BackgroundPostconditioning (PostC) inhibits myocardial apoptosis after ischemia-reperfusion (I/R) injury. The JAK2-STAT3 pathway has anti-apoptotic effects and plays an essential role in the late protection of preconditioning. Our aim was to investigate the anti-apoptotic effect of PostC after prolonged reperfusion and the role of the JAK2-STAT3 pathway in the anti-apoptotic effect of PostC.MethodsWistar rats were subjected to 30 minutes ischemia and 2 or 24 hours (h) reperfusion, with or without PostC (three cycles of 10 seconds reperfusion and 10 seconds reocclusion at the onset of reperfusion). Separate groups of rats were treated with a JAK2 inhibitor (AG490) or a PI3K inhibitor (wortmannin) 5 minutes before PostC. Immunohistochemistry was used to analyze Bcl-2 protein levels after reperfusion. mRNA levels of Bcl-2 were detected by qRT-PCR. TTC staining was used to detect myocardial infarction size. Myocardial apoptosis was evaluated by TUNEL staining. Western-blot was used to detect p-STAT3 and p-Akt levels after reperfusion.ResultsThere was more myocardial apoptosis at 24 h vs 2 h after reperfusion in all groups. PostC significantly reduced myocardial apoptosis and elevated Bcl-2 levels at both 2 and 24 hours after reperfusion. PostC increased p-STAT3 and p-Akt levels after reperfusion. Administration of AG490 reduced p-STAT3 and p-Akt levels and attenuated the anti-apoptotic effect of PostC. Wortmannin also reduced p-Akt levels and attenuated the anti-apoptotic effect of PostC but had no effect on p-STAT3 levels. AG490 abrogated the up-regulation of Bcl-2 by PostC.ConclusionPostC may reduce myocardial apoptosis during prolonged reperfusion via a JAK2-STAT3-Bcl-2 pathway. As a downstream target of JAK2 signaling, activation of PI3K/Akt pathway may be necessary in the protection of PostC.

Atherosclerosis preferentially occurs in atheroprone vasculature where human umbilical vein endothelial cells (HUVECs) are exposed to disturbed flow. Disturbed flow is associated with vascular inflammation and focal distribution. Recent studies have revealed the involvement of epigenetic regulation in atherosclerosis progression. N6-methyladenosine (m6A) is the most prevalent internal modification of eukaryotic mRNA, but its function in endothelial atherogenic progression remains unclear. Here, we show that m6A mediates the EGFR signaling pathway during EC activation to regulate the atherosclerotic process. Oscillatory stress (OS) reduced the expression of METTL3, the primary m6A methyltransferase. Through m6A sequencing and functional studies, we determined that m6A mediates the mRNA decay of the vascular pathophysiology gene EGFR which leads to EC dysfunction. m6A modification of the EGFR 3'UTR accelerated its mRNA degradation. Double mutation of the EGFR 3'UTR abolished METTL3-induced luciferase activity. Adenovirus-mediated METTL3 overexpression significantly reduced EGFR activation and endothelial dysfunction in the presence of OS. Furthermore, TSP-1, an EGFR ligand, was specifically expressed in atheroprone regions without being affected by METTL3. Inhibition of the TSP-1/EGFR axis by using shRNA and AG1478 significantly ameliorated atherogenesis. Overall, our study revealed that METTL3 alleviates endothelial atherogenic progression through m6A-dependent stabilization of EGFR mRNA, highlighting the important role of RNA transcriptomics in atherosclerosis regulation.

Departmental sources Background: Doxorubicin-induced myocardial toxicity is associated with oxidative stress, cardiomyocyte, apoptosis, and loss of contractile function. Previous studies showed that microRNA-375 (miR-375) expression was increased in mouse models of heart failure and clinically, and that inhibition of miR-375 reduced inflammation and increased survival of cardiomyocytes. This study aimed to investigate the effects and mechanisms of inhibition of miR-375 in a mouse model of doxorubicin-induced cardiac toxicity in vivo and in doxorubicin-treated rat and mouse cardiomyocytes in vitro. Material/Methods: The mouse model of doxorubicin-induced cardiac toxicity was developed using an intraperitoneal injection of doxorubicin (15 mg/kg diluted in 0.9% saline) for eight days. Treatment was followed by a single subcutaneous injection of miR-375 inhibitor. H9c2 rat cardiac myocytes and adult murine cardiomyocytes (AMCs) were cultured in vitro and treated with doxorubicin, with and without pretreatment with miR-375 inhibitor. Results: Doxorubicin significantly upregulated miR-375 expression in vitro and in vivo, and inhibition of miR-375 reestablished myocardial redox homeostasis, prevented doxorubicin-induced oxidative stress and cardiomyocyte apoptosis, and activated the PDK1/AKT axis by reducing the direct binding of miR-375 to 3' UTR of the PDK1 gene. Inhibition of PDK1 and AKT abolished the protective role of miR-375 inhibition on doxorubicin-induced oxidative damage. Conclusions: Inhibition of miR-375 prevented oxidative damage in a mouse model of doxorubicin-induced cardiac toxicity in vivo and in doxorubicin-treated rat and mouse cardiomyocytes in vitro through the PDK1/AKT signaling pathway.

BackgroundTo study the influencing factors for different healing patterns of patients with idiopathic macular holes (IMH) after vitrectomy surgery performed with the internal limiting membrane (ILM) flap technique.MethodsThis study was a retrospective, consecutive, observational case series study. We recruited 52 IMH patients who underwent vitrectomy with the ILM flap technique. The participants were divided into 2 groups: group A (25 patients), without significant glial cell proliferation in the macular area on postoperative optical coherence tomography (OCT); and group B (27 patients), with significant glial cell proliferation. The postoperative visual acuity (VA), external limiting membrane (ELM) and ellipsoid zone (EZ) recovery characteristics were compared between the two groups.ResultsThere were statistically significant differences in minimum linear diameter (MLD) of the macular hole and postoperative VA (p = 0.02, 2.81 E-4 respectively) between the two groups. Compared with patients in group A, patients in group B had poorer VA and EZ recovery in the first 12 months after surgery, and a longer ELM recovery period. The OCT results showed that patients in group B had more extensive ILM filling in the macular area after surgery than patients in group A.ConclusionThe presence of aberrant glial cell proliferation was related to a larger MLD of the IMH, and the filling approach for the ILM during the operation was related to the postoperative healing pattern and vision acuity.

Background and aim: Adenosine A2B receptor (A2BAR) agonist reduces myocardial reperfusion injury by acting on inflammatory cells. Recently, a cardiosplenic axis was shown to mediate the myocardial post-ischemic reperfusion injury. This study aimed to explore whether the infarct-squaring effect of A2BAR agonist was primarily due to its action on splenic leukocytes. Methods: C57BL6 (wild-type, WT) mice underwent 40 min of left coronary artery occlusion followed by 60 min of reperfusion. A2BR knockout (KO) and interleukin (IL)- 10KO mice served as donors for splenic leukocytes. Acute splenectomy was performed 30 min prior to ischemia. The acute splenic-leukocyte adoptive transfer was performed by injecting 5 × 106 live splenic leukocytes into splenectomized mice. BAY60–6583, an A2BAR agonist, was injected by i.v. 15 min before ischemia. The infarct size (IS) was determined using TTC and Phthalo blue staining. The expression of p-Akt and IL-10 was estimated by Western blotting. Immunofluorescence staining assessed the localization of IL-10 expression. Results: BAY 60–6583 reduced the myocardial IS in intact mice but failed to reduce the same in splenectomized mice, which had a smaller IS than intact mice. BAY 60–6583 reduced the IS in splenectomized mice with the acute transfer of WT splenic leukocytes; however, it did not protect the heart of splenectomized mice with the acute transfer of A2bRKO splenic leukocytes. Furthermore, BAY 60–6583 increased the levels of p-Akt and IL-10 in WT spleen. Moreover, it did not exert any protective effect in IL-10KO mice. Conclusion: A2BAR activation prior to ischemia stimulated the IL-10 production in splenic leukocytes via a PI3K/Akt pathway, thereby exerting anti-inflammatory effects that limited the myocardial reperfusion injury.