NK Cell-Depleting Anti-Asialo GM1 Antibody Exhibits a Lethal Off-Target Effect on Basophils In Vivo

Abstract

NK cells are innate immune lymphocytes and play a key role in both innate and adaptive immunity. Their pivotal functions in vivo have been illustrated in mice by means of their ablation with NK cell-depleting Abs, particularly anti-asialo GM1 (ASGM1). In this study, we show that the whole population of basophils constitutively expresses ASGM1 as well as CD49b (DX5) as does the NK cell population and was ablated in vivo by anti-ASGM1 as efficiently as by a basophil-depleting anti-FcεRIα Ab. Anti-ASGM1–mediated basophil depletion was operative as for NK cell depletion in various mouse strains, irrespective of NK1 allotype and MHC H2 haplotype, including C57BL/6, BALB/c, C3H, and A/J mice. These results identified basophils as a previously unrecognized target of anti-ASGM1–mediated cell depletion and raised concern about possible contribution of basophils, rather than or in addition to NK cells, to some of phenotypes observed in anti-ASGM1–treated mice. Indeed, regardless of the presence or absence of NK cells in mice, anti-ASGM1 treatment abolished the development of IgE-mediated chronic cutaneous allergic inflammation as efficiently as did the treatment with basophil-depleting Ab. Given the fact that basophils have recently been shown to play crucial roles in a variety of immune responses, our finding of the off-target effect on basophils issues a grave warning about the use of anti-ASGM1 and underscores the need for careful interpretation of phenotypes observed in anti-ASGM1–treated mice.

Natural Killer (NK) Cells in Antibacterial Innate Immunity: Angels or Devils?

“…Divergent results are similarly obtained in vivo depending on the antibody used for NK cell depletion (42). For example, in vivo cell depletion with anti-NK1.1 antibodies leads to the elimination of both NK and NKT cells, whereas the use of the anti-asialo monosialotetrahexosylganglioside (GM1) antibody depletes NK cells and basophils (43). Thus, the purification method (and possible cellular activation) may indeed contribute to the controversial reports published when studying the expression of TLRs in NK cells.…”

Conditional ablation of NKp46+ cells using a novel Ncr1greenCre mouse strain: NK cells are essential for protection against pulmonary B16 metastases

“…Furthermore, studying the role of NK cells in immune responses often involves the use of genetargeted mice lacking NK cells that also harbor other lymphoid defects (e.g. Rag2 −/− Il2rg −/− mice [11] or IL-15 −/− mice [12]), or depletion of NK cells using antibodies that are not NK-cell specific, such as anti-NK1.1 (expressed by NKT cells) or anti-asialo-GM1 (expressed by myeloid cells) [13,14].In order to study gene functions specifically in NKp46 + cells, we and others [15,16] have created mice that allow for conditional gene targeting in Ncr1 expressing cells using the Cre-loxP system [17]. To this end, we generated Ncr1 greenCre mice expressing an EGFPcre fusion under the control of a proximal C57BL/6 Ncr1 promoter fragment.…”

“…Furthermore, studying the role of NK cells in immune responses often involves the use of genetargeted mice lacking NK cells that also harbor other lymphoid defects (e.g. Rag2 −/− Il2rg −/− mice [11] or IL-15 −/− mice [12]), or depletion of NK cells using antibodies that are not NK-cell specific, such as anti-NK1.1 (expressed by NKT cells) or anti-asialo-GM1 (expressed by myeloid cells) [13,14].…”

Natural Killer Cell Reduction and Uteroplacental Vasculopathy

“…This antiserum reduces NK cells in almost every organ in the body, including spleen 16,47 rather than just uNK cells. Moreover, anti-asialo GM1 also reacts with nonparenchymal liver cells, 48 T cells in mice 49 and rats, 50 as well as murine basophils 51 and monocytes 52 and rat granulocytes and macrophages. 50 The effect on various ILC subsets is poorly described.…”