Panoply™ Human CD33 Knockdown Stable Cell Line

3'-sialyllactose is one of the abundant components of human milk oligosaccharides (HMOs) that protect infants from various viral infections during the early stages of immune system development. 3SL is a combination of lactose and sialic acid. Most sialic acids are widely expressed in animal cells and bind to siglec proteins. Here, researchers demonstrated that 3SL specifically binds to CD33. It induces megakaryocyte differentiation of human chronic myeloid leukemia K562 cells by targeting the cell surface protein siglec-3 (CD33) and subsequently causes apoptosis. CD33 bound to 3SL is internalized into the cytosol via vesicle-dependent endocytosis. At the molecular level, 3SL-bound CD33 recruits suppressor of cytokine signaling 3 (SOCS3) and SH2 domain-containing protein tyrosine phosphatase 1 (SHP1). SOCS3 is degraded along with CD33 via the proteasome, while SHP-1 activates extracellular signal-regulated kinase (ERK), inducing megakaryocyte differentiation and subsequent apoptosis. Thus, these studies suggest that 3SL is a potential anti-leukemic agent that can affect both differentiation and apoptosis.

To further explore whether CD33 mediates 3SL-induced megakaryocyte differentiation and apoptosis, the researchers studied HEK293 cells, a cell line derived from human embryonic kidney, which is known to be negative for CD33 expression. HEK293 cells were treated with 3SL for 24 hours, and as shown in Figure 1A, cell viability was not changed, but apoptotic markers were reduced (Figure 1B and C). Comparing the results of HEK293 cells with those of K562 cells, it was found that the expression levels of apoptotic markers were reduced in HEK293 cells, but not in K562 cells. Although there were many other differences between HEK293 cells, the results can infer that 3SL-induced apoptosis occurs through CD33. In addition,  knockdown of CD33 expression in K562 cells inhibited 3SL-induced endocytosis (Figure 1D), megakaryocyte differentiation (Figure 1E), and apoptosis (Figure 1F and 1G). These results indicate that CD33 is a key cellular protein and 3SL receptor involved in 3SL-induced megakaryocyte differentiation and apoptosis.

Figure 1. Knockdown of CD33 gene suppresses internalization of CD33 and reduces 3′-sialyllactose induced-megakaryocyte differentiation and apoptosis. (Ha S H, et al., 2020)