Treatment of hypoxia‐dependent cardiovascular diseases by myo‐inositol trispyrophosphate (ITPP)‐enhancement of oxygen delivery by red blood cells

Figures

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  Figure 1. Experimental protocol. MI‐myocardial infarction; ITPP‐myo‐inositol trispyrophosphate

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  Figure 2. The effect of ITPP on echocardiographic parameters in sham operated (Sh) and myocardial infarction (MI) rats. A, The representative echocardiographic images of LV end‐diastolic area from the long‐axis parasternal view. (a) Sh; (b) Sh + ITPP; (c) MI; (d) MI + ITPP. LV, left ventricle; LA, left atrium. Dashed line marks the endocardial surface. While in both sham images the LV cavity is oblong in shape, it is almost spherical in MI image; B. Left ventricular diastolic area (upper panel) and ejection fraction (lower panel). * P < .05 MI + ITPP vs MI

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  Figure 3. Effect of ITPP on the cardiomyocyte function after myocardial infarction or sham operation. A, representative recordings of contraction in LV cardiomyocytes isolated from sham or infarcted (MI) rats treated or untreated by ITPP. B, schematic presentation of parameters describing cardiomyocyte contraction. Cell shortening (C), time to peak (D), time to 90% of relaxation (E), contraction (F) and relaxation (G) velocities measured in LV cardiomyocytes isolated from rats 8 wk after sham operation or MI induction treated by ITPP or saline. n = 33‐55 cardiomyocytes in each group isolated from at least 3 rats in each group. Only significant differences are shown (* P < .05)

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  Figure 4. Cardiomyocytic cells characterization and response to ITPP‐modified erythrocytes (RBC‐ITPP+). A, Cardiomyocytes MCmCl4 labelling for FTC anti‐α‐actinin antibodies. B, Cardiomyocytes expression of HIF‐1α upon hypoxia treatment. (a) Normoxia control, (b) Hypoxia 1% for 3 hours; (c) control; (d) Hypoxia 1% for 12 hours. C, HIF‐1α kinetics of production by cardiomyocytes upon hypoxia (1%) measured by ELISA. Data represent the mean values ± SEM from experiments performed 4 times (N = 4) in triplicate (n = 3). D, Distribution of HIF‐1α in cardiomyocytes and colocalization with nuclei upon hypoxia (1%) treatment. (a, c) Normoxic controls; (b, d) hypoxia 1%, for 3 hours. Nuclei are labelled by DAPI, and HIF‐1α was labelled by FTC anti‐HIF‐1α antibodies. Microscope is Zeiss Axiovert video microscope equipped with Apotome

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  Figure 5. HIF‐1α in cardiomyocytes is reduced by ITPP‐charged red blood cells (RBC‐ITPP+)‐ mediated oxygen delivery in flow conditions. A, Effect of RBC‐ITPP+ on the modulation of HIF‐1α expression in hypoxia, as compared to RBC‐ITPP−. HIF‐1α was detected by ELISA, experiments were performed in triplicates. Data represent the mean values ± SEM from four experiments, in triplicates. B, Flow chamber design for the in flow modulation of a cardiomyocytic cell culture. C, Modulation of cardiomyocytes production of HIF‐1α under hypoxia by flow borne ITPP‐charged RBC‐ITPP+ as compared to non‐modified RBC‐ITPP− erythrocytes. Controls are performed in the presence of medium with and without applied flow, and in the presence of ITPP (50 μmol/L) or IHP (50 μmol/L). Data represent the mean values ± SEM from three experiments in triplicates

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  Figure 6. Effect of ITPP and RBC‐ITPP on cardiomyocytic cell cultures. A, ITPP and RBC‐ITPP+ effect of MCmCl4 cells upon hypoxia (1%, 3 h). Controls are treated by ITPP (50 μmol/L) and IHP (50 μmol/L). B, Increase of the cardiomyocytes nuclei size was observed upon hypoxia treatment (pO2 1%) for 3 h and effect of RBC‐ITPP treatment in flow (20 µL/min) for 2 h, as compared to normoxia post‐treatment for 22 h. C, Analysis of the cell evolution in hypoxia (3 and 5 h, followed by a 22 h normoxia post‐treatment or a 2 h RBC‐ITPP+ flow treatment). Data are mean values ± SEM from three experiments performed in triplicates