Figure 2.
Adoptive transfer of LPS-primed monocytes isolated from preconditioned donors decreases infarct volume in stroke recipient mice. A, Experimental design for adoptive transfer of monocytes in stroke mice. Mice were injected with either LPS or saline and monocytes were isolated from BM by immunodepletion 24 h later. Saline or LPS monocytes (5 × 105 monocytes/100 μl) were injected intravenously into stroke mice 24 h before transient MCAO or 7 or 24 h after MCAO. Infarct volume was determined by cresyl violet staining 72 h after MCAO. B, Isolated BM cells were stained for CD11b, CD115, Ly6G, CD117, and Ly6C markers before (start) and after (purified) immunodepletion. Stained cells were assayed by flow cytometry. Monocytes were isolated with high purity as evidenced by the expression of CD11b, CD115, and Ly6C markers and lack of expression of the neutrophil marker Ly6G and the hematopoietic stem cell marker CD117. C, Representative images of brain infarcts in saline-injected mice (SAL), LPS-preconditioned mice (LPS), or mice that received monocytes isolated from LPS-preconditioned mice 24 h before MCAO (−24 h) or 7 or 24 h after MCAO (+7 h and +24 h, respectively). Mice that received monocytes from saline-injected mice are indicated as SAL +7 h and mice that received neutrophils isolated from LPS-injected mice are indicated as PMN +7 h. The infarct (pale areas) is outlined in brain coronal sections stained with cresyl violet 72 h after MCAO. AT, Adoptive transfer. D, Infarct volume was significant smaller in either LPS-preconditioned mice (LPS i.p.) or in mice receiving adoptively transferred LPS-monocytes at +7 h (n = 9–12). Statistical analysis: Kruskal–Wallis (H = 15.65, p = 0.0035) followed by Dunn's post hoc test. *p < 0.05; ***p < 0.001. E, Adoptive transfer of monocytes isolated from saline-injected mice or neutrophils (PMN) from LPS-preconditioned mice at +7 h after MCAO did not reduce infarct volumes (n = 8–11). Statistical analysis: one-way ANOVA F(3,32) = 7.747, p = 0.0006, followed by Holm–Sidak's post hoc test, *p < 0.05.