Figure 1 - uploaded by Oswald Steward
Content may be subject to copyright.
Acute induction of Wnt1, Wnt4, Wnt5 and Frizzled1 in adult spinal cord gray matter and the cortex following lateral hemisection
Source publication
Failure of axon regeneration in the mammalian CNS is attributable in part to the presence of various inhibitory molecules, including myelin-associated proteins and proteoglycans enriched in glial scars. Here, we evaluate whether axon guidance molecules also regulate regenerative growth after injury in adulthood. Wnts are a large family of axon guid...
Contexts in source publication
Context 1
... sections were washed in blocking solution (0.3% Triton X-100, and 10% normal serum in 0.01M PBS (pH7.4)) and were incubated in the primary antibodies diluted in the same blocking solution overnight at 4°C followed by fluorescent secondary antibodies for 90 minutes at room temperature. The following primary antibodies were used in this study: mouse anti-GFAP (1:400; Invitrogen) or rabbit anti- GFAP (1:200, chemicon) to detect glial fibrillary acidic protein; mouse anti-SMI-31 (1:500, Sternberger) to detect axons (in Supplemental Figure 1); mouse anti-growth associated protein (GAP-43; 1:1,000, Chemicon) to label regenerating axons; mouse anti-ED-1 (1:300, Chemicon)) to microglia/ macrophages (in Supplemental Figure 2). The staining was visualized with appropriate secondary antibodies conjugated to TRIFC and FITC (Jackson). ...
Context 2
... investigate the role of Wnts in regulating axon regeneration after spinal cord injury, we examined the expression of all Wnt genes over a period of one month after spinal cord hemisection at the cervical level, C5 in mice ( Figure 1M). The uninjured adult spinal cord does not express any of the 19 members ( Figure 1B, 1D, 1G and data not shown). ...
Context 3
... investigate the role of Wnts in regulating axon regeneration after spinal cord injury, we examined the expression of all Wnt genes over a period of one month after spinal cord hemisection at the cervical level, C5 in mice ( Figure 1M). The uninjured adult spinal cord does not express any of the 19 members ( Figure 1B, 1D, 1G and data not shown). However, we found that a small number of Wnt genes were induced following injury. ...
Context 4
... we found that a small number of Wnt genes were induced following injury. Wnt1 and Wnt5a, potent repellents of developing corticospinal tract axons, were rapidly and robustly induced broadly both rostral and caudal to the lesion area one day after injury ( Figure 1A, 1C). Interestingly, Wnt1 and Wnt5a are also induced in the areas on the contralateral side of the injury site. ...
Context 5
... induction appears rapidly one day after injury and persists for seven days. Expression starts to decline after one week and by thirty days the levels of Wnt1 and Wnt5a mRNA decreased to background level ( Figure 1N). In addition to Wnt1 and Wnt5a, which are potent repellents for cortiospinal tract axons, we also observed rapid induction of Wnt4 in the gray matter surrounding the lesion, which decreases in levels within a week, much sooner than the decline of Wnt1 and Wnt5a ( Figure 1E and 1F). ...
Context 6
... starts to decline after one week and by thirty days the levels of Wnt1 and Wnt5a mRNA decreased to background level ( Figure 1N). In addition to Wnt1 and Wnt5a, which are potent repellents for cortiospinal tract axons, we also observed rapid induction of Wnt4 in the gray matter surrounding the lesion, which decreases in levels within a week, much sooner than the decline of Wnt1 and Wnt5a ( Figure 1E and 1F). Wnt4 does not appear to repel corticospinal tract axons (data not shown) ( Liu et al., 2005). ...
Context 7
... the induced Wnt4 is unlikely responsible for regeneration inhibition but may be responsible for axonal sprouting instead. Frizzled1 was found induced in similar regions immediately around the lesion ( Figure 1H) and declined at a similar rate as Wnt4. Frizzled1 mRNA is also rapidly and broadly induced in the cortex, particularly in the contralateral side to the injury, and declined within a week ( Figure 1I and 1N). ...
Context 8
... was found induced in similar regions immediately around the lesion ( Figure 1H) and declined at a similar rate as Wnt4. Frizzled1 mRNA is also rapidly and broadly induced in the cortex, particularly in the contralateral side to the injury, and declined within a week ( Figure 1I and 1N). Neither Wnt4 nor Frizzled1 are expressed in uninjured spinal adult spinal cord ( Figure 1G and 1J). ...
Context 9
... mRNA is also rapidly and broadly induced in the cortex, particularly in the contralateral side to the injury, and declined within a week ( Figure 1I and 1N). Neither Wnt4 nor Frizzled1 are expressed in uninjured spinal adult spinal cord ( Figure 1G and 1J). ...
Context 10
... should be noted that the sprouting of adult regenerating CST collateral branches to the contralateral gray matter following Ryk antibody injection is reminiscent of the reported response of young CST axons when lesion was introduced right after birth (Bregman et al., 1989). We did not observe CST axons within the lesion site (judging by the lack of BDA signals), although GAP-43-positive and SMI-31-positive regenerating axons of a currently unknown source were observed within the lesion site (supplemental Figure 1). We found that antibody treatment did not cause a significant change of the ED-1-positive cells at the lesion area (Supplemental Figure 2). ...
Context 11
... should be noted that after Ryk-antibody treatment, many CST axons were present right at the border of the lesion created by VibraKnife. Although the lesion gap was filled with cells and we observed Neurofilament-positive and GAP-43-positive and SMI-31-positive axons in the lesion site, no BDA-positive CST axons were present inside the lesion (Supplemental Figure 1). It is possible that additional inhibitors within the lesion site/scar tissue, such as Sema3A and CSPG, prevent axons from regenerating across the lesion. ...
Similar publications
Background:
Although constitutive activating mutations in the Wnt/β-catenin signalling pathway are important for colorectal cancer development, canonical signalling through Wnt ligands is essential for β-catenin activation. Here, we investigated the role of (pro)renin receptor ((P)RR), a component of the Wnt receptor complex, in the pathogenesis o...
The modular nature of the Wnt pathway provides multiple potential points of intervention to alter its activity. In this chapter
we review potential opportunities to inhibit Wnt signalling at various levels, including the synthesis and secretion of Wnt
ligands, activation of the Wnt receptor complex, degradation of cytoplasmic β-catenin and finally...
Wnt signaling is a key regulator of bone metabolism and fracture healing. The canonical Wnt/β-catenin pathway is regarded as the dominant mechanism, and targeting this pathway has emerged as a promising strategy for the treatment of osteoporosis and poorly healing fractures. In contrast, little is known about the role of non-canonical Wnt signaling...
Half of lung cancers are diagnosed in former smokers, leading to a significant treatment burden in this population. Chemoprevention in former smokers using the prostacyclin analogue iloprost reduces endobronchial dysplasia, a premalignant lung lesion. Iloprost requires the presence of the WNT receptor Frizzled 9 (Fzd9) for inhibition of transformed...
Surgical removal of the lens from larval Xenopus laevis results in a rapid transdifferention of central corneal cells to form a new lens. The trigger for this process is understood to be an induction event arising from the unprecedented exposure of the cornea to the vitreous humour that occurs following lens removal. The molecular identity of this...
Citations
... (CST) branches around the boundaries of the injured zone [34]. The behavioural consequences of antagonising Wnt5a-Ryk activity were further demonstrated by Miyashita et al., wherein SCI rats treated with anti-Ryk antibody exhibited CST axon growth and enhanced Basso, Beattie, and Bresnahan (BBB) scores indicative of functional motor recovery [35]. ...
Spinal cord injury (SCI) is a complex neurodegenerative pathology that consistently harbours a poor prognostic outcome. At present, there are few therapeutic strategies that can halt neuronal cell death and facilitate functional motor recovery. However, recent studies have highlighted the Wnt pathway as a key promoter of axon regeneration following central nervous system (CNS) injuries. Emerging evidence also suggests that the temporal dysregulation of Wnt may drive cell death post-SCI. A major challenge in SCI treatment resides in developing therapeutics that can effectively target inflammation and facilitate glial scar repair. Before Wnt signalling is exploited for SCI therapy, further research is needed to clarify the implications of Wnt on neuroinflammation during chronic stages of injury. In this review, an attempt is made to dissect the impact of canonical and non-canonical Wnt pathways in relation to individual aspects of glial and fibrotic scar formation. Furthermore, it is also highlighted how modulating Wnt activity at chronic time points may aid in limiting lesion expansion and promoting axonal repair.
... RYK has been observed to have a pathological association with the nervous system in diseases. In a C. elegans model, the intracellular region of the gene was found to inhibit the neuroprotective activity of the transcription factor FOXO in the early phases of Huntington's disease [12]. Furthermore, RYK was found to be upregulated in the motor neurons and ventral white matter of a mouse model of amyotrophic lateral sclerosis [13]. ...
The RYK gene encodes a receptor-like tyrosine kinase crucial for several biological processes, including development, tissue homeostasis, and cancer. This study utilized data from the Cancer Genome Atlas Project (TCGA) to evaluate RYK expression at both mRNA and protein levels in various cancers, determine its prognostic significance, and explore its involvement in cancer-related signaling pathways. Elevated levels of RYK mRNA were identified in cholangiocarcinoma (CHOL), pancreatic adenocarcinoma (PAAD), glioblastoma multiforme (GBM), lung squamous cell carcinoma (LUSC), brain lower grade glioma (LGG), head and neck squamous cell carcinoma (HNSC), liver hepatocellular carcinoma (LICH), esophageal carcinoma (ESCA), and colon adenocarcinoma (COAD), while RYK protein levels were observed to be increased in colon adenocarcinoma (COAD), GBM, LICH, cervical and endocervical adenocarcinoma (CESC), and breast invasive carcinoma (BRCA). Additionally, RYK overexpression correlated with poorer prognosis in several cancers, including PAAD, LICH, BRCA, ESCA, COAD, and CESC. Furthermore, RYK showed a positive correlation with the upregulation of multiple receptors and coreceptors in the WNT signaling pathway in various types of cancer. In terms of cancer-related signaling pathways, RYK was found to potentially interact with DNA damage, TSC/mTOR, PI3K/AKT, EMT, RTK, RAS/MAPK, ER hormone, AR hormone, and the cell cycle. This study provides new and previously unreported insights into the role of RYK in cancer biology.
... A spinal cord transection was performed during electrode implantation. After each spinal cord electrode was implanted, the spinal cord was completely transected at T9 using microscissors as described 68,69 . Complete transection lesions were verified postmortem by confirming the absence of neural tissues throughout the dorsoventral extent of the spinal cord 68,69 . ...
... After each spinal cord electrode was implanted, the spinal cord was completely transected at T9 using microscissors as described 68,69 . Complete transection lesions were verified postmortem by confirming the absence of neural tissues throughout the dorsoventral extent of the spinal cord 68,69 . Each sham operation was performed as follows: the dorsal vertebral lamina at spinal T9 segment was removed and the electrode was implanted into the spinal cord, but the spinal cord itself was left intact. ...
The neural signals produced by varying electrical stimulation parameters lead to characteristic neural circuit responses. However, the characteristics of neural circuits reconstructed by electrical signals remain poorly understood, which greatly limits the application of such electrical neuromodulation techniques for the treatment of spinal cord injury. Here, we develop a dual electrical stimulation system that combines epidural electrical and muscle stimulation to mimic feedforward and feedback electrical signals in spinal sensorimotor circuits. We demonstrate that a stimulus frequency of 10−20 Hz under dual stimulation conditions is required for structural and functional reconstruction of spinal sensorimotor circuits, which not only activates genes associated with axonal regeneration of motoneurons, but also improves the excitability of spinal neurons. Overall, the results provide insights into neural signal decoding during spinal sensorimotor circuit reconstruction, suggesting that the combination of epidural electrical and muscle stimulation is a promising method for the treatment of spinal cord injury.
... The copyright holder for this preprint this version posted November 5, 2023. ; https://doi.org/10.1101/2023.11.04.565649 doi: bioRxiv preprint DEGs of naive DRG with the 3,022 RAGs induced by PL (conditioning lesion) at 1dpi, revealing 61 overlapping genes (25%) that concerned Axonal extension, DNA binding/transcription (e.g., DNA unwinding, TFIIH core complex, 3'-5' DNA helicase activity, Double stranded DNA binding), Negative regulation of Wnt signaling (pro-axon regeneration 46,47 ), as well as Retinoid metabolic process and Phospholipase activity (Fig. 5f, S11a, Table S11). Heatmap highlighted that transcriptional changes of 10 RAGs in the GO terms related to axonogenesis (e.g., Klf8, Ncam1, Hoxd11, Hoxc8) as a result of Ahr ablation mimicked that of conditioning lesion (Fig. 5g). ...
Injured neurons sense environmental cues to balance neural protection and axon regeneration, but the mechanisms are unclear. Here, we unveil aryl hydrocarbon receptor (AhR), a ligand-activated bHLH-PAS transcription factor, as molecular sensor and key regulator of acute stress response at the expense of axon regeneration. We demonstrate responsiveness of DRG sensory neurons to ligand-mediated AhR signaling, which functions to inhibit axon regeneration. Ahr deletion mimics the conditioning lesion in priming DRG to initiate axonogenesis gene programs; upon peripheral axotomy, Ahr ablation suppresses inflammation and stress signaling while augmenting pro-growth pathways. Moreover, comparative transcriptomics revealed signaling interactions between AhR and HIF-1α, two structurally related bHLH-PAS α units that share the dimerization partner Arnt/HIF-1β. Functional assays showed that the growth advantage of AhR-deficient DRG neurons requires HIF-1α; but in the absence of Arnt, DRG neurons can still mount a regenerative response. We further unveil a link between bHLH-PAS transcription factors and DNA hydroxymethylation in response to peripheral axotomy, while neuronal single cell RNA-seq analysis revealed a link of the AhR regulon to RNA polymerase III regulation and integrated stress response (ISR). Altogether, AhR activation favors stress coping and inflammation at the expense of axon regeneration; targeting AhR can enhance nerve repair.
... The canonical Wnt/β-catenin signaling pathway is an essential signaling cascade that plays a key role in the regulation of neuronal differentiation, axonal extension, cell proliferation, and neuronal survival [9][10][11]. Several studies have reported that the activation of Wnt/β-catenin signaling after SCI promotes neuronal differentiation, axonal regeneration, and functional recovery [12][13][14][15]. Therefore, the targeted use of drugs to activate Wnt/β-catenin signaling may provide a novel approach for the recovery of spinal cord function in SCI. ...
Spinal cord injury (SCI) is an overwhelming and incurable disabling condition, for which increasing forms of multifunctional biomaterials are being tested, but with limited progression. The promising material should be able to fill SCI-induced cavities and direct the growth of new neurons, with effective drug loading to improve the local micro-organism environment and promote neural tissue regeneration. In this study, a double crosslinked biomimetic composite hydrogel comprised of acellularized spinal cord matrix (ASCM) and gelatin-acrylated-β-cyclodextrin-polyethene glycol diacrylate (designated G-CD-PEGDA) hydrogel, loaded with WAY-316606 to activate canonical Wnt/β-catenin signaling, and reinforced by a bundle of three-dimensionally printed aligned polycaprolactone (PCL) microfibers, was constructed. The G-CD-PEGDA component endowed the composite hydrogel with a dynamic structure with a self-healing capability which enabled cell migration, while the ASCM component promoted neural cell affinity and proliferation. The diffusion of WAY-316606 could recruit endogenous neural stem cells and improve neuronal differentiation. The aligned PCL microfibers guided neurite elongation in the longitudinal direction. Animal behavior studies further showed that the composite hydrogel could significantly recover the motor function of rats after SCI. This study provides a proficient approach to produce a multifunctional system with desirable physiological, chemical, and topographical cues for treating patients with SCI.
... The Wnt signaling pathway, which has been widely studied, regulates cell proliferation, migration, differentiation and survival during animal development as well as the maintenance of homeostasis and regeneration (Liu et al., 2008;De Robertis, 2010;Clevers and Nusse, 2012). In the trend analysis, we found that the Wnt signaling pathway was significantly enriched in profile 2, a gene set that was decreased at the early stage during exposure to juvenile oysters. ...
As a vital developmental event, metamorphosis controls the population dynamics of most marine invertebrates and affects the breeding of economic shellfish. Rapana venosa is an economically important species in China, but artificial aquaculture has hampered its metamorphosis process. Previous studies have found that juvenile oysters can effectively induce the metamorphosis of R. venosa, but the specific induction mechanism is not clear. Here, we investigated the mechanism underlying the response of R. venosa to juvenile oysters through the RNA-seq analysis. In this study, the gene set responses to metamorphosis cues (juvenile oysters) in R. venosa were identified, and GO and KEGG enrichment analyses were further performed on these gene sets. The results showed that the expression of the prototype of the class of immediate early genes, the transcription factor AP-1, was rapidly and significantly increased, and the molecular chaperone of NOS, HSP90, exhibited lower expression in the M12 group than in the control group. In contrast, the expression of inhibitors of apoptosis (IAPs) was significantly increased upon exposure to juvenile oysters. Additionally, the Wnt signaling pathway and MAPK signaling pathway were enriched in the trend analysis. These pathways may also play critical regulatory roles in the response to juvenile oysters. Taken together, the results show that competent larvae rapidly respond to the inducing effects of oysters via some immediate early genes, such as the transcription factor AP-1, which may further regulate downstream pathways such as the MAPK signaling pathway to cause subsequent changes, including a decrease in HSP90 and an increase in IAPs. These changes together may regulate the metamorphosis of R. venosa. This study provides further evidence that juvenile oysters are the metamorphosis cues of R. venosa, which may enhance our understanding of the metamorphosis mechanism in this marine invertebrate.
... The response to these environmental cues depends on the receptor components and intracellular signalling mediators. Following spinal cord injury, some guidance molecules, such as ephrin B3 (EPHB3), semaphorin 4D (SEMA4D) and netrin, associate with myelin while others, such as WNTs and class 3 semaphorins, are present in a reactive extracellular matrix that forms at or near the injury site [24][25][26][27][28] . Genetic evidence indicated that WNT signalling mediated by the repulsive receptor RYK inhibits CST collateral sprouting, cortical remapping and functional recovery 29 . ...
One hundred years ago, Ramón y Cajal, considered by many as the founder of modern neuroscience, stated that neurons of the adult central nervous system (CNS) are incapable of regenerating. Yet, recent years have seen a tremendous expansion of knowledge in the molecular control of axon regeneration after CNS injury. We now understand that regeneration in the adult CNS is limited by (1) a failure to form cellular or molecular substrates for axon attachment and elongation through the lesion site; (2) environmental factors, including inhibitors of axon growth associated with myelin and the extracellular matrix; (3) astrocyte responses, which can both limit and support axon growth; and (4) intraneuronal mechanisms controlling the establishment of an active cellular growth programme. We discuss these topics together with newly emerging hypotheses, including the surprising finding from transcriptomic analyses of the corticospinal system in mice that neurons revert to an embryonic state after spinal cord injury, which can be sustained to promote regeneration with neural stem cell transplantation. These gains in knowledge are steadily advancing efforts to develop effective treatment strategies for spinal cord injury in humans. The inability of the mammalian central nervous system to functionally regenerate after injury is largely attributable to the limited capacity of injured neurons to regrow axons. In the spinal cord, recent work on the mechanisms restricting axon regrowth suggests new therapeutic avenues to promote functional recovery after damage.
... Wnt ligands target particular receptors to execute their activities; for example, Wnt-4/Ryk ligand interaction is found to inhibit sensory neuron regeneration, whereas Wnt-LRP5/6 interaction is found to promote neurite outgrowth [164]. Consistent with this interaction, downregulation of Wnt-Ryk signalling is an attractive therapeutical approach to promote the plasticity of cortico-spinal motor neurons and robust axonal growth following injury [163,165,166]. ...
Axons in the peripheral nervous system have the ability to repair themselves after damage, whereas axons in the central nervous system are unable to do so. A common and important characteristic of damage to the spinal cord, brain, and peripheral nerves is the disruption of axonal regrowth. Interestingly, intrinsic growth factors play a significant role in the axonal regeneration of injured nerves. Various factors such as proteomic profile, microtubule stability, ribosomal location, and signalling pathways mark a line between the central and peripheral axons’ capacity for self-renewal. Unfortunately, glial scar development, myelin-associated inhibitor molecules, lack of neurotrophic factors, and inflammatory reactions are among the factors that restrict axonal regeneration. Molecular pathways such as cAMP, MAPK, JAK/STAT, ATF3/CREB, BMP/SMAD, AKT/mTORC1/p70S6K, PI3K/AKT, GSK-3β/CLASP, BDNF/Trk, Ras/ERK, integrin/FAK, RhoA/ROCK/LIMK, and POSTN/integrin are activated after nerve injury and are considered significant players in axonal regeneration. In addition to the aforementioned pathways, growth factors, microRNAs, and astrocytes are also commendable participants in regeneration. In this review, we discuss the detailed mechanism of each pathway along with key players that can be potentially valuable targets to help achieve quick axonal healing. We also identify the prospective targets that could help close knowledge gaps in the molecular pathways underlying regeneration and shed light on the creation of more powerful strategies to encourage axonal regeneration after nervous system injury.
... It should be noted that not all Wnt ligands are created equal, and some have been found to serve as repulsive growth cues and inhibitors of neurite outgrowth [151]. Generally, signaling through the canonical pathway is associated with augmented growth, while non-canonical activation of the Wnt/Ryk pathway provides an obstacle to regeneration [151][152][153][154][155]. Specifically, blockage of Wnt signaling by endogenous expression of Wnt inhibitory factor 1 (WIF1) or secreted frizzle related protein (SFRP) in preconditioned sensory neurons improved their regenerative capacity within the normally growth restricted central branch [151]. ...
... The non-canonical Wnt/β-catenin signaling cascade also has implications within the CNS. Wnt molecules are found to be upregulated following a dorsal hemisection of the spinal cord that restricts the recovery of descending corticospinal tract axons [153]. ...
Common mechanisms of peripheral axon regeneration are recruited following diverse forms of damage to peripheral nerve axons. Whether the injury is traumatic or disease related neuropathy, reconnection of axons to their targets is required to restore function. Supporting peripheral axon regrowth, while not yet available in clinics, might be accomplished from several directions focusing on one or more of the complex stages of regrowth. Direct axon support, with follow on participation of supporting Schwann cells is one approach, emphasized in this review. However alternative approaches might include direct support of Schwann cells that instruct axons to regrow, manipulation of the inflammatory milieu to prevent ongoing bystander axon damage, or use of inflammatory cytokines as growth factors. Axons may be supported by a growing list of growth factors, extending well beyond the classical neurotrophin family. The understanding of growth factor roles continues to expand but their impact experimentally and in humans has faced serious limitations. The downstream signaling pathways that impact neuron growth have been exploited less frequently in regeneration models and rarely in human work, despite their promise and potency. Here we review the major regenerative signaling cascades that are known to influence adult peripheral axon regeneration. Within these pathways there are major checkpoints or roadblocks that normally check unwanted growth, but are an impediment to robust growth after injury. Several molecular roadblocks, overlapping with tumour suppressor systems in oncology, operate at the level of the perikarya. They have impacts on overall neuron plasticity and growth. A second approach targets proteins that largely operate at growth cones. Addressing both sites might offer synergistic benefits to regrowing neurons. This review emphasizes intrinsic aspects of adult peripheral axon regeneration, emphasizing several molecular barriers to regrowth that have been studied in our laboratory.
... In this context, the exploration of those molecules that are implicated in neuroinflammation, particularly in the SCI-associated microglia/macrophage response modulation, might help us to shed light on the pathological mechanisms underlying the progress and outcome of this neuropathological condition. Interestingly, over the last years there is growing evidence showing that the Wnt family of proteins play a relevant role in different neuropathologies that are characterized by a dysregulated neuroinflammatory response [17][18][19][20], including SCI [21][22][23][24][25][26][27][28][29][30][31]. However, very few studies have evaluated the expression of the Wnt family of proteins in microglial cells. ...
... Further understanding of molecular signaling pathways that are involved in microglia/MDMs cellular responses to traumatic injuries may help us to pinpoint future therapeutic targets that promote the beneficial effects that are regulated by these cells while preventing the detrimental and neurotoxic ones. Interestingly, it is now increasingly evident that the Wnt family of proteins is involved in different neuropathologies that are characterized by a dysregulated neuroinflammatory response [17][18][19][20], including SCI [21][22][23][24][25][26][27][28][29][30][31]. As a consequence, we decided to assess whether the use of ex vivo and in vitro methods might help to foster the characterization of the gene expression alterations of Wntrelated molecules of microglia/MDMs cells in the traumatically-lesioned rat spinal cord. ...
It is well known that inflammation is crucial in the onset and progression of neurodegenerative diseases and traumatic central nervous system (CNS) injuries, and that microglia and monocyte-derived macrophages (MDMs) play a pivotal role in neuroinflammation. Therefore, the exploration of molecular signaling pathways that are involved in the microglia/macrophage response might help us to shed light on their eventual therapeutic modulation. Interestingly, there is growing evidence showing that the Wnt family of proteins is involved in different neuropathologies that are characterized by a dysregulated neuroinflammatory response, including spinal cord injury (SCI). Here, we aimed to validate a methodology with competence to assess the physiologically relevant Wnt expression patterns of active microglia and MDMs in a rat model of SCI. For that purpose, we have selected and adapted an in vitro system of primary microglia culture that were stimulated with a lesioned spinal cord extract (SCE), together with an ex vivo protocol of flow cytometry sorting of rat microglia/MDMs at different time-points after contusive SCI. Our study demonstrates that the expression profile of Wnt-related genes in microglia/MDM cells exhibit important differences between these particular scenarios which would be in line with previous studies where similar discrepancies have been described for other molecules. Moreover, our results provide for a first experimental report of the Wnt transcriptome in rat microglia and MDMs after SCI which, together with the research platform that was used in the study, and considering its limitations, we expect might contribute to foster the research on Wnt-driven immunomodulatory therapies.
