Development of a Lentivirus Vector-based Vaccine Carrying Follicle-stimulating Hormone Receptor and Assay of Its Immunological Effect

doi:10.13560/j.cnki.biotech.bull.1985.2016.03.024

Abstract

Abstract: This work aims to prepare lentivirus vaccine with a follicle-stimulating hormone receptor(FSHR)and investigate the immunological effect of it on mice. The gene(fshr366)of extracellular region of FSHR was cloned into lentivirus vector. The recombinant plasmids were transfected into the 239T cells by the method of lipidosome transfection, and the virus particles(Lenti-FSHR366)with target gene were produced after encapsulation. The expressions of fshr366 mRNA and FSHR366 protein in 293T cells infected by Lenti-FSHR366 were detected by RT-PCR and Western blot. For evaluating the immunological effects, BALB/c mice were intraperitoneally inoculated with single immunization of fshr366-carring virus, collecting the blood samples from the orbits of mice at day 0, 14, 21 and 28 after immunization, then ELISA method was used to detect the specificity of the sera from immunized mice and determine the titer of the antibody. The RE digestion and DNA sequencing results showed the gene fragment of fshr366 was successfully cloned into lentivirus vector. After transfection to 293T cells with the encapsulated lentivirus particles carrying fshr366, the detection results by RT-PCR and Western blot indicated the expression of gene fshr in the cells at transcription and protein level. The results of ELISA revealed that the humoral immune response in mice rose on day 14 after single immunization through intraperitoneally injection of lentivirus particles carrying fshr366, the titer of antibody was 1:1600.

Key words: follicle-stimulating hormone receptor, lentivirus vector, vaccine, humoral response

MA Xiao-ling, LIU Hong-chun, LI Jiang-wei. Development of a Lentivirus Vector-based Vaccine Carrying Follicle-stimulating Hormone Receptor and Assay of Its Immunological Effect[J]. Biotechnology Bulletin, 2016, 32(3): 148-154.

References

[1]Pierce JG, Parsons TF. Glycoprotein hormones：structure and function[J]. Annual Review of Biochemistry, 1981, 50(1)：465-495.
[2]任春明, 字向东, 张重庆, 等. 卵泡刺激素的研究进展[J]. 科研前沿, 2006, 10(208)：10-13.
[3]Simoni M, Gromoll J, Nieschlag E. The Follicle-stimulating hormone receptor：Biochemistry, molecular biology, physiology, and pathop-hysiology 1[J]. Endocrine Reviews, 1997, 18(6)：739-773.
[4]Li Y, Ganta S, Cheng C, et al. FSH stimulates ovarian cancer cell growth by action on growth factor variant receptor[J]. Molecular and Cellular Endocrinology, 2007, 267(1)：26-37.
[5]Parrott JA, Doraiswamy V, Kim G, et al. Expression and actions of both the follicle stimulating hormone receptor and the luteinizing hormone receptor in normal ovarian surface epithelium and ovarian cancer[J]. Molecular and Cellular Endocrinology, 2001, 172(1)：213-222.
[6]Zheng W, Lu JJ, Luo F, et al. Ovarian epithelial tumor growth promotion by follicle-stimulating hormone and inhibition of the effect by luteinizing hormone[J]. Gynecologic Oncology, 2000, 76(1)：80-88.
[7]Radu A, Pichon C, Camparo P, et al. Expression of follicle-stimulating hormone receptor in tumor blood vessels[J]. New England Journal of Medicine, 2010, 363(17)：1621-1630.
[8]Stilley JA, Guan R, Duffy DM, et al. Signaling through FSH receptors on human umbilical vein endothelial cells promotes angiogenesis[J]. The Journal of Clinical Endocrinology & Metabolism, 2014, 99(5)：E813-E820.
[9]Gartrell BA, Tsao C, Galsky MD. The follicle-stimulating hormone receptor：a novel target in genitourinary malignancies[C]//Urologic Oncology：Seminars and Original Investigations. Elsevier, 2013, 31(8)：1403-1407.
[10] Ghinea N. A novel role for FSH receptor as a tumor endothelial cell marker[J]. Acta Endocrinologica(Buc), 2010, 6(4)：507-512.
[11]柏琦, 那鑫妮, 冯姗姗. 病毒载体的发展历程及现状[J]. 中国医药指南, 2011, 9(23)：224-225.
[12]Palmowski MJ, Lopes L, Ikeda Y, et al. Intravenous injection of a lentiviral vector encoding NY-ESO-1 induces an effective CTL response[J]. The Journal of Immunology, 2004, 172(3)：1582-1587.
[13]李明峰, 候静, 熊伟民. 携带AFP基因慢病毒载体制备并感染树突状细胞的实验研究[J]. 细胞与分子免疫学杂志, 2009, 25(9)：791-793.
[14]罗望, 张泓, 许淼, 等. 慢病毒—基因转移的潜在新载体[J]. 江苏药学与临床研究, 2006, 14(6)：366-371.
[15]Esslinger C, Chapatte L, Finke D, et al. In vivo administration of a lentiviral vaccine targets DCs and induces efficient CD8(+)T cell responses[J]. J Clin Invest, 2003, 111：1673-1681.
[16]Breckpot K, Dullaers M, Bonehill A, et al. Lentivirally transduced dendritic cells as a tool for cancer immunotherapy[J]. J Gene Med, 2003, 5：654-667.
[17]Esslinger C, Romero P, MacDonald HR. Efficient transduction of dendritic cells and induction of a T-cell response by thirdgeneration lentivectors[J]. Hum Gene Ther, 2002, 13：1091-1100.
[18]Palmowski MJ, Lopes L, Ikeda Y, et al. Intravenous injection of a lentiviral vector encoding NY-ESO-1 induces an effective CTL response[J]. J Immunol, 2004, 172：1582-1587.
[19]Iglesias MC, Frenkiel MP, Mollier K, et al. A single immunization with a minute dose of a lentiviral vector-based vaccine is highly effective at eliciting protective humoral immunity against West Nile virus[J]. The Journal of Gene Medicine, 2006, 8(3)：265-274.
[20]Wunseh A, AhdaY, Banaz-Yasar F, et al. Single-nucleotide poly-morphisms in the promoter region influence the expression of the human follicle-stimulating Hormone receptor[J]. Fertil Steril, 2005, 84(2)：446-453．
[21]Zhang X, Chen J, Zheng Y, et al. Follicle-stimulating hormone peptide can facilitate paclitaxel nanoparticles to target ovarian carcinoma in vivo[J]. Cancer Research, 2009, 69(16)：6506-6514.
[22]董俊. 基于卵泡刺激素素受体介导的靶向治疗卵巢癌实验研究[D]. 上海：复旦大学, 2011.
[23]Stilley JA, Guan R, Duffy DM, et al. Signaling through FSH recept-ors on human umbilical vein endothelial cells promotes angiogene-sis[J]. The Journal of Clinical Endocrinology & Metabolism, 2014, 99(5)：E813-E820.