Background <p>Platelet transfusions are frequently given to preterm neonates. However, observational and randomized studies have reported higher morbidity and mortality associated with increased numbers of transfusions. We hypothesized that this was related to pro-inflammatory effects of transfused platelets. This study assessed the effects of platelet transfusions on bleeding, plasma cytokines, and neutrophil extracellular trap (NET) levels in neonates.</p> Methods <p>This was a prospective cohort study of thrombocytopenic neonates conducted from 2020 to 2024. Bleeding was assessed with each platelet count. Blood was collected before and 2 h or 4 h after platelet transfusion for evaluation of a plasma cytokine panel and NETs. Aliquots from transfused units were also collected.</p> Results <p>Forty-two infants with severe thrombocytopenia received 68 platelet transfusions. Transfusion increased the platelet count by 15 ± 3 × 10⁹/L but had no significant effect on bleeding. Four hours post-transfusion, RANTES levels increased 6.5-fold (<i>p</i> = 0.03) and were higher than expected based on the quantity in the units. Plasma NETs increased 1.24-fold (<i>p</i> = 0.007) post-transfusion. The transfused platelet units’ storage time and free mtDNA concentrations correlated with RANTES increases (<i>p</i> = 0.04 and 0.03, respectively).</p> Conclusions <p>Platelet transfusions are associated with increases in inflammatory cytokines and NET in neonates, which may contribute to the negative outcomes associated with platelet transfusions.</p> Impact <p><UnorderedList Mark="Bullet"> <ItemContent> <p>In a recent randomized trial, liberal platelet transfusions were associated with increased bleeding and/or mortality in preterm neonates. We hypothesized that these findings would be related to the pro-inflammatory effects of platelet transfusions.</p> </ItemContent> <ItemContent> <p>In a cohort of thrombocytopenic neonates, platelet transfusions were associated with increases in RANTES, a potent chemokine, and neutrophil extracellular traps (NETs).</p> </ItemContent> <ItemContent> <p>The transfused platelet units’ storage time and free mitochondrial DNA concentration correlated with the RANTES increase.</p> </ItemContent> <ItemContent> <p>This study identified novel transfusion-related inflammatory changes in neonates, which may contribute to the observed outcomes, and Blood Banking factors that might influence these effects.</p> </ItemContent> </UnorderedList></p>