Cowell S, Knäuper V, Stewart M L, D'Ortho M P, Stanton H, Hembry R M, López-Otín C, Reynolds J J, Murphy G
Department of Cell and Molecular Biology, Strangeways Research Laboratory, Worts' Causeway, Cambridge, CB1 4RN, U.K.
Biochem J. 1998 Apr 15;331 ( Pt 2)(Pt 2):453-8. doi: 10.1042/bj3310453.
SW1353 chondrosarcoma cells cultured in the presence of interleukin-1, concanavalin A or PMA secreted procollagenase 3 (matrix metalloproteinase-13). The enzyme was detected in the culture medium by Western blotting using a specific polyclonal antibody raised against recombinant human procollagenase 3. Oncostatin M enhanced the interleukin-1-induced production of procollagenase 3, whereas interleukin-4 decreased procollagenase 3 synthesis. The enzyme was latent except when the cells had been treated with concanavalin A, when a processed form of 48 kDa, which corresponds to the active form, was found in the culture medium and collagenolytic activity was detected by degradation of 14C-labelled type I collagen. The concanavalin A-induced activation of procollagenase 3 coincided with the processing of progelatinase A (matrix metalloproteinase-2) by the cells, as measured by gelatin zymography. In addition, progelatinase B (matrix metalloproteinase-9) was activated when gelatinase A and collagenase 3 were in their active forms. Concanavalin A treatment of SW1353 cells increased the amount of membrane-type-1 matrix metalloproteinase protein in the cell membranes, suggesting that this membrane-bound enzyme participates in an activation cascade involving collagenase 3 and the gelatinases. This cascade was effectively inhibited by tissue inhibitors of metalloproteinases-2 and -3. Tissue inhibitor of metalloproteinases-1, which is a much weaker inhibitor of membrane-type 1 matrix metalloproteinase than tissue inhibitors of metalloproteinases-2 and -3 [Will, Atkinson, Butler, Smith and Murphy (1996) J. Biol. Chem. 271, 17119-17123], was a weaker inhibitor of the activation cascade.
在白细胞介素-1、伴刀豆球蛋白A或佛波酯存在的情况下培养的SW1353软骨肉瘤细胞分泌前胶原酶3（基质金属蛋白酶-13）。使用针对重组人前胶原酶3产生的特异性多克隆抗体，通过蛋白质印迹法在培养基中检测到该酶。制瘤素M增强白细胞介素-1诱导的前胶原酶3的产生，而白细胞介素-4降低前胶原酶3的合成。该酶是无活性的，除非细胞用伴刀豆球蛋白A处理，此时在培养基中发现了48 kDa的加工形式，其对应于活性形式，并且通过14C标记的I型胶原的降解检测到胶原olytic活性。伴刀豆球蛋白A诱导的前胶原酶3的激活与细胞对前明胶酶A（基质金属蛋白酶-2）的加工同时发生，如通过明胶酶谱法所测量。此外，当前明胶酶A和胶原酶3处于活性形式时，前明胶酶B（基质金属蛋白酶-9）被激活。用伴刀豆球蛋白A处理SW1353细胞增加了细胞膜中膜型-1基质金属蛋白酶蛋白的量，表明这种膜结合酶参与了涉及胶原酶3和明胶酶的激活级联反应。该级联反应被金属蛋白酶组织抑制剂-2和-3有效抑制。金属蛋白酶组织抑制剂-1是膜型1基质金属蛋白酶的抑制剂，比金属蛋白酶组织抑制剂-2和-3弱得多[Will，Atkinson，Butler，Smith和Murphy（1996）J.Biol.Chem.271，17119-17123]，是激活级联反应的较弱抑制剂。
