This study shows that liraglutide can affect the proliferation, migration, calcification and apoptosis of VSMCs through its effect on autophagy, thereby achieving anti-AS effects. These data also reveal a new perspective regarding the anti-AS biological mechanisms of liraglutide.
AS is a serious cardiovascular condition associated with complications of diabetes. Vascular remodeling is vital to the process of AS, which affects pathologic processes in VSMCs, including proliferation, migration, calcification and apoptosis[8, 35–37]. High glucose conditions promote proliferation, migration and calcification, but inhibit apoptosis[4, 8, 38–41].
Autophagy in VSMCs also plays an important pathophysiological role in AS[21–23]. Xue et al. showed that the autophagy of gastric smooth muscle cells was significantly increased after 24 hr of culture with high-glucose[42]. Chen et al. showed that high glucose inhibits the autophagy of VSMCs in the superior mesenteric artery[43]. This study indicated that high glucose can induce autophagy in VSMCs and that liraglutide can inhibit autophagy induced by high glucose. This is similar to the results from Zhao et al., which showed that high glucose can induce autophagy in human renal tubular endothelial cells (HK-2 cells), while liraglutide can inhibit this autophagy in a dose-dependent manner[44]. However, another study showed that high glucose can inhibit cardiomyocyte autophagy and that liraglutide can increase the level of cardiomyocyte autophagy[26, 45]. Therefore, autophagy in different cell types might participate in the progression of disease through diverse mechanisms, and high glucose and liraglutide might affect autophagy in different cell types through a variety of mechanisms.
However, the specific mechanism of autophagy in VSMCs is still unclear. The PI3K/AKT signaling pathway can regulate cell proliferation and migration, and inhibit cell apoptosis[46, 47]. Previous studies have found that high glucose can induce dynamic changes in VSMCs through the PI3K/Akt pathway[4, 21–24]. Moreover, our previous data also showed that liraglutide exerts beneficial effects on VSMCs partly through the inhibition of extracellular signal-regulated kinase (ERK)1/2 and PI3K/AKT pathways[4]. Therefore, PI3K/AKT signaling possibly plays a large role in the tissue damage induced by AS, which may be an important pathway for GLP-1 to exert cardiovascular protection. A variety of signaling pathways including mammalian target of rapamycin (mTOR), 5’-AMP-activated protein kinase (AMPK), tumor protein 53 (p53) as well as PI3K/AKT are involved in autophagy[48–50]. Vasopressin inhibits autophagy and promotes cell proliferation of VSMCs cultured with high glucose through the PI3K/AKT/mTOR signaling pathway[51]. In this study, to further investigate the specific mechanisms of the effect of liraglutide on autophagy in VSMCs under high glucose conditions, the PI3K inhibitor LY294002 was used. The results showed that high glucose affects autophagy in VSMCs through the PI3K/AKT signaling pathway. This is consistent with previous studies that have indicated that the effect of high glucose on the biological characteristics of VSMCs is dependent on the PI3K/AKT pathway[4, 41, 52, 53].
Moreover, several cells types, including cardiomyocytes, macrophages, and VSMCs all express the GLP-1R, which mediates the anti-inflammatory and anti-proliferative effects of GLP-1[16, 54, 55]. To investigate if the effects of liraglutide in high glucose-treated cells occur via GLP-1R, VSMCs were pretreated with Exe9–39. Indeed, Exe9–39 treatment abolished the beneficial effects of liraglutide treatment.
How liraglutide protects cardiovascular in VSMCs remains unclear. Previous studies indicated that autophagy is closely related to AS and biological changes of VSMCs. Li et al. found that the secreted protein sonic hedgehog (SHH) can induce autophagy, which can promote the proliferation of VSMCs[56]. Marina et al. found that tumor necrosis factor alpha (TNF-α) promotes the proliferation and migration of VSMCs through autophagy[57]. Salabei et al. found that platelet-derived growth factor (PDGF) can further promote the phenotypic transition and calcification of VSMCs by promoting autophagy[58]. Aleksandar et al. have shown that early autophagy promotes an increase of ALP, Runt-related transcription factor 2 (Runx2), and bone morphogenic protein 2 (BMP2) in cells, thereby promoting the occurrence of cell calcification[59]. Park et al. found that reactive oxygen species (ROS) regulate the autophagy of VSMCs induced by PDGF and exert an anti-apoptotic effect[60]. The present study showed that autophagy induced by high glucose can promote proliferation, migration and calcification but inhibit apoptosis, which is consistent with the literature cited above.
However, there are certain studies that are inconsistent with the findings of the present study. One study showed that high glucose induces the activation of mTOR in VSMCs, thereby inhibiting autophagy[51]. Chronic hyperglycemia inhibits AMPK activation and autophagy, thereby promoting VSMC proliferation and migration[61, 62]. Research by Chen et al. showed that high glucose can upregulate endothelin type B (ETB) receptors and inhibit VSMC autophagy through AMPK and mTOR signaling pathways[43]. These studies have shown that high glucose promotes VSMC proliferation and migration by inhibiting autophagy. In addition, rapamycin (an inhibitor of the mTOR pathway and inducer of autophagy) can prevent phenotypic transition and excessive proliferation of VSMCs, and therefore can prevent restenosis after angioplasty. These data indicate that autophagy in VSMCs might exert an anti-AS effect by inhibiting phenotypic transition and excessive proliferation[63]. Although the results of these studies are different from the current study, all of the outcomes are considered to be related to the degree of autophagy. Specifically, under physiological conditions, autophagy of VSMCs can inhibit cell proliferation, thereby further inhibiting the development of AS. However, excessive autophagy or insufficient VSMC autophagy can promote the development of AS[23, 64]. To study whether autophagy was involved in the anti-AS effect of liraglutide, the inhibitor of autophagy 3-MA was used to pretreat VSMCs under high glucose conditions and cells were then exposed to liraglutide (HG + LIRA). The results indicated that liraglutide can affect the proliferation, migration, calcification and apoptosis of VSMCs by inhibiting autophagy under high glucose conditions. Li et al. showed that liraglutide can inhibit insulin cell apoptosis induced by high glucose by promoting autophagy[25]. Yu et al. showed that liraglutide and exenatide can achieve anti-apoptotic effects by promoting autophagy in cardiomyocytes under high glucose[26]. Those differences are most likely related to the different cell types used in different experimental setttings. These results also suggest that the mechanisms of liraglutide in different tissues and cell types, and in diverse diseases, might be not the same.