Reduction in Hemoglobin–Oxygen Affinity Results in the Improvement of Exercise Capacity in Mice With Chronic Heart Failure
Tetsuya Watanabe, MD, PhD*,
Toshihiro Takeda, MD, PhD*,
Shigemiki Omiya, MD*,
Shungo Hikoso, MD, PhD*,
Osamu Yamaguchi, MD, PhD*,
Yuko Nakano, DDS ,
Yoshiharu Higuchi, MD, PhD*,
Atsuko Nakai, PhD*,
Yusuke Abe, MS ,
Yayoi Aki-Jin, PhD ,
Masayuki Taniike, MD, PhD*,
Isamu Mizote, MD*,
Yasushi Matsumura, MD, PhD||,
Takahiko Shimizu, PhD,
Kazuhiko Nishida, MD, PhD*,
Kiyohiro Imai, PhD,
Masatsugu Hori, MD, PhD, FACC*,
Takuji Shirasawa, MD, PhD and
Kinya Otsu, MD, PhD*,*
* Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, Japan
First Department of Oral and Maxillo-Facial Surgery, Osaka University Graduate School of Dentistry, Osaka, Japan
Department of Molecular Gerontology, Tokyo Metropolitan Institute of Gerontology, Tokyo, Japan
Department of Frontier Bioscience, Faculty of Engineering and Micro-Nano Technology Research Center, Hosei University, Tokyo, Japan
|| Department of Medical Information Science, Osaka University Graduate School of Medicine, Osaka, Japan.
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Figure 1 Characterization of Transplanted BMCs
(A) Detection of Presbyterian β (βPres)-globin. Reversed-phase high-performance liquid chromatography profiles of hemolysate prepared from WT (top) or Presbyterian bone marrow cell (BMC)-transplanted (bottom) mice. The peaks of  -globin, WT (βWT-globin), and βPres-globin are indicated. (B) The O2 dissociation curves of the red blood cells from WT and Presbyterian BMC-transplanted mice measured at pH 7.4 (open circles and triangles) or at pH 6.9 (closed circles and triangles). Circles and triangles indicate WT and Presbyterian BMC transplanted mice, respectively. BMT = bone marrow cell transplantation; Pres = Presbyterian; WT = wild type.
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Figure 2 Characterization of MI and BMT Mouse Hearts
(A) Experimental protocol. The LCA ligation operation was performed on 8-week-old WT mice. Four weeks after LCA ligation, echocardiographic analysis was performed and BMCs were transplanted into the lethally irradiated WT mice. Then, 4 weeks after BMT, the mice were echocardiographically analyzed again and subjected to the treadmill test. (B) Echocardiographic analysis findings before BMT (4 weeks post-operatively) and before the treadmill test (8 weeks post-operatively). (C) Hemodynamic data of the mice 4 weeks after BMT. Light blue, blue, yellow, and red bars represent Sham-WT BMT, MI-WT BMT, Sham-Pres BMT, and MI-Pres BMT, respectively. (D) Morphometric data from the mice 4 weeks after BMT. (E) Hematoxylin-eosin stain of heart section and cross-sectional area of cardiomyocytes at remote area. (F) Cardiac fibrosis in border and remote area with Azan-Mallory staining. BMC = bone marrow cell; HR = heart rate; HW/BW = heart weight to body weight ratio; LCA = left coronary artery; lungW/BW = lung weight-to-body weight ratio; LVSP = left ventricular systolic pressure; MI = myocardial infarction; other abbreviations as in Figure 1.
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Figure 3 Effect of Pres BMT on Exercise Capacity in MI Mice
(A and B) Physical activity of BMT mice on a treadmill. Total rest time is shown in (A). Data represent the means ± SEM (n = 7). *p < 0.05 versus other groups. The distance run is shown in (B). Open triangles and closed circles represent MI-Pres BMT and MI-WT BMT, respectively. Data indicate the means ± SEM (n = 7). *p < 0.05 versus at corresponding time point. (C) Changes in tissue O2 (PtiO2) in BMT mice during hypoxia. Closed circles and open triangles indicate MI-WT and -Pres BMT mice, respectively. Data show the means ± SEM (n = 3). *p < 0.05 versus at corresponding time point. (D) Serum lactate level in BMT mice after exercise. Data represent the means ± SEM (n = 6 for MI-WT BMT mice and n = 8 for MI-Pres BMT mice). *p < 0.05. Abbreviations as in Figures 1 and 2.
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Figure 4 Histological and Biochemical Analyses of Skeletal Muscles in BMT Mice
(A) Transverse sections of the deep region in the tibialis anterior muscle were stained for adenosine triphosphatase activity (top) and for succinate dehydrogenase (SDH) activity (bottom). Scale bars indicate 50 µm. Quantitative analysis results for muscle type distribution and SDH activity are shown in B and C, respectively. (D) Total SDH activity from tibialis anterior muscle measured by enzymological methods (n = 3 for WT-BMT and n = 5 for Pres-BMT mice). (E and F) Endothelial cells were identified by staining with von Willebrand factor antibody. Capillary density was calculated by counting the numbers of capillaries in 20 random high-power fields (x400 power). Scale bars indicate 50 µm. Abbreviations as in Figures 1 and 2.
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Figure 5 Effect of RSR13 on Exercise Capacity of MI Mice
(A) Chemical structure of RSR13. (B) Effect of RSR13 on running distance by normal mice. Thirty minutes after RSR13 or vehicle injection, the exercise test was started. Total distance run for each 30-min period are shown. C = vehicle-treated control group; R = RSR13-treated group. (C and D) Effect of RSR13 on exercise capacity of MI mice. Four weeks after LCA ligation, RSR13 or vehicle was injected into the mice (n = 4 each), and 30 min after the injection, the exercise test was started. Total rest time and distance run for each 30-min period are shown in C and D, respectively. Data show the means ± SEM. *p < 0.05 versus corresponding control subjects. Abbreviations as in Figure 2.
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