Characterization of Immune Cell Infiltration and Collagen Type III Disorganization in Human Secondary Lymphedema: A Case-control Study

Background: Secondary lymphedema (SL) affects 120 million people globally, posing a lifelong burden for up to 37% of cancer survivors. Chronic inflammation and progressive fibrosis are key drivers of SL, yet detailed characterization of immune cell subpopulations across lymphedema stages is lacking. This study aimed to investigate the immunologic profile of lymphedematous skin and its association with extracellular matrix changes, which could serve as clinical biomarkers or therapeutic targets. Methods: This case-control study analyzed the skin from 36 patients with and without SL, using immunofluorescence to quantify T cells, B cells, macrophages, and their subpopulations. Collagen quantity and composition were examined using picrosirius red staining, and mast cell infiltration was assessed with toluidine blue staining. Early and late SL stages were compared to identify histomorphological and immunologic correlates of stage progression. Results: We found a predominance of CD4+ T cells and mast cells in SL skin (1.4/ mm² versus 1.0/mm^2, P < 0.01; 1.2/mm² versus 0.2/mm^2, P < 0.0001) and a higher ratio of collagen III to collagen I fibers (51.6% versus 75.0%, P < 0.001). M2 macrophages were more abundant in late-stage than in early-stage lymphedema (1.7/ mm² versus 1.0/mm^2, P = 0.02). Conclusions: This study demonstrated a shift toward CD4+ T cell and mast cell infiltration in SL skin, correlating with extracellular matrix disorganization and an altered collagen III/I ratio. These findings enhance our understanding of the cellular and morphological changes in SL, potentially guiding future diagnostic and therapeutic strategies.