The production of the recombinant human TNFSF15 protein includes gene cloning, plasmid construction, protein expression, protein purification, and protein analysis. Design appropriate primers to amplify the gene fragment corresponding to 1-192 amino acids (aa) of the human TNFSF15 protein. The amplified target gene fragment is cloned into the plasmid together with the N-terminal 10xHis-tag gene. The recombinant plasmid is transfected into mammalian cells using a transfection reagent. After 24 hours of transfection, the medium is replaced with a selective antibiotic to screen the successfully transfected cells for protein expression. After that, the cells are lysed to release the TNFSF15 protein. The recombinant TNFSF15 protein is purified from the cell culture supernatant through Ni-NTA affinity chromatography. The purity of the recombinant TNFSF15 protein is analyzed by SDS-PAGE, which is greater than 95%. The endotoxin content of the TNFSF15 protein is detected by the LAL method, which is less than 1.0 EU/ug. The protein can bind to the TNFSF15 recombinant antibody (CSB-RA023992MA1HU) in a functional ELISA, with the EC50 of 0.8389-0.9731 ng/mL.
Human TNFSF15, also known as TL1A, is a cytokine involved in immune regulation, inflammation, and vascular homeostasis. It is part of the TNF superfamily and is primarily expressed in macrophages and T cells, particularly under pro-inflammatory conditions [1]. TNFSF15 is known to interact with the death receptor 3 (DR3), providing co-stimulatory signals that enhance T-helper 17 cell differentiation and proliferation, thereby amplifying the inflammatory response [1][2]. This interaction is crucial for the modulation of immune responses, particularly in conditions like inflammatory bowel disease (IBD) and various cancers [3].
TNFSF15 is implicated in various pathological conditions, including cancer and autoimmune diseases. Studies have shown that TNFSF15 promotes lymphangiogenesis and metastasis in cancer models, particularly through the upregulation of vascular endothelial growth factor C (VEGF-C), suggesting TNFSF15's role in tumor progression and metastasis [4].
TNFSF15 has been identified as a negative regulator of neovascularization, which is critical for maintaining vascular homeostasis [5]. It inhibits the production of VEGF, a key pro-angiogenic factor, thereby contributing to the stability of established blood vessels [6].
References:
[1] M. Zucchelli, M. Camilleri, A. Andréasson, F. Bresso, A. Dlugosz, J. Halfvarson, et al. Association of tnfsf15 polymorphism with irritable bowel syndrome, Gut, vol. 60, no. 12, p. 1671-1677, 2011. https://doi.org/10.1136/gut.2011.241877
[2] R. Sun, M. Hedl, & C. Abraham. Tnfsf15 promotes antimicrobial pathways in human macrophages and these are modulated by tnfsf15 disease-risk variants, Cellular and Molecular Gastroenterology and Hepatology, vol. 11, no. 1, p. 249-272, 2021. https://doi.org/10.1016/j.jcmgh.2020.08.003
[3] A. Richard, J. Peters, N. Savinykh, J. Lee, E. Hawley, F. Meylan, et al. Reduced monocyte and macrophage tnfsf15/tl1a expression is associated with susceptibility to inflammatory bowel disease, Plos Genetics, vol. 14, no. 9, p. e1007458, 2018. https://doi.org/10.1371/journal.pgen.1007458
[4] T. Qin, D. Huang, Z. Liu, Y. Jia, X. Chen, & K. Li. Tumor necrosis factor superfamily 15 promotes lymphatic metastasis via upregulation of vascular endothelial growth factor‐c in a mouse model of lung cancer, Cancer Science, vol. 109, no. 8, p. 2469-2478, 2018. https://doi.org/10.1111/cas.13665
[5] F. Jiang, Q. Chen, L. Huang, Y. Wang, Z. Zhang, X. Meng, et al. Tnfsf15 inhibits blood retinal barrier breakdown induced by diabetes, International Journal of Molecular Sciences, vol. 17, no. 5, p. 615, 2016. https://doi.org/10.3390/ijms17050615
[6] P. Liang, F. Tian, Y. Lü, B. Duan, D. Stolz, & L. Li. Vascular endothelial growth inhibitor (vegi; tnfsf15) inhibits bone marrow-derived endothelial progenitor cell incorporation into lewis lung carcinoma tumors, Angiogenesis, vol. 14, no. 1, p. 61-68, 2010. https://doi.org/10.1007/s10456-010-9195-8
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