General Obstetrics and Gynecology: Obstetrics
Mechanisms of abruption-induced premature rupture of the fetal membranes: Thrombin enhanced decidual matrix metalloproteinase-3 (stromelysin-1) expression

https://doi.org/10.1016/j.ajog.2004.08.003Get rights and content

Objective

The aim of the study was to evaluate thrombin and progestin effects on matrix metalloproteinase-3 expression in term decidual cells as a mechanism of abruption-related preterm delivery.

Study design

Decidual cells were isolated by standard techniques, purified to homogeneity, grown to confluence, and passaged. Cultures were primed with 10−8 M estradiol or estradiol plus 10−7 progestin and then incubated in a defined medium with corresponding steroid(s) plus or minus thrombin or the protease-activated thrombin receptor-1 agonist for 24 hours. Secreted matrix metalloproteinase-3 levels were assessed by enzyme-linked immunosorbent assay, and immunoblotting and messenger RNA levels were measured by Northern blotting and quantitative reverse transcription-polymerase chain reaction.

Results

Immunoreactive matrix metalloproteinase-3 levels were inhibited 66% by estradiol plus progestin versus estradiol (P < .05). Thrombin elicited a dose-dependent reversal in this progestin inhibition, producing a 2.5-fold increase at 2.5 U/mL (P < .05) that attained 33% of matrix metalloproteinase-3 levels in parallel incubations with estradiol plus thrombin. Protease-activated thrombin receptor-1 agonist mimicked 60% of thrombin-enhanced matrix metalloproteinase-3 output. Immunoblotting validated the enzyme-linked immunosorbent assay results. Northern blotting and quantitative reverse transcription-polymerase chain reaction demonstrated corresponding effects on steady-state messenger RNA levels.

Conclusion

Abruption-generated thrombin promotes preterm delivery by mediating fetal membrane extracellular matrix degradation via enhanced decidual cell matrix metalloproteinase-3 expression, whereas progesterone blunts this thrombin-induced effect.

Section snippets

Isolation of DCs

After receiving written informed consent, fetal membrane specimens were obtained after repeat cesarean deliveries from patients with uncomplicated pregnancies delivering at term either at Bellevue Hospital or Yale-New Haven Hospital under investigational review board and human investigation committee approval, respectively. The decidua was scraped from the maternal surface of the chorion and then minced and digested in Ham's F-10 plus 10% stripped calf serum (SCS) containing 25 mg/mL of

Statistical analysis

Comparisons of control versus treatment groups were performed using the analysis of variance test with P < .05 representing statistical significance.

Effects of steroids and thrombin on MMP-3 protein expression in term DCs

Figure 1 indicates that in 7 separate DC preparations, secreted levels of MMP-3 were reduced by more than 3- fold in cultures incubated in E2 plus MPA, compared with cultures maintained in E2 (n = 7; P < .05). Thrombin (2.5 U/mL) elevated MMP-3 output by 2.6-fold (P < .05) in both E2 and E2 plus MPA-treated cultures. The addition of MPA inhibited the effects of thrombin on MMP-3 output by reducing MMP-3 levels by about 70% (P < .05), compared with parallel cultures in which thrombin was added

Comment

The current study revealed that MMP-3 mRNA and protein expression in cultured term DCs is progestin inhibited and that thrombin reverses this inhibition. Previously, we found that constitutively expressed PAR1 was involved in mediating thrombin-enhanced plasminogen activator expression in decidualized stromal cells from cycling endometrium.22 To determine whether PAR1 also plays a role in thrombin-enhanced MMP-3 expression in term DCs, the current study evaluated the effects of TRAP-14, which

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