Hydrogen Sulfide Reduces Recruitment of CD11b+Gr-1+ Cells in Mice With Myocardial Infarction

Abstract

The present study aimed to elucidate the mechanisms by which hydrogen sulfide (H2S) attenuates left ventricular remodeling after myocardial infarction (MI). MI was created in mice by left coronary artery ligation. One group of mice received injections of the H2S donor sodium hydrosulfide (NaHS) immediately before and 1 h after ligation, while the control group received saline alone. During both the subacute and chronic stages (1 and 4 weeks postinfarction, respectively), NaHS-treated mice demonstrated attenuation of cardiac dilation in the infarcted myocardium. Furthermore, fewer CD11b+Gr-1+ myeloid cells were detected in the infarct myocardium and peripheral blood from NaHS-treated mice, while more CD11b+Gr-1+ cells remained in the spleen and bone marrow in these animals. NaHS-treated mice also exhibited reduction in cardiomyocyte apoptosis, interstitial fibrosis, cardiac hypertrophy, and pulmonary edema, as well as overall better survival rates, when compared to controls. Thus, exogenous H2S has favorable effects on cardiac remodeling after MI. These observations further support the emerging concept that H2S treatment might have therapeutic benefits in the setting of ischemia-induced heart failure.

Figure 1.

Kaplan-Meier survival analysis. Percentages of surviving mice after sham operation or coronary occlusion were plotted. The survival curves were statistically different (^*p < 0.05) by log-rank test. NaHS, sodium hydrosulfide; MI, myocardial infarction.

Figure 2.

Cardiac hypertrophy and pulmonary edema 4 weeks after coronary occlusion or sham surgery. The ratio of heart weight (HW; in mg) to body weight (BW; in g) was determined in the four experimental groups (n = 5 per group) as a measure of cardiac hypertrophy (A). The ratio of lung weight (LW; in mg) to BW (in g) was determined in the four experimental groups (n = 5 per group) as a measure of pulmonary edema (B). ^*p < 0.05, MI + saline versus sham + saline; #p < 0.05, MI + NaHS versus sham + NaHS. NaHS, sodium hydrosulfide; MI, myocardial infarction; NS, not significant; wk, week.

Figure 3.

Echocardiographic parameters of mice with ischemia-induced heart failure. Representative M-mode measurement (A) and echocardiographic results of left ventricular end-diastolic diameter (LVEDD) (B), end-systolic diameter (LVESD) (C), and ejection fraction (LVEF) (D) in the various treatment groups. Measurements made 1 week after surgery for sham groups and 1 week and 4 weeks after surgery for the MI groups. n = 8-10 for all treatment groups. ^*p<0.01, MI+saline versus sham + saline; #p<0.05, ##p<0.01, MI+NaHS versus sham+NaHS. NaHS, sodium hydrosulfide; MI, myocardial infarction; NS, not significant; w, week.

Figure 4.

Cardioprotective effects of NaHS as measured by PET/CT and TUNEL apoptosis assay. PET/CT was performed 1 week after surgery. Representative PET/CT images of coronal whole-body slice at the level of left ventricle are shown. Left: horizontal view of the heart; middle: side view of the heart; right: front view of the heart (A). PET/CT standard uptake values (SUV) determined by manual delineation (n = 4) (B). Representative TUNEL assay images (200x) (C). Quantification of TUNEL-positive cells (percentage of total nuclei; n = 5 hearts per group) (D). Scale bars: 100 μm. &p < 0.05, 1 week post-MI + saline versus sham. ^*p < 0.05, MI + saline versus sham + saline; ^**p < 0.01, MI + saline versus sham + saline; #p < 0.01, MI + NaHS versus sham + NaHS. NaHS, sodium hydro-sulfide; MI, myocardial infarction; NS, not significant; PET/CT, positron emission tomography-computed tomography; TUNEL, terminal deoxynucleotidyl transferase 2′-deoxyuridine 5′-triphosphate (dUTP) nick-end labeling; w, week.

Figure 5.

Collagen deposition and fibrosis in cardiac muscle 4 weeks after surgery. Transverse sections of hearts stained with H&E (A; 200x; scale bars: 100 μm) and Masson trichrome (blue staining indicates collagen) (B; 200x; scale bars: 100 μm). Relative area of collagen, as indicated by blue stain, was measured among various groups. Whole section area was set as 100% (four to six animals per group) (C). Representative TEM images of left ventricular muscle collected 4 weeks after surgery. Scale bars: 2 μm (D). ^*p<0.01, MI+saline versus sham+saline; #p<0.01, MI+NaHS versus sham +NaHS. &p<0.05, MI+NaHS versus MI+saline. NaHS, sodium hydrosulfide; MI, myocardial infarction; NS, not significant; H&E, hematoxylin and eosin; w, week.

Figure 6.

NaHS inhibits recruitment and migration of CD11b⁺Gr-1⁺ myeloid cells. Representative flow cytometry dot plots of CD11b⁺Gr-1⁺ myeloid cells in myocardium (A), blood (C), spleen (E), and bone marrow (G) 1 week and 4 weeks after surgery. Quantification of CD11b⁺Gr-1⁺ cells is also shown for myocardium (B; n = 6), blood (D; n = 6), spleen (F; n = 6), and bone marrow (H; n = 6). ^*p < 0.01, MI + saline versus sham + saline; #p < 0.01, MI + NaHS versus sham + NaHS. &p < 0.01, 1 week post-MI + NaHS versus 1 week post-MI + saline. $p < 0.05, $$p < 0.01, 4 weeks post-MI + NaHS versus 4 weeks post-MI + saline. NaHS, sodium hydrosulfide; MI, myocardial infarction; NS, not significant; w, week.

Figure 7.

NaHS mitigates inflammatory response in the subacute phase of infarction. Serum TNF-α (A) and IL-1β (B) levels were measured by ELISA 1 week and 4 weeks after occlusion for MI + saline and MI + NaHS groups and 1 week after sham surgery for sham groups (n = 6). ^*p < 0.01, MI + saline versus sham + saline; #p < 0.01, MI + NaHS versus sham + NaHS. NaHS, sodium hydrosulfide; MI, myocardial infarction; NS, not significant; TNF, tumor necrosis factor; IL, interleukin; w, week.