The tumor cells in classic Hodgkin lymphoma(HL) are surrounded by CD4+ T-cell rosettes that provide support and form a physical shield against anti-tumor immune responses. To study the mechanisms behind rosette formation, we previously developed an in vitro co-culture model and found that CD2-CD58 interactions increase rosetting and activation of CD4+ T cells. In the current study, we explore the role of CD59, an alternative CD2 ligand, in rosetting. In 45/62 diagnostic cases, tumor cells showed increased expression of CD59. HL cell lines L428(CD58+/CD59+), L1236(CD58+/CD59-) and KMH2(CD58-/CD59+) and peripheral blood mononuclear cells (PBMCs) from healthy donors were co-cultured. After 30 minutes, L428 showed the highest percentage of rosette formation, followed by L1236 and KMH2, respectively. Accordingly, the production of IL-2 after 24 hours, as a marker of T cell activation, showed the same pattern. CD59 knockout in L428 reduced the rosette formation, strongly indicating that CD59 expression is involved. During rosette formation, CD58 firstly redistributed along the HRS cell membrane towards the T cell interaction site, and CD59 followed CD58 to the same sites within 30 minutes. Proximity ligation assays showed that the CD58-CD59 interaction was only present at the interface between tumor cells and T cells. There were more CD2-CD58 signals in L428 than in L1236, while CD2-CD59 signals were present in L428 and missing in KMH2. In conclusion, CD59 expression is upregulated by tumor cells and helps to enhance the interaction between CD58 and CD2 during the formation of rosettes in HL.
Yajie Lei, Rodrigo Martinez Alcala, Annika Seitz, Lydia Visser, Arjan Diepstra