Background: Levels of breath methane, together with breath hydrogen, are determined by means of repeated collections of both, following ingestion of a carbohydrate substrate, at 15–20 minutes intervals, until 10 samples have been obtained. The frequent sampling is required to capture a rise of hydrogen emissions, which typically occur later in the test: in contrast, methane levels are typically elevated at baseline. If methane emissions represent the principal objective of the test, a spot methane test (i.e. a single-time-point sample taken after an overnight fast without administration of substrate) may be sufficient.
Methods: We analysed 10-sample lactulose breath test data from 11 674 consecutive unique subjects who submitted samples to Commonwealth Laboratories (Salem, MA, USA) from sites in all of the states of the USA over a one-year period. The North American Consensus (NAC) guidelines criteria for breath testing served as a reference standard.
Results: The overall prevalence of methane-positive subjects (by NAC criteria) was 20.4%, based on corrected methane results, and 18.9% based on raw data. In our USA dataset, the optimal cut-off level to maximize sensitivity and specificity was ≥4 ppm CH4, 94.5% [confidence interval (CI): 93.5–95.4%] and 95.0% (CI: 94.6–95.5%), respectively. The use of a correction factor (CF) (5% CO2 as numerator) led to reclassifications CH4-high to CH4-low in 0.7 % and CH4-low to CH4-high in 2.1%.
Conclusions: A cut-off value for methane at baseline of either ≥4 ppm, as in our USA dataset, or ≥ 5 ppm, as described in a single institution study, are both highly accurate in identifying subjects at baseline that would be diagnosed as ‘methane-positive’ in a 10-sample lactulose breath test for small intestinal bacterial overgrowth.
Aberrant ceramide build-up in preeclampsia, a serious disorder of pregnancy, causes exuberant autophagy-mediated trophoblast cell death. The significance of ceramide accumulation for lysosomal biogenesis in preeclampsia is unknown. Here we report that lysosome formation is markedly increased in trophoblast cells of early-onset preeclamptic placentae, in particular in syncytiotrophoblasts. This is accompanied by augmented levels of transcription factor EB (TFEB). In vitro and in vivo experiments demonstrate that ceramide increases TFEB expression and nuclear translocation and induces lysosomal formation and exocytosis. Further, we show that TFEB directly regulates the expression of lysosomal sphingomyelin phosphodiesterase (L-SMPD1) that degrades sphingomyelin to ceramide. In early-onset preeclampsia, ceramide-induced lysosomal exocytosis carries L-SMPD1 to the apical membrane of the syncytial epithelium, resulting in ceramide accumulation in lipid rafts and release of active L-SMPD1 via ceramide-enriched exosomes into the maternal circulation. The SMPD1-containing exosomes promote endothelial activation and impair endothelial tubule formation in vitro. Both exosome-induced processes are attenuated by SMPD1 inhibitors. These findings suggest that ceramide-induced lysosomal biogenesis and exocytosis in preeclamptic placentae contributes to maternal endothelial dysfunction, characteristic of this pathology.
Preeclampsia is a serious pregnancy disorder that lacks effective treatments other than delivery.Improper sensing of oxygen changes during placentation by prolyl hydroxylases (PHD), specifically PHD2, causes placental Hypoxia-Inducible Factor-1 (HIF1) buildup and abnormal downstream signaling in early-onset preeclampsia; yet therapeutic targeting of HIF1 has never been attempted. Here we generated a conditional (placenta-specific) knockout of Phd2 in mice (Phd2 -/-cKO) to reproduce HIF1 excess and to assess anti-HIF therapy. Conditional deletion of Phd2 in the junctional zone (JZ) during pregnancy increased placental HIF1 content, resulting in abnormal placentation, impaired remodeling of the uterine spiral arteries, and fetal growth restriction. Pregnant dams developed newonset hypertension at mid-gestation (E9.5) in addition to proteinuria and renal and cardiac pathology, hallmarks of severe preeclampsia in humans. Daily injection of acriflavine, a small-molecule inhibitor of HIF1, to pregnant Phd2 -/-cKO mice from E7.5 (prior to hypertension) or E10.5 (after hypertension has been established) to E14.5 corrected placental dysmorphologies and improved fetal growth.Moreover, it reduced maternal blood pressure and reverted renal and myocardial pathology. Thus, therapeutic targeting of the HIF pathway may improve placental development and function, as well as maternal and fetal health, in preeclampsia.
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