Tissue resident cell activation leads to Increase, Pro-Inflammatory Mediators

Modulating factor  

Details  

Effects on the KER  

References  

Drug 

Flavonoids 

Flavonoids can inhibit NF-κB, preventing transcription of pro-inflammatory mediators in active glial cells. 

Wang et al., 2020 

Drug 

Tamoxifen (estrogen receptor 

modulator commonly used in breast cancer treatment) 

Treatment with Tamoxifen decreased the radiation-induced activation of glial cells. It also consistently decreased the amount of TNF-α and IL-1β and blood-brain barrier permeability after irradiation at various doses. 

Liu et al., 2010 

Drug 

RA (modulates inflammatory effects in different cell types) 

RA treatment completely inhibited the increase in pro-inflammatory mediators after LPS-induced glial activation. 

van Neerven et al., 2010 

Drug 

SP (JNK, c-jun N-terminal kinase, inhibitor) 

AP-1 DNA binding (glial activation) was reduced by SP treatment after irradiation. TNF-α, Cox-2 and IL-1β were reduced by SP treatment after irradiation or viral infection. 

Ramanan et al., 2008  

Drug 

Q (NF-κB inhibitor) 

NF-κB DNA binding (glial activation) was reduced by Q treatment after irradiation. IL-1β was also reduced by Q treatment after irradiation. 

Ramanan et al., 2008 

Drug 

NS-398 (Cox-2 inhibitor) 

Treatment with NS-398 reduced TNF-α, IL-1β, IL-6, ICAM-1 and MCP-1 expression after irradiation. 

Kyrkanides et al., 2002 

Age 

Increased age 

Aging tissue becomes more sensitive to immune signals and increases inflammation. In the aging brain, microglia will produce more pro-inflammatory mediators. 

Patterson, 2015 

Drug 

NAC 

NAC treatment inhibited pro-inflammatory mediator production in macrophages. 

Komatsu et al., 2017 

Reference 

Experiment Description 

Result 

Liu et al., 2010  

In vivo. BV2 murine microglia were irradiated with 0, 2, 4, 6, 8 or 10 Gy X-rays to measure microglial activation, and OX-42 and GFAP staining was performed on 15 Gy irradiated rat brains to measure microglial and astrocyte activation, respectively.  

In vitro. BV-2 murine microglia were irradiated with 0, 2, 4, 6, 8 or 10 Gy X-rays to determine cytokines IL-1β and TNF-α production. Glial activation was identified by light microscopy and immunohistochemistry. ELISA was used to assess cytokine levels of IL-1β and TNF-α. 

Irradiation of in vitro microglia cultures caused a dose-dependent increase in microglial activation from 0 to 10 Gy. At 15 Gy, in vivo astrocyte activation increased 5-fold, while microglial activation increased 3-fold. Levels of IL-1β and TNF-α production were also dose-dependently increased following 0 to 10 Gy irradiated microglia, resulting in an 8.6-fold increase for IL-1β and a 6.8-fold increase for TNF-α after 10 Gy. 

Welser-Alves & Milner, 2013  

In vitro. Cultures of microglia and astrocytes from postnatal mouse central nervous system were stimulated with 1 µg/mL LPS. ELISA and immunocytochemistry were used to measure glial cytokine (TNF-α) production with Mac-1 as a microglial marker. 

Microglia activation with 1 µg/mL LPS led to increased TNF-α production from 40.6 pg/ml to 1875.0 pg/ml, and TNF-α showed co-localization with Mac-1 positive microglia. TNF-α was not present in astrocytes. 

Lee et al., 2010 

In vivo. Rats received whole-brain gamma ray irradiation at 10 Gy. Levels of AP-1 and NF-κB (microglial activation) as well as pro-inflammatory mediators TNF-α, IL-1β, IL-6, and MCP-1 were determined in the hippocampus and cortex. AP-1 and NF-κB DNA binding was determined through electrophoretic mobility shift assay (EMSA), and pro-inflammatory mediator levels were determined using enzyme-linked immunosorbent assay (ELISA). 

After 10 Gy, DNA binding of NF-κB and AP-1 increased a maximum of 3.6-fold and 2.8-fold, respectively. 

Hippocampus: 

After 10 Gy at maximum, TNF-α was increased 23-fold, IL-1β increased 10-fold, IL-6 did not significantly change, and MCP-1 increased 1.6-fold. 

Cortex: 

After 10 Gy at maximum, TNF-α increased 30-fold, IL-1β increased 7-fold. IL-6 did not significantly change, and MCP-1 increased 2.2-fold. 

Chen et al., 2016 

In vitro. Human CHME5 microglia were irradiated with various doses of 137Cs gamma radiation delivered acutely over 1-3 minutes. Microglial activation markers CR3/43 and Glut-5 were determined by Western blot, morphology of microglia was determined through fluorescence microscopy, and expression of cytokines IL-1α and TNF-α were determined through RT-PCR. 

After 8 Gy, microglia showed a characteristic activated morphology, but not after 0.5 Gy. CR3/43 and Glut-5 were both expressed after 8 Gy, but not 0.5 Gy. mRNA levels measured at 8 Gy were found to increase a maximum of 7.8-fold for IL-1α and 5.8-fold for TNF-α. 

Dong et al., 2015 

In vivo and in vitro. BV2 mouse microglial cells and C57BL/6J mice brains were irradiated with various doses of X-rays. Iba1 staining was performed to determine cell morphology, while anti-F4-80 antibodies were used to determine microglial activation. TNF-α and IL-1β levels were determined through RT-PCR, ELISA (in vitro), and Western blot (in vivo). 

In vitro: 

At 16 Gy, microglia adopted a characteristic activated morphology while F4-80 was greatly upregulated. Also at 16 Gy, IL-1β expression increased a maximum of 23-fold, TNF-α expression increased a maximum of 13-fold, IL-1β increased from 0 to a maximum of 530 pg/mL, and TNF-α increased from almost 0 to a maximum of 115 pg/mL. 

In vivo: 

The number of F4-80 positive cells/mm2 increased from 9 to a maximum of 40 after 10 Gy. Also at 10 Gy, IL-1β expression increased a maximum of 7-fold, TNF-α expression increased a maximum of 5-fold, IL-1β levels increased a maximum of 10-fold, and TNF-α levels increased a maximum of 5-fold compared to controls. 

Komatsu et al., 2017 

In vitro. Murine macrophage RAW264 cell line activation was induced by 1 µg/mL LPS treatment. Activation was determined through NF-κB nuclear translocation measured by western blot. Pro-inflammatory mediators TNF-α and IL-6 were measured by ELISA. 

After LPS treatment, cytosolic NF-κB decreased 0.64-fold, nuclear NF-κB increased 7.2-fold and IκB (NF-κB inhibitor) decreased 0.21-fold. Both TNF-α and IL-6 were increased from around 0 ng/mL to about 70 and 30 ng/mL, respectively. 

Lodermann et al., 2012 

In vitro. Human monocytic leukaemia cell lines were irradiated with X-rays at 0.1, 0.3, 0.5, 0.7 and 1 Gy. Activation was determined through NF-κB nuclear translocation by western blot. IL-1β was measured by ELISA. 

NF-κB and IL-1β both showed linear decreases from 0.1 to 0.7 Gy, resulting in maximum decreases of 0.6- to 0.7-fold. Although, the decrease for NF-κB was nonsignificant. No changes were observed at 1 Gy. 

Scharpfenecker et al., 2012 

In vitro. Mice kidneys were irradiated with 16 Gy of X-rays. Immunofluorescence stainings were performed for IL-6, IL-1β and macrophage marker F4-80. 

F4-80 positive area increased by 6.7 and 3.8-fold in Eng+/+ and Eng+/- irradiated mice respectively. Macrophages in irradiated mouse kidney led to IL-6 and IL-1β production.  

Reference 

Experiment Description 

Result 

Parihar et al., 2018 

In vivo. C57BL/6 J mice were irradiated with 4He particles 1 year after irradiation. The level of CCL-3 was determined in the brain, and microglial activation was determined by immunohistochemistry in the perirhinal cortex. 

At maximum, there was a 3.8-fold increase in activated microglia, CCL-3 increased 2.2-fold. 

Liu et al., 2010  

In vitro. BV2 murine microglia were irradiated with X-rays to measure microglial activation after 24h, and OX-42 and GFAP staining was performed on irradiated rat brains to measure microglial and astrocyte activation, respectively, after 3 days. BV-2 murine microglia were irradiated with X-rays to determine cytokines IL-1β and TNF-α production after 24h. Glial activation was identified by light microscopy and immunohistochemistry. ELISA was used to assess cytokine levels of IL-1β and TNF-α. 

10 Gy irradiation of  microglia cultures caused an increase in microglial activation after 24h. At 15 Gy after 3 days, astrocyte activation increased 5-fold, while microglial activation increased 3-fold. Levels of IL-1β and TNF-α were also increased in 10 Gy irradiated microglia, resulting in an 8.6-fold increase for IL-1β and a 6.8-fold increase for TNF-α after 24h. 

Lee et al., 2010 

In vivo. Rats received whole-brain gamma ray irradiation at 10 Gy. Levels of AP-1 and NF-κB (microglial activation) as well as pro-inflammatory mediators TNF-α, IL-1β, IL-6, and MCP-1 were determined 4, 8, and 24h after irradiation in the hippocampus and cortex. AP-1 and NF-κB DNA binding was determined through electrophoretic mobility shift assay (EMSA), and pro-inflammatory mediator levels were determined using enzyme-linked immunosorbent assay (ELISA). 

DNA binding of NF-κB and AP-1 increased a maximum of 3.6-fold and 2.8-fold, respectively, after 8h. Binding activity returned to control levels after 24h. 

Hippocampus: 

At control, TNF-α was 3.6 pg/mg protein, which increased 23-fold after 4 hours, 8.3-fold after 8h, and 3.6-fold after 24h. IL-1β increased linearly from 4 to 24h, reaching a 10-fold maximum increase. IL-6 did not significantly change other than a non-significant decrease over 24h, even though an increase in IL-6 mRNA was observed at 4h with RT-qPCR. MCP-1 showed a maximum increase of 1.6-fold after 8h. 

Cortex: 

At control, TNF-α was 4.4 pg/mg protein, which increased 30-fold after 4 hours, 13-fold after 8h, and 4.1-fold after 24h. IL-1β showed a maximum increase of 7-fold after 4h. IL-6 did not significantly change other than a non-significant decrease over 24h, even though an increase in IL-6 mRNA was observed at 4h with RT-qPCR. MCP-1 showed a maximum increase of 2.2-fold after 8h. 

Chen et al., 2016 

In vitro. Human CHME5 microglia were irradiated with 8 Gy gamma radiation (137Cs source) delivered acutely over 1-3 minutes. Microglial activation markers CR3/43 and Glut-5 were determined by Western blot, morphology of microglia was determined through fluorescence microscopy, and expression of cytokines IL-1α and TNF-α were determined through RT-PCR. 

Beginning after 7 days, microglia showed a characteristic activated morphology. CR3/43 was expressed after 2 weeks, while Glut-5 was expressed after 1 week, 10 days, and 2 weeks. After 7 days, mRNA levels were found to increase a maximum of 7.8-fold for IL-1α and 5.8-fold for TNF-α. mRNA levels dropped slightly but were still above controls after 2 weeks. 

Zhou et al., 2017 

In vivo. Juvenile rats were irradiated with 4MV nominal photon energy and a single dose of 6 Gy (2.3 Gy/min). At 6 or 24h, the molecular and cellular changes in the EGL of the cerebellum was studied. Immunohistochemistry staining was used to measure Iba1 (microglia marker) with morphometry analysis performed on microglia. Luminex assay measured cytokines, chemokines and growth factors for the inflammatory response. 

Microglia density increased by 2.3-fold after 6 h and 6.77-fold 24 h post-irradiation. Most of the iba1-positive cells had a bushy or amoeboid morphology, signifying an activated state. 

At 6 h of irradiation, IL-1α and CCL-2 increased by 2-2.8-fold. IL-1β decreased at 6 h then slightly increased at 24 h post-irradiation. IL-6, IL-18, GRO/KC, VEGF, and GM-CSF all increased significantly at 24 h compared to the control group. 

Dong et al., 2015 

In vivo and in vitro. BV2 mouse microglial cells and C57BL/6J mice brains were irradiated with X-rays. Iba1 staining was performed to determine cell morphology, while anti-F4-80 antibodies were used to determine microglial activation. TNF-α and IL-1β levels were determined through RT-PCR, ELISA (in vitro), and Western blot (in vivo) from 3h to 6 weeks after irradiation. 

In vivo, the number of F4-80 positive cells/mm2 increased from 9 to a maximum of 40 after 24h, decreased over 2 weeks, and returned to control levels at 4 and 6 weeks. IL-1β expression increased to a maximum after 72h and was significantly increased from 6h to 6 weeks. TNF-α expression was a maximum after 3 and 6h, was at controls from 24h to 1 week, and was increased again from 2 to 6 weeks. IL-1β levels were high after 3h, lower at 6h, increased to a maximum at 2 weeks, then decreased at 4 and 6 weeks. TNF-α levels increased to a maximum after 72h, then decreased until 4 weeks, where they increased again after 6 weeks. 

In vitro. IL-1β levels reached a maximum at 3h, but then decreased at 6h, before rising again at 12h. TNF-α levels remained elevated up to 24h, although its peak was at 6h. 

Reference 

Experimental Description 

Result 

Parihar et al., 2018 

In vivo. C57BL/6 J mice were irradiated with 4He particles at either 5 or 30 cGy (5 cGy/min). The level of CCL-3 was determined in the brain, and microglial activation was determined by ED-1 immunohistochemistry in the perirhinal cortex. 

Microglial activation increased 3.5-fold after 5 cGy and 3.8-fold after 30 cGy. CCL-3 increased 1.4-fold (non-significant, ns) after 5 cGy and 2.2-fold after 30 cGy. 

Zhou et al., 2017 

In vivo. Juvenile rats were irradiated with 4 MV nominal photon energy and a single dose of 6 Gy (2.3 Gy/min). At 6 or 24h, the molecular and cellular changes in the EGL of the cerebellum was studied. Immunohistochemistry staining was used to measure Iba1 (microglia marker) with morphometry analysis performed on microglia. Luminex assay measured cytokines, chemokines and growth factors for the inflammatory response. 

Microglia density increased by 2.3-fold after 6 h.and 6.77-fold 24 h post-irradiation. Most of the iba1-positive cells had a bushy or amoeboid morphology, signifying an activated state. 

At 6 h of irradiation, IL-1α and CCL-2 increased by 2-2.8-fold. IL-1β decreased at 6 h then slightly increased at 24 h post-irradiation. IL-6, IL-18, GRO/KC, VEGF, and GM-CSF all increased significantly at 24 h compared to the control group.