Supplementary Material for: Investigating the Mechanisms of Reduced Blood Transfusions after Delayed Umbilical Cord Clamping: The TITANS causal mediation analysis

Introduction
Delaying clamping of the umbilical cord (DCC) in preterm infants reduces mortality and need for blood transfusions. The mechanisms leading to these benefits are not well understood. The TITANS study investigates potential mediators of the reduction in blood transfusions in infants who received DCC.

Materials and Methods
Additional patient data was sourced from Australian and New Zealand sites from the Australian Placental Transfusion Study (APTS). APTS randomized preterm infants <30 weeks’ gestation to receive DCC (60 seconds) or immediate-cord-clamping (ICC). We examined whether placental transfusion or initial severity of illness mediated the reduced requirement for blood transfusions for infants randomized to DCC. Peak haematocrit in the first 7 days (Hct) was used as an indicator of placental transfusion quantity. Cumulative blood sampled, mechanical ventilation, and arterial sampling lines were used as indicators of severity if illness. We quantified the natural indirect effect of peak Hct and then for all mediators in a joint model with sequential mediation.

Results
Data from 1260 (of 1401) Australian and New Zealand APTS infants were obtained. The effect of DCC on subsequent blood transfusion was mediated through peak Hct (indirect effect OR = 0.85, 95%CI:0.79-0.93; p<0.001), which accounted for 37% of the total effect. Indicators of severity of illness did not mediate the effect independently of peak Hct.

Discussion/Conclusion
Peak Hct mediated some, but not all, of the effect of DCC on blood transfusion, whereas markers of severity of illness were not independent mediators.