The use of a method for determining the levels of cellular phosphatidic acid in detecting the Aspergillus fumigatus activating phospholipase D activity in pulmonary epithelial cells

Chinese Journal of Mycology　2014, Vol. 9 　Issue (1): 16-19.

Previous Articles Next Articles

The use of a method for determining the levels of cellular phosphatidic acid in detecting the Aspergillus fumigatus activating phospholipase D activity in pulmonary epithelial cells

WANG Shuo¹, HAN Xue-lin², GAO Huan², ZONG Hao¹, HAN Li²

1. College of Life Science, Sichuan Normal University, Chengdu 610101, China;
2. Center for Hospital Infection Control, CDC of PLA, Beijing 100071, China

Received:2013-11-21 Online:2014-02-28 Published:2014-02-28

Abstract

Abstract: Objective To establish a method to detect Phospholipase D (PLD) activity in the A549 cells stimulated by A.fumigatus by measuring the levels of cellular phosphatidic acid (PA).Method Using the interaction model of epithelial cells stimulated by A.fumigatus ATCC13073;Chloroform-methanol method was used to extrat the intracellular lipid. A modified method was established to measure PA standard and the levels of cellular PA.Results The calibration curve of PA standard was linear between 5 and 250 μmol/L. Swollen conidia, but not resting conidia increased the level of PA in A549 cells.Conclusion This improved method for determining the levels of cellular PA was high sensitive, stable and convenient for measuring indirectly the intracellular PLD activity. Our data indicated that the PLD activity in the pulmonary epithelial cells could be significantly activated by A.fumigatus swollen conidia.

Key words: phospholipase D, phosphotidic acid, detection method

CLC Number:

R379.6

WANG Shuo, HAN Xue-lin, GAO Huan, ZONG Hao, HAN Li. The use of a method for determining the levels of cellular phosphatidic acid in detecting the Aspergillus fumigatus activating phospholipase D activity in pulmonary epithelial cells[J]. Chinese Journal of Mycology, 2014, 9(1): 16-19.

References

[1] Filler S G, Sheppard D C. Fungal invasion of normally non-phagocytic host cells[J]. PLoS pathogens, 2006, 2(12): e129.
[2] Han X, Yu R, Zhen D, et al. β-1, 3-Glucan-induced host phospholipase D activation is involved in Aspergillus fumigatus internalization into type Ⅱ human pneumocyte A549 cells[J]. PloS one, 2011, 6(7): e21468.
[3] 李平, 周汉良. 磷脂酶D和炎症的关系[J]. 生理科学进展, 1999, 30(2): 118-122.
[4] Morita S, Ueda K, Kitagawa S. Enzymatic measurement of phosphatidic acid in cultured cells[J]. J Lipid Res, 2009, 50(9): 1945-1952.
[5] Antonescu C N, Danuser G, Schmid S L. Phosphatidic acid plays a regulatory role in clathrin-mediated endocytosis[J]. Mol Biol Cell, 2010, 21(16): 2944-2952.
[6] Bligh E G, Dyer W J. A rapid method of total lipid extraction and purification[J]. Can J Biochem Physiol, 1959, 37(8): 911-917.
[7] Serrander L, Fllman M, Stendahl O. Activation of phospholipase D is an early event in integrin-mediated signalling leading to phagocytosis in human neutrophils[J]. Inflammation, 1996, 20(4): 439-450.
[8] Han X, Yu R, Ji L, et al. InlB-mediated Listeria monocytogenes internalization requires a balanced phospholipase D activity maintained through phospho-cofilin[J]. Mol Microbiol, 2011, 81(4): 860-880.
[9] Moreno-García ME, López-Bojórques LN, Zentella A, et al. CD38 signaling regulates B lymphocyte activation via a phospholipase C (PLC)-γ2-independent, protein kinase C, phosphatidylcholine-PLC, and phospholipase D-dependent signaling cascade[J]. J Immunol, 2005, 174(5): 2687-2695.
[10] Vinggaard A, Jensen T, Morgan C, et al. Didecanoyl phosphatidylcholine is a superior substrate for assaying mammalian phospholipase D[J]. Biochem J, 1996, 319 (Pt 3): 861-864.
[11] 韩丽, 毕阳, 魏晋梅, 等. 应用荧光高效液相测定麦芽根中磷脂酶 D 的活性[J]. 天然产物研究与开发, 2009, 21(2): 343-345.
[12] English D, Cui Y, Siddiqui RA. Messenger functions of phosphatidic acid[J]. Chem Phys Lipids, 1996, 80(1): 117-132.

The use of a method for determining the levels of cellular phosphatidic acid in detecting the Aspergillus fumigatus activating phospholipase D activity in pulmonary epithelial cells

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 2

Recommended Articles

Metrics

Comments

[1]	LIU Xiao-yu, ZHANG Chang-jian, HU Ying-song, CHEN Fang-yan, HAN Li. Loss of phospholipase D in mice against Aspergillus fumigatus infection [J]. Chinese Journal of Mycology, 2018, 13(3): 129-133.
[2]	TONG Jian-bo, ZENG Rong, LI Min. Research progress of Aspergillus fumigatus biofilms [J]. Chinese Journal of Mycology, 2018, 13(1): 61-64.