Astrocyte-Mediated Myelin Phagocytosis in Ischemia

The myelin sheath-supporting cells, oligodendrocytes and Schwann cells, are susceptible to ischemia. Therefore, ischemic stroke is usually characterized by evident demyelination [1]. Although oligodendrocytes are the major player in myelination, glial cells, including microglia and astrocytes, also contribute to myelination and demyelination [2]. In the early stages of ischemic stroke, microglia-mediated myelin debris removal through phagocytosis has been widely studied [3]. However, the role of astrocytes in myelin phagocytosis, especially in demyelination at the later phase of brain injury, remains unclear. In a recent study, Wan et al. elucidated the mechanisms underlying astrocytic phagocytosis-mediated demyelination in ischemic stroke [4] (Fig. 1).

Fig. 1.

Cytosolic LCN2/LRP1 pathway-mediated astrocytic myelin phagocytosis in ischemia. After an ischemic injury, cytosolic LCN2 and LRP1 were markedly increased in the non-ischemic corpus callosum. The increased LCN2 and LRP1 improved astrocyte-mediated myelin phagocytosis, and Lcn2 ablation or Lrp1 knockdown attenuated demyelination and reversed white matter damage.

The authors hypothesized that lipocalin-2 (LCN2) plays an essential role in astrocyte-mediated myelin phagocytosis after ischemic injury. To test their hypothesis, Wan et al. first analyzed whether ischemic stroke causes demyelination in the non-ischemic corpus callosum using a mouse model of focal cortical ischemia [4]. Immunostaining and immunoblotting showed significantly lower levels of myelinated fiber markers following ischemic stroke. In addition, significant thinning of myelin sheaths and a remarkably decreased length of myelinated axons were found using electron microscopy in mice with stroke. These findings demonstrated the significant compromise of myelination in the non-ischemic areas of the corpus callosum following ischemic injury.

Reactive astrocytes play an indispensable role in several myelination-related processes, including synaptic pruning during development and myelin damage in many brain disorders. Therefore, the authors next examined the role of astrocytes in the demyelinating corpus callosum following ischemic injury [4]. Their results showed that ischemic injury shifted astrocytes from a quiescent to an activated state. In addition, co-staining using markers of lysosomes, astrocytes, and demyelinated/myelinated fibers revealed engulfed myelin debris within activated astrocytes, displaying that reactive astrocytes have phagocytic effects in ischemic stroke. The electron microscope analysis also showed myelin-like structures within the cytoplasm of astrocytes. Moreover, astrocyte culture validated astrocytic myelin phagocytosis after applying LPS. In detail, LPS treatment induced an acute inflammatory response in astrocytes and increased myelin-laden within LPS-treated astrocytes.

Because LCN2 is a strong marker of active astrocytes and contributes to the acute damage of white matter in other brain disorders, the authors then examined the role of LCN2 in astrocyte-mediated myelin phagocytosis [4]. They found that LCN2 is mainly expressed in astrocytes and is associated with the activation status of astrocytes. Moreover, significantly increased LCN2 levels occur in astrocytes containing myelin debris. To better understand the role of LCN2 in astrocyte activation and myelin phagocytosis, Lcn2^-/- mice were investigated. Interestingly, the astrocytic reactivity and astrocytic phagocytosis were significantly suppressed in Lcn2-knockout mice. These results were substantiated in LPS-exposed astrocytes with Lcn2 deficiency. Furthermore, consistent with these findings, Lcn2 knockout alleviated demyelination in the non-ischemic areas of the corpus callosum and attenuated the cognitive deficits following focal cortical ischemia. These findings suggest that LCN2 is necessary for reactive astrogliosis and astrocyte-mediated myelin phagocytosis both in vitro and in vivo. To exclude the effects of secreted LCN2, the authors re-expressed cytosolic LCN2 by inducing a ∆2-20 Lcn2 mutation in cultured astrocytes and Lcn2-knockout mice. Intriguingly, they found that the cytosolic re-expression of LCN2 promoted the activation of astrocytes and progressively increased astrocyte-mediated myelin phagocytosis, indicating that astrocytic LCN2 is involved in a specific mechanism for astrocyte-mediated myelin phagocytosis in ischemia.

Because LRP1 is an essential receptor promoting macrophage survival and mediating phagocytosis [5], the authors proceeded to its role in astrocytic LCN2-mediated myelin phagocytosis. Immunostaining and co-immunoprecipitation results confirmed the co-colocalization and interaction between LRP1 and LCN2 within astrocytes. Moreover, both focal ischemia in vivo and LPS treatment in vitro elevated the expression of LRP1 and promoted the interaction between LRP1 and LCN2, indicating that LCN2/LRP1 signaling is essential in astrocyte-mediated myelin phagocytosis.

To further confirm the role of LCN2/LRP1 signaling in astrocyte-mediated myelin phagocytosis, Lrp1 was knocked down before ischemia surgery [4]. Interestingly, the phagocytic function of astrocytes and the levels of myelin debris were significantly reduced in the Lrp1-knockdown group. Furthermore, to understand the role of LRP1 in astrocyte-mediated myelin phagocytosis under LPS treatment in vitro, Lrp1 was knocked down in primary astrocytes. Consistent with the in vivo experiments, LRP1 repression blocked astrocyte-mediated myelin phagocytosis in both primary WT astrocytes and Lcn2^-/- astrocytes with resumed cytosolic LCN2 levels. Finally, the authors also confirmed that the demyelination was significantly suppressed by Lrp1 knockdown, as evidenced by significantly increased myelinated area, myelin sheath thickness, and myelinated axons.

In summary, Wan et al. found that LCN2 and LRP1 were markedly increased following ischemic injury and confirmed astrocytic LCN2/LRP2 signaling as essential in mediating astrocytic astrocyte-mediated myelin phagocytosis [4]. Their study fills a critical knowledge- gap in understanding astrocyte-mediated myelin phagocytosis and uncovers the underlying mechanism of LCN2/LRP1 signaling-mediated myelin phagocytosis. Based on their exciting findings, several important questions are worth further investigation. First, axonal loss and demyelination are critical aspects of the pathogenesis of several neurodegenerative diseases, including Alzheimer’s disease and multiple sclerosis [6, 7]. However, little is known about the astrocyte LCN2/LRP1 signaling in these neurological diseases. Therefore, it would be interesting to explore the role of LCN2/LRP1 signaling in demyelination-related brain disorders. Second, astrocyte-mediated myelin phagocytosis and microglia-mediated pruning play essential roles in brain remodeling [8, 9]. The microglia-mediated synapse pruning plays a crucial role in regulating synaptic remodeling by removing unnecessary neural connections during early brain development [8]. However, astrocyte-mediated myelin phagocytosis may contribute to myelin damage and lesion pathology in neurodegenerative disorders and brain injury during postnatal development and adulthood [4]. The role of LCN2/LRP1 signaling in these processes and the interplay between astrocyte-mediated myelin phagocytosis and microglia-mediated pruning deserve further investigation. Finally, according to a previous study, LCN2 can also be released by injured neurons and serve as a “help-me” signal to protect against neuronal injury [10]. Therefore, the role of neuronal LCN2 in astrocyte-mediated myelin phagocytosis should be clarified. Unraveling these mechanisms related to neuronal or astrocytic LCN2 may help identify potential therapeutic targets for neurodegenerative and neurological development-related diseases.

Conflict of interest

The authors declare no competing financial interests.