ArticleLipids in health and disease2017
The purification of reduced β2-glycoprotein I showed its native activity in vitro.
Article in Lipids in health and disease, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 3 papers.
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3 citing papers in PubMed, 4 citations in OpenAlex.
- Article
- β2 glycoprotein I participates in phagocytosis of apoptotic neurons and in vascular injury in experimental brain stroke.Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism · 2021Article
- ReducedBioMed research international · 2018Article
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8 authors at 1 institution in 1 country.
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No grant is acknowledged in the PubMed record.
Abstract
backgroundNew evidence has shown that reduced β2-glycoprotein I (β2GPI) has anti-oxidative stress and anti-inflammatory activity. However, the details are still poorly understood. This study aims to prepare stable reduced β2GPI with its native bioactivity in vitro.
methodsHuman β2GPI was purified from plasma first with perchloric acid precipitation and then purified with a series of chromatography methods including Sephadex G-25 desalting, SP HP, AF-heparin HC-650 M, and Sephacryl S-200. The purified human β2GPI was reduced with thioredoxin-1 (TRX-1) activated by DL-dithiothreitol (DTT). Glutathione (GSH) was selected to block the free thiols in reduced β2GPI. LC/MS was used to verify the location of free thiols. Western blot analysis was used to detect β2GPI immunoreactivity. MTS and flow cytometry were conducted to investigate its biological effect on oxidative-stress-induced death of human umbilical vein endothelial cells (HUVECs). The levels of tumour necrosis factor-alpha (TNF-α),interleukin-6 (IL-6) interleukin-10 (IL-10),interleukin-12P70 (IL-12P70),interferon-gamma (IFN-γ) and monocyte chemoattractant protein -1(MCP-1) in mouse serum were quantified to assess its anti-inflammatory activity in lipopolysaccharide (LPS)-mediated systemic inflammation.
resultsWe obtained approximately 10 mg β2GPI (purity 98.7%) from 200 ml plasma. The protein yield was 0.05 mg/ml plasma. β2GPI was then reduced by TRX-1/DTT in vitro; the free thiols were detected on Cys288 and Cys326 in domain V of β2GPI. The GSH blockage stabilized the reduced β2GPI in vitro. This reduced β2GPI can be recognized by the anti-β2GPI antibody, can significantly reduce the death of HUVECs after H
conclusionStable reduced β2GPI can be obtained in vitro by TRX-1 deoxidation followed by the blockage of thiols with GSH. This reduced β2GPI maintains the same immunological activity as oxidized β2GPI and has the ability to counter the oxidative stress induced by H
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