Endothelia inflammation damage is vital to the development and progression of chronic venous disease. In the present study, we explored the protective effect of melatonin on endothelia apoptosis induced by LPS, particularly focusing on the mitochondrial fission. We demonstrated that human umbilical vein endothelial cells (HUVEC) subjected to LPS for 12 h exhibited a higher apoptotic rate. However, melatonin (1-20 μM) treatment 12 h before LPS had the ability to protect HUVEC cell against LPS-mediated apoptosis in a dose-dependent manner. Furthermore, LPS induced the cytoplasmic calcium overload which was responsible for the upregulation of calcium-dependent xanthine oxidase (XO). Higher XO expression was associated with reactive oxygen species (ROS) overproduction, leading to the Drp1 phosphorylation at the Ser616 site and migration on the surface of mitochondria. Furthermore, phosphorylated Drp1 initiated the mitochondrial fission contributing to the caspase9-dependent mitochondrial apoptosis as evidenced by lower membrane potential, more cyt-c leakage into the nuclear, and higher expression of proapoptotic proteins. However, melatonin treatment could trigger the AMPK pathway, which was followed by the increased SERCA2a expression. Activation of AMPK/SERCA2a by melatonin inhibited the calcium overload, XO-mediated ROS outburst, Drp1-required mitochondrial fission, and final mitochondrial apoptosis. In summary, this study confirmed that LPS induced HUVEC apoptosis through Ca-XO-ROS-Drp1-mitochondrial fission axis and that melatonin reduced the apoptosis of HUVEC through activation of the AMPK/SERCA2a pathway.

Angiotensin II (Ang II) has been reported as key in inducing endothelial cell injury, and endothelial cells may produce nitric oxide (NO) to protect themselves. However, the underlying mechanism remains elusive. Human umbilical vein endothelial cells (HUVECs) were divided into five treatment groups as follows: Normal control, Ang II, Ang II + sodium hydrosulfide [NaHS; hydrogen sulfide (H2S) donor], Ang II + Akt inhibitors + NaHS, and Ang II + endothelial nitric oxide synthases (eNOS) inhibitors + NaHS. Subsequently, cell viability, apoptosis, migration, proliferation and adhesion ability were determined. In addition, tubular structure formation was observed, and the NO and phosphorylation levels of Akt and eNOS were evaluated. Compared with the normal control group, Ang II treatment reduced the viability of HUVECs and increased the level of cell apoptosis (P<0.05). Furthermore, Ang II treatment inhibited the phosphorylation level of eNOS and Akt, as well as the generation of NO (P<0.05). H2S reversed the above‑mentioned effects significantly and increased cell proliferation, adhesion ability and promoted tubular structure formation (P<0.05); however, H2S did not reverse the impact of eNOS and Akt phosphorylation levels after being processed with Akt and eNOS inhibitors, which indicates that H2S is capable of protecting HUVECs via the eNOS/Akt signaling pathway (P<0.05). Thus, H2S stimulates the production of NO and protects HUVECs via inducing the Akt/eNOS signaling pathway.