Cytotoxicity of a GalNAc-specific C-type lectin CEL-I toward various cell lines.
We found that CEL-I was a potent cytotoxic lectin. MDCK, HeLa, and XC cells were highly sensitive to CEL-I cytotoxicity and killed in a dose-dependent manner, whereas CHO, L929, and RAW264.7 cells were relatively resistant to CEL-I, and no significant toxicity was observed up to 10 microg/ml. Among these cell lines, MDCK cells showed the highest susceptibility to CEL-I cytotoxicity. A binding study using FITC-labeled CEL-I (F-CEL-I) revealed that the amounts of bound F-CEL-I on the sensitive cell lines were evidently greater than those on the resistant cell lines, suggesting that the different susceptibility of the cell lines to CEL-I cytotoxicity is partly explained by different efficiencies of binding of CEL-I to these cell lines. Interestingly, the cytotoxicity of CEL-I toward MDCK cells was more potent than those of other lectins such as WGA, PHA-L, and Con A, even though these lectins were capable of binding to MDCK cells at comparable levels to CEL-I. Since the cytotoxicity of CEL-I was strongly inhibited by GalNAc, the binding to cell surface specific carbohydrates is essential for the CEL-I cytotoxicity. The trypan blue dye exclusion test indicated that CEL-I caused a disorder of plasma membrane integrity as a relatively early event. CEL-I failed to induce the release of carboxyfluorescein (CF) from CF-loaded MDCK cells as seen for pore-forming hemolytic isolectin CEL-III, suggesting that the primary cellular target of CEL-I may be the plasma membrane, but its action mechanism differs from that of CEL-III. Although CEL-I induced dramatic cellular morphological changes in MDCK cells, neither typical apoptotic nuclear morphological changes nor DNA fragmentation was observed in CEL-I-treated MDCK cells even after such cellular changes. Our results demonstrated that CEL-I showed a potent cytotoxic effect, especially on MDCK cells, by causing plasma membrane disorder without induction of apoptosis.
Kuramoto
Takuya
T
Division of Biochemistry, Faculty of Fisheries, and Department of Applied Chemistry, Faculty of Engineering, Nagasaki University, 1-14 Bunkyo-machi, Nagasaki 852-8521, Japan.
Uzuyama
Hitomi
H
Hatakeyama
Tomomitsu
T
Tamura
Tadashi
T
Nakashima
Takuji
T
Yamaguchi
Kenichi
K
Oda
Tatsuya
T
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D048016:Cucumaria / Q000737:chemistry*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D037181:Lectins, C-Type / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice; D051381:Rats
D037181:Lectins, C-Type