Infiltrating Blood-Derived Macrophages Are Vital Cells Playing an Anti-inflammatory Role in Recovery from Spinal Cord Injury in Mice

Using a mouse model of spinal injury, Michal Schwartz and colleagues tested the effect of macrophages on the recovery process and demonstrate an important anti-inflammatory role for a subset of infiltrating monocyte-derived macrophages that is dependent upon their expression of interleukin 10

Selective killing of spinal cord neural stem cells impairs locomotor recovery in a mouse model of spinal cord injury

Abstract Background Spinal cord injury (SCI) is a devastating condition mainly deriving from a traumatic damage of the spinal cord (SC). Immune cells and endogenous SC-neural stem cells (SC-NSCs) play a critical role in wound healing processes, although both are ineffective to completely restore tissue functioning. The role of SC-NSCs in SCI and, in particular, whether such cells can interplay with the immune response are poorly investigated issues, although mechanisms governing such interactions might open new avenues to develop novel therapeutic approaches. Methods We used two transgenic mouse lines to trace as well as to kill SC-NSCs in mice receiving SCI. We used Nestin CreERT2 mice to trace SC-NSCs descendants in the spinal cord of mice subjected to SCI. While mice carrying the suicide gene thymidine kinase (TK) along with the GFP reporter, under the control of the Nestin promoter regions (NestinTK mice) were used to label and selectively kill SC-NSCs. Results We found that SC-NSCs are capable to self-activate after SCI. In addition, a significant worsening of clinical and pathological features of SCI was observed in the NestinTK mice, upon selective ablation of SC-NSCs before the injury induction. Finally, mice lacking in SC-NSCs and receiving SCI displayed reduced levels of different neurotrophic factors in the SC and significantly higher number of M1-like myeloid cells. Conclusion Our data show that SC-NSCs undergo cell proliferation in response to traumatic spinal cord injury. Mice lacking SC-NSCs display overt microglia activation and exaggerate expression of pro-inflammatory cytokines. The absence of SC-NSCs impaired functional recovery as well as neuronal and oligodendrocyte cell survival. Collectively our data indicate that SC-NSCs can interact with microglia/macrophages modulating their activation/responses and that such interaction is importantly involved in mechanisms leading tissue recovery

Additional file 6: of Selective killing of spinal cord neural stem cells impairs locomotor recovery in a mouse model of spinal cord injury

Movie S2 Video of Catwalk gait analysis of a representative GCV-NestinTK mouse. NestinTK mice were treated with GCV, subjected to SCI and followed for locomotor recovery by BMS score when behavioral amelioration reached a plateau value, mice were subjected to Catwalk gait analysis. The video shows recording of a representative GCV-NestinTK animal (BMS score = 1). Free ambulation along an illuminated glass plate in a darkened room has been recorded for 3.22 s. Video has been captured at day 18 after SCI. (AVI 104621 kb

Transplanted neural stem/precursor cells instruct phagocytes and reduce secondary tissue damage in the injured spinal cord

Transplanted neural stem/precursor cells possess peculiar therapeutic plasticity and can simultaneously instruct several therapeutic mechanisms in addition to cell replacement. Here, we interrogated the therapeutic plasticity of neural stem/precursor cells after their focal implantation in the severely contused spinal cord. We injected syngeneic neural stem/precursor cells at the proximal and distal ends of the contused mouse spinal cord and analysed locomotor functions and relevant secondary pathological events in the mice, cell fate of transplanted neural stem/precursor cells, and gene expression and inflammatory cell infiltration at the injured site. We used two different doses of neural stem/precursor cells and two treatment schedules, either subacute (7 days) or early chronic (21 days) neural stem/precursor cell transplantation after the induction of experimental thoracic severe spinal cord injury. Only the subacute transplant of neural stem/precursor cells enhanced the recovery of locomotor functions of mice with spinal cord injury. Transplanted neural stem/precursor cells survived undifferentiated at the level of the peri-lesion environment and established contacts with endogenous phagocytes via cellular-junctional coupling. This was associated with significant modulation of the expression levels of important inflammatory cell transcripts in vivo. Transplanted neural stem/precursor cells skewed the inflammatory cell infiltrate at the injured site by reducing the proportion of 'classically-activated' (M1-like) macrophages, while promoting the healing of the injured cord. We here identify a precise window of opportunity for the treatment of complex spinal cord injuries with therapeutically plastic somatic stem cells, and suggest that neural stem/precursor cells have the ability to re-programme the local inflammatory cell microenvironment from a 'hostile' to an 'instructive' role, thus facilitating the healing or regeneration past the lesion

Additional file 7: of Selective killing of spinal cord neural stem cells impairs locomotor recovery in a mouse model of spinal cord injury

Figure S5. macrophages/macrophages activation affects both GCV-WT and GCV-NestinTK mice. (A) Histological analysis of a SC region located 3 mm far from the injury site from a GCV-WT mice (18 days post injury). Microglia/macrophages are labeled for Iba1 and F4/80. Arrowhead indicates cells that are shown at high magnification in panel A’. Panel B shows the site of the injury in the SC of GCV-WT mouse. Arrowhead indicates cells that are shown at high magnification in panel B′. A representative section of the SC located 3 mm far from the site of the injury from a GCV-NestinTK mouse (18 days post injury) is shown in panel C. Arrowhead indicates cells that are shown at high magnification in panel C′. Panel D shows the site of the injury while the arrowhead indicates cells that are shown at high magnification in D’ (n = 3 for each group). Scale bar 50 μm (TIFF 4124 kb

Additional file 3: of Selective killing of spinal cord neural stem cells impairs locomotor recovery in a mouse model of spinal cord injury

Figure S3. GCV treatment ablates proliferating SC-eNSCs in NestinTK mice. Panels A and B show representative confocal images of VIM (red) and Brdu (green) in the ependymal layer of GCV-WT (A) and GCV-NestinTK (B) mice (n = 3 for each group). Mice were sacrificed at the end of the GCV treatment. Quantifications (means ± S.E.M.) are shown in panel C. Two-way ANOVA followed by Bonferroni’s multiple Comparison test has been used to analyze data. ** p = 0.011 and p = 0.045 in thoracic and lumbar segments, respectively. Scale bar 20 μm. (TIFF 2754 kb

Additional file 2: of Selective killing of spinal cord neural stem cells impairs locomotor recovery in a mouse model of spinal cord injury

Figure S2. Sorted GFP+ cells from Nestin floxGFPflox-TK mice give rise to neurospheres. Panel A and B show the gating strategy for sorting GFP+ cells from SCs bulk cultures obtained from Nestin floxGFPflox-TK mice. WT litters (A) were used to set up the gating strategy that we used to sort GFP+ cells (B). GFP+ cells were plated at the density of 8000 cells/cm2 and daily examined for the presence of neurospheres. Small spheres were observed after 3 days (C), while spheres with diameters larger than 100 μm were easily observed after 7 days (D). Scale bar 50 μm (n = 3 independent preparations). (TIFF 9717 kb

Additional file 5: of Selective killing of spinal cord neural stem cells impairs locomotor recovery in a mouse model of spinal cord injury

Movie S1 Video of Catwalk gait analysis of a representative GCV- WT mouse. WT mice were treated with GCV, subjected to SCI and followed for locomotor recovery by BMS score when behavioral amelioration reached a plateau value mice, were subjected to Catwalk gait analysis. The video shows recording of a representative GCV-WT animal (BMS score = 3). Free ambulation along an illuminated glass plate in a darkened room has been recoded for 4.30 s. Video has been captured at day 18 after SCI. (AVI 139494 kb