Silencing MEKK3 attenuates cardiomyocyte injury caused by hypoxia/reoxygenation via the sonic hedgehog pathway

Abstract

MEKK3 is a member of MAP3K, which plays a pivotal role in cardiac diseases. In this study, we aimed to investigate the effects and potential mechanisms of MEKK3 on hypoxia/reoxygenation (H/R) injury of cardiomyocytes. After exposing H9C2 cells to H/R insult, real‐time polymerase chain reaction and western blot analysis showed that MEKK3 was highly expressed. Cell viability, cell apoptosis, caspase 3/7 activity, and cleaved‐caspase 3 expression were tested using a CCK‐8 assay, Cell Death Detection PLUS ELISA, Caspase‐Glo 3/7 Assay Kit and western blot analysis, respectively. Mitochondrial membrane potential, cytochrome C expression, adenosine triphosphate (ATP), and reactive oxygen species also were measured using JC‐1 staining, western blot analysis, an ATP Assay Kit, and DCFH 2‐DA staining, respectively. The messenger RNA (mRNA) levels and secretions of TNF‐α, IL‐6, and IL‐1β were evaluated. The results revealed that MEKK3 silencing promoted cell survival and attenuated lactate dehydrogenase leakage, cell apoptosis, caspase 3/7 activity, and the protein level of cleaved‐caspase 3. Moreover, knockdown of MEKK3 blocked mitochondrial impairment by inhibiting the loss of mitochondrial membrane potential and cytochrome C expression as well as promoting ATP synthesis. MEKK3 deficiency led to a decrease in reactive oxygen species and malondialdehyde (MDA) generation and an increase in superoxide dismutase (SOD) activity. Deletion of MEKK3 led to reduced inflammatory cytokines in mRNA level and secretion. MEKK3 suppression activated the sonic hedgehog (Shh) signaling pathway in H9C2 cells. After blocking the Shh signaling pathway with a specific inhibitor, cyclopamine, the cardioprotective functions of MEKK3 downregulation were partly abolished. In conclusion, downregulation of MEKK3 prevented apoptosis and inflammation in H9C2 cells via the Shh signaling pathway.

RETRACTED: MicroRNA-377 Alleviates Myocardial Injury Induced by Hypoxia/Reoxygenation via Downregulating LILRB2 Expression

“…5 It has indicated that ischemia-induced hypoxia and reoxygenation (H/R) were related to apoptosis, production of reactive oxygen species, inflammation, and so on. 6,7 However, the mechanism of myocardial ischemia remains ill-defined. Therefore, it is important to figure out potential mechanisms of action to develop novel therapeutic regimen for myocardial ischemia injury.…”

Alantolactone suppresses inflammation, apoptosis and oxidative stress in cigarette smoke-induced human bronchial epithelial cells through activation of Nrf2/HO-1 and inhibition of the NF-κB pathways

“…Beas-2B and NHBE cells were plated into 96-well plates and pretreated for 2 h with 1, 5 and 10 μM ALT and then treated with 5% CSE for 24 h. Caspase-3 activity was measured by using caspase-Glo 3/7 Assay (Promega, Madison, WI, USA) kit in accordance with a previous report and the manufacturer's instructions [23]. Capase-Glo 3/7 Reagent (100 μL) was added to cells for another 2 h at room temperature and the luminescence intensity was recorded at 570 nm by using a microplate reader.…”

miR‐181b regulates vascular endothelial aging by modulating an MAP3K3 signaling pathway

“…MAP3K3 knockdown blocked mitochondrial impairment by inhibiting the loss of mitochondrial membrane potential and cytochrome C expression as well as promoting ATP synthesis and reduced apoptosis and mitochondrial damage in cardiomyocytes in response to hypoxia/reoxygenation (H/R). 34 MAP3K3 deficiency also increased the life span in Caenorhabditis elegans and decreased the levels of ROS. 71 In the current study, we also found that MAP3K3 knockdown reduced apoptosis and senescence markers in ECs responding to H 2 O 2 through the MAPK signaling pathway (Figure 6A-D).…”

“…31,32 Some studies demonstrate key effects of MAP3K3 in apoptosis and senescence in diverse cell types. 14,[33][34][35][36] In this study, we identify that miR-181b regulates endothelial aging by modulating the MAPK signaling pathway. We explore the mechanistic basis for miR-181b regulation of apoptosis and senescence in ECs that is dependent on a MAP3K3/MAPK signaling cascade.…”

“…MAP3K3 (mitogen‐activated protein kinase kinase kinase 3), a key upstream regulator in the MAPK cascade, directly modulates the phosphorylation of MKK3/MKK6 or MKK4, which then regulates substrates, for example, p38, JNK 31,32 . Some studies demonstrate key effects of MAP3K3 in apoptosis and senescence in diverse cell types 14,33–36 …”