Matrix metalloproteinase-2 activity, protein, mRNA, and tissue inhibitors in small arteries from pregnant and relaxin-treated nonpregnant rats

Vascular gelatinase activity is essential for pregnancy- and relaxin (R1x)-induced renal vasodilation and hyperfiltration in rats. The objective of this study was to further elucidate the mechanisms for the increase in vascular matrix metalloproteinase (MMP)-2 activity caused by pregnancy and R1x. We first corroborated our earlier work by showing that pro- and active forms of MMP-2 were increased in small renal arteries from pregnant compared with virgin rats and R1x-treated compared with vehicle-treated nonpregnant rats. We next investigated other artery types and showed that MMP-2 activity was upregulated in mesenteric arteries from pregnant rats (pro-MMP-2 by 50% and active MMP-2 by 40%, both P < 0.05) and from R1x-treated nonpregnant rats (pro-MMP-2 by 50% and active MMP-2 by 90%, both P < 0.005) compared with their respective controls. To corroborate these results obtained by gelatin zymography, pro-MMP-2 protein was determined by Western analysis in the same small arteries. Pro-MMP-2 protein was increased in small renal arteries from pregnant compared with virgin rats and from R1x- compared with vehicle-treated nonpregnant rats: pro-MMP-2-to-beta- actin ratio = 0.29 vs. 0.21 (P < 0.01) and 0.43 vs. 0.32 (P < 0.005). Findings were similar for mesenteric arteries. MMP- 2 mRNA as measured by real-time PCR was increased in small renal arteries from pregnant and R1x-treated nonpregnant rats compared with their respective controls. There were no significant differences in tissue inhibitor of metalloproteinase (TIMP-1 or TIMP-2) activity by reverse zymography in small renal arteries. Thus increases in MMP-2 mRNA and protein expression are major factors contributing to increased MMP- 2 activity in small arteries from pregnant and R1x-treated nonpregnant rats.