Purification of high-quality micro RNA from the heart tissue

Faragó, Nóra; Zvara, Ágnes; Varga, Zoltán V.; Ferdinándy, Péter; Puskás, László G.

doi:10.1556/abiol.62.2011.4.7

Cited by 12 publications

(13 citation statements)

References 21 publications

(22 reference statements)

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“…Statistical analysis of microRNA microarrays were done as previously described using Feature Extraction software (Agilent Technologies) (14). All individual microRNAs were represented by 20 different probes on the array.…”

Section: Discussionmentioning

confidence: 99%

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MicroRNAs associated with ischemia-reperfusion injury and cardioprotection by ischemic pre- and postconditioning: protectomiRs

Varga

Zvara

Faragó

et al. 2014

American Journal of Physiology-Heart and Circulatory Physiology

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117

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We aimed to characterize early changes in microRNA expression in acute cardioprotection by ischemic pre- and postconditioning in rat hearts. Hearts isolated from male Wistar rats were subjected to 1) time-matched nonischemic perfusion, 2) ischemia-reperfusion (30 min of coronary occlusion and 120 min of reperfusion), 3) preconditioning (3 × 5 min of coronary occlusion) followed by ischemia-reperfusion, or 4) ischemia-reperfusion with postconditioning (6 × 10 s of global ischemia-reperfusion at the onset of reperfusion). Infarct size was significantly reduced by both interventions. Of 350 different microRNAs assessed by microarray analysis, 147-160 microRNAs showed detectable expression levels. Compared with microRNA alterations induced by ischemia-reperfusion versus time-matched nonischemic controls, five microRNAs were significantly affected by both pre- and postconditioning (microRNA-125b*, microRNA-139-3p, microRNA-320, microRNA-532-3p, and microRNA-188), four microRNAs were significantly affected by preconditioning (microRNA-487b, microRNA-139-5p, microRNA-192, and microRNA-212), and nine microRNAs were significantly affected by postconditioning (microRNA-1, microRNA let-7i, microRNA let-7e, microRNA let-7b, microRNA-181a, microRNA-208, microRNA-328, microRNA-335, and microRNA-503). Expression of randomly selected microRNAs was validated by quantitative real-time PCR. By a systematic comparison of the direction of microRNA expression changes in all groups, we identified microRNAs, specific mimics, or antagomiRs that may have pre- and postconditioning-like cardioprotective effects (protectomiRs). Transfection of selected protectomiRs (mimics of microRNA-139-5p, microRNA-125b*, microRNA let-7b, and inhibitor of microRNA-487b) into cardiac myocytes subjected to simulated ischemia-reperfusion showed a significant cytoprotective effect. This is the first demonstration that the ischemia-reperfusion-induced microRNA expression profile is significantly influenced by both pre- and postconditioning, which shows the involvement of microRNAs in cardioprotective signaling. Moreover, by analysis of microRNA expression patterns in cardioprotection by pre- and postconditioning, specific protectomiRs can be revealed as potential therapeutic tools for the treatment of ischemia-reperfusion injury.

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Section: Discussionmentioning

confidence: 99%

“…MicroRNA isolation was performed according to the protocol for isolation of microRNA from tissues as supplied in the manufacturer's kit (Roche) with some modifications (14). RNA quality and quantity were assessed spectrophotometrically (Nanodrop) with a 2100 Bioanalyzer (Agilent).…”

Section: Microrna Isolationmentioning

confidence: 99%

MicroRNAs associated with ischemia-reperfusion injury and cardioprotection by ischemic pre- and postconditioning: protectomiRs

Varga

Zvara

Faragó

et al. 2014

American Journal of Physiology-Heart and Circulatory Physiology

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117

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“…Impedance based real-time cell electronic sensing (RT-CES) assay was performed with xCELLigence RTCA instrument [24]. Cells were plated in 16-well e-plates at 2000 cells/well density.…”

Section: Methodsmentioning

confidence: 99%

Combination of unsaturated fatty acids and ionizing radiation on human glioma cells: cellular, biochemical and gene expression analysis

Antal

Hackler²,

Shen

et al. 2014

Lipids Health Dis

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BackgroundBased on previous observations a potential resort in the therapy of the particularly radioresistant glioma would be its treatment with unsaturated fatty acids (UFAs) combined with irradiation.MethodsWe evaluated the effect of different UFAs (arachidonic acid (AA), docosahexaenoic acid (DHA), gamma-linolenic acid (GLA), eicosapentaenoic acid (EPA) and oleic acid (OA)) on human U87 MG glioma cell line by classical biochemical end-point assays, impedance-based, real-time cellular and holographic microscopic analysis. We further analyzed AA, DHA, and GLA at morphological, gene and miRNA expression level.ResultsCorresponding to LDH-, MTS assays and real-time cytoxicity profiles AA, DHA, and GLA enhanced the radio sensitivity of glioma cells. The collective application of polyunsaturated fatty acids (PUFAs) and irradiation significantly changed the expression of EGR1, TNF-α, NOTCH1, c-MYC, TP53, HMOX1, AKR1C1, NQO1, while up-regulation of GADD45A, EGR1, GRP78, DDIT3, c-MYC, FOSL1 were recorded both in response to PUFA treatment or irradiation alone. Among the analyzed miRNAs miR-146 and miR-181a were induced by DHA treatment. Overexpression of miR-146 was also detected by combined treatment of GLA and irradiation.ConclusionsBecause PUFAs increased the radio responsiveness of glioma cells as assessed by biochemical and cellular assays, they might increase the therapeutic efficacy of radiation in treatment of gliomas. We demonstrated that treatment with DHA, AA and GLA as adjunct to irradiation up-regulated the expression of oxidative-stress and endoplasmic reticulum stress related genes, and affected NOTCH1 expression, which could explain their additive effects.Electronic supplementary materialThe online version of this article (doi:10.1186/1476-511X-13-142) contains supplementary material, which is available to authorized users.

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“…For loading controls, we used reactions containing 50 ng of the genomic DNA, 0.25 mM of murine-specific tumor necrosis factor-a (TNF-a) gene primers TNF-α-F: 5′-CCTGGAGGAGAAGAGGAAAGAGA-3′ and TNF-α-R: 5′-TTGAGGACCTCTGTGTATTTGTCAA-3′. To determine the intracardiac levels of mir-34a expression, high-purity miRNA was obtained from heart tissue by using standard protocol (Faragó et al, 2011 ). Quantitative real-time PCR analysis were performed using miScript SYBR Green PCR Kit for mouse mir-34a (Quiagen) at the same conditions as described above.…”

Section: Methodsmentioning

confidence: 99%

Circulating Plasma MicroRNA-208a as Potential Biomarker of Chronic Indeterminate Phase of Chagas Disease

et al. 2018

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Chagas cardiomyopathy is the most severe clinical manifestation of chronic Chagas disease. The disease affects most of the Latin American countries, being considered one of the leading causes of morbidity and death in the continent. The pathogenesis of Chagas cardiomyopathy is very complex, with mechanisms involving parasite-dependent cytopathy, immune-mediated myocardial damage and neurogenic disturbances. These pathological changes eventually result in cardiac myocyte hypertrophy, arrhythmias, congestive heart failure and stroke during chronic infection phase. Herein, we show that miR-208a, a microRNA that is a key factor in promoting cardiovascular dysfunction during cardiac hypertrophy processes of heart failure, has its circulating levels increased during chronic indeterminate phase when compared to cardiac (CARD) clinical forms in patients with Chagas disease. In contrast, we have not found altered serum levels of miR-34a, a microRNA known to promote pro-apoptotic role in myocardial infarction during degenerative process of cardiac injuries thus indicating intrinsic differences in the nature of the mechanisms underlying the heart failure triggered by Trypanosoma cruzi infection. Our findings support that the chronic indeterminate phase is a progressive phase involved in the genesis of chagasic cardiopathy and point out the use of plasma levels of miR-208a as candidate biomarker in risk-prediction score for the clinical prognosis of Chagas disease.

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Purification of high-quality micro RNA from the heart tissue

Cited by 12 publications

References 21 publications

MicroRNAs associated with ischemia-reperfusion injury and cardioprotection by ischemic pre- and postconditioning: protectomiRs

MicroRNAs associated with ischemia-reperfusion injury and cardioprotection by ischemic pre- and postconditioning: protectomiRs

Combination of unsaturated fatty acids and ionizing radiation on human glioma cells: cellular, biochemical and gene expression analysis

Circulating Plasma MicroRNA-208a as Potential Biomarker of Chronic Indeterminate Phase of Chagas Disease

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