Monitoring Astrocyte Reactivity and Proliferation &lt;em&gt;in Vitro&lt;/em&gt; Under Ischemic-Like Conditions

Ferrer-Acosta, Yancy; Gonzalez-Vega, Maxine N.; Rivera-Aponte, David E.; Martinez-Jimenez, Solianne M.; Martins, Ana

doi:10.3791/55108-v

Cited by 3 publications

(6 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…As Ki-67 is the most cited proliferation marker that can be determined by immunofluorescence staining with highest levels during G2 phase and mitosis ( 106 , 107 ), the fraction of Ki-67 positive cells after exposure of astrocytes to X-rays was quantified and found to be largely unaffected by X-irradiation at around 10 %. A similar low percentage of proliferating cells of 10–25 % was also observed in primary rat astrocytes ( 66 ) and in adult mouse astrocytes ( 108 ). As the proliferation rate decreased in higher passages ( 108 ), only passage 1 astrocytes were used in this work.…”

Section: Discussionsupporting

confidence: 71%

“…Astrocyte proliferation is in general a measure of their reactivity ( 64 – 66 ). Furthermore, cell proliferation can be influenced by NF-κB pathway activation ( 67 – 69 ).…”

Section: Resultsmentioning

confidence: 99%

“…The radiation response of astrocytes is an important piece of the puzzle in understanding the effects of different ionizing radiation qualities on the brain as they represent the predominant cell type in the mammalian brain ( 22 ) and are crucial to normal brain function ( 83 ). Although the isolation of primary astrocytes is laborious and the results are sometimes variable, primary astrocytes from rodent embryos are recognized as a potent tool to study astrocytes, biology and mechanistic features ( 66 , 84 ). As alternative cell models such as immortalized astrocytes and C6 glioma cells showed major differences in morphology, protein expression and functionality in comparison to primary astrocytes ( 84 ), we chose primary murine astrocytes for our studies.…”

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Unraveling astrocyte behavior in the space brain: Radiation response of primary astrocytes

Roggan

Kronenberg

Wollert

et al. 2023

Front. Public Health

View full text Add to dashboard Cite

IntroductionExposure to space conditions during crewed long-term exploration missions can cause several health risks for astronauts. Space radiation, isolation and microgravity are major limiting factors. The role of astrocytes in cognitive disturbances by space radiation is unknown. Astrocytes' response toward low linear energy transfer (LET) X-rays and high-LET carbon (12C) and iron (56Fe) ions was compared to reveal possible effects of space-relevant high-LET radiation. Since astronauts are exposed to ionizing radiation and microgravity during space missions, the effect of simulated microgravity on DNA damage induction and repair was investigated.MethodsPrimary murine cortical astrocytes were irradiated with different doses of X-rays, 12C and 56Fe ions at the heavy ion accelerator GSI. DNA damage and repair (γH2AX, 53BP1), cell proliferation (Ki-67), astrocytes' reactivity (GFAP) and NF-κB pathway activation (p65) were analyzed by immunofluorescence microscopy. Cell cycle progression was investigated by flow cytometry of DNA content. Gene expression changes after exposure to X- rays were investigated by mRNA-sequencing. RT-qPCR for several genes of interest was performed with RNA from X-rays- and heavy-ion-irradiated astrocytes: Cdkn1a, Cdkn2a, Gfap, Tnf, Il1β, Il6, and Tgfβ1. Levels of the pro inflammatory cytokine IL-6 were determined using ELISA. DNA damage response was investigated after exposure to X-rays followed by incubation on a 2D clinostat to simulate the conditions of microgravity.ResultsAstrocytes showed distinct responses toward the three different radiation qualities. Induction of radiation-induced DNA double strand breaks (DSBs) and the respective repair was dose-, LET- and time-dependent. Simulated microgravity had no significant influence on DNA DSB repair. Proliferation and cell cycle progression was not affected by radiation qualities examined in this study. Astrocytes expressed IL-6 and GFAP with constitutive NF-κB activity independent of radiation exposure. mRNA sequencing of X-irradiated astrocytes revealed downregulation of 66 genes involved in DNA damage response and repair, mitosis, proliferation and cell cycle regulation.DiscussionIn conclusion, primary murine astrocytes are DNA repair proficient irrespective of radiation quality. Only minor gene expression changes were observed after X-ray exposure and reactivity was not induced. Co-culture of astrocytes with microglial cells, brain organoids or organotypic brain slice culture experiments might reveal whether astrocytes show a more pronounced radiation response in more complex network architectures in the presence of other neuronal cell types.

show abstract

Section: Discussionsupporting

confidence: 71%

“…Astrocyte proliferation is in general a measure of their reactivity ( 64 – 66 ). Furthermore, cell proliferation can be influenced by NF-κB pathway activation ( 67 – 69 ).…”

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Unraveling astrocyte behavior in the space brain: Radiation response of primary astrocytes

Roggan

Kronenberg

Wollert

et al. 2023

Front. Public Health

View full text Add to dashboard Cite

show abstract

Section: Methodsmentioning

confidence: 99%

“…Stretch-induced cell injury is an in vitro injury model, which is used to simulate cell injury during TBI (21). It has become the most recognized cell damage mode by researchers, and its reliability has been verified in our previous study and also by other groups (19,21,22). The PC12 cell and astrocytes stretch injury (SI) model was applied to reproduce the in vitro injury process of nerve cells after TBI.…”

Section: Methodsmentioning

confidence: 99%

Polydatin ameliorates TBI induced secondary brain injury by inhibiting NLRP3-induced neuroinflammation associated with SOD2 acetylation

Zhao

et al. 2022

Shock

View full text Add to dashboard Cite

Traumatic brain injury (TBI) is a kind of disease with high morbidity, mortality, and disability, and its pathogenesis is still unclear. Research shows that nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) activation in neurons and astrocytes is involved in neuroinflammatory cascades after TBI. What is more, polydatin (PD) has been shown to have a protective effect on TBI-induced neuroinflammation, but the mechanisms remain unclear. Here, we speculated that PD could alleviate TBI-induced neuroinflammatory damage through the superoxide dismutase (SOD2)-NLRP3 signal pathway, and SOD2 might regulate NLRP3 inflammasome activation. The model of lateral fluid percussion for in vivo and cell stretching injury for in vitro were established to mimic TBI. NLRP3 chemical inhibitor MCC950, SOD2 inhibitor 2-methoxyestradiol, and PD were administered immediately after TBI. As a result, the expression of SOD2 acetylation (SOD2 Ac-K122), NLRP3, and cleaved caspase-1 were increased after TBI both in vivo and in vitro, and using SOD2 inhibitor 2-methoxyestradiol significantly promoted SOD2 Ac-K122, NLRP3, and cleaved caspase-1 expression, as well as exacerbated mitochondrial ROS (mtROS) accumulation and mitochondrial membrane potential (MMP) collapse in PC12 cells. However, using NLRP3 inhibitor MCC950 significantly inhibited cleaved caspase-1 activation after TBI both in vivo and in vitro; meanwhile, MCC950 inhibited mtROS accumulation and MMP collapse after TBI. More importantly, PD could inhibit the level of SOD2 Ac-K122, NLRP3, and cleaved caspase-1 and promote the expression of SOD2 after TBI both in vivo and in vitro. Polydatin also inhibited mtROS accumulation and MMP collapse after stretching injury. These results indicated that PD inhibited SOD2 acetylation to alleviate NLRP3 inflammasome activation, thus acting a protective role against TBI neuroinflammation.

show abstract

Vascular endothelial growth factor promotes transdifferentiation of astrocytes into neurons via activation of the MAPK/Erk‐Pax6 signal pathway

Lei

Chen

et al. 2023

Glia

View full text Add to dashboard Cite

Reactive astrocytes can be transformed into new neurons. Vascular endothelial growth factor (VEGF) promotes the transformation of reactive astrocytes into neurons in ischemic brain. Therefore, in this study, the molecular mechanism of VEGF's effect on ischemia/hypoxia-induced astrocyte to neuron transformation was investigated in the models of rat middle cerebral artery occlusion (MCAO) and in astrocyte culture with oxygen and glucose deprivation (OGD). We found that VEGF enhanced ischemia-induced Pax6, a neurogenic fate determinant, expression and Erk phosphorylation in reactive astrocytes and reduced infarct volume of rat brain at 3 days after MCAO, which effects could be blocked by administration of U0126, a MAPK/Erk inhibitor. In cultured astrocytes, VEGF also enhanced OGD-induced Erk phosphorylation and Pax6 expression, which was blocked by U0126, but not wortmannin, a PI3K/Akt inhibitor, or SB203580, a MAPK/p38 inhibitor, suggesting VEGF enhanced Pax6 expression via activation of MAPK/Erk pathway. OGD induced the increase of miR365 and VEGF inhibited the increase of OGD-induced miR365 expression. However, miR365 agonists blocked VEGF-enhanced Pax6 expression in hypoxic astrocytes, but did not block VEGF-enhanced Erk phosphorylation. We further found that VEGF promoted OGD-induced astrocyte-converted to neuron. Interestingly, both U0126 and Pax6 RNAi significantly reduced enhancement of VEGF on astrocytes-toneurons transformation, as indicated Dcx and MAP2 immunopositive signals in reactive astrocytes. Moreover, those transformed neurons become mature and functional. We concluded that VEGF enhanced astrocytic neurogenesis via the MAPK/Erk-miR-365-Pax6 signal axis. The results also indicated that astrocytes play important roles in the reconstruction of neurovascular units in brain after stroke.

show abstract

Monitoring Astrocyte Reactivity and Proliferation <em>in Vitro</em> Under Ischemic-Like Conditions

Cited by 3 publications

References 0 publications

Unraveling astrocyte behavior in the space brain: Radiation response of primary astrocytes

Unraveling astrocyte behavior in the space brain: Radiation response of primary astrocytes

Polydatin ameliorates TBI induced secondary brain injury by inhibiting NLRP3-induced neuroinflammation associated with SOD2 acetylation

Vascular endothelial growth factor promotes transdifferentiation of astrocytes into neurons via activation of the MAPK/Erk‐Pax6 signal pathway

Contact Info

Product

Resources

About