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A newly-synthesized compound CP-07 alleviates microglia-mediated neuroinflammation and ischemic brain injury via
Mengdi Guo1,2,3,4, Qian Cao5, Shengnan Xia1,2,3,4
1Department of Neurology, Nanjing Drum Tower Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing 210008, Jiangsu Province, China.
Background And Objectives:
Overactivated glial cells, especially microglia, are core components in the progression of pathologic neuroinflammation, and the application of anti-inflammatory reagents has been regarded as a potential therapy in the management of infarction/reperfusion (I/R) brain injury. This research aims to clarify the anti-inflammatory efect of a novel lipophilic compound N-(2-[4-tert-butylphenyl]-2-[pyrrolidine-1-yl]ethyl)-7-methyl-4-oxo-4H-chromene-2-carboxamide (named CP-07 in this study) in LPS-stimulated BV2 cell line and primary mouse microglia, and its therapeutic effect on I/R brain injury.
Method:
Cell Counting Kit-8 assay was used to determine the maximal nontoxic dose of CP-07. The mRNA levels of representative proinflammatory cytokines were determined by quantitative real-time polymerase chain reaction both in vitro and in vivo. TTC staining was performed to calculate infarct volumes while behavioral tests were used to assess the neurological deficits at 24 h after middle cerebral artery occlusion (MCAO). Flow cytometry analysis and immunofluorescence staining were performed to calculate the percentage of pro-inflammatory microglia in vivo.A selective JAK2/STAT3 pathway inhibitor, AG490 was used to block STAT3 phosphorylation before the CP-07 anti-inflammation tests in vitro.
Results:
CP-07 could effectively suppress the mRNA levels of IL-6, IL-1β, iNOS and TNF-α induced by lipopolysaccharide (LPS) in vitro, and markedly block the evaluation of the fluorescence intensity of Iba-1 in primary mouse microglia. In middle cerebral arteryocclusion models, intraperitoneal injection with 1 mg/kg CP-07 significantly reduced cerebral infarct volumes at 24 h after surgery compared with vehicle treatment group, and promoted the recovery of neurological functions in MCAO mice. Further studies validated that CP-07 administration reduced the percentage of CD86 positive microglia after I/R injury, and the expression level of p-STAT3 was also markedly reduced in both microglial cells and the penumbra tissues. Blocking STAT3 phosphorylation with AG490 could completely eliminate the anti-inflammatory effects of CP-07, at least in vitro.
Conclusion:
We showed that a newly synthesized compound, CP-07, could effectively reduce the inflammatory responses in LPS-stimulated BV2 cells and primary mouse microglia, and overproduction of cytokines in middle cerebral artery occlusion mouse models by inhibiting STAT3 phosphorylation, leading to a neuroprotective effect on I/R brain injury.
Insights
A novel compound, CP-07, reduces neuroinflammation by inhibiting STAT3 phosphorylation. This compound offers a potential therapy for brain injury caused by infarction/reperfusion (I/R).
Area of Science:
- Neuroscience
- Pharmacology
- Immunology
Background:
- Overactivated glial cells, particularly microglia, drive neuroinflammation and brain injury.
- Anti-inflammatory reagents are explored for treating infarction/reperfusion (I/R) brain injury.
Purpose of the Study:
- To investigate the anti-inflammatory effects of a novel lipophilic compound, CP-07.
- To evaluate CP-07's therapeutic potential in I/R brain injury models.
Main Methods:
- CP-07's efficacy was tested in LPS-stimulated BV2 cells and primary mouse microglia.
- In vivo studies involved middle cerebral artery occlusion (MCAO) models to assess infarct volume and neurological deficits.
- Mechanisms were explored using JAK2/STAT3 pathway inhibition (AG490).
Main Results:
- CP-07 suppressed pro-inflammatory cytokines (IL-6, IL-1β, iNOS, TNF-α) in vitro.
- CP-07 reduced cerebral infarct volume and improved neurological function in MCAO mice.
- CP-07 inhibited STAT3 phosphorylation, a key inflammatory pathway.
Conclusions:
- CP-07 demonstrates significant anti-inflammatory and neuroprotective effects.
- Inhibition of STAT3 phosphorylation is the likely mechanism for CP-07's action.
- CP-07 represents a promising therapeutic candidate for I/R brain injury.
