In vitro secretion and activity profiles of matrix metalloproteinases, MMP-9 and MMP-2, in human term extra-placental membranes after exposure to Escherichia coli

Abstract

BackgroundPremature rupture of fetal membranes (PROM) complicated with intrauterine infection has been associated to alterations of the extracellular matrix (ECM) homeostasis. The aim of this work was to evaluate the integral/functional response of the amnion (AMN) and choriodecidua (CHD) to synthesis, secretion, and activity of MMP-2 and MMP-9 and of their inhibitors TIMP-1, -2, and -4, after stimulation with Escherichia coli.MethodsFull-thickness membranes were mounted on a Transwell device, constituting two independent chambers, Escherichia coli (1×10 (6) CFU/mL) were added to either the amniotic or the choriodecidual face or to both. Secretion profiles of MMP-2, MMP-9, TIMP-1, TIMP-2, and TIMP-4 were quantified by ELISA and gelatinolytic activity by zymography. Immunoreactivity for MMP-2 and MMP-9 was revealed by immunohistochemistry and the collagen content was assessed by the hydroxyproline assay.ResultsLevels of MMP-9 in CHD and AMN increased 4- and 8-fold, respectively, after simultaneous infection. MMP-2 secreted to the medium by CHD increased a mean of 3 times after direct stimulation. Secretion profiles of TIMP-1, TIMP-2, and TIMP-4 remained without significant changes. Collagen content was significantly decreased (4-fold) in infected membranes, and was associated with loss of structural continuity and co-localization with immunoreactive forms of MMP-2 and MMP-9.ConclusionsInfection of chorioamniotic membranes with E. coli induces an increase in the secretion of inactive forms and an association to ECM of active forms of MMP-2 and MMP-9 without changes in TIMP-1, -2, and -4. These changes could explain the significant decrease of collagen content and loss of structural continuity.

In Vitro Secretion Profile of Pro‐Inflammatory Cytokines IL‐1β, TNF‐α, IL‐6, and of Human Beta‐Defensins (HBD)‐1, HBD‐2, and HBD‐3 from Human Chorioamniotic Membranes After Selective Stimulation with Gardnerella vaginalis

“…[26][27][28] Therefore, the IL-1b increase in the amniotic compartment, after this modality of stimulation, could be the result of different mechanisms, including contact of the bacterium with other cell populations such as fibroblasts and macrophages in the compact layer of the AMN, and alteration of the trans-membranal diffusion of cytokines, which has been characterized in fetal membranes and is directly related with membrane integrity. [26][27][28] Therefore, the IL-1b increase in the amniotic compartment, after this modality of stimulation, could be the result of different mechanisms, including contact of the bacterium with other cell populations such as fibroblasts and macrophages in the compact layer of the AMN, and alteration of the trans-membranal diffusion of cytokines, which has been characterized in fetal membranes and is directly related with membrane integrity.…”

Carbon nanotubes stimulate synovial inflammation by inducing systemic pro-inflammatory cytokines

“…Afterwards, the activities of MMP-2 and MMP-9 were determined by gelatin zymography, as described previously. 34 …”

Burkholderia multivorans acts as an antagonist against the growth of Burkholderia pseudomallei in soil

“…A Transwell culture system (6.5 mm diameter with a pore size of 0.4 μm; Costar, Cambridge, MA, USA) was used to test the release of factors by the isolates (26). First, 10 6 CFU/mL of B. multivorans NKI379 or one of the other antagonistic strains was seeded into the bottom chamber and incubated for 48 hr in LB broth at 37 • C. Next, 10 6 CFU/mL of B. pseudomallei KN07 was added to the upper chamber of the same Transwell system at 0 hr or 24 hr of an initial culture.…”