Posts Tagged ‘Metabolism

Thirty hours after infection, the medium surrounding amnions infected with AdMMP-9 contained a prominent proMMP-9 signal detected as a 92-kDa lysis band by zymog-raphy and an 83-kDa band beneath it, reflecting processed proenzyme (Fig. 4), results that are consistent with transduction of cultured cells with this vector. MMP-2 activity was present in the media from […]

In agreement with our previous zymo-graphic studies, MMP-2 (66-kDa lysis band) and MmP-9 (92- and 180-kDa lysis bands) activities were increased on Day 21 compared to Day 19 (Fig. 3D). A lysis band at 68 kDa, possibly reflecting the activated form of MMP-9, was also detected on Day 21. A faint lysis band at 31 […]

Loss of Type IV Collagen from Rat Amnions Before Labor and Increased MMP Expression Western blot analyses of extracts of rat amnions collected on Days 18-21 of pregnancy using an antibody recognizing a1(IV)/a2(IV) chains of type IV collagen demonstrated a marked reduction in the content of type IV col-lagens per unit protein by Day 21 […]

Type IV Collagen Chains a1(IV) and a2(IV) Predominated in Rat Amnion Extracts of amnions collected on Day 19 of pregnancy were treated with bacterial collagenase and then subjected to SDS PAGE to resolve the NC1 domains. The proteins were transferred to membranes, which were subsequently probed with polyclonal antibodies recognizing specific NC1 sequences in the […]

This concentration of recombinant adenovirus was selected on the basis of previous experience with transduction of cultured cells and experiments in which rat amnions were infected with lower concentrations of virus. In some experiments, the MMP inhibitor batimas-tat (BB-94: 4-[Af-hydroxylamino]-2R-isobutyl-3S-[thienyl-thiomethyl]-succinyl-L-phenylalanine-A/-methylamide; British Biotech, Oxford, UK) was added to the culture medium at a concentration of 3 […]

In Vitro Basement Membrane Degradation Assays Basement membranes were prepared from rat amnions and maternal kidney as previously described. The basement membrane extracts (1 mg dry weight) were incubated with bacterial collagenase (Worthington, Freehold, NJ; 18.5 U/mg) or active MMP-9 enzyme (Calbiochem, San Diego, CA; 1 ^g of enzyme/mg of detergent-extracted basement membrane) over a […]

Animals Timed pregnant Sprague-Dawley rats (Zivic-Miller, Port-ersville, PA) were killed on Days 18-21 of pregnancy. The animals usually deliver on the afternoon of Day 21. Amnions were collected as previously described. Tissue extracts prepared in a buffer consisting of 50 mM Tris, 1% Nonidet NP-40, 0.1% deoxycholate, 0.1% SDS, 150 mM NaCl, 10 ^g/ml aprotinin, […]