Phytohaemagglutinin stimulates the proliferation of peripheral blood mononuclear cells and expression of secretory cytokines

doi:10.3969/j.issn.2095-4344.2014.23.017

Abstract

Abstract:

BACKGROUND: Phytohemagglutinin (PHA) can stimulate the peripheral blood mononuclear cells (PBMCs) into cell cycle, and cause their immune activation, which is a common immune proliferation model. However, the role of non-PBMC ingredient of peripheral blood is unclear, as well as the expression of endothelial cells related cytokines.

OBJECTIVE: To study the effect of whole blood culture and PBMCs alone culture with PHA on the PBMC proliferation and apoptosis, expression of inflammatory cytokine and endothelial cell secreted cytokine markers.

METHODS: Morphological changes of PBMCs separated from normal karyotype human peripheral blood individually cultured with or without PHA were observed. The PBMCs were collected by whole blood culture or PBMC separated culture. mRNA was extracted for the fluorescence quantitative RT-PCR, which was applied to detect the cell proliferation, apoptosis, and expression of inflammatory cytokine and endothelial cell secreted cytokines. The statistic analysis was used for the significance explication.

RESULTS AND CONCLUSION: PBMCs alone cultured ere different from those undergoing whole blood culture. The PHA could up-regulate the gene expression of Ki67, proliferating cell nuclear antigen, Caspase 3, interferon-γ, tumor necrosis factor-β and interleukin-6, but down-regulate Protein C. This indicted that PHA could promote the proliferation and apoptosis of PBMCs and up-regulate the expression of inflammatory cytokines, but down-regulate the expression of endothelial cells secreted coagulation cytokines.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

全文链接：

Key words: stem cells, plant lectins, cytokines, cell proliferation, apoptosis

CLC Number:

R394.2

Wang Ding, Song Bing, Zhong Xuan, Sun Xiao-fang, Fan Yong . Phytohaemagglutinin stimulates the proliferation of peripheral blood mononuclear cells and expression of secretory cytokines[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(23): 3707-3714.

Figures/Tables 4

References

[1] 许树成.人微量全血培养与染色体标本制作技术研究[J].生物学杂志, 2005,22(4):46-47.
[2] 谭雷涛,王凌燕,常维山,等.全血培养与酶标仪联用检测淋巴细胞转化的研究[J].中国农学通报,2006,22(1): 1-3.
[3] Tsai WJ, Chen YC, Wu MH, et al. Seselin from Plumbago zeylanica inhibits phytohemagglutinin (PHA)-stimulated cell proliferation in human peripheral blood mononuclear cells. J Ethnopharmacol.2008; 119(1):67-73.
[4] Katiyar S, Awasthi SK, Sahu RK. Suppression of IL-6 level in human peripheral blood mononuclear cells stimulated with PHA/LPS after occupational exposure to chromium. Sci Total Environ. 2008; 390(2-3):355-361.
[5] Lei P, He Y, Ye Q, et al. Antigen-binding characteristics of AbCD71 and its inhibitory effect on PHA-induced lymphoproliferation. Acta Pharmacologica Sinica. 2007; 28(10):1659-1664.
[6] Ide K, Momoi Y, Iwasaki T. Canine PHA-stimulated adherent cell enhance interferon-gamma production and proliferation of autologous peripheral blood mononuclear cells. Vet Comp Oncol. 2005; 3(1):25-31.
[7] 王悦.人全血培养方法筛选抗类风湿性关节炎青藤碱衍生物[J]. 中国实用医药, 2013, 8(30):255.
[8] Janefjord CK, Jenmalm MC. PHA-induced IL-12R beta(2) mRNA expression in atopic and non-atopic children. Clin Exp Allergy. 2001;31(10):1493-1500.
[9] Dabrowski MP, Stankiewicz W, Plusa T, et al. Competition of IL-1 and IL-1ra determines lymphocyte response to delayed stimulation with PHA. Mediators of inflammation. 2001; 10(3): 101-107.
[10] Chen YC, Chang SC, Wu MH, et al. Norcantharidin reduced cyclins and cytokines production in human peripheral blood mononuclear cells. Life Sci. 2009;84(7-8):218-226.
[11] Javadian A, Salehi E, Bidad K,et al. Effect of Estrogen on Th1, Th2 and Th17 Cytokines Production by Proteolipid Protein and PHA Activated Peripheral Blood Mononuclear Cells Isolated from Multiple Sclerosis Patients. Arch Med Res. 2014; 45(2):177-182.
[12] Valor L, Sarmiento E, Navarro J, et al. Evaluation of lymphoproliferative responses by carboxy fluorescein succinimidyl ester assay in heart recipients with infections. Transplant Proc. 2012; 44(9):2649-2652.
[13] Scavella A, Leiva L, Monjure H, et al. Effect of L-arginine supplementation on immune responsiveness in patients with sickle cell disease. Pediatr Blood Cancer. 2010; 55(2): 318-323.
[14] Meier V, Burger E, Mihm S, et al. Ribavirin inhibits DNA, RNA, and protein synthesis in PHA-stimulated human peripheral blood mononuclear cells: possible explanation for therapeutic efficacy in patients with chronic HCV infection. J Med Virol. 2003; 69(1):50-58.
[15] Morgan R, Chen Z, Richkind K, et al. PHA/IL2: an efficient mitogen cocktail for cytogenetic studies of non-Hodgkin lymphoma and chronic lymphocytic leukemia. Cancer Geneti Cytogenet. 1999; 109(2):134-137.
[16] Martin-Ramirez J, Hofman M, van den Biggelaar M, et al. Establishment of outgrowth endothelial cells from peripheral blood. Nat Protocol. 2012; 7(9):1709-1715.
[17] Estes ML, Mund JA, Ingram DA, et al. Identification of endothelial cells and progenitor cell subsets in human peripheral blood.Curr Protoc Cytom. 2010;Chapter 9:Unit 9.33.1-11.
[18] Fidan I, Yesilyurt E, Kalkanci A, et al. Immunomodulatory Effects of Voriconazole and Caspofungin on Human Peripheral Blood Mononuclear Cells Stimulated by Candida albicans and Candida krusei. Am J Med Sci. 2014.
[19] Bellik L, Musilli C, Vinci MC, et al. Human mature endothelial cells modulate peripheral blood mononuclear cell differentiation toward an endothelial phenotype. Exp Cell Res. 2008;314(16):2965-2974.
[20] MacCarthy EP, Ooi YM, Hsu A, et al. Evidence for an endothelial cell-derived factor which stimulates the growth of human peripheral blood mononuclear cells. Cell Immunol. 1985;91(2):492-497.
[21] Ulmer AJ, Scholz W, Ernst M, et al. Response of human T lymphocytes to phytohemagglutinin (PHA) after sequential depletion of monocytes, HLA-DR+, Leu11a+, and Leu7+ cells. Immunobiology. 1985;170(5):419-433.
[22] Luquetti A, Janossy G. Lymphocyte activation. VIII. The application of a whole blood test to the quantitative analysis of PHA responsive T cells. J Immunol Methods. 1976; 10(1): 7-25.
[23] Scappaticci S, Danesino C, Rossi E, et al. Cytogenetic abnormalities in PHA-stimulated lymphocytes from patients with Langerhans cell histocytosis. AIEOP-Istiocitosi Group. Br J Haematol. 2000;111(1):258-262.
[24] Vermaelen K, Barbieri D, Van den Berghe H. Indirect stimulation of B-cell proliferation in vitro by T cells, as evidenced by cytogenetic analysis of PHA-stimulated cell cultures of B-cell lymphomas. Cancer Genet Cytogenet. 1984; 11(4):425-428.
[25] Zhang Z, Ding Y, Li W, et al. Interleukin-17A- or tumor necrosis factor alpha-mediated increase in proliferation of T cells cocultured with synovium-derived mesenchymal stem cells in rheumatoid arthritis. Arthritis Res Ther. 2013;15(5): R169.
[26] Ribeiro A, Laranjeira P, Mendes S, et al. Mesenchymal stem cells from umbilical cord matrix, adipose tissue and bone marrow exhibit different capability to suppress peripheral blood B, natural killer and T cells. Stem Cell Res Ther. 2013;4(5):125.
[27] Gupta D, Rani M, Khan N, et al. HIV-1 Infected Peripheral Blood Mononuclear Cells Modulate the Fibrogenic Activity of Hepatic Stellate Cells through Secreted TGF-beta and JNK Signaling. PloS one. 2014;9(3):e91569.
[28] Glantz Y, Sabag O, Lichtenstein M, et al. Eliminating the six N-terminal amino acids of the caspase 3 large subunit improved production of a biologically active IL2-Caspase3 chimeric protein. Biotechnol Prog. 2012;28(2):573-580.
[29] Shing CM, Peake JM, Suzuki K, et al. Bovine colostrum modulates cytokine production in human peripheral blood mononuclear cells stimulated with lipopolysaccharide and phytohemagglutinin. J Interferon Cytokine Res. 2009;29(1): 37-44.
[30] Vongsakul M, Aksomboon A. Cyclins D1, E, A expression and synergistic cytotoxicity to a cholangiocarcinoma cell line from recombinant TNF-alpha and PHA supernate. Asian Pac J Allergy Immunol. 2002;20(1):57-60.
[31] Eichmann A, Simons M. VEGF signaling inside vascular endothelial cells and beyond. Curr Opin Cell Biol. 2012; 24(2): 188-193.
[32] Trincavelli ML, Da Pozzo E, Ciampi O, et al. Regulation of Erythropoietin Receptor Activity in Endothelial Cells by Different Erythropoietin (EPO) Derivatives: An in Vitro Study. Int J Molecular Sci. 2013;14(2):2258-2281.
[33] Castellino FJ, Ploplis VA. The protein C pathway and pathologic processes. J Thromb Haemost. 2009;7 Suppl 1:140-145.