CCN family member 1 deregulates cholesterol metabolism and aggravates atherosclerosis.

AIM: CCN family member 1 (CCN1) is an extracellular matrix cytokine and appears in atherosclerotic lesions. However, we have no evidence to support the role of CCN1 in regulating cholesterol metabolism and atherosclerosis. METHODS: Apolipoprotein E-deficient (apoE-/- ) mice were used as in vivo model. Oxidized low-density lipoprotein (oxLDL)-induced macrophage-foam cells were used as in vitro model. RT-PCR and western blot analysis were used for evaluating gene and protein expression, respectively. Conventional assay kits were used for assessing the levels of cholesterol, triglycerides, and cytokines. RESULTS: We show predominant expression of CCN1 in foamy macrophages in atherosclerotic aortas of apoE-/- mice. In apoE-/- mice, CCN1 treatment worsened hyperlipidaemia, systemic inflammation, and the progression of atherosclerosis. In addition, CCN1 decreased the capacity of reverse cholesterol transport and downregulated the protein expression of ATP-binding cassette transporter A1 (ABCA1) and ABCG1 in atherosclerotic aortas. Notably, CCN1 decreased the protein expression of cholesterol clearance-related proteins, including ABCG5, ABCG8, liver X receptor α (LXRα), cholesterol 7α-hydrolase and LDL receptor in liver, and exacerbated hepatic lipid accumulation. In macrophages, treatment with oxLDL increased CCN1 expression. Inhibition of CCN1 activity by neutralizing antibody or small interfering RNA attenuated the oxLDL-induced lipid accumulation. In contrast, cotreatment with CCN1 or overexpression of CCN1 augmented oxLDL-induced lipid accumulation by impairing apolipoprotein AI- and high-density lipoprotein-dependent cholesterol efflux, which was attributed to downregulation of LXRα-dependent expression of ABCA1 and ABCG1. CONCLUSION: Our findings suggest that CCN1 plays a pivotal role in regulating cholesterol metabolism and the development of atherosclerosis.

Novel effect of paeonol on the formation of foam cells: promotion of LXRα-ABCA1-dependent cholesterol efflux in macrophages.

Paeonol, a phenolic component purified from Paeonia suffruticosa (Cortex Moutan), is used in traditional Chinese medicine to treat inflammatory diseases. However, little is known about the effect of paeonol on cholesterol metabolism. We investigated the efficacy of paeonol on cholesterol metabolism and the underlying mechanism in macrophages and apolipoprotein E deficient (apoE(-/-)) mice. Treatment with paeonol markedly attenuated cholesterol accumulation induced by oxidized LDL in macrophages, which was due to increased cholesterol efflux. Additionally, paeonol enhanced the mRNA and protein expression of ATP-binding membrane cassette transport protein A1 (ABCA1) but did not alter the protein level of ABCG1 or other scavenger receptors. Inhibition of ABCA1 activity with a pharmacological inhibitor, neutralizing antibody or small interfering RNA (siRNA), negated the effects of paeonol on cholesterol efflux and cholesterol accumulation. Furthermore, paeonol induced the nuclear translocation of liver X receptor α (LXRα) by increasing its activity. siRNA knockdown of LXRα abolished the paeonol-induced upregulation of ABCA1, promotion of cholesterol efflux and suppression of cholesterol accumulation. Moreover, atherosclerotic lesions, hyperlipidemia and systemic inflammation were reduced and the protein expression of ABCA1 was increased in aortas of paeonol-treated apoE(-/-) mice. Paeonol may alleviate the formation of foam cells by enhancing LXRα-ABCA1-dependent cholesterol efflux.

A novel anti-CCN1 monoclonal antibody suppresses Rac-dependent cytoskeletal reorganization and migratory activities in breast cancer cells.

CCN1, a secreted matrix-associated molecule, is involved in multiple cellular processes. Accumulating evidence supports that CCN1 plays an important role in tumorigenesis and progression of breast cancer. In this study, we have developed a novel CCN1 function-blocking monoclonal antibody (mAb), designated YM1B. YM1B binds to human CCN1 with high specificity, recognizing the native CCN1 structure with undisturbed disulfide linkages. Our analyses have mapped the YM1B recognition region to domain IV of CCN1, likely in proximity to the DM site. In breast cancer cells, CCN1 can induce actin reorganization, formation of lamellipodia, and cell migration/invasion through the αV integrins/Rac1/ERK signaling axis; these CCN1-dependent activities can be effectively suppressed by YM1B. Our results also suggest that YM1B may exert its CCN1-blocking effect by perturbing the interaction of CCN1 with vitronectin and fibronectin, which are ligands of αV integrins and instrumental for integrin activation. This CCN1-specific mAb may open a new potential avenue for therapeutic intervention of breast cancer progression.

The matricellular protein CCN1 suppresses lung cancer cell growth by inducing senescence via the p53/p21 pathway.

CCN1, a secreted matrix-associated molecule, is involved in multiple cellular processes. Previous studies have indicated that expression of CCN1 correlates inversely with the aggressiveness of non-small-cell lung carcinoma (NSCLC); however, the underlying mechanisms remain elusive. Using three NSCLC cell line systems, here we show that long-term treatment of cells with the recombinant CCN1 protein led to a permanent cell cycle arrest in G1 phase; cells remained viable as judged by apoptotic assays. CCN1-treated NSCLC cells acquired a phenotype characteristic of senescent cells, including an enlarged and flattened cell shape and expression of the senescence-associated ß-galactosidase. Immunoblot analysis showed that addition of CCN1 increased the abundance of hypo-phosphorylated Rb, as well as accumulation of p53 and p21. Silencing the expression of p53 or p21 by lentivirus-mediated shRNA production in cells blocked the CCN1-induced senescence. Furthermore, a CCN1 mutant defective for binding integrin α6ß1 and co-receptor heparan sulfate proteoglycans was incapable of senescence induction. Our finding that direct addition of CCN1 induces senescence in NSCLC cells provides a potential novel strategy for therapeutic intervention of lung cancers.