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Intravital Imaging of Axonal Interactions with Microglia and Macrophages in a Mouse Dorsal Column Crush Injury
Published on: November 23, 2014
A Combination of Ex vivo Diffusion MRI and Multiphoton to Study Microglia/Monocytes Alterations after Spinal Cord
Harun N Noristani1,2, Hassan Boukhaddaoui1, Guillaume Saint-Martin2,3
1Institut National de la Santé et de la Recherche Médicale, U1051Montpellier, France.
Abstract:
Central nervous system (CNS) injury has been observed to lead to microglia activation and monocytes infiltration at the lesion site. Ex vivo diffusion magnetic resonance imaging (diffusion MRI or DWI) allows detailed examination of CNS tissues, and recent advances in clearing procedures allow detailed imaging of fluorescent-labeled cells at high resolution. No study has yet combined ex vivo diffusion MRI and clearing procedures to establish a possible link between microglia/monocytes response and diffusion coefficient in the context of spinal cord injury (SCI). We carried out ex vivo MRI of the spinal cord at different time-points after spinal cord transection followed by tetrahydrofuran based clearing and examined the density and morphology of microglia/monocytes using two-photon microscopy. Quantitative analysis revealed an early marked increase in microglial/monocytes density that is associated with an increase in the extension of the lesion measured using diffusion MRI. Morphological examination of microglia/monocytes somata at the lesion site revealed a significant increase in their surface area and volume as early as 72 hours post-injury. Time-course analysis showed differential microglial/monocytes response rostral and caudal to the lesion site. Microglia/monocytes showed a decrease in reactivity over time caudal to the lesion site, but an increase was observed rostrally. Direct comparison of microglia/monocytes morphology, obtained through multiphoton, and the longitudinal apparent diffusion coefficient (ADC), measured with diffusion MRI, highlighted that axonal integrity does not correlate with the density of microglia/monocytes or their somata morphology. We emphasize that differential microglial/monocytes reactivity rostral and caudal to the lesion site may thus coincide, at least partially, with reported temporal differences in debris clearance. Our study demonstrates that the combination of ex vivo diffusion MRI and two-photon microscopy may be used to follow structural tissue alteration. Lesion extension coincides with microglia/monocytes density; however, a direct relationship between ADC and microglia/monocytes density and morphology was not observed. We highlighted a differential rostro-caudal microglia/monocytes reactivity that may correspond to a temporal difference in debris clearance and axonal integrity. Thus, potential therapeutic strategies targeting microglia/monocytes after SCI may need to be adjusted not only with the time after injury but also relative to the location to the lesion site.
Insights
Central nervous system (CNS) injury triggers microglia and monocyte responses. Combining diffusion MRI and clearing techniques revealed lesion expansion linked to cell density, but not directly to cell morphology, in spinal cord injury (SCI).
Area of Science:
- Neuroscience
- Biomedical Imaging
- Cellular Biology
Background:
- Central nervous system (CNS) injury, particularly spinal cord injury (SCI), leads to microglia activation and monocyte infiltration.
- Ex vivo diffusion MRI and tissue clearing offer advanced methods for examining CNS tissue and cellular responses at high resolution.
Purpose of the Study:
- To investigate the relationship between microglia/monocyte response and diffusion coefficient changes after SCI.
- To combine ex vivo diffusion MRI with tissue clearing and two-photon microscopy to analyze cellular responses and tissue alterations.
Main Methods:
- Ex vivo diffusion MRI of rat spinal cords post-transection at various time points.
- Tetrahydrofuran-based tissue clearing for high-resolution imaging.
- Two-photon microscopy to quantify microglia/monocyte density, morphology, and distribution.
- Analysis of apparent diffusion coefficient (ADC) in relation to cellular changes.
Main Results:
- Early increase in microglia/monocyte density correlated with increased lesion extension measured by diffusion MRI.
- Significant increase in microglia/monocyte soma surface area and volume observed as early as 72 hours post-injury.
- Differential microglia/monocyte reactivity observed rostral and caudal to the lesion site, suggesting distinct temporal responses.
Conclusions:
- The combination of ex vivo diffusion MRI and two-photon microscopy effectively tracks structural tissue alterations after SCI.
- Lesion extension is associated with microglia/monocyte density, but not directly with their morphology or ADC values.
- Therapeutic strategies targeting microglia/monocytes in SCI may require adjustments based on both time post-injury and location relative to the lesion site.
