Dynamic changes of CX3CL1/CX3CR1 axis during microglial activation and motor neuron loss in the spinal cord of ALS mouse model

Background: Neuron-microglia communication plays a crucial role in the motor neurons (MNs) death in amyotrophic lateral sclerosis (ALS). Neurons can express chemokine (C-X3-C motif) ligand 1 (CX3CL1), which mediates microglial activation via interacting with its sole receptor CX3CR1 in microglia. In the present study, we aimed to investigate the dynamic changes of CX3CL1/CX3CR1 axis during microglial activation and MNs loss in SOD1(G93A) mouse model of ALS. Methods: qPCR, western blot and immunofluorescent staining were used to examine the mRNA and protein levels and localization of CX3CL1/CX3CR1 in the anterior horn region of spinal cord in both SOD1(G93)(A) mice and their age-matched wild type (WT) littermates at 40, 60, 90 and 120 days of age. The M1/M2 microglial activation in the spinal cord tissues of SOD1(G93A) mice and WT mice were evaluated by immunofluorescent staining of M1/M2 markers and further confirmed by qPCR analysis of M1/M2-related cytokines. Results: The immunofluorescent staining revealed that CX3CL1 was predominately expressed in MNs, while CX3CR1 was highly expressed in microglia in the anterior horn region of spinal cord. Compared with age-matched WT mice, CX3CL1 mRNA level was elevated at 40 days but decreased at 90 and 120 days in the anterior horn region of spinal cords in ALS mice. Consistently, CX3CR1 mRNA level was increased at 90 and 120 days. Western blot assay further confirmed the dynamic changes of CX3CL1/CX3CR1 axis in ALS mice. Additionally, the levels of M1/M2 markers of microglia and their related cytokines in the anterior horn region of spinal cord in ALS mice were increased at 90 and 120 days. Moreover, while M1-related cytokines in ALS mice were persistently increased at 120 days, the upregulated M2-related cytokines started to decline at 120 days, suggesting an altered microglial activation. Conclusions: Our data revealed the dynamic changes of CX3CL1/CX3CR1 axis and an imbalanced M1/M2 microglial activation during ALS pathological progression. These findings may help identify potential molecular targets for ALS therapy.