Role of CD11b+Gr-1+ myeloid cells in AGEs-induced myocardial injury in a mice model of acute myocardial infarction - PubMed (original) (raw)

. 2015 Mar 1;8(3):3238-49.

eCollection 2015.

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Role of CD11b+Gr-1+ myeloid cells in AGEs-induced myocardial injury in a mice model of acute myocardial infarction

Tongqing Yao et al. Int J Clin Exp Pathol. 2015.

Abstract

Aims: Polymorph neutrophils are the predominant inflammatory cells and play a crucial role on the pathogenesis of myocardial injury at the early stage of acute myocardial infarction (AMI). However, the precursors and the differentiation of neutrophils are not fully understood. Here we explored the role of CD11b+Gr-1+ myeloid-derived suppressor cells (MDSCs) on myocardial injury in the absence and presence of advanced glycation end-products (AGEs) in a mice model of AMI.

Methods and results: Male C57BL/6J mice were selected. Fluorescent actived cell sortor (FACS) data demonstrated significantly increased CD11b+Gr-1+ MDSCs both in peripheral blood circulation and in the ischemic myocardium at 24 hours post AMI. Quantitative-real-time PCR results also revealed significantly upregulated CD11b and Ly6G mRNA expression in the ischemic myocardium. AGEs treatment further aggravated these changes in AMI mice but not in sham mice. Moreover, AGEs treatment also significantly increased infarction size and enhanced cardiomyocyte apoptosis. The mRNA expression of pro-inflammatory cytokine IL-6 and iNOS2 was also significantly increased in AMI + AGEs group compared to AMI group.

Conclusion: These data suggest enhanced infiltration of MDSCs by AGEs contributes to aggravated myocardial injury in AMI mice, which might be one of the mechanisms responsible for severer myocardial injury in AMI patients complicating diabetes.

Keywords: Acute myocardial infarction (AMI); CD11b+Gr-1+ myeloid cells; advanced glycation end-products (AGEs); inflammation; myeloid-derived suppressor cells (MDSCs); pro-inflammatory cytokines.

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Figures

Figure 1

Figure 1

Mouse Myeloid cells were examined in ischemic myocardium of AMI mice. A. The immunofluorescence image of CD11b+Gr1+ MDSC at marginal zone along infarction area of 24 h post AMI mice heart (n = 6). Arrows indicate where infarct area was. Arrows indicate where infarct area was detected by immune-fluorescence staining. B. CD11b and Ly6G mRNA expression in the ischemic myocardium at 24 h post operation (n = 6) compared with sham (n = 6) (student t-tests, *P < 0.05 compared with control groups). AMI = acute myocardial infarction; MI = myocardial infarction.

Figure 2

Figure 2

Mouse Myeloid cells expressing cell surface markers CD11b+ and Gr1+ were examined in blood, spleen and bone marrow of AMI mice and sham mice. A. The representative FACS graphs of CD11b+Gr-1+ in mice blood, spleen and bone marrow by flow cytometer 24 h after AMI compared to the sham group. B. Bars are divided to display percentage of CD11b+Gr1+ myeloid cells by FACS. (AMI group n = 6, sham group n = 6, student t-tests, *P < 0.05 compared with sham groups). AMI = acute myocardial infarction

Figure 3

Figure 3

AGEs enhanced infiltrating of CD11b+Gr1+ Myeloid cells in mice. A. The representative FACS graphs of CD11b+Gr-1+ MDSC in mice blood, spleen and bone marrow by flow cytometer 72 hours after AGEs injection compared to the control group. B. Bars are divided to display percentage of CD11b+Gr1+ myeloid cells changes between control group (n = 6) and AGEs injection group (n = 6) by FACS (student t-tests, *P < 0.05 compared with control). C. The representative FACS graphs of CD11b+Gr1+ MDSC in mice blood of sham (n = 6), sham+AGEs (n = 6), AMI (n = 6) and AMI+AGEs (n = 6) groups by flow cytometer one day after AMI. D. Bars are divided to display percentage of CD11b+Gr1+ myeloid cells changes among the four groups by FACS (one-way ANOVA followed by a Tukey’s test, *P < 0.05 compared with sham, †P < 0.05 compared with AMI). E. mRNA levels of RAGE in CD11b+Ly6G+ and CD11b+Ly6G- cells of AGEs treated sham mice (n = 6) detected by real time RCR (student t-tests, *P < 0.05 compared with CD11b+Ly6G-). AMI = acute myocardial infarction; AGEs = advanced glycation end products; RAGE = receptor for advanced glycation end-product.

Figure 4

Figure 4

AGEs enhanced infiltrating of CD11b positive cells in mice. A. CD11b positive cells count by immunohistochemistry in the ischemic myocardium of sham (n = 6), sham+AGEs (n = 6), AMI (n = 6) and AMI+AGEs (n = 6) groups. B. Statistical of bar chart from immunohistochemistry (one-way ANOVA followed by a Tukey’s test, *P < 0.05 VS sham group, †P < 0.05 VS AMI group). AMI = acute myocardial infarction; AGEs = advanced glycation end products.

Figure 5

Figure 5

AGEs aggravate myocardial ischemic injury. A. Ultrasonic data showed worse ventricular Systolic function in AMI + AGEs group compared with AMI group. Images are representative of 6 mice analyzed. B. Left Ventricular Ejection Fraction was detected in sham (n = 6), sham + AGEs (n = 6), AMI (n = 6) and AMI + AGEs (n = 6) groups. (one-way ANOVA followed by a Tukey’s test, EF%: 36.2 ± 9.1 vs. 43.7 ± 8.6, *P < 0.05, compared with sham group, †P < 0.05, compared with AMI group). C. TUNEL results: apoptotic cardiomyocytes were no obvious change between sham and sham + AGEs groups. markedly increased apoptotic cardiomyocytes in the infarcted myocardium in the AMI + AGEs injection group. D. Bars are divided to display the positive cells by TUNEL. Images are representative of 6 mice analyzed. (one-way ANOVA followed by a Tukey’s test, *P < 0.05, compared with sham group, †P < 0.05, compared with AMI group). E. Caspase3+ cells count by immunohistochemistry sham, sham + AGEs, AMI and AMI + AGEs groups. The microscope was 400 times. F. Statistical of bar chart display the number of caspase3+ cells. Images are representative of 6 mice analyzed. (one-way ANOVA followed by a Tukey’s test, *P < 0.05, compared with sham group, †P < 0.05, compared with AMI group). AMI = acute myocardial infarction; AGEs = advanced glycation end products; EF = Ejection Fraction; TUNEL = Terminal deoxynucleotidy ltransferase-mediated dUTP nick-end labeling.

Figure 6

Figure 6

AGEs increased pro-inflammatory cytokines IL-6 and iNOS releasing. A and B. mRNA levels of IL-6 in myocardium of sham (n = 6) and sham + AGEs (n = 6) mice, or in infarction myocardium of AMI (n = 6) and AMI + AGEs (n = 6) mice, detected by real time RCR. mRNA levels of IL-6 in infarction heart of AMI mice detected by real time RCR (one-way ANOVA followed by a Tukey’s test, *P < 0.05, compared with sham group, †P < 0.05, compared with AMI group). AMI = acute myocardial infarction; AGEs = advanced glycation end products; iNOS = inducible nitricoxide synthase.

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