Platelet-specific P2Y1 receptor deficient mice have suppressed pulmonary leukocyte recruitment in response to lipopolysaccharide

A role for the P2Y1 receptor (P2Y1R) in platelet-driven inflammation has been established using a pharmacological approach, limited to an acute 4 h time span. Nucleotide-structure P2Y1R antagonists have restricted experimental use due to inadequate pharmacokinetics, and an inability to decipher global versus cell specific effects in vivo. The creation of a conditional knock out (platelet) P2Y1R transgenic mouse model was designed to overcome these restrictions. A homozygous P2Y1 LoxP mouse colony was created using CRISPR/Cas9 technology, and crossed with a hemizygous P2Y1 LoxP with PF4-cre to provide offspring that are homozygous for P2Y1 LoxP flanked allele, and hemizygous for the PF4cre (platelet P2Y1−/−) and offspring homozygous for P2Y1 LoxP flanked allele, but non-carriers for PF4cre (control mice). Animals were intranasally administered LPS to induce pulmonary inflammation to assess the influence of phenotype on leukocyte recruitment. 24 h post intranasal LPS administration; pulmonary neutrophil and platelet recruitment were significantly suppressed, despite the fact that neutrophils retained the ability to migrate to fMLP ex vivo. Circulating platelet and leukocyte numbers were not different between control and platelet P2Y1−/− animals. Tail bleeding times revealed the platelet P2Y1−/− mice had a severe bleeding phenotype. The platelet specific P2Y1−/− mouse model confirms the importance of platelet P2Y1R in the regulation of inflammatory responses. A 60–70% inhibition of leukocyte recruitment over an extended time period was observed compared to previous pharmacological studies. Platelet P2Y1−/− mice will help further elucidate the mechanisms by which P2Y1R regulate platelet activation during inflammation. • Mice selectively deficient in the platelet P2Y1R have suppressed pulmonary leukocyte and platelet recruitment during inflammation. • We provide a methodology to determine mechanistic relevance that is otherwise limited by pharmacological approaches.