNK, PLC-c2 and LCK), the B cell activating factor-receptor (BAFF-R), a B cell-associated transcription factor (Oct2) and two pan-B cell markers (CD37 and CD19). The t(14;18) chromosomal translocations were investigated by fluorescence in situ hybridization. Immunostaining was assessed together with CD20 labelling, which gave an accurate estimate of the number of neoplastic cells present. BAFF-R, BLNK, CD19, LCK, Oct2 and Syk were expressed heterogeneously and labelling varied between cases, ranging from negative to strongly positive. In addition, cell-tocell variation was noted within individual cases, and this was particularly striking for CD19, LCK, Oct2 and Syk (Figure 1A). The expression of BAFF-R, BLNK, CD19 and Oct2 was confined to the neoplastic follicles, with very few positive cells in the interfollicular region (Figure 1B). BLNK was expressed strongly in both follicles and in mantle zone cells (Figure 1C), but labelling in the interfollicular region was generally weak. In addition, immunostaining revealed an additional unexpected pattern of expression, as scattered cells, some of which aggregated around high endothelial venules, were strongly positive. B cells were not present in these areas. These cells had a moderately abundant cytoplasm with round to oval nuclei similar to plasmacytoid dendritic cells ⁄ monocytes (Figure 1C). Double immunofluorescence labelling for BLNK and CD68 confirmed their coexpression (Figure 1C). Another interesting observation is the expression of Oct2, where staining was generally restricted to the neoplastic follicles (Figure 2A). In three cases, the positive cells were located frequently at the periphery of the neoplastic follicles, where the cells were confirmed to be proliferating (Figure 2B). It was also interesting that in one case in which the biopsy also contained a number of reactive follicles, the expression of Oct2 was strong in reactive germinal centres but was down-regulated in the neoplastic follicles (Figure 2C). It was also clear in this case that its expression was related inversely to BCL2, which is in contrast to a previous report which suggested that BCL2 protein expression is up-regulated by Oct2. The staining for CD37 and PLCc2 did not show any significant heterogeneity or down-regulation in the interfollicular cells. A summary of these findings is shown schematically in Figure 2C. This report throws new light on the molecular heterogeneity of follicular lymphoma. A number of molecules that are expressed in a homogeneous fashion by normal B cells ⁄ germinal centre cells (CD19, LCK, Oct2 and Syk) were often expressed heterogeneously in follicular lymphoma. In addition, the expression of BAFF-R, BLNK, CD19 and Oct2 tended to be confined to the neoplastic follicles, with very few positive cells in the interfollicular region. In the past, Dogan et al. showed that neoplastic B cells in the interfollicular region are phenotypically different from those in the follicles despite their shared clonal origin. They noted that germinal centre-associated markers CD10 and CD38 and the CD95 ⁄ Fas molecule were down-regulated in the interfollicular region. It can be concluded that the tumour cells in follicular lymphoma are not as homogeneous as is generally thought. Secondly, the down-regulation of several B cell-associated molecules in the extrafollicular neoplastic cells in this report reinforces the findings of Dogan et al. that these cells are phenotypically different from the follicular component of the neoplasm.