Retinoic Acid Family Compounds Influence Secreted ApoE Level In Astrocytes And The Study Of Related Mechanism

Alzheimer’s disease (AD) is the most common form of dementia in the elderly population. The accumulation and deposition of amyloid-β (Aβ) peptides cleaved from amyloid precursor protein (APP) in the brain is thought to be a central event in AD pathogenesis. Whilst multiple genetic and environmental factors are involved in AD pathogenesis, the ε4 allele of the apolipoprotein E (APOE) gene is the strongest genetic risk factor for late-onset AD compared to the common allele APOE3 or the protective allele ApoE2. ApoE isoforms differentially regulate AD pathogenesis through Aβ-dependent and Aβ-independent pathways. ApoE4 suppresses Aβ clearance in the brain compared with apoE3, which results in accelerated Aβ deposition as senile plaques and cerebral amyloid angiopathy. Interestingly, apoE protein levels in cerebrospinal fluid (CSF) and plasma are reduced in ApoE4 carriers and AD patients. Furthermore, low levels of total cholesterol also increase the risk of AD. Therefore, increasing apoE and cholesterol levels in the brain may be an effective therapeutic strategy for AD. In fact, treatment of AD mice with liver X receptor (LXR) agonists or retinoid X receptor (RXR) agonists consistently improves memory and reduces Aβ levels by increasing apoE level and lipidation in amyloid model mice. However, despite their efficacy and tolerability in rodents, the controversy on drug toxicity still precludes their translation into human clinical trials. To identify novel compounds that can regulate apoE production, we performed a drug screen using a Known Bioactives Library from Institute of Chemistry and Cell Biology Bioactives (ICCB), which contains 480 compounds with defined biological activity and a culture system of immortalized astrocytes from human ApoE3 targeted replacement (TR) mice. Then we made further verification and analyze to the screening result, followed by the mechanism exploration on some compounds, and finally got these data:1. Through the primary screening of all 480 compounds,14 compounds were found to increase secreted apoE level, while 27 compounds decreased the apoE level in the medium. The 14 compounds that increase apoE secretion contained all-trans retinoic acid (all-trans RA, ATRA),9-cis retinoic acid (9-cis RA,9RA) and 13-cis retinoic acid (13-cis RA,13RA), which are RA family members. Through further verification, we found that all-trans RA,9-cis RA,13- cis RA and bexarotene significantly increased apoE secretion in a concentration-dependent manner. Then we analyzed the apoE mRNA levels by qRT-PCR. RA isomers and bexarotene facilitate apoE production at least partially by increasing apoE expression. RA isomers increase apoE secretion in primary culture of astrocytes from apoE3- and apoE4-TR mice.2. In the following mechanism study, we found RA isomers and bexarotene increase ApoE lipidation by stimulating ABCA1 and ABCG1 mRNA and protein expression. We also found LDLR and LRP1 expression at both protein and mRNA levels were not changed by tested compounds, indicating that cellular apoE clearance pathways are unlikely involved in the mechanisms. RA isomers and bexarotene facilitate apoE production through activation of the RXR/RAR pathways, which may require LXR. RA isomers suppress cellular Aβ uptake in an apoE-dependent manner in immortalized astrocytes and protect neurons from Abeta toxicity.3. Finally, we checked the effect of RA isomers in the ApoE3-TR mice. We found bexarotene and 9-cis RA increase ABCA1, ABCG1 and apoE levels in the brains of apoE3-TR mice. In addition, we found that there was no significant toxicity in the drug treated groups.Taken together, our results demonstrate that RXR/RAR agonists, including several RA isomers, are effective modulators of ApoE secretion and lipidation, and may be explored as potential drugs for AD therapy.