MiR-21a-5p Promotes Inflammation following Traumatic Spinal Cord Injury through Upregulation of Neurotoxic Reactive Astrocyte (A1) Polarization by Inhibiting the CNTF/STAT3/Nkrf Pathway.
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ABSTRACT:
SUBMITTER: Zhang Y
PROVIDER: S-EPMC8326122 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Astrocyte-derived ciliary neurotrophic factor (CNTF) promotes adult subventricular zone (SVZ) neurogenesis. We found that focal adhesion kinase (FAK) and JNK, but not ERK or P38, repress CNTF in vitro. Here, we defined the FAK-JNK pathway and its regulation of CNTF in mice, and the related leukemia inhibitory factor (LIF) and interleukin-6 (IL-6), which promote stem cell renewal at the expense of neurogenesis. Intrastriatal injection of FAK inhibitor, FAK14, in adult male C57BL/6 mice reduced pJNK and increased CNTF expression in the SVZ-containing periventricular region. Injection of a JNK inhibitor increased CNTF without affecting LIF and IL-6, and increased SVZ proliferation and neuroblast formation. The JNK inhibitor had no effect in CNTF-/- mice, suggesting that JNK inhibits SVZ neurogenesis by repressing CNTF. Inducible deletion of FAK in astrocytes increased SVZ CNTF and neurogenesis, but not LIF and IL-6. Intrastriatal injection of inhibitors suggested that P38 reduces LIF and IL-6 expression, whereas ERK induces CNTF and LIF. Intrastriatal FAK inhibition increased LIF, possibly through ERK, and IL-6 through another pathway that does not involve P38. Systemic injection of FAK14 also inhibited JNK while increasing CNTF, but did not affect P38 and ERK activation, or LIF and IL-6 expression. Importantly, systemic FAK14 increased SVZ neurogenesis in wild-type C57BL/6 and CNTF+/+ mice, but not in CNTF-/- littermates, indicating that it acts by upregulating CNTF. These data show a surprising differential regulation of related cytokines and identify the FAK-JNK-CNTF pathway as a specific target in astrocytes to promote neurogenesis and possibly neuroprotection in neurological disorders.
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Project description:Purpose:Ciliary neurotrophic factor (CNTF) is a well-characterized neurotrophic factor currently in clinical trials for the treatment of macular telangiectasia type II. Our previous work showed that CNTF-induced STAT3 signaling is a potent inhibitor of pathologic preretinal neovascular tuft formation in the mouse model of oxygen-induced retinopathy. In this study, we investigated the effect of CNTF on outer retinal and choroidal angiogenesis and the mechanisms that underpin the observed decrease in outer retinal neovascularization following CNTF treatment. Methods:In the Vldlr-/- and laser-CNV mouse models, mice received a one-time injection (on postnatal day [P] 12 in the Vldlr-/- model and 1 day after laser in the Choroidal Neovascularization (CNV) model) of recombinant CNTF or CxCl10, and the extent of neovascular lesions was assessed 6 days posttreatment. STAT3 downstream targets affected by CNTF treatment were identified using quantitative PCR analysis. A proteome array was used to compare media conditioned by CNTF-treated and control-treated primary Müller cells to screen for CNTF-induced changes in secreted angiogenic factors. Results:Intravitreal treatment with recombinant CNTF led to significant reduction in neovascularization in the Vldlr-/- and laser-CNV mouse models. Treatment effect in the Vldlr-/- was long-lasting but time sensitive, requiring intravitreal treatment before P19. Mechanistic workup in vitro as well as in vivo confirmed significant activation of the STAT3-signaling pathway in Müller cells in response to CNTF treatment and upregulation of CxCl10. Intravitreal injections of recombinant CxCl10 significantly reduced outer retinal neovascularization in vivo in both the Vldlr-/- and laser-CNV mouse models. Conclusions:CNTF treatment indirectly affects outer retinal and choroidal neovascularization by inducing CxCl10 secretion from retinal Müller cells.
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Project description:Chronic consumption of β-sitosterol-β-D-glucoside (BSSG), a neurotoxin contained in cycad seeds, leads to Parkinson's disease in humans and rodents. Here, we explored whether a single intranigral administration of BSSG triggers neuroinflammation and neurotoxic A1 reactive astrocytes besides dopaminergic neurodegeneration. We injected 6 μg BSSG/1 μL DMSO or vehicle into the left substantia nigra and immunostained with antibodies against tyrosine hydroxylase (TH) together with markers of microglia (OX42), astrocytes (GFAP, S100β, C3), and leukocytes (CD45). We also measured nitric oxide (NO), lipid peroxidation (LPX), and proinflammatory cytokines (TNF-α, IL-1β, IL-6). The Evans blue assay was used to explore the blood-brain barrier (BBB) permeability. We found that BSSG activates NO production on days 15 and 30 and LPX on day 120. Throughout the study, high levels of TNF-α were present in BSSG-treated animals, whereas IL-1β was induced until day 60 and IL-6 until day 30. Immunoreactivity of activated microglia (899.0 ± 80.20%) and reactive astrocytes (651.50 ± 11.28%) progressively increased until day 30 and then decreased to remain 251.2 ± 48.8% (microglia) and 91.02 ± 39.8 (astrocytes) higher over controls on day 120. C3(+) cells were also GFAP and S100β immunoreactive, showing they were neurotoxic A1 reactive astrocytes. BBB remained permeable until day 15 when immune cell infiltration was maximum. TH immunoreactivity progressively declined, reaching 83.6 ± 1.8% reduction on day 120. Our data show that BSSG acute administration causes chronic neuroinflammation mediated by activated microglia, neurotoxic A1 reactive astrocytes, and infiltrated immune cells. The severe neuroinflammation might trigger Parkinson's disease in BSSG intoxication.
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Project description:Accumulating evidence indicates that cryptochrome circadian regulatory (CRY) proteins have emerged as crucial regulators of osteogenic differentiation. However, the associated mechanisms are quite elusive. In this study, we show that knockdown of CRY2 downregulated the expression of runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), osteocalcin (OCN), and osteopontin (OPN) to facilitate osteoblast differentiation. Further study identified that CRY2 was directly targeted by microRNA (miR)-7-5p, which was highly induced during osteoblast differentiation. The expression of Runx2, ALP, collagen type I alpha 1 (Col1a1), and OCN was upregulated by overexpression of miR-7-5p and induction of osteoblast differentiation. Moreover, signal transducer and activator of transcription 3 (STAT3) transcriptionally activated miR-7-5p to significantly enhance the expression of above osteogenic marker genes and mineral formation. However, overexpression of CRY2 abolished the osteogenic differentiation induced by miR-7-5p overexpression. Silencing of CRY2 unraveled the binding of CRY2 with the circadian locomotor output cycles kaput (CLOCK)/brain and muscle ARNT-like 1 (BMAL1) complex to release CLOCK/BMAL1, which facilitated the binding of CLOCK/BMAL1 to the promoter region of the P300 E-box to stimulate the transcription of P300. P300 subsequently promoted the acetylation of histone 3 and the formation of a transcriptional complex with Runx2 to enhance osteogenesis. Taken together, our study revealed that CRY2 is repressed by STAT3/miR-7-5p to promote osteogenic differentiation through CLOCK/BMAL1/P300 signaling. The involved molecules may be potentially targeted for treatment of osteoporosis.
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