Indomethacin Modulation of Hematopoietic Stem Cells

For the HSC Replicate 1 experiment, we used the following mouse strain (#016617) that was obtained from Jackson labs but bred in-house. For external stimulant treatments, male and female mice (8–10 weeks) were ordered from Jackson labs (strain CD 45.2 [Ly5.2], #00664). Mice were kept for at least 1 week in the animal facility before initiating experiments and allocated at random (by cage) into experimental groups. Indomethacin (Sigma, 6 mg/l) was administered for 7 days in acidified drinking water to maintain stability (Curry et al., 1982 (link); Praticò et al., 2001 (link)). Indomethacin supplemented drinking water was changed every other day. Mice were injected with the following drugs and euthanized after 2 hr: poly(I:C) HMW (Invivogen), IP injection 10 mg/kg (Pietras et al., 2014 (link)). G-CSF Recombinant Human Protein (Thermo Fisher), IP injection, 0.25 mg/kg (Morrison et al., 1997 (link)). dmPGE₂ (Cayman), SC injection, 2 mg/kg (Hoggatt et al., 2013 (link)). Mice were weighed before injection and injection volume was adjusted to ensure equal dose between individual mice. The ‘control’ condition from the external stimulant treatments was also used as the second independent biological replicate of unperturbed HSCs (HSC Replicate 2). All animal procedures were approved by the Harvard University Institutional Animal Care and Use Committee.