Tissue inhibitor of metalloproteinases-3 inhibits shedding of L-selectin from leukocytes

Although the Enzyme or enzymes mediating shedding of L-selectin have not yet been identified, this activity can be blocked by synthetic hydroxamic acid-based inhibitors of metalloproteinases such as Ro 31-9790. However, the endogenous matrix metalloproteinase inhibitor tissue inhibitor of metalloproteinases (TIMP)-1 does not block L-selectin shedding. Here, we report that TIMP-3, but not TIMP-2, inhibits L-selectin shedding from mouse and human lymphocytes, Jurkat T cells, and human monocytes. TIMP-3 has an IC50 of 0.3-0.4 microM on these cell types compared with 0.7-4.8 microM for Ro 31-9790. A metalloproteinase (tumor necrosis factor-alpha (TNF-alpha)-converting enzyme; ADAM17) has recently been identified which cleaves the pro-form of TNF-alpha to produce soluble cytokine. We compared inhibition of L-selectin shedding by TIMPs and Ro 31-9790 with inhibition of TNF-alpha shedding from human monocytes. TIMP-3 inhibited TNF-alpha shedding (IC50 of 0.1 microM), as did Ro 31-9790 (IC50 of 0.4 microM). TIMP-2 had a partial effect, and TIMP-1 did not inhibit. This study confirms that L-selectin sheddase is a metalloproteinase, but not a matrix metalloproteinase, and investigates the relationship between shedding of L-selectin and TNF-alpha.