[Chiral selectivity in differentiation of lung cancer A549 cell to vascular endothelial cells after drug resistance induced by D-or L-methotrexate enantiomers].
OBJECTIVE
To study the chiral selectivity in vascular endothelial differentiation in drug resistant lung cancer cells induced by high-dose L- or D-methotrexate (MTX) enantiomer.


METHODS
Human lung cancer cells of the line A549 were co-cultured with high-dose (15 micromol/L) L- or D-MTX enantiomer so as to develop cancer cells resistant to MTX. MTT method was used to detect the drug resistant index. Flow cytometry was used to detect the expression of CD44, a transmembrane glycoprotein reflecting the migration ability of cells, CD31, a marker of vascular endothelium, and P-170 protein. Fifteen BALB/c nude mice were inoculated with the parent A549 cells, L-MTX-resistant A549 cells induced by L-MTX enantiomer, and D-MTX-resistant A549 cells induced by D-MTX enantiomer. Four weeks later the mice were killed to take out the tumor masses. Immunohistochemistry with CD34 staining was used to detect the microvascular density (MVD).


RESULTS
The drug resistant index of the D-MTX induced drug resistant A549 cells was 20.1 +/- 2.3, significantly higher than that of the L-MTX-induced cells (12.4 +/- 1.2, P = 0.000). The CD44 positive rate of the D-MTX induced A549 cells was 97.0% +/- 0.9%, not significantly different from that of the L-MTX-induced A549 cells (96.7% +/- 1.4%, P = 0.544). The CD31 positive rate of the D-MTX induced A549 cells was 91.9% +/- 3.2%, significantly higher than that of the L-MTX-induced A549 cells (32.9% +/- 8.0%, P = 0.000). The P-170 protein positive rate of the parent cells was 85.5% +/- 4.6%, and the P-170 protein positive rate of the D-MTX-induced A549 cells was 87.0% +/- 8.9%, significantly higher than that of the L-MTX-induced cells (71.5% +/- 8.2%, P = 0.002). The MVD of the D-MTX-induced cells was 55.9 +/- 11.9, significantly higher than that of the L-MTX-induced cells (7.2 +/- 1.7, P = 0.000). MVD was significantly positively correlated with the CD31 level (r = 0.462, P = 0.007), and not correlated with P-170 protein and CD34 levels.


CONCLUSIONS
The MTX enantiomers have different chiral selectivity on human lung cancer cell, D-MTX resistant cells shows a potential of differentiation from cancer cells to vascular endothelial cells. D-MTX is not be regarded just as a pollutant in the drug MTX, MTX with single enantiomer (L-MTX) should be selected clinically so as to decrease the side effects of D-MTX.
He
Xiao-Dong
XD
Anhui Provincial Center for Clinical Laboratories, Anhui Provincial Hospital Affiliated to Anhui Medical University, Hefei 230001, China.
Hu
Shi-Lian
SL
Shen
Zuo-Jun
ZJ
Tao
Shao-Neng
SN
Dong
Lin
L
Zhu
Yuan-Yuan
YY
Li
Ming
M
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019008:Drug Resistance, Neoplasm; D042783:Endothelial Cells / Q000166:cytology*; D006801:Humans; D008727:Methotrexate / Q000737:chemistry / Q000494:pharmacology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude
D008727:Methotrexate