Comparison of LPS-stimulated release of cytokines in punch versus transwell tissue culture systems of human gestational membranes.
BACKGROUND
Cytokine 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.


METHODS
Tissue 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.


RESULTS
Levels 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.


CONCLUSIONS
These 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.
Miller
Mark F
MF
Department of Environmental Health Science, School of Public Health, University of Michigan, Ann Arbor, Michigan 48109, USA. miller.mark@epa.gov
Loch-Caruso
Rita
R
D001706:Biopsy; D016207:Cytokines / Q000378:metabolism*; D005321:Extraembryonic Membranes / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D005260:Female; D005865:Gestational Age; D006801:Humans; D007249:Inflammation / Q000276:immunology / Q000378:metabolism; D016753:Interleukin-10 / Q000378:metabolism; D053583:Interleukin-1beta; D015850:Interleukin-6 / Q000378:metabolism; D016209:Interleukin-8 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D011247:Pregnancy; D011263:Pregnancy Trimester, Third / Q000378:metabolism; D046509:Tissue Culture Techniques / Q000295:instrumentation* / Q000379:methods*; D014409:Tumor Necrosis Factor-alpha
D016207:Cytokines; D053583:Interleukin-1beta; D015850:Interleukin-6; D016209:Interleukin-8; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha; D016753:Interleukin-10