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Intracardiac injection of erythropoietin induces stem cell recruitment and improves cardiac functions in a rat myocardial infarction model.

Klopsch C, Furlani D, Gäbel R, Li W, Pittermann E, Ugurlucan M, Kundt G, Zingler C, Titze U, Wang W, Ong LL, Wagner K, Li RK, Ma N, Steinhoff G - J. Cell. Mol. Med. (2009)

Bottom Line: MI-EPO hearts exhibited smaller infarction size (20.1 +/- 1.1% versus 27.8 +/- 1.2%; n = 6-8, P < 0.001) and greater capillary density (338.5 +/- 14.7 versus 259.8 +/- 9.2 vessels per mm2; n = 6-8, P < 0.001) than MIC hearts.The chemoattractant SDF-1 was up-regulated significantly assessed by quantitative realtime PCR and immunohistology. c-Kit(+) and CD34(+) stem cells were significantly more numerous in MI-EPO than in MIC at 24 hrs in peripheral blood (n = 7, P < 0.05) and 48 hrs in the infarcted hearts (n = 6, P < 0.001).Further, the mRNAs of Akt, eNOS and EPO receptor were significantly enhanced in MI-EPO hearts (n = 7, P < 0.05).

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiac Surgery, University of Rostock, Germany.

ABSTRACT
Erythropoietin (EPO) protects the myocardium from ischaemic injury and promotes beneficial remodelling. We assessed the therapeutic efficacy of intracardiac EPO injection and EPO-mediated stem cell homing in a rat myocardial infarction (MI) model. Following MI, EPO (3000 U/kg) or saline was delivered by intracardiac injection. Compared to myocardial infarction control group (MIC), EPO significantly improved left ventricular function (n =11-14, P < 0.05) and decreased right ventricular wall stress (n = 8, P < 0.05) assessed by pressure-volume loops after 6 weeks. MI-EPO hearts exhibited smaller infarction size (20.1 +/- 1.1% versus 27.8 +/- 1.2%; n = 6-8, P < 0.001) and greater capillary density (338.5 +/- 14.7 versus 259.8 +/- 9.2 vessels per mm2; n = 6-8, P < 0.001) than MIC hearts. Direct EPO injection reduced post-MI myocardial apoptosis by approximately 41% (0.27 +/- 0.03% versus 0.42 +/- 0.03%; n = 6, P= 0.005). The chemoattractant SDF-1 was up-regulated significantly assessed by quantitative realtime PCR and immunohistology. c-Kit(+) and CD34(+) stem cells were significantly more numerous in MI-EPO than in MIC at 24 hrs in peripheral blood (n = 7, P < 0.05) and 48 hrs in the infarcted hearts (n = 6, P < 0.001). Further, the mRNAs of Akt, eNOS and EPO receptor were significantly enhanced in MI-EPO hearts (n = 7, P < 0.05). Intracardiac EPO injection restores myocardial functions following MI, which may attribute to the improved early recruitment of c-Kit(+) and CD34(+) stem cells via the enhanced expression of chemoattractant SDF-1.

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Direct EPO injection increases c-Kit+ and CD34+ signals. (A, B) Quantitative real-time PCR analysis for c-Kit (A) and CD34 (B) genes in IZ and NIZ of MI-EPO (n= 7), MIC (n= 7) and Sham (n= 7) hearts. The average mRNA expression level of c-Kit and CD34 in the sham hearts was arbitrarily given a value of 1 (2°). *P< 0.05. (C, D) Representative immunostaining for c-Kit in an EPO-treated heart. c-Kit+ cells (square) were identified in NIZ (C) and IZ (D) at 48 hrs after treatment. Bar = 5 μm. Blue, TOPRO3 in nuclei. (E) The number of c-Kit+ cells per high power field (HPF) in IZ of MI-EPO (n= 6) hearts was significantly higher than in MIC (n= 6) hearts. **P< 0.01. (F, G) Representative images for CD34+ (green) cells (square) in NIZ (F, Bar = 5 μm) and IZ (G, Bar = 10 μm) at 48 hrs after EPO treatment. Red, TOPRO3 in nuclei. (H) The number of CD34+ cells per HPF in IZ of MI-EPO (n= 6) was significantly higher than in MIC (n= 6) hearts. **P< 0.01.
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fig05: Direct EPO injection increases c-Kit+ and CD34+ signals. (A, B) Quantitative real-time PCR analysis for c-Kit (A) and CD34 (B) genes in IZ and NIZ of MI-EPO (n= 7), MIC (n= 7) and Sham (n= 7) hearts. The average mRNA expression level of c-Kit and CD34 in the sham hearts was arbitrarily given a value of 1 (2°). *P< 0.05. (C, D) Representative immunostaining for c-Kit in an EPO-treated heart. c-Kit+ cells (square) were identified in NIZ (C) and IZ (D) at 48 hrs after treatment. Bar = 5 μm. Blue, TOPRO3 in nuclei. (E) The number of c-Kit+ cells per high power field (HPF) in IZ of MI-EPO (n= 6) hearts was significantly higher than in MIC (n= 6) hearts. **P< 0.01. (F, G) Representative images for CD34+ (green) cells (square) in NIZ (F, Bar = 5 μm) and IZ (G, Bar = 10 μm) at 48 hrs after EPO treatment. Red, TOPRO3 in nuclei. (H) The number of CD34+ cells per HPF in IZ of MI-EPO (n= 6) was significantly higher than in MIC (n= 6) hearts. **P< 0.01.

Mentions: Given the potential role of the stem cell recruitment by EPO, c-Kit+ and CD34+ cell number and mRNA level in the heart were assessed by immunostaining and quantitative real-time PCR. At 24 hrs c-Kit mRNA in MI-EPO hearts was significantly up-regulated in IZ. In both IZ and NIZ, c-Kit mRNA in MI-EPO hearts was markedly higher compared with MIC at 48 hrs (P< 0.05, Fig. 5A). There was an 80% increase of c-Kit+ cells after injection at 48 hrs in IZ (P< 0.001, Fig. 5E) detected by immunostaining. Similarly, CD34 mRNA in both IZ and NIZ of MI-EPO rats was up-regulated compared with MIC at 24 hrs (P< 0.05, Fig. 5B). Immunostaining, which was devoid of CD31 reactivity, revealed 72% increase of CD34+ cells in MI-EPO at 48 hrs in IZ (P= 0.001; Fig. 5H). These observations therefore revealed that EPO may enhance stem cell recruitment on the acute phase post-MI.


Intracardiac injection of erythropoietin induces stem cell recruitment and improves cardiac functions in a rat myocardial infarction model.

Klopsch C, Furlani D, Gäbel R, Li W, Pittermann E, Ugurlucan M, Kundt G, Zingler C, Titze U, Wang W, Ong LL, Wagner K, Li RK, Ma N, Steinhoff G - J. Cell. Mol. Med. (2009)

Direct EPO injection increases c-Kit+ and CD34+ signals. (A, B) Quantitative real-time PCR analysis for c-Kit (A) and CD34 (B) genes in IZ and NIZ of MI-EPO (n= 7), MIC (n= 7) and Sham (n= 7) hearts. The average mRNA expression level of c-Kit and CD34 in the sham hearts was arbitrarily given a value of 1 (2°). *P< 0.05. (C, D) Representative immunostaining for c-Kit in an EPO-treated heart. c-Kit+ cells (square) were identified in NIZ (C) and IZ (D) at 48 hrs after treatment. Bar = 5 μm. Blue, TOPRO3 in nuclei. (E) The number of c-Kit+ cells per high power field (HPF) in IZ of MI-EPO (n= 6) hearts was significantly higher than in MIC (n= 6) hearts. **P< 0.01. (F, G) Representative images for CD34+ (green) cells (square) in NIZ (F, Bar = 5 μm) and IZ (G, Bar = 10 μm) at 48 hrs after EPO treatment. Red, TOPRO3 in nuclei. (H) The number of CD34+ cells per HPF in IZ of MI-EPO (n= 6) was significantly higher than in MIC (n= 6) hearts. **P< 0.01.
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fig05: Direct EPO injection increases c-Kit+ and CD34+ signals. (A, B) Quantitative real-time PCR analysis for c-Kit (A) and CD34 (B) genes in IZ and NIZ of MI-EPO (n= 7), MIC (n= 7) and Sham (n= 7) hearts. The average mRNA expression level of c-Kit and CD34 in the sham hearts was arbitrarily given a value of 1 (2°). *P< 0.05. (C, D) Representative immunostaining for c-Kit in an EPO-treated heart. c-Kit+ cells (square) were identified in NIZ (C) and IZ (D) at 48 hrs after treatment. Bar = 5 μm. Blue, TOPRO3 in nuclei. (E) The number of c-Kit+ cells per high power field (HPF) in IZ of MI-EPO (n= 6) hearts was significantly higher than in MIC (n= 6) hearts. **P< 0.01. (F, G) Representative images for CD34+ (green) cells (square) in NIZ (F, Bar = 5 μm) and IZ (G, Bar = 10 μm) at 48 hrs after EPO treatment. Red, TOPRO3 in nuclei. (H) The number of CD34+ cells per HPF in IZ of MI-EPO (n= 6) was significantly higher than in MIC (n= 6) hearts. **P< 0.01.
Mentions: Given the potential role of the stem cell recruitment by EPO, c-Kit+ and CD34+ cell number and mRNA level in the heart were assessed by immunostaining and quantitative real-time PCR. At 24 hrs c-Kit mRNA in MI-EPO hearts was significantly up-regulated in IZ. In both IZ and NIZ, c-Kit mRNA in MI-EPO hearts was markedly higher compared with MIC at 48 hrs (P< 0.05, Fig. 5A). There was an 80% increase of c-Kit+ cells after injection at 48 hrs in IZ (P< 0.001, Fig. 5E) detected by immunostaining. Similarly, CD34 mRNA in both IZ and NIZ of MI-EPO rats was up-regulated compared with MIC at 24 hrs (P< 0.05, Fig. 5B). Immunostaining, which was devoid of CD31 reactivity, revealed 72% increase of CD34+ cells in MI-EPO at 48 hrs in IZ (P= 0.001; Fig. 5H). These observations therefore revealed that EPO may enhance stem cell recruitment on the acute phase post-MI.

Bottom Line: MI-EPO hearts exhibited smaller infarction size (20.1 +/- 1.1% versus 27.8 +/- 1.2%; n = 6-8, P < 0.001) and greater capillary density (338.5 +/- 14.7 versus 259.8 +/- 9.2 vessels per mm2; n = 6-8, P < 0.001) than MIC hearts.The chemoattractant SDF-1 was up-regulated significantly assessed by quantitative realtime PCR and immunohistology. c-Kit(+) and CD34(+) stem cells were significantly more numerous in MI-EPO than in MIC at 24 hrs in peripheral blood (n = 7, P < 0.05) and 48 hrs in the infarcted hearts (n = 6, P < 0.001).Further, the mRNAs of Akt, eNOS and EPO receptor were significantly enhanced in MI-EPO hearts (n = 7, P < 0.05).

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiac Surgery, University of Rostock, Germany.

ABSTRACT
Erythropoietin (EPO) protects the myocardium from ischaemic injury and promotes beneficial remodelling. We assessed the therapeutic efficacy of intracardiac EPO injection and EPO-mediated stem cell homing in a rat myocardial infarction (MI) model. Following MI, EPO (3000 U/kg) or saline was delivered by intracardiac injection. Compared to myocardial infarction control group (MIC), EPO significantly improved left ventricular function (n =11-14, P < 0.05) and decreased right ventricular wall stress (n = 8, P < 0.05) assessed by pressure-volume loops after 6 weeks. MI-EPO hearts exhibited smaller infarction size (20.1 +/- 1.1% versus 27.8 +/- 1.2%; n = 6-8, P < 0.001) and greater capillary density (338.5 +/- 14.7 versus 259.8 +/- 9.2 vessels per mm2; n = 6-8, P < 0.001) than MIC hearts. Direct EPO injection reduced post-MI myocardial apoptosis by approximately 41% (0.27 +/- 0.03% versus 0.42 +/- 0.03%; n = 6, P= 0.005). The chemoattractant SDF-1 was up-regulated significantly assessed by quantitative realtime PCR and immunohistology. c-Kit(+) and CD34(+) stem cells were significantly more numerous in MI-EPO than in MIC at 24 hrs in peripheral blood (n = 7, P < 0.05) and 48 hrs in the infarcted hearts (n = 6, P < 0.001). Further, the mRNAs of Akt, eNOS and EPO receptor were significantly enhanced in MI-EPO hearts (n = 7, P < 0.05). Intracardiac EPO injection restores myocardial functions following MI, which may attribute to the improved early recruitment of c-Kit(+) and CD34(+) stem cells via the enhanced expression of chemoattractant SDF-1.

Show MeSH
Related in: MedlinePlus