Trimetazidine Preconditioning Potentiates the Effect of Mesenchymal Stem Cells Secretome on the Preservation of Rat Pancreatic Islet Survival and Function In Vitro.

Islet transplantation offers improved glycemic control in individuals with type 1 diabetes mellitus. However, in vitro islet culture is associated with islet apoptosis and eventually will lose their functionality prior to transplantation. In this study, we examined the effects of mesenchymal stem cells (MSCs) secretome preconditioned with diazoxide (DZ) and trimetazidine (TMZ) on rat islet cells during pre-transplant culture. With and without preconditioned hAD-MSCs' concentrated conditioned media (CCM) were added to the culture medium containing rat islets every 12 h for 24 and 48 h, after testing for selected cytokine concentrations (interleukin (IL)-4, IL-6, IL-13). Insulin content, glucose-stimulated insulin secretion, islet cell apoptosis, and mRNA expression of pro-apoptotic (BAX, BAK-1, and PUMA) and anti-apoptotic factors (BCL-2, BCL-xL, and XIAP) in rat islets were assessed after 24 and 48 h of culture. The protein level of IL-6 and IL-4 was significantly higher in TMZ-MSC-CM compared to MSC-non-CM. In rat isolated islets, normalized secreted insulin in the presence of 16.7 mM glucose was significantly higher in treated islet groups compared to control islets at both 24 and 48 h cultivation. Also, the percentage of apoptotic islet cells TMZ-MSC-CCM-treated islets was significantly lower compared to MSC-CM and MSC-CCM-treated islets in both 24 and 48 h cultivation. Consistent with the number of apoptotic cells, after 24 h culture, the expression of BCL-2 and BCL-xL genes in the control islets was lower than all treatment islet groups and in 48 h was lower than only TMZ-MSC-CM-treated islets. Also, the expression of the XIAP gene in control islets was significantly lower compared to the TMZ-MSC-CCM-treated islets at both at 24 and 48 h. In addition, mRNA level of the BAX gene in TMZ-MSC-CCM-treated islets was significantly lower compared to other groups at 48 h. Our findings revealed that TMZ proved to be more effective than DZ and could enhance the potential of hAD-MSCs-CM to improve the function and viability of islets prior to transplantation.

Fatty acid oxidation fuels agonist-induced platelet activation and thrombus formation: Targeting ß-oxidation of fatty acids as an effective anti-platelet strategy.

Platelet mitochondria possess remarkable plasticity for oxidation of energy substrates, where metabolic dependency on glucose or fatty acids is higher than glutamine. Since platelets metabolize nearly the entire pool of glucose to lactate rather than fluxing through mitochondrial tricarboxylic acid cycle, we posit that majority of mitochondrial ATP, which is essential for platelet granule secretion and thrombus formation, is sourced from oxidation of fatty acids. We performed a comprehensive analysis of bioenergetics and function of stimulated platelets in the presence of etomoxir, trimetazidine and oxfenicine, three pharmacologically distinct inhibitors of ß-oxidation. Each of them significantly impaired oxidative phosphorylation in unstimulated as well as thrombin-stimulated platelets leading to a small but consistent drop in ATP level in activated cells due to a lack of compensation from glycolytic ATP. Trimetazidine and oxfenicine attenuated platelet aggregation, P-selectin externalization and integrin αIIb ß3 activation. Both etomoxir and trimetazidine impeded agonist-induced dense granule release and platelet thrombus formation on collagen under arterial shear. The effect of inhibitors on platelet aggregation and dense granule release was dose- and incubation time- dependent with significant inhibition at higher doses and prolonged incubation times. Neither of the inhibitors could protect mice from collagen-epinephrine-induced pulmonary embolism or prolong mouse tail bleeding times. However, mice pre-administered with etomoxir, trimetazidine and oxfenicine were protected from ferric chloride-induced mesenteric thrombosis. In conclusion, ß-oxidation of fatty acids sustains ATP level in stimulated platelets and is therefore essential for energy-intensive agonist-induced platelet responses. Thus, fatty acid oxidation may constitute an attractive therapeutic target for novel antiplatelet agents.

Trimetazidine mitigates high glucose-induced retinal endothelial dysfunction by inhibiting PI3K/Akt/mTOR pathway-mediated autophagy.

Trimetazidine (TMZ), as a metabolic regulator, has been widely testified to exhibit positive therapeutic effects on various disease models, but its role in diabetic retinopathy has not been reported. Therefore, this study was designed with the purpose of exploring the effects of TMZ on high-glucose (HG)-induced retinal endothelial dysfunction and its underlying mechanism. To establish DR model in vitro, 30 mM glucose was applied to induce human retinal endothelial cells (HRECs). Cell proliferation, invasion, and migration were examined by means of Cell Counting Kit-8, transwell, and wound healing assays, respectively. The tubule formation experiment was used to test the tubulogenesis ability and fluorescein isothiocyanate (FITC)-albumin was utilized to measure the permeability of monolayer HRECs. In addition, immunofluorescence and Western blot were employed to detect protein expression. Compared with the HG-induced group, TMZ concentration dependently inhibited the proliferation, migration, and angiogenesis of HG-induced HRECs, decreased the permeability of monolayer HRECs, and increased the protein expression levels of Claudin-5 and VE-cadherin. In addition, TMZ intervention increased the expression of p-PI3K, p-AKT, and p-mTOR but decreased the expression of LC3I, LC3II, and Beclin 1, which were then partially reversed by P13 K inhibitor (LY294002). Moreover, the autophagy agonist rapamycin (RAPA) was also testified to reverse the inhibitory effects of TMZ on the proliferation, migration, and angiogenesis of HG-induced HRECs. In summary, TMZ inhibited excessive autophagy by activating PI3K/Akt/mTOR pathway, thereby improving retinal endothelial dysfunction induced by HG.

Trimetazidine enhances myocardial angiogenesis in pressure overload-induced cardiac hypertrophy mice through directly activating Akt and promoting the binding of HSF1 to VEGF-A promoter.

Latest clinical research shows that trimetazidine therapy during the perioperative period relieves endothelial dysfunction in patients with unstable angina induced by percutaneous coronary intervention. In this study we investigated the effects of TMZ on myocardial angiogenesis in pressure overload-induced cardiac hypertrophy mice. Cardiac hypertrophy was induced in mice by transverse aortic constriction (TAC) surgery. TAC mice were administered trimetazidine (2.8 mg/100 µL, i.g.) for 28 consecutive days. We showed that trimetazidine administration significantly increased blood vessel density in the left ventricular myocardium and abrogated cardiac dysfunction in TAC mice. Co-administration of a specific HSF1 inhibitor KRIBB11 (1.25 mg/100 µL, i.h.) abrogated the angiogenesis-promoting effects of trimetazidine in TAC mice. Using luciferase reporter and electrophoretic mobility shift assays we demonstrated that the transcription factor HSF1 bound to the promoter region of VEGF-A, and the transcriptional activity of HSF1 was enhanced upon trimetazidine treatment. In molecular docking analysis we found that trimetazidine directly bound to Akt via a hydrogen bond with Asp292 and a pi-pi bond with Trp80. In norepinephrine-treated HUVECs, we showed that trimetazidine significantly increased the phosphorylation of Akt and the synergistic nuclear translocation of Akt and HSF1, as well as the binding of Akt and HSF1 in the nucleus. These results suggest that trimetazidine enhances myocardial angiogenesis through a direct interaction with Akt and promotion of nuclear translocation of HSF1, and that trimetazidine may be used for the treatment of myocardial angiogenic disorders in hypertensive patients.