Development of In Situ Zymography to Localize Active Matrix Metalloproteinase-7 (Matrilysin-1)

Matrix metalloproteinase-7 (MMP-7) is upregulated during carcinogenesis and its expression correlates with metastasis of human endometrial and gastrointestinal carcinomas. In the present study, we have developed a new method to localize the activity of MMP-7 within tissues. Polyethylene terephthalate films were uniformly coated with crosslinked carboxymethylated transferrin (CCm-Tf) as a substrate and incubated with frozen tissue sections mounted on the films. CCm-Tf on the films was degraded selectively by MMP-7, but showed little or no susceptibility to MMP-1, -2, -3, -9, or -13; MT1-MMP; MT3-MMP; or ADAMTS4. Although some serine proteinases such as elastase also digested CCm-Tf, CCm-Tf films impregnated with serine proteinase inhibitors prevented the digestion. When frozen sections of human endometrial carcinoma and lung carcinoma tissues were incubated on CCm-Tf films or those treated with proteinase inhibitors, the activity was detected in the carcinoma cell nests, where MMP-7 was immunolocalized. Th...