Identification of naturally secreted soluble form of TL1A, a TNF-like cytokine

TL1A, a TNF-like ligand, mediates signaling via its cognate receptor DR3, a death receptor whose activation was known to induce both death and survival factors. TL1A, like TNF, is also presumed to circulate as a homotrimeric soluble form. To identify soluble TL1A in the immune system, we have developed a quantitative ELISA by pairing a monoclonal antibody (MAb) with a biotinylated anti-TL1A polyclonal antibody (PAb) as capture and detection antibodies, respectively. The assay had a detection limit of 32 pg/ml. Overnight culture of human umbilical vein endothelial cells (HUVEC) expressed up to 160 pg/ml of TL1A, and that proinflammatory cytokines, IL-1 and TNF, significantly increased TL1A mRNA and soluble protein up to several folds in contrast to IL-6 and IL-11, which induced neither protein nor mRNA in the cells. IL-1 appeared to be a better stimulator of TL1A than TNF-α, and the induction of soluble TL1A in HUVEC in response to IL-1 was dose and time-dependent. We have also purified soluble TL1A from a large volume of IL-1 stimulated HUVEC conditioned medium using an anti-TL1A PAb-coupled affinity column. The protein eluted from the column was further reacted with anti-TL1A MAbs in Western blot: 30-kDa and 32-kDa under non-reducing and reducing conditions, respectively. Taken together, our results indicated that ELISA might be useful in studying soluble TL1A regulation in certain inflammatory conditions.