Abstract
BackgroundCytokine signaling within the amnionic, chorionic and decidual extraplacental gestational membranes plays an important role in membrane rupture and the timing of birth. The predominant in vitro explant culture system for evaluating cytokine induction in human gestational membranes has been the free-floating biopsy punch culture. Punch systems have been used to investigate the impact of various toxicants, pharmaceuticals and genetic variation on expression of pro-inflammatory cytokines. More recently, a dual compartment transwell culture system has been developed that more closely mimics the intrauterine compartment. The current study compares these two systems with respect to release of pro- and anti-inflammatory cytokines in response to lipopolysaccharide (LPS), a model stimulant.MethodsTissue samples were exposed to 100 ng/ml LPS for 12 h and cytokines were measured by ELISA. Data are expressed as increase relative to non-treated controls.ResultsLevels of interleukin-6 increased in punch culture medium samples to a significantly greater extent (34.2 fold) compared with medium from transwell cultures in the amnion (6.6 fold) or choriodecidual (7.1 fold) compartments. Interleukin-8 also showed a significantly greater induction in punch (4.8 fold) than transwell amnion (1.6 fold) or choriodecidual (1.7 fold) samples. The anti-inflammatory interleukin-10 showed a significant difference between punch (36.5 fold) and transwell amnion (15.4 fold) samples, but no difference was observed between punch and transwell choriodecidual (28.5 fold) samples. Neither interleukin-1beta nor tumor necrosis factor-alpha (TNF-alpha) showed a significant difference between the punch and transwell samples.ConclusionsThese results indicate that the pattern of LPS-stimulated cytokine release from gestational membranes in vitro depends on the culture system used, confounding comparisons of studies that use different gestational membrane culture systems to study inflammatory responses.
Highlights
Cytokine signaling within the amnionic, chorionic and decidual extraplacental gestational membranes plays an important role in membrane rupture and the timing of birth
In contrast to the free floating biopsy punch system, dual-compartment systems employ gestational membrane explants attached to rigid frames and suspended in culture medium, with discrete amniotic and choriodecidual chambers
Dual-compartment systems have used tissues mounted on a transwell insert [13,14,15,16], Ussing chamber [17], or other apparatus [18,19,20] to study cytokine and prostaglandin production induced by bacteria or bacterial toxins [13,14,16,17,21,22], yeast [15] or oxygen tension [18], as well as meconium interactions with the gestational membrane [19]
Summary
Cytokine signaling within the amnionic, chorionic and decidual extraplacental gestational membranes plays an important role in membrane rupture and the timing of birth. One model system used extensively to study stimulated production and release of cytokines in human gestational membranes in vitro involves explant culture of a biopsy punch, with the gestational tissue punch explant free-floating in culture medium. Biopsy punch explant cultures may use full-thickness membranes [1,2] as well as separated amnion [3,4] or choriodecidua [5,6] This single-compartment explant culture system has been used to investigate cytokine [2,3,4,5,7,8,9,10], prostaglandin [4,5,6,8,9,10,11], adipokine [12] and protease [1,8] regulation. The two-compartment system has provided a model system in which investigators can expose and sample medium from each side of the membranes independently
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