In utero topographic analysis of astrocytes and neuronal cells in the spinal cord of mutant mice with myelomeningocele

Abstract Background Spinal cord injury (SCI) is a severe traumatic disease which causes high disability and mortality rates. The molecular pathological features after spinal cord injury mainly involve the inflammatory response, microglial and neuronal apoptosis, abnormal proliferation of astrocytes, and the formation of glial scars. However, the microenvironmental changes after spinal cord injury are complex, and the interactions between glial cells and nerve cells remain unclear. Small extracellular vesicles (sEVs) may play a key role in cell communication by transporting RNA, proteins, and bioactive lipids between cells. Few studies have examined the intercellular communication of astrocytes through sEVs after SCI. The inflammatory signal released from astrocytes is known to initiate microglial activation, but its effects on neurons after SCI remain to be further clarified. Methods Electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blotting were applied to characterize sEVs. We examined microglial activation and neuronal apoptosis mediated by astrocyte activation in an experimental model of acute spinal cord injury and in cell culture in vitro. Results Our results indicated that astrocytes activated after spinal cord injury release CCL2, act on microglia and neuronal cells through the sEV pathway, and promote neuronal apoptosis and microglial activation after binding the CCR2. Subsequently, the activated microglia release IL-1β, which acts on neuronal cells, thereby further aggravating their apoptosis. Conclusion This study elucidates that astrocytes interact with microglia and neurons through the sEV pathway after SCI, enriching the mechanism of CCL2 in neuroinflammation and spinal neurodegeneration, and providing a new theoretical basis of CCL2 as a therapeutic target for SCI.

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Multiple developmental defects in Engrailed-1 mutant mice: an early mid-hindbrain deletion and patterning defects in forelimbs and sternum

Development ◽

10.1242/dev.120.7.2065 ◽

1994 ◽

Vol 120 (7) ◽

pp. 2065-2075 ◽

Cited By ~ 48

Author(s):

W. Wurst ◽

A.B. Auerbach ◽

A.L. Joyner

Keyword(s):

Spinal Cord ◽

Embryonic Development ◽

Normal Development ◽

Cranial Nerves ◽

Mutant Mice ◽

Mouse Development ◽

Developmental Defects ◽

Targeted Deletion ◽

The Third ◽

A Cell

During mouse development, the homeobox-containing gene En-1 is specifically expressed across the mid-hindbrain junction, the ventral ectoderm of the limb buds, and in regions of the hindbrain, spinal cord, somites and somite-derived tissues. To address the function of En-1 during embryogenesis, we have generated mice homozygous for a targeted deletion of the En-1 homeobox. En-1 mutant mice died shortly after birth and exhibited multiple developmental defects. In the brains of newborn mutants, most of the colliculi and cerebellum were missing and the third and fourth cranial nerves were absent. A deletion of midhindbrain tissue was observed as early as 9.5 days of embryonic development and the phenotype resembles that previously reported for Wnt-1 mutant mice. In addition, patterning of the forelimb paws and sternum was disrupted, and the 13th ribs were truncated. The results of these studies suggest a cell autonomous role for En-1 in generation and/or survival of mid-hindbrain precursor cells and also a non-cell autonomous role in signalling normal development of the limbs and possibly sternum.

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Exosome derived from mesenchymal stem cells enhance autophagy and inhibit iNOS/TXNIP/NLRP3 inflammasome axis in injured spinal cord

10.21203/rs.3.rs-16805/v1 ◽

2020 ◽

Author(s):

Bin Lv ◽

Lei Wang ◽

Anquan Huang ◽

Tianming Zou ◽

Jishan Yuan

Keyword(s):

Nitric Oxide ◽

Spinal Cord ◽

Nlrp3 Inflammasome ◽

Neuroprotective Effect ◽

Neuronal Cells ◽

Tissue Loss ◽

Protective Effects ◽

Sprague Dawley ◽

Cord Injury

Abstract Background: Neuroinflammation, autophagy, NLRP3 inflammasome, and microglia polarizationhave been implicated in spinal cord injury (SCI).Moreover, exosomes, a classic nanovesicles secreted by MSCs, may have a neuroprotective effect on transformation of microglia from the M1 state to the M2 phenotype. However, the effect of MSCs derived exosomes on neuroinflammation is still unclear. Here, we investigated the mechanisms of MSCs derived exosomes mediated NLRP3 inflammasome signaling cascades and its protective effects in SCI. Methods:The SCI model was performed by weight-drop impact in adult male Sprague-Dawley (SD) rats. Control andexosome rats were randomly subjecttoexosomeadminister (20 mg/kg) or placebo via intraperitoneal route 1 h after SCI.Autophagy inhibitor(3-MA) was administered intraperitoneally 20 min before experiment.Neurological function was measured by Basso-Beattie-Bresnahan (BBB) scoring and an open-field test.Neuronal death was measured by HE stainingandNisslstaining.Inducible nitric oxide synthase (iNOS) levels were determined using fluorescent probes. The autophagy and TXNIP and its downstream signaling pathways-mediated polarization of macrophages/microglia was assessed by immunohistochemistry. Results:Exosome significantly downregulated intracellular iNOS and inhibited TXNIP, pyrin domain-containing 3 (NLRP3) inflammasome pathway activation by activating autophagy. Additionally, Exosomepromoted expression of autophagy markers, such as LC3A/B and beclin1,and abrogated the expression of p62. Autophagy inhibitor, 3-MA, blockage of autophagy flux abolished the inhibition of apoptosis and iNOS/TXNIP/NLRP3 inflammasome axisafterSCI. Here, we demonstrated that exosomeadministration in spinal cord markedly reduced tissue loss, attenuate pathological morphology of the injuredregion, and promoted tissue recovery. Moreover. our resultshowed that exosome administration alleviated neuronal cells apoptosis, and inhibited nitric oxide release in microglia.The activation of inflammatoryresponse in neuronal cells facilitates interactions of iNOS‐NLRP3 andTXNIP‐NLRP3and inhibited NLRP3 inflammasome where neuronal cells apoptosis was induced.Further, we found that exosome could suppress macrophages/microglia polarized to M1 phenotype in vivo and in vitro.Taken together, exosome administration exerts protective effects in neuronal cells through inhibiting iNOS production, and exosome administration could inhibit iNOS/TXNIP/NLRP3 inflammasome axis via enhancing autophagy and both in vitro and in vivo. Conclusions:These resultsreveal that exosometreatment alleviatedneuroinflammation and mitigates neuronal apoptosis via autophagy-mediate inhibition of the iNOS/TXNIP/NLRP3 inflammasome axis. Our findings suggest that exosome may be a novel therapeutic target for treating SCI.

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