Gw2580

GW2580 (LC Laboratories) was administered once daily by oral gavage at a dose of 80 mg/kg in 0.1% Hydroxipropylmethylcellulose/0.1% Tween-20 as described previously (62 (link)). Treatment started 4 days prior HLI induction. Body weight was measured daily to exclude weight loss. Anti-CSF-1 (aCSF-1) antibody treatment (Clone 5A1, BioXCell) or PBS control was subcutaneously injected in the ischemic limb, at a dose of 50 μg, immediately after surgery or up to 3 days (for details see Figure 3H).

Oral treatment started immediately after SCI and ended 6 weeks post-lesion when the last behavioral analysis protocol was completed (Figure 1A). Mice were fed with a regular chow (A04, maintenance diet, SAFE diet, Augy, France) or with the same diet containing 0.1% GW2580 (LC Laboratories, Woburn, MA, USA). GW2580 food pellets were prepared as previously described [10 (link),11 (link)]. Food intake was monitored daily throughout the study duration.

The CSF1R kinase inhibitor GW2580 (LC Laboratories) was suspended in 0.5% hydroxypropylmethylcellulose and 0.1% Tween 20 using a Teflon glass homogenizer. Diluent control or 160 mg/kg GW2580 was administered daily for 4 d by oral gavage before mice were culled on day 5.

For gain of function experiments, NBH (control) and ME7 (prion) macgreen mice were treated at 12 weeks post-injection with 50 ng of recombinant murine CSF1 (Merck Chemicals; n = 4), murine IL34 (R&D Systems; n = 4) or saline (vehicle; n = 4) by stereotactic injection in the dorsal hippocampus (CA1 field; anteroposterior, −2.0 mm; lateral, −1.7 mm; depth, 1.6 mm) with a Hamilton syringe. Mice received 5 injections of i.p. BrdU (Sigma–Aldrich; 7.5 mg/ml, 0.1 ml/10 g weight in sterile saline), before the end of the experiment (+1 week), as previously described (Gomez-Nicola et al., 2013 (link)).
Inhibition of the tyrosine kinase activity of CSF1R was achieved by the administration of GW2580, as previously described (Gomez-Nicola et al., 2013 (link)). GW2580 (LC Laboratories) was suspended in 0.5% hydroxypropylmethylcellulose and 0.1% Tween 80 and was dosed by oral gavage, using gavaging needles, at 0.2 ml per mouse (75 mg/kg), daily for 4 consecutive weeks (from 14th to 18th week post-injection), to NBH and ME7 mice (n = 8), using the vehicle as control (n = 8). Mice weight was monitored during the experiment. Mice received 2 daily injections of i.p. BrdU (Sigma–Aldrich; 7.5 mg/ml, 0.1 ml/10 g weight in sterile saline), before the end of the experiment (18th week).

The GW2580 (LC Laboratories) was used as an inhibitor to inhibit the CSF1R cascade as described by previous studies (34 (link), 35 (link)). GW2580 was suspended in 0.5% hydroxypropyl methylcellulose and 0.1% Tween 80. The solution was dosed orally to mouse (at the body weight of 75 mg/kg) after ischemia at a daily interval for 3 days.