[Tanshinone IIA alleviates monocrotaline-induced pulmonary hypertension in rats through the PI3K/Akt-eNOS signaling pathway].

OBJECTIVE: To explore the therapeutic mechanism of tanshinone IIA in the treatment of pulmonary arterial hypertension (PAH) in rats. METHODS: A total of 100 male SD rats were randomized into 5 groups (n=20), and except for those in the control group with saline injection, all the rats were injected with monocrotaline (MCT) on the back of the neck to establish models of pulmonary hypertension. Two weeks after the injection, the rat models received intraperitoneal injections of tanshinone IIA (10 mg/kg), phosphatidylinositol 3 kinase (PI3K) inhibitor (1 mg/kg), both tanshinone IIA and PI3K inhibitor, or saline (model group) on a daily basis. After 2 weeks of treatment, HE staining and α-SMA immunofluorescence staining were used to evaluate the morphology of the pulmonary vessels of the rats. The phosphorylation levels of PI3K, protein kinase B (PKB/Akt) and endothelial nitric oxide synthase (eNOS) in the lung tissue were determined with Western blotting; the levels of eNOS and NO were measured using enzyme-linked immunosorbent assay (ELISA). RESULTS: The results of HE staining and α-SMA immunofluorescence staining showed that tanshinone IIA effectively inhibited MCT-induced pulmonary artery intimamedia thickening and muscularization of the pulmonary arterioles (P < 0.01). The results of Western blotting showed that treatment with tanshinone IIA significantly increased the phosphorylation levels of PI3K, Akt and eNOS proteins in the lung tissue of PAH rats; ELISA results showed that the levels of eNOS and NO were significantly decreased in the rat models after tanshinone IIA treatment (P < 0.01). CONCLUSION: Treatment with tanshinone IIA can improve MCT-induced pulmonary hypertension in rats through the PI3K/Akt-eNOS signaling pathway.

Intelligent Nanodelivery System-Generated 1 O2 Mediates Tumor Vessel Normalization by Activating Endothelial TRPV4-eNOS Signaling.

Tumor microenvironment (TME)-responsive intelligent photodynamic therapy (PDT) systems have attracted increasing interest in anticancer therapy, due to their potential to address significant and unsatisfactory therapeutic issues, such as limited tissue penetration, inevitable normal tissue damage, and excessive impaired vessels. Here, an H2 O2 -triggered intelligent LCL/ZnO PDT nanodelivery system is elaborately designed. LCL/ZnO can selectively regulate tumor-derived endothelial cells (TECs) and specifically kill tumor cells, by responding to different H2 O2 gradients in TECs and tumor cells. The LCL/ZnO is able to normalize tumor vessels, thereby resulting in decreased metastases, and ameliorating the immunosuppressive TME. Further analysis demonstrates that singlet oxygen (1 O2 )-activated transient receptor potential vanilloid-4-endothelial nitric oxide synthase signals generated in TECs by LCL/ZnO induce tumor vascular normalization, which is identified as a novel mechanism contributing to the increased ability of PDT to promote cancer therapy. In conclusion, designing an intelligent PDT nanodelivery system response to the TME, that includes both selective TECs regulation and specific tumor-killing, will facilitate the development of effective interventions for future clinical applications.

Effect of miR-134 against myocardial hypoxia/reoxygenation injury by directly targeting NOS3 and regulating PI3K/Akt pathway.

PURPOSE: To reveal the function of miR-134 in myocardial ischemia. METHODS: Real-time PCR and western blotting were performed to measure the expression of miR-134, nitric oxide synthase 3 (NOS3) and apoptotic-associated proteins. Lactic dehydrogenase (LDH) assay, cell counting kit-8 (CCK-8), Hoechst 33342/PI double staining and flow cytometry assay were implemented in H9c2 cells, respectively. MiR-134 mimic/inhibitor was used to regulate miR-134 expression. Bioinformatic analysis and luciferase reporter assay were utilized to identify the interrelation between miR-134 and NOS3. Rescue experiments exhibited the role of NOS3. The involvement of PI3K/AKT was assessed by western blot analysis. RESULTS: MiR-134 was high regulated in the myocardial ischemia model, and miR-134 mimic/inhibitor transfection accelerated/impaired the speed of cell apoptosis and attenuated/exerted the cell proliferative prosperity induced by H/R regulating active status of PI3K/AKT signaling. LDH activity was also changed due to the different treatments. Moreover, miR-134 could target NOS3 directly and simultaneously attenuated the expression of NOS3. Co-transfection miR-134 inhibitor and pcDNA3.1-NOS3 highlighted the inhibitory effects of miR-134 on myocardial H/R injury. CONCLUSION: This present work puts insights into the crucial effects of the miR-134/NOS3 axis in myocardial H/R injury, delivering a potential therapeutic technology in future.

Effect of miR-134 against myocardial hypoxia/reoxygenation injury by directly targeting NOS3 and regulating PI3K/Akt pathway

Abstract Purpose To reveal the function of miR-134 in myocardial ischemia. Methods Real-time PCR and western blotting were performed to measure the expression of miR-134, nitric oxide synthase 3 (NOS3) and apoptotic-associated proteins. Lactic dehydrogenase (LDH) assay, cell counting kit-8 (CCK-8), Hoechst 33342/PI double staining and flow cytometry assay were implemented in H9c2 cells, respectively. MiR-134 mimic/inhibitor was used to regulate miR-134 expression. Bioinformatic analysis and luciferase reporter assay were utilized to identify the interrelation between miR-134 and NOS3. Rescue experiments exhibited the role of NOS3. The involvement of PI3K/AKT was assessed by western blot analysis. Results MiR-134 was high regulated in the myocardial ischemia model, and miR-134 mimic/inhibitor transfection accelerated/impaired the speed of cell apoptosis and attenuated/exerted the cell proliferative prosperity induced by H/R regulating active status of PI3K/AKT signaling. LDH activity was also changed due to the different treatments. Moreover, miR-134 could target NOS3 directly and simultaneously attenuated the expression of NOS3. Co-transfection miR-134 inhibitor and pcDNA3.1-NOS3 highlighted the inhibitory effects of miR-134 on myocardial H/R injury. Conclusion This present work puts insights into the crucial effects of the miR-134/NOS3 axis in myocardial H/R injury, delivering a potential therapeutic technology in future.

Inhibition of MAPK signaling by eNOS gene transfer improves ventricular remodeling after myocardial infarction through reduction of inflammation.

Endothelial nitric oxide synthase (eNOS) and nitric oxide (NO) may play an important role in attenuating cardiac remodeling and apoptosis after myocardial infarction. However, the anti-inflammation effects of eNOS in infarcted myocardium and the role of MAPK signaling in eNOS/NO mediated cardiac remodeling have not yet been elucidated. Adenovirus carrying Human eNOS gene was delivered locally into heart 4 days prior to induction of myocardial infarction (MI) by left anterior descending coronary artery ligation. Monocyte/macrophage infiltration was detected by ED-1 immunohistochemistry. Western blot was employed to examine the activation of MAPK. eNOS gene transfer significantly reduced myocardial infarct size and improved cardiac contractility as well as left ventricle (LV) diastolic function at 7 days after MI. In addition, eNOS gene transfer decreased monocyte/macrophage infiltration in the infarct region of the heart. Phosphorylation of MAPK after MI were also dramatically reduced by eNOS gene transfer. All the protective effects of eNOS were blocked by N(ω)-nitro-L-arginine methyl ester (L-NAME) administration, indicating a NO-mediated event. These results demonstrate that the eNOS/NO system provides cardiac protection after MI injury through inhibition of inflammation and suppression of MAPK signaling.

Clinicopathological evaluation of immunohistochemical Ki-67 and endothelial nitric oxide synthase expression in intracranial ependymoma.

PURPOSE: To analyze the association between Ki-67 and eNOS expression with the pathological grades of patients with intracranial ependymomas, and to determine its value in distinguishing the progression of the disease. METHODS: A clinicopathological study was undertaken in 82 patients with intracranial ependymomas. Tissue samples, obtained by tumour resection, were divided into three groups: low-grade, mid-grade and high-grade ependymomas. Tissue samples obtained from 15 patients with brain contusion were used as control. Immuno-histochemical staining was performed to analyze the association between Ki-67 and eNOS expression with various tumour grades. The cell proliferating marker Ki-67 was assessed by positive cell count. The levels of eNOS positive expression were evaluated as slight, moderate and intense. RESULTS: 48 of 82 cases (58.54%) expressed Ki-67 protein. Expression of Ki-67 and eNOS was negative in all control samples. Positive cell rates were 2.65+/-0.83 % in the low-grade, 9.63+/-0.08 % in the mid-grade, and 28.41+/-0.71 % in the high-grade ependymoma groups. In low-grade ependymomas there were 8 and 12 cases that expressed eNOS slightly or moderately. In the mid-grade ependymoma group eNOS was expressed moderately in 10 cases and intensely in 15. In the high-grade group 20 cases showed intense positive expression of eNOS. The Ki-67 positive cell counts for slight, moderate and intense eNOS expression were 2.20, 6.07 and 22.25, respectively. CONCLUSION: Ki-67 and eNOS expression in intracranial ependymoma tissue was associated with the histopathological grade and malignant degree.