Protective Effects of Hydrogen Sulfide in Hypoxic Human Umbilical Vein Endothelial Cells: A Possible Mitochondria-Dependent Pathway

Abstract

The aim of the study was to investigate the protective effects of sodium hydrosulfide (NaHS), a H2S donor, against hypoxia-induced injury in human umbilical vein endothelial cells (HUVECs) and also to look into the possible mechanisms by which H2S exerts this protective effect. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and scratch wound healing assay were chosen to measure the cell viability and migration-promoting effects. The fluorescent probe, DCFH-DA and 5,5′,6,6′-Tetrachloro-1,1′,3,3′-tetraethyl-imidacarbocyanine iodide (JC-1) were applied to detect the reactive oxygen species (ROS) level and mitochondrial membrane potential (ΔΨm). Furthermore, western blots were used to measure the expressions of the apoptosis-related proteins. Under hypoxic conditions, 300 μM and 600 μM of H2S could protect HUVECs against hypoxia-induced injury, as determined by MTT assay. Following the treatment of 60 μM NaHS for 18 h, scratch wound healing assays indicated that the scratch became much narrower than control group. After treatment with 60 μM, 120 μM, and 600 μM NaHS, and hypoxia for 30 min, flow cytometry demonstrated that the ROS concentrations decreased to 95.08% ± 5.52%, 73.14% ± 3.36%, and 73.51% ± 3.05%, respectively, compared with the control group. In addition, the JC-1 assay showed NaHS had a protective effect on mitochondria damage. Additionally, NaHS increased Bcl-2 expression and decreased the expression of Bax, Caspase-3 and Caspase-9 in a dose-dependent way. Our results suggest that H2S can protect endothelial cells and promote migration under hypoxic condition in HUVECs. These effects are partially associated with the preservation of mitochondrial function mediated by regulating the mitochondrial-dependent apoptotic pathway.

Plasma-activated medium induces A549 cell injury via a spiral apoptotic cascade involving the mitochondrial–nuclear network

“…3c and d). The fluorescence characteristics of JC-1 changed in accordance with ǻȥm; green fluorescence indicated a decreased membrane potential in injured cells while red fluorescence reflected a normal membrane potential [26]. As shown in Figure 4 …”

ROS-Dependent Neuroprotective Effects of NaHS in Ischemia Brain Injury Involves the PARP/AIF Pathway

“…Interestingly, H 2 S has been demonstrated to protect cells from apoptosis by removing cellular ROS [24], which prompted us to examine the ROS status in the presence or absence of NaHS. HT22 cells were pretreated with OGD and followed by treatment with increasing dosage of NaHS.…”

Hypoxia and hydrogen sulfide differentially affect normal and tumor-derived vascular endothelium

“…Moreover, evidence provided in this work indicates that H 2 S is not protective against hypoxia and oxidative stress for normal and altered vascular endothelium, differently from its well-known role on the heart [25], [26].…”