Flow-cytometry Assessment of DNA content Analysis and Immunophenotyping of Immune-cells in Lymph-node-specimens as Potential Diagnostic Signature of Aggressiveness in B-Non-Hodgkin Lymphomas

Introduction: Flow-cytometry (FC) is a powerful tool that can assist in lymphoma diagnosis in lymph node (LN) specimens. Although lymphoma diagnosis and classification are mainly based on tumor cell characteristics, surrounding cells are less employed in this process. Methods: We retrospectively investigated alterations in the ploidy status, proliferative cell fraction (PF) and the percentages of surrounding immune cells in 62 consecutive LN specimens with B-Cell Non-Hodgkin Lymphoma (B-NHL) that were submitted for FC evaluation between 2019-2022. Results: Compared with indolent B-NHLs, aggressive B-NHLs show increased DNA aneuploidy and PF, increased monocytes, immature-granulocytes, mature granulocytes, CD8+ T-cells, Double-Negative-T-cells and Double-Positive-T-cells, and decreased total CD45+ cells, total lymphocytes, CD4+ T-cells and CD4/CD8 ratio. Receiver operating characteristic analysis determined PF > 6.8% and immature-granulocytes > 0.9% as optimal cutoffs with highest specificity and sensitivity in differentiating aggressive and indolent B-NHLs. Conclusions: These findings further strength the diagnostic value of DNA content analysis by FC and suggest the utilization of tumor surrounding immune cells in NHL diagnosis and classification.