Rapamycin promotes vascular smooth muscle cell differentiation through insulin receptor substrate-1/phosphatidylinositol 3-kinase/Akt2 feedback signaling. Martin Kathleen A,Merenick Bethany L,Ding Min,Fetalvero Kristina M,Rzucidlo Eva M,Kozul Courtney D,Brown David J,Chiu Helen Y,Shyu Maureen,Drapeau Bethany L,Wagner Robert J,Powell Richard J The Journal of biological chemistry The phenotypic plasticity of mature vascular smooth muscle cells (VSMCs) facilitates angiogenesis and wound healing, but VSCM dedifferentiation also contributes to vascular pathologies such as intimal hyperplasia. Insulin/insulin-like growth factor I (IGF-I) is unique among growth factors in promoting VSMC differentiation via preferential activation of phosphatidylinositol 3-kinase (PI3K) and Akt. We have previously reported that rapamycin promotes VSMC differentiation by inhibiting the mammalian target of rapamycin (mTOR) target S6K1. Here, we show that rapamycin activates Akt and induces contractile protein expression in human VSMC in an insulin-like growth factor I-dependent manner, by relieving S6K1-dependent negative regulation of insulin receptor substrate-1 (IRS-1). In skeletal muscle and adipocytes, rapamycin relieves mTOR/S6K1-dependent inhibitory phosphorylation of IRS-1, thus preventing IRS-1 degradation and enhancing PI3K activation. We report that this mechanism is functional in VSMCs and crucial for rapamycin-induced differentiation. Rapamycin inhibits S6K1-dependent IRS-1 serine phosphorylation, increases IRS-1 protein levels, and promotes association of tyrosine-phosphorylated IRS-1 with PI3K. A rapamycin-resistant S6K1 mutant prevents rapamycin-induced Akt activation and VSMC differentiation. Notably, we find that rapamycin selectively activates only the Akt2 isoform and that Akt2, but not Akt1, is sufficient to induce contractile protein expression. Akt2 is required for rapamycin-induced VSMC differentiation, whereas Akt1 appears to oppose contractile protein expression. The anti-restenotic effect of rapamycin in patients may be attributable to this unique pattern of PI3K effector regulation wherein anti-differentiation signals from S6K1 are inhibited, but pro-differentiation Akt2 activity is promoted through an IRS-1 feedback signaling mechanism. 10.1074/jbc.M703914200

Rapamycin‑induced miR‑30a downregulation inhibits senescence of VSMCs by targeting Beclin1. Tan Pan,Wang Haiqin,Zhan Junkun,Ma Xinyu,Cui Xingjun,Wang Yanjiao,Wang Yi,Zhong Jiayu,Liu Youshuo International journal of molecular medicine Vascular senescence is considered to be an independent risk factor for cardiovascular diseases. The present study aimed to investigate the effects of rapamycin on miR‑30a and its relationship with autophagy and senescence in vascular smooth muscle cells (VSMCs). Young and aging VSMCs were treated with rapamycin or transfected with miR‑30a mimics. Measurement of cellular senescence was conducted using senescence‑associated (SA)‑β‑Galactosidase (gal) staining. Dual luciferase reporter assay was used to confirm binding for miR‑30a and Beclin1. The expression levels of miR‑30a and Beclin1 were determined with reverse transcription‑quantitative polymerase chain reaction analysis. Autophagy‑related protein levels were determined using immunofluorescence or western blot assays. The results demonstrated that rapamycin treatment significantly decreased miR‑30a expression and increased Beclin1 expression in both young and aging cells, as well as promoted autophagy in VSMCs. In addition, rapamycin inhibited senescence in VSMCs and could also alleviate the aging VSMC cycle arrest. Dual luciferase reporter assay confirmed that miR‑30a could directly bind the 3'untranslated region of Beclin1 and inhibit its expression. Furthermore, miR‑30a inhibited autophagy and promoted senescence of VSMCs. In conclusion, the present results indicated that rapamycin could inhibit the senescence of VSMCs by downregulating miR‑30a, which resulted in upregulation of Beclin1 and activation of autophagy. The current study is the first to demonstrate an inhibitory role of rapamycin on VSMC senescence and might provide novel insights and potential new molecular targets in senescence treatment. 10.3892/ijmm.2019.4074

Rapamycin limits the growth of established experimental abdominal aortic aneurysms. Rouer M,Xu B H,Xuan H J,Tanaka H,Fujimura N,Glover K J,Furusho Y,Gerritsen M,Dalman R L European journal of vascular and endovascular surgery : the official journal of the European Society for Vascular Surgery OBJECTIVES:Abdominal aortic aneurysm (AAA) is a chronic inflammatory disease affecting 4-8% of men older than 60 years. No pharmacologic strategies limit disease progression, aneurysm rupture, or aneurysm-related death. We examined the ability of rapamycin to limit the progression of established experimental AAAs. METHODS:AAAs were created in 10-12-week-old male C57BL/6J mice via the porcine pancreatic elastase (PPE) infusion method. Beginning 4 days after PPE infusion, mice were treated with rapamycin (5 mg/kg/day) or an equal volume of vehicle for 10 days. AAA progression was monitored by serial ultrasound examination. Aortae were harvested for histological analyses at sacrifice. RESULTS:Three days after PPE infusion, prior to vehicle or rapamycin treatment, aneurysms were enlarging at an equal rate between groups. In the rapamycin group, treatment reduced aortic enlargement by 38%, and 53% at 3 and 10 days, respectively. On histological analysis, medial elastin and smooth muscle cell populations were relatively preserved in the rapamycin group. Rapamycin treatment also reduced mural macrophage density and neoangiogenesis. CONCLUSION:Rapamycin limits the progression of established experimental aneurysms, increasing the translational potential of mechanistic target of rapamycin-related AAA inhibition strategies. 10.1016/j.ejvs.2014.02.006

A Targeting Nanotherapy for Abdominal Aortic Aneurysms. Cheng Juan,Zhang Runjun,Li Chenwen,Tao Hui,Dou Yin,Wang Yuquan,Hu Houyuan,Zhang Jianxiang Journal of the American College of Cardiology BACKGROUND:Abdominal aortic aneurysm (AAA) is a leading cause of mortality and morbidity in the elderly. Currently, there remain no effective drugs that can prevent the growth of aneurysms and delay aneurysm rupture in the clinical setting. OBJECTIVES:The aim of this study was to develop a nanotherapy that can target aneurysms and release drug molecules in response to the inflammatory microenvironment. METHODS:Using a reactive oxygen species (ROS)-responsive nanoparticle and a candidate drug rapamycin, in combination with a peptide ligand for integrin and biomimetic cloaking with macrophage cell membrane, a nanotherapy was developed. Its effectiveness was demonstrated by in vitro and in vivo studies. RESULTS:Based on a facile and translational method, a rapamycin-loaded responsive nanotherapy was successfully prepared, which could release drug molecules upon triggering by the high level of ROS. In cells associated with the development of AAAs, the nanotherapy significantly inhibited calcification and attenuated ROS-mediated oxidative stress and apoptosis. By passively targeting aneurysms and releasing drug molecules in response to the inflammatory microenvironment, the intravenously injected ROS-responsive nanotherapy more effectively prevented aneurysm expansion in AAA rats than a nonresponsive control nanotherapy. After decoration with a peptide ligand cRGDfK and macrophage cell membrane, the aneurysmal targeting capability and therapeutic effects of a ROS-responsive nanotherapy with a mean diameter of 190 nm were further enhanced. Moreover, the nanotherapy showed a good safety profile in a preliminary safety test. CONCLUSIONS:The multifunctional nanotherapy can be further studied as a promising targeted drug for treatment of aneurysms. The underlying design principles enable the development of a broad range of nanomedicines for targeted therapy of other vascular diseases. 10.1016/j.jacc.2018.08.2188

Rapamycin-Loaded Biomimetic Nanoparticles Reverse Vascular Inflammation. Boada Christian,Zinger Assaf,Tsao Christopher,Zhao Picheng,Martinez Jonathan O,Hartman Kelly,Naoi Tomoyuki,Sukhoveshin Roman,Sushnitha Manuela,Molinaro Roberto,Trachtenberg Barry,Cooke John P,Tasciotti Ennio Circulation research RATIONALE:Through localized delivery of rapamycin via a biomimetic drug delivery system, it is possible to reduce vascular inflammation and thus the progression of vascular disease. OBJECTIVE:Use biomimetic nanoparticles to deliver rapamycin to the vessel wall to reduce inflammation in an in vivo model of atherosclerosis after a short dosing schedule. METHODS AND RESULTS:Biomimetic nanoparticles (leukosomes) were synthesized using membrane proteins purified from activated J774 macrophages. Rapamycin-loaded nanoparticles were characterized using dynamic light scattering and were found to have a diameter of 108±2.3 nm, a surface charge of -15.4±14.4 mV, and a polydispersity index of 0.11 +/ 0.2. For in vivo studies, ApoE mice were fed a high-fat diet for 12 weeks. Mice were injected with either PBS, free rapamycin (5 mg/kg), or rapamycin-loaded leukosomes (Leuko-Rapa; 5 mg/kg) once daily for 7 days. In mice treated with Leuko-Rapa, flow cytometry of disaggregated aortic tissue revealed fewer proliferating macrophages in the aorta (15.6±9.79 %) compared with untreated mice (30.2±13.34 %) and rapamycin alone (26.8±9.87 %). Decreased macrophage proliferation correlated with decreased levels of MCP (monocyte chemoattractant protein)-1 and IL (interleukin)-b1 in mice treated with Leuko-Rapa. Furthermore, Leuko-Rapa-treated mice also displayed significantly decreased MMP (matrix metalloproteinases) activity in the aorta (mean difference 2554±363.9, =9.95122×10). No significant changes in metabolic or inflammation markers observed in liver metabolic assays. Histological analysis showed improvements in lung morphology, with no alterations in heart, spleen, lung, or liver in Leuko-Rapa-treated mice. CONCLUSIONS:We showed that our biomimetic nanoparticles showed a decrease in proliferating macrophage population that was accompanied by the reduction of key proinflammatory cytokines and changes in plaque morphology. This proof-of-concept showed that our platform was capable of suppressing macrophage proliferation within the aorta after a short dosing schedule (7 days) and with a favorable toxicity profile. This treatment could be a promising intervention for the acute stabilization of late-stage plaques. 10.1161/CIRCRESAHA.119.315185