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Profiling of cell stress protein expression in cardiac tissue of cardiosurgical patients undergoing remote ischemic preconditioning: implications for thioredoxin in cardioprotection.

Zitta K, Meybohm P, Gruenewald M, Cremer J, Zacharowski KD, Scholz J, Steinfath M, Albrecht M - J Transl Med (2015)

Bottom Line: Array results for thioredoxin were verified by semi-quantitative Westernblotting studies which showed a significant up-regulation of thioredoxin protein levels in cardiac tissue samples of RIPC patients taken before CPB (RIPC: 5.36 ± 0.85 a.u.; control: 3.23 ± 0.39 a.u.; P < 0.05).Quantification of thioredoxin levels in tissue of RIPC and control patients by ELISA experiments further confirmed the Westernblotting results (RIPC: 0.30 ± 0.02 ng/mg protein; control: 0.24 ± 0.02 ng/mg protein; P < 0.05).We provide evidence for thioredoxin as a RIPC-induced factor in heart tissue of cardiosurgical patients and identified several cell stress associated proteins that are regulated by RIPC and may play a role in RIPC-mediated cardioprotection.

View Article: PubMed Central - PubMed

Affiliation: Department of Anaesthesiology and Intensive Care Medicine, University Hospital Schleswig-Holstein, Schwanenweg 21, 24105, Kiel, Germany. karina.zitta@uksh.de.

ABSTRACT

Background: Transient episodes of ischemia in a remote organ (remote ischemic preconditioning, RIPC) can attenuate myocardial ischemia/reperfusion injury but the underlying mechanisms of RIPC in the target organ are still poorly understood. Recent animal studies suggested that the small redox protein thioredoxin may be a potential candidate for preconditioning-induced organprotection. Here we employed a human proteome profiler array to investigate the RIPC regulated expression of cell stress proteins and particularly of thioredoxin in heart tissue of cardiosurgical patients with cardiopulmonary bypass (CPB).

Methods: RIPC was induced by four 5 minute cycles of transient upper limb ischemia/reperfusion using a blood pressure cuff. Right atrial tissue was obtained from patients receiving RIPC (N = 19) and control patients (N = 19) before and after CPB. Cell stress proteome profiler arrays as well as Westernblotting and ELISA experiments for thioredoxin (Thio-1) were performed employing the respective tissue samples.

Results: Protein arrays revealed an up-regulation of 26.9% (7/26; CA IX, Cyt C, HSP-60, HSP-70, pJNK, SOD2, Thio-1) of cell stress associated proteins in RIPC tissue obtained before CPB, while 3.8% (1/26; SIRT2) of the proteins were down-regulated. Array results for thioredoxin were verified by semi-quantitative Westernblotting studies which showed a significant up-regulation of thioredoxin protein levels in cardiac tissue samples of RIPC patients taken before CPB (RIPC: 5.36 ± 0.85 a.u.; control: 3.23 ± 0.39 a.u.; P < 0.05). Quantification of thioredoxin levels in tissue of RIPC and control patients by ELISA experiments further confirmed the Westernblotting results (RIPC: 0.30 ± 0.02 ng/mg protein; control: 0.24 ± 0.02 ng/mg protein; P < 0.05).

Conclusion: We provide evidence for thioredoxin as a RIPC-induced factor in heart tissue of cardiosurgical patients and identified several cell stress associated proteins that are regulated by RIPC and may play a role in RIPC-mediated cardioprotection.

No MeSH data available.


Related in: MedlinePlus

Semi-quantitative evaluation of thioredoxin protein expression. A: Representative Westernblotting experiment performed with cardiac tissue samples of 3 control and 3 RIPC patients. B: Evaluation of the relative protein expression levels of thioredoxin in control and RIPC patients. Numbers in the columns display the numbers of patients employed in the respective experiment. MW, molecular weight; kDa, kiloDalton; a.u., arbitrary units; columns display the mean; bars denote SEM; *, P < 0.05, two way ANOVA with Bonferroni post-test.
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Fig4: Semi-quantitative evaluation of thioredoxin protein expression. A: Representative Westernblotting experiment performed with cardiac tissue samples of 3 control and 3 RIPC patients. B: Evaluation of the relative protein expression levels of thioredoxin in control and RIPC patients. Numbers in the columns display the numbers of patients employed in the respective experiment. MW, molecular weight; kDa, kiloDalton; a.u., arbitrary units; columns display the mean; bars denote SEM; *, P < 0.05, two way ANOVA with Bonferroni post-test.

Mentions: One of the proteins that showed a strong expression within the cardiac tissue and that was also regulated by RIPC is thioredoxin, a small redox protein and at least in animal studies, a possible candidate for cardioprotection [32,33,37,38]. Based on our results of the human proteome profiling approach which was performed with pooled tissue samples of the respective patients, we decided to further focus on thioredoxin as a potential molecule conferring RIPC-mediated cardioprotection and investigated the expression of thioredoxin in each patient separately. Westernblotting results mainly confirmed the outcome of the protein arrays, showing an increased expression of thioredoxin in cardiac tissue of RIPC patients that was obtained before CPB (RIPC: 5.36 ± 0.85 a.u.; control: 3.23 ± 0.39 a.u.; P < 0.05, Figure 4), while no statistically significant differences in thioredoxin protein expression levels were detected in samples taken after CPB (RIPC, 3.94 ± 0.73 a.u.; control, 2.79 ± 0.31 a.u.; P > 0.05; Figure 4).Figure 4


Profiling of cell stress protein expression in cardiac tissue of cardiosurgical patients undergoing remote ischemic preconditioning: implications for thioredoxin in cardioprotection.

Zitta K, Meybohm P, Gruenewald M, Cremer J, Zacharowski KD, Scholz J, Steinfath M, Albrecht M - J Transl Med (2015)

Semi-quantitative evaluation of thioredoxin protein expression. A: Representative Westernblotting experiment performed with cardiac tissue samples of 3 control and 3 RIPC patients. B: Evaluation of the relative protein expression levels of thioredoxin in control and RIPC patients. Numbers in the columns display the numbers of patients employed in the respective experiment. MW, molecular weight; kDa, kiloDalton; a.u., arbitrary units; columns display the mean; bars denote SEM; *, P < 0.05, two way ANOVA with Bonferroni post-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4316390&req=5

Fig4: Semi-quantitative evaluation of thioredoxin protein expression. A: Representative Westernblotting experiment performed with cardiac tissue samples of 3 control and 3 RIPC patients. B: Evaluation of the relative protein expression levels of thioredoxin in control and RIPC patients. Numbers in the columns display the numbers of patients employed in the respective experiment. MW, molecular weight; kDa, kiloDalton; a.u., arbitrary units; columns display the mean; bars denote SEM; *, P < 0.05, two way ANOVA with Bonferroni post-test.
Mentions: One of the proteins that showed a strong expression within the cardiac tissue and that was also regulated by RIPC is thioredoxin, a small redox protein and at least in animal studies, a possible candidate for cardioprotection [32,33,37,38]. Based on our results of the human proteome profiling approach which was performed with pooled tissue samples of the respective patients, we decided to further focus on thioredoxin as a potential molecule conferring RIPC-mediated cardioprotection and investigated the expression of thioredoxin in each patient separately. Westernblotting results mainly confirmed the outcome of the protein arrays, showing an increased expression of thioredoxin in cardiac tissue of RIPC patients that was obtained before CPB (RIPC: 5.36 ± 0.85 a.u.; control: 3.23 ± 0.39 a.u.; P < 0.05, Figure 4), while no statistically significant differences in thioredoxin protein expression levels were detected in samples taken after CPB (RIPC, 3.94 ± 0.73 a.u.; control, 2.79 ± 0.31 a.u.; P > 0.05; Figure 4).Figure 4

Bottom Line: Array results for thioredoxin were verified by semi-quantitative Westernblotting studies which showed a significant up-regulation of thioredoxin protein levels in cardiac tissue samples of RIPC patients taken before CPB (RIPC: 5.36 ± 0.85 a.u.; control: 3.23 ± 0.39 a.u.; P < 0.05).Quantification of thioredoxin levels in tissue of RIPC and control patients by ELISA experiments further confirmed the Westernblotting results (RIPC: 0.30 ± 0.02 ng/mg protein; control: 0.24 ± 0.02 ng/mg protein; P < 0.05).We provide evidence for thioredoxin as a RIPC-induced factor in heart tissue of cardiosurgical patients and identified several cell stress associated proteins that are regulated by RIPC and may play a role in RIPC-mediated cardioprotection.

View Article: PubMed Central - PubMed

Affiliation: Department of Anaesthesiology and Intensive Care Medicine, University Hospital Schleswig-Holstein, Schwanenweg 21, 24105, Kiel, Germany. karina.zitta@uksh.de.

ABSTRACT

Background: Transient episodes of ischemia in a remote organ (remote ischemic preconditioning, RIPC) can attenuate myocardial ischemia/reperfusion injury but the underlying mechanisms of RIPC in the target organ are still poorly understood. Recent animal studies suggested that the small redox protein thioredoxin may be a potential candidate for preconditioning-induced organprotection. Here we employed a human proteome profiler array to investigate the RIPC regulated expression of cell stress proteins and particularly of thioredoxin in heart tissue of cardiosurgical patients with cardiopulmonary bypass (CPB).

Methods: RIPC was induced by four 5 minute cycles of transient upper limb ischemia/reperfusion using a blood pressure cuff. Right atrial tissue was obtained from patients receiving RIPC (N = 19) and control patients (N = 19) before and after CPB. Cell stress proteome profiler arrays as well as Westernblotting and ELISA experiments for thioredoxin (Thio-1) were performed employing the respective tissue samples.

Results: Protein arrays revealed an up-regulation of 26.9% (7/26; CA IX, Cyt C, HSP-60, HSP-70, pJNK, SOD2, Thio-1) of cell stress associated proteins in RIPC tissue obtained before CPB, while 3.8% (1/26; SIRT2) of the proteins were down-regulated. Array results for thioredoxin were verified by semi-quantitative Westernblotting studies which showed a significant up-regulation of thioredoxin protein levels in cardiac tissue samples of RIPC patients taken before CPB (RIPC: 5.36 ± 0.85 a.u.; control: 3.23 ± 0.39 a.u.; P < 0.05). Quantification of thioredoxin levels in tissue of RIPC and control patients by ELISA experiments further confirmed the Westernblotting results (RIPC: 0.30 ± 0.02 ng/mg protein; control: 0.24 ± 0.02 ng/mg protein; P < 0.05).

Conclusion: We provide evidence for thioredoxin as a RIPC-induced factor in heart tissue of cardiosurgical patients and identified several cell stress associated proteins that are regulated by RIPC and may play a role in RIPC-mediated cardioprotection.

No MeSH data available.


Related in: MedlinePlus