Contribution of trypsin-sensitive proteins to binding of cationic liposomes to the mouse macrophage-like cell line RAW264.7.
We studied the binding of cationic liposomes, including didodecyl N-(alpha-(trimethylammonio)acetyl)-D-glutamate chloride (TMAG), to a mouse macrophage-like cell line RAW264.7 to clarify which molecules contribute to the binding of TMAG liposomes to the cell surface. Several types of TMAG liposomes encapsulating [3H]inulin, intra-aqueous markers of liposomes, were prepared and their binding characteristics were compared with those of neutral and negatively charged liposomes. The binding of TMAG liposomes to cells was superior to those of neutral and negatively charged liposomes and increased with increasing TMAG content. Scatchard plots for the binding of TMAG liposomes to the cells were approximately linear, indicating a single class of binding sites. Pretreatment of the cell surface with heparinase, heparitiase, chondroitinase ABC, or neuraminidase did not reduce the binding of TMAG liposomes. These results suggested that neuraminic acid and glycosaminoglycan on the cell surface have little contribution to TMAG liposome binding. Pretreatment of the cells with trypsin reduced the binding of TMAG liposomes in a concentration-dependent manner but did not detach the cells from the culture plates. In addition, alpha-chymotrypsin pretreatment had no effect even up to 5 micrograms/mL. Post-treatment with trypsin enhanced the release of TMAG liposomes from the cell surface in a concentration-dependent manner. These results demonstrated that TMAG liposomes bind to trypsin-sensitive proteins on the cell surface.
Arima
H
H
School of Pharmacy, Tokyo University of Pharmacy and Life Science, Japan.
Aramaki
Y
Y
Tsuchiya
S
S
D000818:Animals; D002460:Cell Line; D002808:Chondroitin Lyases / Q000378:metabolism; D002918:Chymotrypsin / Q000378:metabolism; D005971:Glutamates / Q000378:metabolism*; D019763:Heparin Lyase; D007444:Inulin / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D009439:Neuraminidase / Q000378:metabolism; D011133:Polysaccharide-Lyases / Q000378:metabolism; D011485:Protein Binding; D014357:Trypsin / Q000378:metabolism*
D005971:Glutamates; D008081:Liposomes; D008565:Membrane Proteins; C084512:didodecyl N-(alpha-trimethylammonioacetyl)glutamate; D007444:Inulin; D009439:Neuraminidase; D002918:Chymotrypsin; D014357:Trypsin; D002808:Chondroitin Lyases; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase; C021349:heparitinsulfate lyase