Comprehensive Physiology Wiley Online Library

Trans‐System Mechanisms Against Ischemic Myocardial Injury

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Abstract

A mammalian organism possesses a hierarchy of naturally evolved protective mechanisms against ischemic myocardial injury at the molecular, cellular, and organ levels. These mechanisms comprise regional protective processes, including upregulation and secretion of paracrine cell‐survival factors, inflammation, angiogenesis, fibrosis, and resident stem cell‐based cardiomyocyte regeneration. There are also interactive protective processes between the injured heart, circulation, and selected remote organs, defined as trans‐system protective mechanisms, including upregulation and secretion of endocrine cell‐survival factors from the liver and adipose tissue as well as mobilization of bone marrow, splenic, and hepatic cells to the injury site to mediate myocardial protection and repair. The injured heart and activated remote organs exploit molecular and cellular processes, including signal transduction, gene expression, cell proliferation, differentiation, migration, mobilization, and/or extracellular matrix production, to establish protective mechanisms. Both regional and trans‐system cardioprotective mechanisms are mediated by paracrine and endocrine messengers and act in coordination and synergy to maximize the protective effect, minimize myocardial infarction, and improve myocardial function, ensuring the survival and timely repair of the injured heart. The concept of the trans‐system protective mechanisms may be generalized to other organ systems—injury in one organ may initiate regional as well as trans‐system protective responses, thereby minimizing injury and ensuring the survival of the entire organism. Selected trans‐system processes may serve as core protective mechanisms that can be exploited by selected organs in injury. These naturally evolved protective mechanisms are the foundation for developing protective strategies for myocardial infarction and injury‐induced disorders in other organ systems. © 2015 American Physiological Society. Compr Physiol 5:167‐192, 2015.

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Figure 1. Figure 1. Genome‐wide profiling of mRNA transcription demonstrating upregulation of AGP2, BMPER, CXCL13, FGF21, NRG4, PRG4, and TFF3 mRNAs in hepatocytes from mice with sham operation (time 0) or myocardial ischemia. The “Relative level” is defined as a fold‐change in reference to the sham control level. SAA1 and SAA2, acute phase protein genes, were used as positive controls for the presence of liver inflammation. The dashed lines represent transcription of the β actin gene (202).
Figure 2. Figure 2. Identification of liver‐derived cardioprotective proteins. (A) TTC‐stained left ventricular specimens from mice with 24‐hr myocardial ischemia administered with PBS or recombinant AGP2, BMPER, CXCL13, FGF21, NRG4, PRG4, or TFF3 immediately post myocardial ischemia (50 ng/gm each, single dose, IV). A combination of AGP2, BMPER, FGF21, NRG4, and TFF3, identified as cardioprotective proteins, was administered with a similar strategy (50 ng/gm each). The intact myocardium was stained red and the infarct remains unstained. Scale bar: 1 mm. (B) Graphic representation of the influence of PBS, AGP2, BMPER, CXCL13, FGF21, NRG4, PRG4, or TFF3 administration on the degree of myocardial infarcts at 24 hrs. The influence of AGP2, BMPER, FGF21, NRG4, and TFF3 in combination is also presented (5 SPs or secreted proteins). Means and SDs are presented (n = 8 each group). * P < 0.05, ** P < 0.01, and *** P < 0.001 for comparisons between hepatic protein(s) and PBS. # P < 0.05 and ### P < 0.001 for comparisons between a single hepatic protein and the combination of AGP2, BMPER, FGF21, NRG4, and TFF3 (196).
Figure 3. Figure 3. Mobilization of hepatic cells into the circulatory system in response to myocardial ischemia. (A) A fluorescence micrograph showing circulating EYFP‐positive cells (green) from an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. (B) Expression of CK19 (red) in a circulating EYFP‐positive cell (green) from an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. Blue: cell nuclei for panels A and B. Scale: 10 μm. (C) Graphic representation of circulating EYFP‐positive cells with reference to the total nucleated blood cells in Alb‐Cre;Stopflox‐EYFP mice with sham‐operation (open circles) and myocardial ischemia (solid circles) as well as in C57BL/6 mice with myocardial ischemia (solid squares) measured by fluorescence microscopy. Means and standard deviations are presented (P < 0.001 for changes in Alb‐Cre;Stopflox‐EYFP mice with myocardial ischemia by ANOVA, n = 6 each time). (D) Flow cytometry analysis of EYFP‐positive cells from the liver of Alb‐Cre;Stopflox‐EYFP mice with sham operation and from the blood of Alb‐Cre;Stopflox‐EYFP mice with sham operation or myocardial ischemia (MI). A standard level of fluorescence intensity (the left side of the red rectangle) was established based on hepatic cell samples from Alb‐Cre;Stopflox‐EYFP mice and used to assess the population size of blood‐borne EYFP‐positive cells. The fraction shown in each panel represents the mean and standard deviation of the EYFP‐positive cell population size from six tests (199).
Figure 4. Figure 4. Characterization of the Alb‐Cre;Stopflox‐EYFP mouse model. (A) Expression of EYFP (green) in a liver specimen from an Alb‐Cre;Stopflox‐EYFP mouse. H: hepatocyte. BE: biliary epithelial cell. Red: cytokeratin‐19, an epithelial cell marker expressed in biliary epithelial cells. (B) Expression of CD45 (red) in periductular cells of a liver specimen from an Alb‐Cre;Stopflox‐EYFP mouse. Blue: cell nuclei for both panels. Scale: 100 μm (199).
Figure 5. Figure 5. Leukocyte migration into the liver parenchyma in myocardial ischemia and the role of IL‐6 in mediating leukocyte migration. (A) Fluorescence micrographs showing the lack of CD45‐positive leukocytes (red) in the liver parenchyma of an Alb‐Cre;Stopflox‐EYFP mouse with sham operation at day 5. Panel A2 is a magnified image of the selected area in A1 (white rectangle). (B) CD45‐positive leukocytes (red) retained in the liver parenchyma of an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. Panel B2 is a magnified image of the selected area in B1. (C) CD45‐positive leukocytes (red) with coexpression of MMP2 (green) in the liver parenchyma of a C57BL/6J mouse with 5‐day myocardial ischemia. (D) Association of CD45‐positive leukocytes (red) with mobilized EYFP‐positive hepatic cells (green) within a central vein of the liver of an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. Panel D2 and D3 are magnified images of the left and right white rectangles, respectively, from D1. (E) Fluorescence micrographs showing the lack of CD45‐positive leukocytes (red) in the liver of an Alb‐Cre;Stopflox‐EYFP;IL6−/− mouse with 5‐day myocardial ischemia. Panel E2 is a magnified image of the selected area in E1. (F) CD45‐positive leukocytes (red) retained in the liver parenchyma of an Alb‐Cre;Stopflox‐EYFP;IL6−/− mouse with 5‐day myocardial ischemia with IL‐6 administration. Panel F2 is a magnified image of the selected area in F1. For panel A, B, D‐F, the green color represents EYFP. For panel A‐F, the blue color is for cell nuclei and the scale bars are 10 μm. (G) Graphic representation of the relative density of CD45‐positive leukocytes migrated into the liver parenchyma of mice with sham‐operation (open circles) and myocardial ischemia (solid circles) by fluorescence microscopy. Means and SDs are presented (P < 0.001 for changes in myocardial ischemia by ANOVA, n = 6 each time). Specimens at time zero were prepared from healthy mice. (H) Influence of IL6 on CD45‐positive leukocyte migration into the liver parenchyma of Alb‐Cre;Stopflox‐EYFP (Cre‐EYFP) and Alb‐Cre;Stopflox‐EYFP;IL6−/− (IL6−/−) mice with sham‐operation or myocardial ischemia. In panel G and H, the fraction of liver‐retained CD45‐positive leukocytes was calculated with reference to the total liver cells. (I) Flow cytometry analysis of liver cells derived from Alb‐Cre;Stopflox‐EYFP mice with sham operation or myocardial ischemia at day 5, showing CD45‐positive leukocyte retention in the liver parenchyma in myocardial ischemia (199).
Figure 6. Figure 6. AZAN‐stained left ventricular sections from mice with ischemic myocardial injury administered with PBS or hepatic proteins, including AGP2, BMPER, FGF21, NRG4, and TFF3, showing the cardioprotective effect of the liver‐derived proteins. Scale bar: 1 mm (196).
Figure 7. Figure 7. Left ventricular dp/dt (A) and echocardiographs (B) from mice with 5‐day sham operation or ischemic myocardial injury administrated with PBS or hepatic proteins, including AGP2, BMPER, FGF21, NRG4, and TFF3, showing the beneficial effect of the liver‐derived proteins on left ventricular function. MI: Myocardial ischemia. Hep: Hepatic (196).
Figure 8. Figure 8. Influence of siRNA‐mediated FGFR1, β‐Klotho, PI3K p110, or Akt1 gene silencing on the degree of myocardial infarcts. (A) AZAN‐stained left ventricular sections from FGF21−/− mice administered with a control, FGFR1, β‐Klotho, PI3K p110, or Akt1 siRNA (siRNA injected into the myocardium at 3 days prior to myocardial ischemia/reperfusion injury, specimens collected at 5 days following myocardial injury). These mice were administered intravenously with recombinant FGF21 immediately following myocardial injury for 3 days with a 12 hr interval to establish FGF21‐based myocardial protection. Scale bar: 1 mm. (B) Graphic representation of the influence of siRNA‐mediated FGFR1, β‐Klotho, PI3K p110, or Akt1 gene silencing on the degree of myocardial infarcts at day 5. Means and SDs are presented (n = 7) (197).


Figure 1. Genome‐wide profiling of mRNA transcription demonstrating upregulation of AGP2, BMPER, CXCL13, FGF21, NRG4, PRG4, and TFF3 mRNAs in hepatocytes from mice with sham operation (time 0) or myocardial ischemia. The “Relative level” is defined as a fold‐change in reference to the sham control level. SAA1 and SAA2, acute phase protein genes, were used as positive controls for the presence of liver inflammation. The dashed lines represent transcription of the β actin gene (202).


Figure 2. Identification of liver‐derived cardioprotective proteins. (A) TTC‐stained left ventricular specimens from mice with 24‐hr myocardial ischemia administered with PBS or recombinant AGP2, BMPER, CXCL13, FGF21, NRG4, PRG4, or TFF3 immediately post myocardial ischemia (50 ng/gm each, single dose, IV). A combination of AGP2, BMPER, FGF21, NRG4, and TFF3, identified as cardioprotective proteins, was administered with a similar strategy (50 ng/gm each). The intact myocardium was stained red and the infarct remains unstained. Scale bar: 1 mm. (B) Graphic representation of the influence of PBS, AGP2, BMPER, CXCL13, FGF21, NRG4, PRG4, or TFF3 administration on the degree of myocardial infarcts at 24 hrs. The influence of AGP2, BMPER, FGF21, NRG4, and TFF3 in combination is also presented (5 SPs or secreted proteins). Means and SDs are presented (n = 8 each group). * P < 0.05, ** P < 0.01, and *** P < 0.001 for comparisons between hepatic protein(s) and PBS. # P < 0.05 and ### P < 0.001 for comparisons between a single hepatic protein and the combination of AGP2, BMPER, FGF21, NRG4, and TFF3 (196).


Figure 3. Mobilization of hepatic cells into the circulatory system in response to myocardial ischemia. (A) A fluorescence micrograph showing circulating EYFP‐positive cells (green) from an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. (B) Expression of CK19 (red) in a circulating EYFP‐positive cell (green) from an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. Blue: cell nuclei for panels A and B. Scale: 10 μm. (C) Graphic representation of circulating EYFP‐positive cells with reference to the total nucleated blood cells in Alb‐Cre;Stopflox‐EYFP mice with sham‐operation (open circles) and myocardial ischemia (solid circles) as well as in C57BL/6 mice with myocardial ischemia (solid squares) measured by fluorescence microscopy. Means and standard deviations are presented (P < 0.001 for changes in Alb‐Cre;Stopflox‐EYFP mice with myocardial ischemia by ANOVA, n = 6 each time). (D) Flow cytometry analysis of EYFP‐positive cells from the liver of Alb‐Cre;Stopflox‐EYFP mice with sham operation and from the blood of Alb‐Cre;Stopflox‐EYFP mice with sham operation or myocardial ischemia (MI). A standard level of fluorescence intensity (the left side of the red rectangle) was established based on hepatic cell samples from Alb‐Cre;Stopflox‐EYFP mice and used to assess the population size of blood‐borne EYFP‐positive cells. The fraction shown in each panel represents the mean and standard deviation of the EYFP‐positive cell population size from six tests (199).


Figure 4. Characterization of the Alb‐Cre;Stopflox‐EYFP mouse model. (A) Expression of EYFP (green) in a liver specimen from an Alb‐Cre;Stopflox‐EYFP mouse. H: hepatocyte. BE: biliary epithelial cell. Red: cytokeratin‐19, an epithelial cell marker expressed in biliary epithelial cells. (B) Expression of CD45 (red) in periductular cells of a liver specimen from an Alb‐Cre;Stopflox‐EYFP mouse. Blue: cell nuclei for both panels. Scale: 100 μm (199).


Figure 5. Leukocyte migration into the liver parenchyma in myocardial ischemia and the role of IL‐6 in mediating leukocyte migration. (A) Fluorescence micrographs showing the lack of CD45‐positive leukocytes (red) in the liver parenchyma of an Alb‐Cre;Stopflox‐EYFP mouse with sham operation at day 5. Panel A2 is a magnified image of the selected area in A1 (white rectangle). (B) CD45‐positive leukocytes (red) retained in the liver parenchyma of an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. Panel B2 is a magnified image of the selected area in B1. (C) CD45‐positive leukocytes (red) with coexpression of MMP2 (green) in the liver parenchyma of a C57BL/6J mouse with 5‐day myocardial ischemia. (D) Association of CD45‐positive leukocytes (red) with mobilized EYFP‐positive hepatic cells (green) within a central vein of the liver of an Alb‐Cre;Stopflox‐EYFP mouse with 5‐day myocardial ischemia. Panel D2 and D3 are magnified images of the left and right white rectangles, respectively, from D1. (E) Fluorescence micrographs showing the lack of CD45‐positive leukocytes (red) in the liver of an Alb‐Cre;Stopflox‐EYFP;IL6−/− mouse with 5‐day myocardial ischemia. Panel E2 is a magnified image of the selected area in E1. (F) CD45‐positive leukocytes (red) retained in the liver parenchyma of an Alb‐Cre;Stopflox‐EYFP;IL6−/− mouse with 5‐day myocardial ischemia with IL‐6 administration. Panel F2 is a magnified image of the selected area in F1. For panel A, B, D‐F, the green color represents EYFP. For panel A‐F, the blue color is for cell nuclei and the scale bars are 10 μm. (G) Graphic representation of the relative density of CD45‐positive leukocytes migrated into the liver parenchyma of mice with sham‐operation (open circles) and myocardial ischemia (solid circles) by fluorescence microscopy. Means and SDs are presented (P < 0.001 for changes in myocardial ischemia by ANOVA, n = 6 each time). Specimens at time zero were prepared from healthy mice. (H) Influence of IL6 on CD45‐positive leukocyte migration into the liver parenchyma of Alb‐Cre;Stopflox‐EYFP (Cre‐EYFP) and Alb‐Cre;Stopflox‐EYFP;IL6−/− (IL6−/−) mice with sham‐operation or myocardial ischemia. In panel G and H, the fraction of liver‐retained CD45‐positive leukocytes was calculated with reference to the total liver cells. (I) Flow cytometry analysis of liver cells derived from Alb‐Cre;Stopflox‐EYFP mice with sham operation or myocardial ischemia at day 5, showing CD45‐positive leukocyte retention in the liver parenchyma in myocardial ischemia (199).


Figure 6. AZAN‐stained left ventricular sections from mice with ischemic myocardial injury administered with PBS or hepatic proteins, including AGP2, BMPER, FGF21, NRG4, and TFF3, showing the cardioprotective effect of the liver‐derived proteins. Scale bar: 1 mm (196).


Figure 7. Left ventricular dp/dt (A) and echocardiographs (B) from mice with 5‐day sham operation or ischemic myocardial injury administrated with PBS or hepatic proteins, including AGP2, BMPER, FGF21, NRG4, and TFF3, showing the beneficial effect of the liver‐derived proteins on left ventricular function. MI: Myocardial ischemia. Hep: Hepatic (196).


Figure 8. Influence of siRNA‐mediated FGFR1, β‐Klotho, PI3K p110, or Akt1 gene silencing on the degree of myocardial infarcts. (A) AZAN‐stained left ventricular sections from FGF21−/− mice administered with a control, FGFR1, β‐Klotho, PI3K p110, or Akt1 siRNA (siRNA injected into the myocardium at 3 days prior to myocardial ischemia/reperfusion injury, specimens collected at 5 days following myocardial injury). These mice were administered intravenously with recombinant FGF21 immediately following myocardial injury for 3 days with a 12 hr interval to establish FGF21‐based myocardial protection. Scale bar: 1 mm. (B) Graphic representation of the influence of siRNA‐mediated FGFR1, β‐Klotho, PI3K p110, or Akt1 gene silencing on the degree of myocardial infarcts at day 5. Means and SDs are presented (n = 7) (197).
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Shu Q. Liu, Xin‐Liang Ma, Gangjian Qin, Qingping Liu, Yan‐Chun Li, Yu H. Wu. Trans‐System Mechanisms Against Ischemic Myocardial Injury. Compr Physiol 2014, 5: 167-192. doi: 10.1002/cphy.c140026