BACKGROUND: Numerous genome-wide association studies revealed that SNPs at the PHACTR1 (phosphatase and actin regulator 1) locus strongly correlate with coronary artery disease. However, the biological function of PHACTR1 remains poorly understood. Here, we identified the proatherosclerotic effect of endothelial PHACTR1, contrary to macrophage PHACTR1. METHODS: We generated global ( Phactr1 −/ − ) and endothelial cell (EC)–specific ( Phactr1 ECKO ) Phactr1 KO (knockout) mice and crossed these mice with apolipoprotein E–deficient ( Apoe −/− ) mice. Atherosclerosis was induced by feeding the high-fat/high-cholesterol diet for 12 weeks or partially ligating carotid arteries combined with a 2-week high-fat/high-cholesterol diet. PHACTR1 localization was identified by immunostaining of overexpressed PHACTR1 in human umbilical vein ECs exposed to different types of flow. The molecular function of endothelial PHACTR1 was explored by RNA sequencing using EC-enriched mRNA from global or EC-specific Phactr1 KO mice. Endothelial activation was evaluated in human umbilical vein ECs transfected with siRNA targeting PHACTR1 and in Phactr1 ECKO mice after partial carotid ligation. RESULTS: Global or EC-specific Phactr1 deficiency significantly inhibited atherosclerosis in regions of disturbed flow. PHACTR1 was enriched in ECs and located in the nucleus of disturbed flow areas but shuttled to cytoplasm under laminar flow in vitro. RNA sequencing showed that endothelial Phactr1 depletion affected vascular function, and PPARγ (peroxisome proliferator-activated receptor gamma) was the top transcription factor regulating differentially expressed genes. PHACTR1 functioned as a PPARγ transcriptional corepressor by binding to PPARγ through the corepressor motifs. PPARγ activation protects against atherosclerosis by inhibiting endothelial activation. Consistently, PHACTR1 deficiency remarkably reduced endothelial activation induced by disturbed flow in vivo and in vitro. PPARγ antagonist GW9662 abolished the protective effects of Phactr1 KO on EC activation and atherosclerosis in vivo. CONCLUSIONS: Our results identified endothelial PHACTR1 as a novel PPARγ corepressor to promote atherosclerosis in disturbed flow regions. Endothelial PHACTR1 is a potential therapeutic target for atherosclerosis treatment.

The pathogenesis of Alzheimer’s disease (AD) involves multiple pathophysiological processes. Oxidative stress is a major cause of AD-associated neuronal injury. The current research was designed to examine whether a novel (-)-meptazinol-serotonin hybrid (Mep-S) with potent antioxidant activity and additional inhibitory properties for acetylcholinesterase (AChE) activity could attenuate oxidative neuronal damage and cognitive deficits. In human SH-SY5Y cells, Mep-S suppressed H2O2-induced apoptosis by restoring mitochondrial membrane potential and inhibiting caspase-3 activation. Meanwhile, it attenuated oxidative stress elicited by H2O2 through lessening generation of reactive oxygen species as well as enhancing production of glutathione (GSH) and activity of superoxide dismutase (SOD). Mechanistically, Mep-S promoted nuclear translocation of a transcription factor nuclear factor E2-related factor-2 (Nrf2) in H2O2-challenged cells. This effect was accompanied by reduction in Kelch-like ECH-associated protein-1 (Keap1) levels as well as augmentation of Akt phosphorylation and expression of heme oxygenase-1 (HO-1) and NAD(P)H quinine oxidoreductase-1 (NQO-1). Molecular docking analysis revealed that Mep-S may disrupt the protein-protein interactions between Keap1 and Nrf2. In an in vivo mouse model, Mep-S attenuated scopolamine-caused cognitive deficits with inhibition of apoptotic neuronal death and brain AChE activity. Furthermore, the scopolamine-induced impairment of total antioxidant capacity and reduction in SOD1, SOD2, and γ-glutamate-cysteine ligase expression in the brain were counteracted by Mep-S, accompanied by decreased Keap1 levels, increased Akt catalytic subunit and Nrf2 phosphorylation, and decreased Nrf2, HO-1, and NQO-1 expression. Collectively, our results suggest that Mep-S ameliorates apoptotic neuronal death and memory dysfunction associated with oxidative stress by regulating the Nrf2/antioxidant enzyme pathway through inactivating Keap1 and phosphorylating Nrf2 via Akt activation. Therefore, Mep-S may be a potential lead for multitarget neuroprotective agents to treat AD-like symptoms.

Heterotypic interaction between tumor cells and adjacent stromal cells mediates tumor development. However, how tumor heterogeneity commits tumors to the malignant transformation and evasion of immunity against metastasis is poorly understood. Here, we have investigated the fusogenicity of human invasive glioblastoma, triple negative breast cancer and gallbladder cancer cells that are all characterized by mesenchymal cell plasticity. These cells displayed the rigorous ability to fuse with macrophages and augment epithelial-mesenchymal transition (EMT), transforming the fused cells into highly invasive hybrids. YKL-40 (Chitinase-3-like-1), known to promote inflammation and serve as an EMT marker, was essential and sufficient for both cell fusion and the invasiveness of tumor cells that express EMT and tumor-associated macrophage markers. Intriguingly, differential gene profiling of single clones from the hybrids demonstrated that YKL-40 and immune checkpoint protein B7-2 (CD86) were elevated and functioned to independently suppress anti-tumor immune factor levels of CD8+-cytotoxic T lymphocytes (CTL); thus resulting in escape of immune surveillance. YKL-40 and B7-2 dual shRNA abrogated YKL-40-mediated cell fusion and restored CTL anti-tumor immunity, compromising tumor development in xenografts. Clinically, we found tumor hybrids were present in mesenchymal types of glioblastomas, gallbladder cancer and breast cancer. In addition, YKL-40 expression in glioblastoma was correlated with decreased disease-free survival in patients. Collectively, these data offer novel cellular and molecular mechanisms underpinning immune evasion and tumor malignancy, and suggest a new immunotherapeutic intervention strategy by targeting both YKL-40 and B7-2 in cancer.

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