Myocardial ischemia-reperfusion (I/R) injury is the oxidative stress and inflammatory response that occurs when a tissue is reperfused following a prolonged period of ischemic injury. Growing evidence has demonstrated that microRNAs (miRs) are essential in the development of myocardial I/R injury. Salidroside, a phenylpropanoid glycoside isolated from a traditional Chinese medicinal plant, Rhodiola rosea, possesses multiple pharmacological functions and protects against myocardial I/R injury in vitro and in vivo. However, the role of miRs in the cardioprotective effects of salidroside against myocardial I/R injury has not been studied, to the best of our knowledge. In the present study, the role of miR21 in the underlying mechanism of salidroside-induced protection against oxidative stress and inflammatory injuries in hypoxia/reoxygenation (H/R)-treated H9c2 cardiomyocytes was determined. The cell viability was assessed with an MTT assay. Lactate dehydrogenase (LDH) release, caspase-3 activity, malondialdehyde (MDA) level, superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were determined by commercial kits. Cell apoptosis was measured by flow cytometry. Intracellular reactive oxygen species (ROS) generation was monitored by DCFH-DA. The miR-21 level was quantified by reverse transcription-quantitative (RT-q)PCR. The interleukin (IL)-6, IL-1β and tumor necrosis factor (TNF)-α levels were measured by RT-qPCR and ELISA. The results showed that salidroside pretreatment significantly increased cell viability and decreased the release of LDH, accompanied by an increase in miR-21 expression in H/R-treated H9c2 cells and a miR-21 inhibitor reversed these effects. In addition, the miR-21 inhibitor also abrogated the inhibition of salidroside on H/R-induced increases in apoptosis and caspase-3 activity in H9c2 cells. Salidroside mitigated H/R-induced oxidative stress as illustrated by the downregulation of ROS generation and MDA level and increased the activities of the antioxidant enzymes, SOD and GSH-Px, all of which were abrogated in cells transfected with the miR-21 inhibitor. Salidroside induced a decrease in the expression and levels of the pro-inflammatory cytokines, IL-6, IL-1β and TNF-α, which were prevented by the miR-21 inhibitor. Together, these results provide evidence of the beneficial effects of salidroside against myocardial I/R injury by reducing myocardial oxidative stress and inflammation which are enhanced by increasing miR-21 expression.
Background:Leukemia inhibitory factor (LIF) has been reported to possess various pharmacological effects, including displaying vascular and neuroprotective properties, during retinal disease. The aim of this study was to investigate the vascular and structural changes in the retina of diabetic mice and to explore whether LIF prevents experimental diabetes-induced retinal injury in the early stages.Methods:Diabetes was induced in C57Bl/6J mice with streptozotocin (STZ) injections. Successful diabetic animal models were randomly separated into two groups: the diabetic group (n = 15) and the LIF-treated group (n = 15). Normal C57BL/6 mice served as the normal control group (n = 14). Recombinant human LIF was intravitreally injected 8 weeks after the diabetic model was successfully established. Retinas were collected and evaluated using histological and immunohistochemical techniques, and flat-mounted retinas and Western blotting were performed at 18 weeks after the induction of diabetes and 2 days after the intravitreal injection of LIF. The analysis of variance test were used.Results:Histological analysis showed that there were fewer retinal ganglion cells (RGCs) and the inner nuclear layer (INL) became thinner in the diabetic model group (RGC 21.8 ± 4.0 and INL 120.2 ± 4.6 μm) compared with the normal control group (RGC 29.0 ± 6.7, t = −3.02, P = 0.007; INL 150.7 ± 10.6 μm, t = −8.88, P < 0.001, respectively). After LIF treatment, the number of RGCs (26.9 ± 5.3) was significantly increased (t = 3.39, P = 0.030) and the INL (134.5 ± 14.2 μm) was thicker compared to the diabetic group (t = 2.75, P = 0.013). In the anti-Brn-3a-labeled retinas, the number of RGCs in the LIF-treated group (3926.0 ± 143.9) was obviously increased compared to the diabetic group (3507.7 ± 286.1, t = 2.38, P = 0.030), while no significance was found between the LIF-treated group and the control group (4188.3 ± 114.7, t = −2.47, P = 0.069). Flat-mounted retinas demonstrated that a disorganized, dense distribution of the vessel was prominent in the diabetic model group. Vessel distribution in the LIF-treated mouse group was typical and the thickness was uniform. The levels of phosphosignal transducer and activator of transcription 3 activation were obviously higher in the LIF-injected retinas than those in the diabetic control group (t = 3.85, P = 0.019) and the normal control (t = −3.20, P = 0.019).Conclusion:The present study provides evidence that LIF treatment protects the integrity of the vasculature and prevents retinal injury in the early stages of diabetic retinopathy in STZ-induced diabetic models.
The renin-angiotensin system (RAS) serves an essential role in hypertension. MicroRNAs (miRs) have been reported to be important regulators in angiotensin (Ang) II-dependent hypertension. We aimed to explore the roles of Ang II and miR-133a in the mechanism underlying hypertension. Human umbilical vein endothelial cells (HUVECs) were identified by immunofluorescence staining. Cell viability and miR-133a expression under the inhibition of Ang II of various concentrations were determined by an MTT assay and reverse transcription-quantitative polymerase chain reaction (RT-qPCR), respectively. The effects of HUVECs transfected with miR-133a mimic or inhibitor on Ang II-induced apoptosis were measured using flow cytometry. The potential targeting of miR-133a to the 3′ untranslated region of (pro) renin receptor (PRR) was assessed using TargetScan and a dual-luciferase assay. The effects of PRR interference using small interfering (si)RNA on PRR expression and the rate of apoptosis were determined by RT-qPCR, western blotting and flow cytometry, respectively. Ang II at a concentration of 10 −5 M significantly inhibited the cell viability (P<0.05) and miR-133a expression (P<0.01); Downregulation of miR-133a suppressed cell viability. HUVECs transfected with miR-133a mimic reduced the rate of Ang II-induced apoptosis from 21.99 to 12.38%, but miR-133a inhibitor promoted Ang II-induced apoptosis (apoptosis rate, 28.9%). PRR was predicted to be a target gene of miR-133a. Transfection with siPRR decreased the apoptotic rate in Ang II + negative control and Ang II + miR-133a inhibitor group to 11.39 and 12.94%, respectively. Our findings also suggested that Ang II promoted PRR expression to enhance the apoptotic rate of HUVECs via the suppression of miR-133a. Furthermore, siPRR efficiently decreased the Ang II-induced apoptosis.
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