The expression of the histo-blood group carbohydrate structures T-nouvelle (Tn, CD175), sialylated Tn (CD175s) and the Thomsen-Friedenreich disaccharide (TF, CD176) on human leukemia cell lines was analyzed by their reactivity with specific monoclonal antibodies in flow cytometry, immunohistology and immunoprecipitation. Expression of sialylated CD176 was evaluated by comparative immunostaining with anti-CD176 antibodies before and after sialidase treatment. While only few cell lines expressed unmasked CD176, sialylated CD176 was present on all hematopoietic cell lines and native lymphocytes examined. CD175 and CD175s are preferentially expressed on erythroblastic leukemia cell lines. CD175s expression in these cells is consistent with the transcription of the gene encoding the key enzyme alpha2,6-sialyltransferase (hST6GalNAc1). The staining intensity was reduced after methanol pretreatment of cells, indicating that these glycans are partially expressed as constituents of glycosphingolipids. Immunoprecipitation and subsequent Western blotting revealed a series of distinct high molecular glycoproteins as carriers for these carbohydrate antigens. CD34 was identified as major carrier of CD176 by immunoprecipitation and microsequencing on a KG-1 subline enriched for CD176 expression. Incubation of several CD176-positive cell lines with anti-CD176 antibodies induced apoptosis of these cells, an effect not observed with anti-CD175/CD175s antibodies. Since the presence of naturally occurring anti-CD176 antibodies may represent a mechanism of immunosurveillance against CD176-positive tumor cells, we propose that sialylation of surface-expressed CD176--among other functions--protects against apoptosis.
(c) 2008 Wiley-Liss, Inc.