Spectramesh

In order to verify the enhancement of bone marrow stem cell mobilization and homing into brain, a bone marrow niche sample was removed from the long bones of adult male donor GFP mice as previously reported [41 (link)]. Both ends of the femur and tibia were penetrated using a syringe with a 25-gauge needle, and the marrow was flushed out with sterile saline. Total marrow from one femur was diluted to 1 mL then strained through a 30-µm Spectramesh (Fisher Scientific). Before bone marrow transplantation, recipient wild type (C57BL/6 mice-PAC1^+/+ mice) and PAC1^−/− mice [a generous gift provided by DKFZ Dr. Schutz [42 (link)]] underwent whole body gamma irradiation with ¹³⁷Cs using a Gammacell 40 irradiator (MDS Nordion). A total dose of 9 Gy (900 rads) was administered to ablate the whole bone marrow. The mice received rescuing bone marrow transplantations within 24 hours of irradiation. Donor bone marrow was injected into the recipient animal's tail vein as an 80 µL cell suspension containing 3 × 10⁶ cells. At 3 weeks after transplantation, mice were anesthetized with chloral hydrate (0.3 g/kg, i.p.) and subjected to right MCA ligation and right CCAs clamping for 120 minutes, as previously described with modification [24 (link)]. Then, experimental mice were injected intraperitoneally with PACAP38 (50 μg/kg) and vehicle control.