Comparative Analysis of O-glycans from Human Hepatocellular Carcinoma HepG2 and Normal Liver Cells L02†

doi:10.7503/cjcu20150062

Abstract

Abstract:

HepG2(a primary hepatocellular carcinoma cell line) and L02(ones derived from normal liver tissue) cells were chosen as model cell lines for research. The O-glycans of the total proteins extracted from HepG2 and L02 cells were released by Carlson reductive β-elimination. The released O-glycans previously purified by Dowex 50 WX8-400 cation exchange resin and C₁₈ cartridges were identified by electrospray ionization mass spectrometry(ESI-MS)and MS/MS. For comparision studies, β-cyclodextrin was used as the internal standard for relative quantitative analysis of the O-glycans derived from HepG2 and L02 cells by MS. As results, 10 O-glycans were observed in HepG2 cell line and 9 O-glycans were detected in L02 cell line. Moreover, 9 O-glycans were observed in both HepG2 and L02 cells, wherears 1 truncated O-glycan assigned as H1A1(NeuAc-GalNAc, sialyl Tn antigen, ubiquitous in cancer cells), was only found in HepG2 cells. t-Test results show that 5 and 2 O-glycans in HepG2 cells have significant differences(P<0.01 and P<0.05, recpectively), when compared to those of L02 cells. Our studies show methodological significance in structural investigation of O-glycans expressed in hepatocellular carcinoma and early biomarker discovery in clinical diagnose.

Key words: HepG2 cell, L02 cell, O-glycans, ESI-MS

CLC Number:

TrendMD:

PAN Liying, GU Xiao, WANG Chengjian, QIANG Shan, HUANG Linjuan, ZHANG Ying, WANG Zhongfu. Comparative Analysis of O-glycans from Human Hepatocellular Carcinoma HepG2 and Normal Liver Cells L02^†[J]. Chem. J. Chinese Universities, 2015, 36(7): 1275.

Figures/Tables 7

Fig.1 ESI-MS spectra of O-glycans derived from HepG2(A, C) and L02(B, D) cells under positive ion mode(A, B) and negative ion mode(C, D), respectively * Impurities.

Fig.2 MS/MS spectrum of O-glycans with m/z 1355.33(A) and the proposed structures(B)

Fig.3 MS/MS spectra(A, C) and proposed structures(B, D) of O-glycans with m/z 657.17(A,B) and m/z 699.25(C, D)

Fig.4 MS/MS spectrum(A) and proposed structure(B) of O-glycans with m/z 515.25

Fig.5 MS/MS spectra of O-glycans at m/z 1353(A) and its proposed structure(B)

Fig.6 Peak intensity ratio of O-glycans to β-cyclodextrin from HepG2 and L0210 O-glycans: a. H1N1; b. N1A1; c. H1N2; d. H1N1A1([M+H]+) and H1N1A1([M+Na]+); e. H2N2; f. H1N1A2; g. H2N2A1; h. H2N2A2; i. H1N1A1F1; j. H1N2A2F2. Significant difference: ** P<0.01; * P<0.05.

References 21

[1]	Brockhausen I., Biochim. Biophys. Acta(BBA)——Gen. Subj., 1999, 1473(1), 67—95
[2]	Wang Y. Y., Liu J. W., SIMA Z. H., Song H. P., Li R. L., Cai J. Z., Chen W. W., Chem. J. Chinese Universities, 2015, 36(2), 299—305
	(王一寓, 刘嘉炜, 司马贞华, 宋厚盼, 李茹柳, 蔡佳仲, 陈蔚文. 高等学校化学学报,2015, 36(2), 299—305)
[3]	Isailovic D., Kurulugama R. T., Plasencia M. D., Stokes S. T., Kyselova Z. K., Goldman R., Mechref Y., Novotny M. V., Clemmer D. E., J. Proteome Res., 2008, 7(3), 1109—1117
[4]	Bai C., Wu G., Zhao L., Sci. China Chem., 2010, 53(4), 807—811
[5]	Pan L. Y., Wang C. J., Yuan J. B., Zhang Y., Huang L. J., Wang Z. F., Chem. J. Chinese Universities, 2014, 35(2), 237—243
	(潘丽英, 王承健, 袁江北, 张英, 黄琳娟, 王仲孚. 高等学校化学学报,2014, 35(2), 237—243)
[6]	Brockhausen I., Schutzbach J., Kuhns W., Cells Tissues Organs., 1998, 161(1—4), 36—78
[7]	Schulenberg B., Beechem J. M., Patton W. F., J. Chromatography B, 2003, 793(1), 127—139
[8]	Blomme B., Van S. C., Callewaert N., J. Hepatology, 2009, 50(3), 592—603
[9]	Vanhooren V., Liu X. E., Franceschi C., Mech. Ageing. Dev., 2009, 130(1/2), 92—97
[10]	Liu X. E., Desmyter L., Gao C. F., Hepatology, 2009, 46(5), 1426—1435
[11]	Debruyne E. N., Vanderschaeghe D., Van V., Clin. Chem., 2010, 56(5), 823—831
[12]	Carlson D. M., J. Biol. Chem., 1968, 243, 616—626
[13]	Xu S., Zhang P., Huang L. J., Wang Z. F., Chem. J. Chinese Universities, 2010, 31(10), 1992—1998
	(徐莎, 张萍, 黄琳娟, 王仲孚. 高等学校化学学报,2010, 31(10), 1992—1998
[14]	Aoki K., Porterfield M., Lee S. S., J. Biol. Chem., 2008, 283(44), 30385—30400
[15]	Yang X., Tao S., Orlando R., Carbohyd. Res., 2012, 358, 47—55
[16]	Brockhausen I., Biochim. Soc. Trans., 2003, 31, 318—325
[17]	Brockhausen I., Adv. Exp. Med. Biol., 2003, 535, 163—188
[18]	Wang C., Fan W., Zhang P., Wang Z., Huang L., Proteomics, 2011, 11(21), 4229—4242
[19]	Maniatis S., Zhou H., Reinhold V., Anal. Chem., 2010, 82(6), 2421—2425
[20]	Kim Y. J., Varki A., Glycoconjugate J., 1997, 14(5), 569—576
[21]	Taylor M.E., Drickamer K., Introduction to Glycobiology, Translated by Ma Y. J., Chemical Industry Press, Beijing, 2006, 193—195
	(Taylor M.E., Drickamer K., 马毓甲[译]. 糖生物学导论, 北京: 化学工业出版社, 2006, 193—195)