Matrix metalloproteinase-3 (MMP-3) over-expression is associated with tissue destruction in the context of chronic inflammation. Previous studies showed that IL-4 inhibits induction of MMP-3 by IL-1β, and suggested that AP-1 might be involved. Here we show that IL-1 induced binding of transcription factor AP-1 to the MMP-3 promoter consists primarily of c-Jun, JunB, and c-Fos and that binding of c-Jun and c-Fos is inhibited by the combination of cytokines while binding of Jun B is not. Mutation of the AP-1 site in the MMP-3 promoter decreased the ability of IL-4 to inhibit its transcription in transfected MG-63 cells. Western blotting showed that both cytokines activate Jun N-terminal kinase (JNK), but with somewhat different kinetics, and that activation of JNK by both cytokines individually is inhibited by the combination. These results indicate that IL-4 inhibition of MMP-3 expression is associated with reduction of IL-1 induced binding of active forms of the AP-1 dimer, while less active JunB-containing dimers remain, and suggest that these changes are associated with decreased activation of JNK.

Cardiac enlargement that develops during pregnancy is an important yet under‐studied adaptation to physiologic stress. In light of recent evidence of resident stem cell pools in the heart, we tested the hypothesis that pregnancy‐induced cardiac enlargement may involve both myocyte hypertrophy and new myocyte formation. 6 nonpregenant age matched controls (C) and 6 mice in late pregnancy (LP, 17–19 days) were studied. Hearts were formalin fixed and 5μ sections were examined. Laminin staining was used to identify cell perimeter and determine myocyte cross‐sectional area (MCSA). Wet weights of LP hearts were 23% greater than C (240±9 vs 195±13mg). Increases in MCSA were not proportional to increases in heart mass. MCSA of LP (262±3μ2, n=225) was 11% greater than of C (236± 3μ2, n=225). Analysis of sections from different regions of the heart showed different degrees of MCSA hypertrophy. Apex, mid and basal sections were analyzed from all hearts. No difference in MCSA among sections was found in C. MCSA of all sections were greater in LP than C, but MCSA of the mid‐heart in LP (275 ±4μ2, n=75) were greater than MCSA of apex (252± 5μ2, n=75) and base (259 ±4μ2, n=75). Myocyte hypertrophy during pregnancy is not sufficient to account for the whole‐heart enlargement suggesting that new myocytes are generated during pregnancy. Hypertrophy is not evenly distributed during pregnancy suggesting an unequal distribution of wall stress.

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MMP‐3 plays an important role in normal and pathological tissue remodeling, and its expression of is regulated by a variety of cytokines, growth factors, and hormones, mainly at the level of transcription. Transcription factor ZBP‐89 has been shown to interact with a polymorphic promoter sequence and activate transcription of the MMP‐3 gene when over‐expressed. To examine the role of ZBP‐89 in basal and IL‐1‐induced expression of MMP3 we knocked down ZBP‐89 in the MG63 cell line. MMP‐3 mRNA expression was measured by real‐time PCR and compared with expression in a negative control cell line. The basal expression of MMP‐3 was significantly lower in the knockdown cell line when compared to negative control. This is consistent with ZBP‐89 as an activator of basal expression. IL‐1 fold induction was significantly higher at 6 hours in the knockdown cell line, which is consistent with a role of ZBP‐89 as a repressor of IL‐1 induced expression of MMP‐3.