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A New Cell Model Overexpressing sTGFBR3 for Studying Alzheimer's Disease In vitro
Jiangxia Chen1, Lijun Zhou1, Qingchun Zhao1
1General Hospital of Northern Theatre Command, Bei Fang Hospital of Shenyang Pharmaceutical University, Shenyang, China.
Background:
Recent studies have suggested that abnormal microglial hyperactivation has an important role in the progression of Alzheimer's disease (AD). sTGFBR3 (a shed extracellular domain of the transforming growth factor type III receptor) is a newly identified target of microglia polarization dysregulation, whose overexpression can cause abnormal accumulation of transforming growth factor β1 (TGF-β1), promoting Aβ, tau, and neuroinflammatory pathology.
Objective:
The objective of this study is to develop and validate a new cell model overexpressing sTGFBR3 for studying AD in vitro.
Methods:
BV2 cells (a microglial cell derived from C57/BL6 murine) were used as a cell model. Cells were then treated with different concentrations of lipopolysaccharide (LPS) (0, 1, or 0.3 μg/mL) for 12, 24, or 48h and then with or without sodium pervanadate (100 μM) for 30 min. Next, the effect surface optimization method was used to determine optimal experimental conditions. Finally, the optimized model was used to assess the effect of ZQX series compounds and vasicine on cell viability and protein expression. Expression of TGFBR3 and TNF-α was assessed using Western blot. MTT assay was used to assess cell viability, and enzyme- linked immunosorbent assay (ELISA) was employed to evaluate extracellular TGF-β1 and sTGFBR3.
Results:
LPS (0.3 μg/mL) treatment for 11 h at a cell density of 60% and pervanadate concentration (100 μM) incubation for 30 min were the optimal experimental conditions for increasing membrane protein TGFBR3 overexpression, as well as extracellular sTGFBR3 and TGF-β1. Applying ZQX-5 and vasicine reversed this process by reducing extracellular TGF-β1, promoting the phosphorylation of Smad2/3, a protein downstream of TGF-β1, and inhibiting the release of the inflammatory factor TNF-α.
Conclusion:
This new in vitro model may be a useful cell model for studying Alzheimer's disease in vitro.
Insights
This study developed a novel cell model for Alzheimer's disease (AD) research by overexpressing sTGFBR3 in microglia. The model helps study how sTGFBR3 influences AD pathology and tests potential therapeutic compounds.
Area of Science:
- Neuroscience
- Cell Biology
- Pharmacology
Background:
- Microglial hyperactivation is implicated in Alzheimer's disease (AD) progression.
- Overexpression of sTGFBR3 (shed transforming growth factor type III receptor) in microglia leads to TGF-β1 accumulation, exacerbating AD pathology.
Purpose of the Study:
- To develop and validate a new in vitro cell model overexpressing sTGFBR3 for Alzheimer's disease research.
Main Methods:
- Utilized BV2 microglial cells, optimizing conditions with LPS and sodium pervanadate.
- Assessed cell viability (MTT assay) and protein expression (Western blot, ELISA) of TGFBR3, sTGFBR3, TGF-β1, and TNF-α.
- Evaluated the effects of ZQX-5 and vasicine compounds on the optimized model.
Main Results:
- Optimal conditions (0.3 μg/mL LPS for 11h, 60% cell density, 100 μM pervanadate for 30 min) induced sTGFBR3 and TGF-β1 overexpression.
- ZQX-5 and vasicine treatments reduced TGF-β1, promoted Smad2/3 phosphorylation, and inhibited TNF-α release.
Conclusions:
- A novel in vitro cell model overexpressing sTGFBR3 was successfully developed and validated.
- This model serves as a valuable tool for investigating Alzheimer's disease mechanisms and evaluating potential therapeutics.
