TSG attenuates LPC-induced endothelial cells inflammatory damage through notch signaling inhibition.

Lysophosphatidylcholine (LPC) induces inflammation in endothelial cells (ECs) but the mechanism is not fully understood. The Notch signaling pathway is involved in chronic EC inflammation, but its functions in LPC-induced endothelial inflammatory damage and 2,3,5,4'-tetrahydroxystilbene-2-O-ß-d-glucoside's (TSG) protective effect during LPC-induced inflammatory damage in human umbilical vein endothelial cells (HUVECs) is largely unknown. We report that Notch signaling activation contributed to LPC-induced injury in HUVECs, and that TSG protected HUVECs from LPC-induced injury by antagonizing Notch signaling activation by LPC. γ-secretase inhibitor (DAPT), a specific inhibitor of the Notch signaling pathway, and Notch1 siRNA were used to inhibit Notch activity. HUVECs were exposed to LPC in the presence or absence of TSG, DAPT, and Notch1 siRNA. LPC treatment of HUVECs resulted in reduced cell viability, and Notch1 and Hes1 upregulation. Either silencing of Notch1 by siRNA or pharmacological inhibition of Notch signaling by DAPT prevented the loss of cell viability, and induction of apoptosis, and enhanced expression Notch1, Hes1 and MCP-1 by LPC in HUVECs. Similarly, TSG reduced LPC stimulation of Notch1, Hes1, and MCP-1 expression, prevented the release of IL-6 and CRP and rescued HUVECs from LPC-induced cell damage. Our data indicate that the Notch signaling pathway is a crucial mediator of endothelial inflammatory damage and that TSG protects against endothelial inflammatory damage by inhibiting the Notch signaling pathway. Our findings suggest that targeting Notch signaling by natural products such as TSG is a promising strategy for the prevention and treatment of chronic inflammation associated diseases, including atherosclerosis. © 2015 IUBMB Life, 68(1):37-50, 2016.

Proliferation of rat cardiac stem cells is induced by 2, 3, 5, 4'-tetrahydroxystilbene-2-O-ß-D-glucoside in vitro.

AIM: To study the effects of 2, 3, 5, 4'-tetrahydroxystilbene-2-O-ß-d-glucoside (THSG) on proliferation of rat cardiac stem cells (CSCs) in vitro. MATERIALS AND METHODS: C-kit(+) cells were isolated from neonatal (1 day old) Sprague-Dawley rats by using flow cytometry. Optimal THSG treatment times and doses for growth of CSCs were analyzed. CSCs were treated with various THSG doses (0, 1, 10, and 100 µM) for 12h. RESULTS: Sorted c-kit(+) cells exhibited self-renewing and clonogenic capabilities. Cell Counting Kit (CCK-8) and Proliferating Cell Nuclear Antigen (PCNA) ELISA test positive cells were significantly increased in THSG-treated groups compared with untreated controls. The percentage of S-phase cells also increased after THSG treatment. Moreover, we show that some c-kit(+) cells spontaneously express vascular endothelial growth factor (VEGF), T-box transcription factor (Tbx5), hyperpolarization-activated cyclic nucleotide-gated 2 (HCN2), hyperpolarization-activated cyclic nucleotide gated 4 (HCN4), alpha myosin heavy chain (αMHC), and beta myosin heavy chain (ßMHC) mRNA, and stem cell antigen 1 (Sca-1), cardiac troponin-I, GATA-4, Nkx2.5, and connexin 43 protein were also assessed in CSCs. However, their expression was significantly increased with THSG treatment when compared to untreated controls. CONCLUSION: THSG can increase proliferation of rat CSCs in vitro and thus, shows promise as a potential treatment strategy for stimulating endogenous stem cells to help repair the injured heart after myocardial infarction in patients.

2,3,5,4'-tetrahydroxystilbene-2-O-ß-D-glucoside protects human umbilical vein endothelial cells against lysophosphatidylcholine-induced apoptosis by upregulating superoxide dismutase and glutathione peroxidase.

2,3,5,4'-Tetrahydroxystilbene-2-O-ß-d-glucoside (TSG) has been shown to protect human umbilical vein endothelial cells (HUVECs) from lysophosphatidylcholine (LPC)-induced injury; however, the underlying molecular mechanism remains to be determined. The aim of this study was to investigate the protective mechanism of TSG against LPC-induced injury in HUVECs. We established a stable LPC-induced cell model by treating HUVECs with various concentrations of LPC and found 10.0 µg/mL of LPC to be optimal for inducing HUVECs injury. The effects of TSG on LPC-induced cell injury were assessed by cell counting kit-8, apoptosis assay, transmission electron microscope, and measurement of malondialdehyde (MDA), the antioxidant enzymes superoxide dismutase (SOD), reactive oxygen species (ROS), glutathione peroxidase, and mitochondrial membrane potential. The mRNA and protein levels of caspase-3, Bax, Bcl-2, PARP-1, and cytochrome C were assayed by real-time reverse transcriptase-polymerase chain reaction and immunoblotting, respectively. TSG pretreatment was able to prevent LPC-induced HUVECs injury and restore cell viability in a concentration-dependent manner. LPC treated cells showed typical apoptotic morphological changes including cytoplasmic vacuolation, swollen mitochondria, and characteristic biochemical hallmarks of apoptosis including loss of mitochondrial membrane potential, activation of caspase-3, decrease of Bcl-2, increase of PARP-1, upregulation of Bax, and release of cytochrome C, all of which were apparently inhibited by TSG pretreatment. Treatment of HUVECs with LPC led to decrease of SOD and glutathione peroxidase in addition to rapid increase of MDA and ROS levels. Pretreatment with TSG restored SOD and glutathione peroxidase levels to that of normal levels, and significantly decreased ROS and MDA levels. Our data indicate that TSG inhibits apoptosis of HUVECs mediated by LPC through blocking the mitochondrial apoptotic pathway and suggest that the mechanisms underlying the protective effects of TSG are related to the activation of SOD and glutathione peroxidase, the clearance of intracellular ROS, and reduction of lipid peroxidation.