Fig 1.
Representative images for qualitative scoring of airspace infiltration.
Images are representative of the indicated inflammatory scores assigned to each field assessed (× 200 total magnification). Scale bar = 100 μm.
Table 1.
The comparison between the Control and the EVE groups.
Fig 2.
Changes in fetal parameters over time in EVE group.
The horizontal axes represent the time after induction of EVE therapy (hours). The grey dotted lines show mean arterial pressure (mm Hg), the grey solid lines show total circuit blood flow (mL·kg-1·min-1), the black dotted lines show arterial oxygen saturation (%), and the diamonds show blood lactate level (mmol/L). Only the blood lactate levels use the right scale bars. Individual case information is as follows: A) The blood lactate level started to elevate at 19 hours, and continued increasing although other parameters kept stable. Congestive heart failure resulted in a sudden drop of circuit blood flow at 24 hours and euthanasia. B) All the parameters remained within the reference range after stabilisation; however the circuit corrupted (embolism of artificial amniotic fluid) at 12.5 hours, resulting in euthanasia. C) All the parameters remained within the reference range before sudden fatal arrhythmia occurred at 12.5 hours, resulting in euthanasia. D) One membranous oxygenator was blocked with a blood clot at 23 hours (arrow). Despite gradual fetal deterioration including increased the blood lactate levels and decreased circuit blood flow, the fetus completed the pre-determined 40-hour study period. E) Although some fluctuations of parameters were observed in the former period, the fetus completed the pre-determined 40-hour study period with stable physiological parameters.
Fig 3.
Inflammatory protein concentration in fetal plasma measured with ELISA.
H: hours after induction of EVE therapy. Data are presented as box plots with the group median, with whiskers representing maximum and minimum values. Fig 3A shows the data for TNF-α and Fig 3B shows the data for MCP-1. Significant differences vs. value for Control group are indicated: *, p<0.05.
Fig 4.
Relative expression of inflammatory cytokine/chemokine mRNA levels in fetal circulating immunocytes measured with quantitative PCR.
H: hours after induction of EVE therapy. Values of IL-1β, TNF-α, and MCP-2 are presented as bar charts with the group mean normalised expression vs. the value for Control group, with error bars representing SD. Values of IL-6 and IL-8 are presented as box plots with the group median normalised expression vs. the value for Control group, with whiskers representing maximum and minimum values. Significant difference vs. value for Control group is indicated: ‡, p<0.01; significant difference vs. value for 0 H group is indicated: ^, p<0.01.
Fig 5.
Relative expression of fetal liver acute phase protein mRNA levels measured with quantitative PCR.
CRP, C-reactive protein; SAA3, Serum amyloid A 3; HEP, Hepcidin. Values of CRP and SAA3 are presented as bar charts with the group mean normalised expression vs. the value for Control group, with error bars representing SD. Values of HEP are presented as box plots with the group median normalised expression vs. the value for Control group, with whiskers representing maximum and minimum values. Significant differences vs. value for Control group are indicated: ‡, p<0.01.
Fig 6.
Relative expression of inflammatory cytokine/chemokine mRNA levels in fetal tissues measured with quantitative PCR.
All values are presented as bar charts with the group mean normalised expression vs. the value for Control group, with error bars representing SD. Significant differences vs. value for Control group are indicated: ‡, p<0.01.
Fig 7.
Qualitative scoring of histological inflammation in fetal airspace.
Data are presented as box plots with the group median, with whiskers representing maximum and minimum values. Significant difference vs. value for Control group is indicated: ‡, p<0.01.
Table 2.
Comparison of MCP-1 protein concentration in fetal plasma measured with ELISA between the case E and the others.
Table 3.
Comparison of TNF-α protein concentration in fetal plasma measured with ELISA between the case E and the others.
Table 4.
Comparison of relative expression of inflammatory cytokine/chemokine mRNA levels in fetal circulating immunocytes immediately after induction of EVE therapy measured with quantitative PCR between the case E and the others.
Table 5.
Comparison of relative expression of inflammatory cytokine/chemokine mRNA levels in fetal circulating immunocytes 12 hours after induction of EVE therapy measured with quantitative PCR between the case E and the others.
Table 6.
Comparison of relative expression of fetal liver acute phase protein mRNA levels measured with quantitative PCR between the case E and the others.
Table 7.
Comparison of relative expression of fetal pulmonary inflammatory cytokine/chemokine mRNA levels measured with quantitative PCR between the case E and the others.
Table 8.
Comparison of relative expression of fetal brain inflammatory cytokine/chemokine mRNA levels measured with quantitative PCR between the case E and the others.
Table 9.
Comparison of relative expression of fetal skin inflammatory cytokine/chemokine mRNA levels measured with quantitative PCR between the case E and the others.
Table 10.
Comparison of qualitative scoring of histological inflammation in fetal airspace.
Fig 8.
Relative expression of surfactant protein (SP) mRNA levels measured with quantitative PCR.
All values are presented as bar charts with the group mean normalised expression vs. the value for Control group, with error bars representing SD. Significant difference was observed in no SP mRNA expression level between the two groups.