Tumor-infiltrating macrophages influence the glycosphingolipid composition of murine brain tumors

Abstract

A procedure was developed to analyze glycosphingolipids (GSLs) in tumor-infiltrating macrophages (TIMs). The procedure entailed dissociating tumors into single cell suspensions with a concurrent metabolic labeling of GSLs using [ 14 C]galactose. TIMs were then separated from tumor cells and other host cells by magnetic activated cell sorting and CD11b (Mac-1) microbeads. Gangliosides and neutral glycosphingolipids were analyzed in the TIMenriched and TIM-depleted fractions in two different murine brain tumors (EPEN and CT-2A). The TIM gangliosides consisted of over 30 structures as assessed by twodimensional high performance thin-layer chromatography. GSLs enriched in TIMs, relative to the tumors, included Gg4Cer (asialo GM1), GM1b, and GD1 ␣ . TIM GSLs were similar in EPEN and CT-2A despite their differences in growth and morphology. TIM GSLs were similar whether TIMs were isolated from tumors grown intracranially or subcutaneously. TIM GSLs were also similar to activated peritoneal macrophage GSLs, although differences in the ceramide structure were observed. Knowledge of TIM GSLs will be important in determining the function of these molecules in macrophage-tumor interactions. In addition, these methods will be helpful in determining the cellular origin of human brain tumor GSLs and in identifying tumorassociated GSLs for immunotherapy.-Ecsedy, J. A., H. C. Yohe, A. J. Bergeron, and T. N. Seyfried. Tumor-infiltrating macrophages influence the glycosphingolipid composition of murine brain tumors.

Increased glycosphingolipid levels in serum and aortae of apolipoprotein E gene knockout mice

“…Analogous observations have been made where tumor-infiltrating macrophages were found to alter dramatically the ganglioside composition of murine brain (57). Of possible significance, murine macrophages have the capacity to produce a spectrum of gangliosides, including glycolyl GM2, GM1b, and GD1a (57,58). It is also likely that lipoproteinderived glycolyl GM2 contributes to the increased ganglioside levels observed in the atherosclerotic lesion (20).…”

“…Because there is little evidence that vessel wall cells constitutively synthesize glycolyl neuraminic acid, and because this ganglioside is only present in trace amounts in healthy aortae, one interpretation is that the presence of glycolyl GM2 in the diseased tissue may reflect the GSL composition of macrophage foam cells. Analogous observations have been made where tumor-infiltrating macrophages were found to alter dramatically the ganglioside composition of murine brain (57). Of possible significance, murine macrophages have the capacity to produce a spectrum of gangliosides, including glycolyl GM2, GM1b, and GD1a (57,58).…”

“…Infiltrating monocytes that have differentiated to macrophages could utilize both lipoprotein-derived and locally synthesized GSLs to form new gangliosides. An example of the latter could be the formation of GD1a from GM3 via the following pathway: GM3¨GM2¨GM1a¨GD1a (57,58).…”

Personalized Tumor RNA Loaded Lipid-Nanoparticles Prime the Systemic and Intratumoral Milieu for Response to Cancer Immunotherapy

“…16 B16F0 and LLC were purchased from ATCC. CT-2A tumors were originally generated as previously described 25 and maintained in culture. Culture media utilized to grow tumor cells consisted of DMEM with sodium pyruvate (KR158B-luc and CT-2A were grown without sodium pyruvate), supplemented with 10% fetal bovine serum (FBS), and 5.5 mL of penicillin/streptomycin.…”