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Photoluminescent Mesoporous Silicon Nanoparticles with siCCR2 Improve the Effects of Mesenchymal Stromal Cell Transplantation after Acute Myocardial Infarction.

Lu W, Xie Z, Tang Y, Bai L, Yao Y, Fu C, Ma G - Theranostics (2015)

Bottom Line: The therapeutic effects of PMSNs-siCCR2 for MSC transplantation were determined at the mRNA, protein and functional levels.In addition, significant amelioration of left ventricular (LV) remodeling (thickness of the LV posterior walls) (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) was also observed at the same time compared with the control group.These results suggest that PMSNs-siCCR2 could potentially be used to develop an anti-inflammatory therapy for post-AMI MSC transplantation.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Cardiology, ZhongDa Hospital affiliated with Southeast University, China; ; 2. Department of Cardiology, the Second Hospital affiliated with Southeast University, China;

ABSTRACT

Background: Despite the benefits of mesenchymal stromal cell (MSC) transplantation in cardiac tissue, detailed in vivo observations have shown that MSCs only survive for a brief period after transplantation due to harsh microenvironmental conditions, including ischemia, inflammation and anoikis, in the infarcted myocardium. Thus, new strategies are needed to enhance MSC survival and inhibit cardiac remodeling. Studies have now demonstrated that chemokine [C-C motif] ligand 2 (CCL2) and its cognate receptor C-C chemokine receptor 2 (CCR2) promote excessive Ly6C(high) inflammatory monocyte infiltration at the infarct in response to ischemic myocardial injury. Therefore, decreasing the activities of these monocytes immediately after acute myocardial infarction (AMI) could be beneficial for AMI patients.

Objectives: This study tested the hypothesis that therapeutic siRNA-loaded photoluminescent mesoporous silicon nanoparticles (PMSNs) targeting CCR2 expression in Ly6C(high) inflammatory monocytes decrease the accumulation of these cells in the infarct, improve the efficacy of MSC transplantation and attenuate myocardial remodeling.

Methods: PMSNs carrying therapeutic siCCR2 were first synthesized without the inclusion of fluorescent materials or dyes. After AMI BALB/c mice were established, 10(5) 5-ethynyl-2'- deoxyuridine (EdU)-labeled MSCs suspended in 100 µl of phosphate buffered saline (PBS) were injected into the border zone of the infarct of each mouse. PMSNs-siCCR2 (25 µg/g) were also intravenously injected via the tail vein immediately following AMI induction. Control mice were injected with an equal amount of PMSNs without siCCR2. PMSNs-siCCR2 were examined in vivo using near-infrared imaging technology. The therapeutic effects of PMSNs-siCCR2 for MSC transplantation were determined at the mRNA, protein and functional levels.

Results: PMSNs-siCCR2 circulated freely in vivo and were cleared in a relatively short period of time (t(½)=37 min) with no evidence of toxicity. The therapeutic PMSNs-siCCR2 showed higher levels of cellular accumulation in Ly6C(high) monocytes in the spleen and more efficient degradation of CCR2 compared with the control (8.04%±2.17% vs. 20.02%±4.55%, p<0.001). Subsequently, the PMSNs-siCCR2 decreased the accumulation of CD11b-positive monocytes at the infarct (49.3%±17.34% vs. 61.32%±22.43%, p<0.001) on day 1. Increased survival of transplanted MSCs (13±3/mm(2) vs. 4±1/mm(2), p<0.001) and significantly decreased TdT-mediated dUTP nick end labeling (TUNEL)(+) cardiac myocytes (17.44%±6.26% vs. 39.49%±13.28%, p<0.001) were then identified in the infarct zone three days after AMI induction in the PMSNs-siCCR2 group. Three weeks after MSC injection, significant increases were observed in the vascular density (235.5±39.6/mm(2) vs. 147.4±20.3/mm(2), p<0.001) and the cardiac myosin-positive area (21.7%±8.4% vs. 13.2%±4.4%, p<0.001) of the infarct border zone. In addition, significant amelioration of left ventricular (LV) remodeling (thickness of the LV posterior walls) (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) was also observed at the same time compared with the control group.

Conclusions: PMSNs-siCCR2-mediated CCR2 gene silencing in Ly6C(high) monocytes improved the effectiveness of MSC transplantation and selectively ameliorated myocardial remodeling after AMI. These results suggest that PMSNs-siCCR2 could potentially be used to develop an anti-inflammatory therapy for post-AMI MSC transplantation.

No MeSH data available.


Related in: MedlinePlus

PMSNs-siCCR2-PEI enhanced the efficiency of MSC transplantation and attenuated myocardial remodeling (n=12). (a), The surviving MSCs in the infarct zone following cell therapy are shown by representative immunofluorescent staining for EdU (5-Ethynyl-2'-deoxyuridine, green) and visualization of the nuclei (DAPI, blue) within the infarct zone 3 days after cell transplantation. Scale bar=50 μm. (b), SDF-1 and Ang-1 mRNA levels in the infarct zone after MSC transplantation. The data represent the mean±SD, *p<0.05 vs. the corresponding group at 1 day post-AMI; #p<0.05 vs. the control group at 3 days after cell transplantation. (c), Representative image of immunofluorescent staining for TUNEL (Alexa Fluor 488, green) and DAPI in the infarct border zone 3 days after AMI; the number of TUNEL-positive cardiomyocytes was calculated. Scale bar=50 μm. (d), The vascular density was determined by immunofluorescent staining for wheat germ agglutinin (WGA, red), endothelial cells (staining of CD31, green) and DAPI (blue) in the infarct zone at 21 days after MSC transplantation. (e), Confocal image of representative immunofluorescent staining for cardiac myosin heavy chain (eFluor® 660, red) and DAPI (blue) staining of the nuclei within the infarct zone at 21 days after MSC transplantation. (f), Infarct size at baseline and at 21 days after MSC transplantation, Arrowheads (blue) indicate the diastolic thickness of the LV posterior walls (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) over time in the two groups. *p<0.05 vs. the corresponding 1-day-post-AMI group; #p<0.05 vs. the control group at 21 days after cell transplantation.
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Figure 5: PMSNs-siCCR2-PEI enhanced the efficiency of MSC transplantation and attenuated myocardial remodeling (n=12). (a), The surviving MSCs in the infarct zone following cell therapy are shown by representative immunofluorescent staining for EdU (5-Ethynyl-2'-deoxyuridine, green) and visualization of the nuclei (DAPI, blue) within the infarct zone 3 days after cell transplantation. Scale bar=50 μm. (b), SDF-1 and Ang-1 mRNA levels in the infarct zone after MSC transplantation. The data represent the mean±SD, *p<0.05 vs. the corresponding group at 1 day post-AMI; #p<0.05 vs. the control group at 3 days after cell transplantation. (c), Representative image of immunofluorescent staining for TUNEL (Alexa Fluor 488, green) and DAPI in the infarct border zone 3 days after AMI; the number of TUNEL-positive cardiomyocytes was calculated. Scale bar=50 μm. (d), The vascular density was determined by immunofluorescent staining for wheat germ agglutinin (WGA, red), endothelial cells (staining of CD31, green) and DAPI (blue) in the infarct zone at 21 days after MSC transplantation. (e), Confocal image of representative immunofluorescent staining for cardiac myosin heavy chain (eFluor® 660, red) and DAPI (blue) staining of the nuclei within the infarct zone at 21 days after MSC transplantation. (f), Infarct size at baseline and at 21 days after MSC transplantation, Arrowheads (blue) indicate the diastolic thickness of the LV posterior walls (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) over time in the two groups. *p<0.05 vs. the corresponding 1-day-post-AMI group; #p<0.05 vs. the control group at 21 days after cell transplantation.

Mentions: A current limitation of MSC therapy is the impact of the inflammatory microenvironment on cell behavior 25. We next tested the therapeutic potential of MSCs in the context of PMSN-siCCR2 treatment. After infarction, the hearts were harvested and embedded in paraffin, and the sections were incubated with azide-conjugated Alexa Fluor 488. We then observed a significant difference in the number of EdU-positive cells (Fig. 5a) in the infarction (13±3/mm2 vs. 4±1/mm2, p<0.001) between the two groups three days after MSC transplantation. In addition, increased levels of SDF-1 (0.047±0.023 vs. 0.025±0.014, p<0.001) and Ang-1 (1.05±0.51 vs. 0.54±0.29, p<0.001) mRNA were identified in the infarct zone of the siCCR2 group (Fig. 5b).


Photoluminescent Mesoporous Silicon Nanoparticles with siCCR2 Improve the Effects of Mesenchymal Stromal Cell Transplantation after Acute Myocardial Infarction.

Lu W, Xie Z, Tang Y, Bai L, Yao Y, Fu C, Ma G - Theranostics (2015)

PMSNs-siCCR2-PEI enhanced the efficiency of MSC transplantation and attenuated myocardial remodeling (n=12). (a), The surviving MSCs in the infarct zone following cell therapy are shown by representative immunofluorescent staining for EdU (5-Ethynyl-2'-deoxyuridine, green) and visualization of the nuclei (DAPI, blue) within the infarct zone 3 days after cell transplantation. Scale bar=50 μm. (b), SDF-1 and Ang-1 mRNA levels in the infarct zone after MSC transplantation. The data represent the mean±SD, *p<0.05 vs. the corresponding group at 1 day post-AMI; #p<0.05 vs. the control group at 3 days after cell transplantation. (c), Representative image of immunofluorescent staining for TUNEL (Alexa Fluor 488, green) and DAPI in the infarct border zone 3 days after AMI; the number of TUNEL-positive cardiomyocytes was calculated. Scale bar=50 μm. (d), The vascular density was determined by immunofluorescent staining for wheat germ agglutinin (WGA, red), endothelial cells (staining of CD31, green) and DAPI (blue) in the infarct zone at 21 days after MSC transplantation. (e), Confocal image of representative immunofluorescent staining for cardiac myosin heavy chain (eFluor® 660, red) and DAPI (blue) staining of the nuclei within the infarct zone at 21 days after MSC transplantation. (f), Infarct size at baseline and at 21 days after MSC transplantation, Arrowheads (blue) indicate the diastolic thickness of the LV posterior walls (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) over time in the two groups. *p<0.05 vs. the corresponding 1-day-post-AMI group; #p<0.05 vs. the control group at 21 days after cell transplantation.
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Figure 5: PMSNs-siCCR2-PEI enhanced the efficiency of MSC transplantation and attenuated myocardial remodeling (n=12). (a), The surviving MSCs in the infarct zone following cell therapy are shown by representative immunofluorescent staining for EdU (5-Ethynyl-2'-deoxyuridine, green) and visualization of the nuclei (DAPI, blue) within the infarct zone 3 days after cell transplantation. Scale bar=50 μm. (b), SDF-1 and Ang-1 mRNA levels in the infarct zone after MSC transplantation. The data represent the mean±SD, *p<0.05 vs. the corresponding group at 1 day post-AMI; #p<0.05 vs. the control group at 3 days after cell transplantation. (c), Representative image of immunofluorescent staining for TUNEL (Alexa Fluor 488, green) and DAPI in the infarct border zone 3 days after AMI; the number of TUNEL-positive cardiomyocytes was calculated. Scale bar=50 μm. (d), The vascular density was determined by immunofluorescent staining for wheat germ agglutinin (WGA, red), endothelial cells (staining of CD31, green) and DAPI (blue) in the infarct zone at 21 days after MSC transplantation. (e), Confocal image of representative immunofluorescent staining for cardiac myosin heavy chain (eFluor® 660, red) and DAPI (blue) staining of the nuclei within the infarct zone at 21 days after MSC transplantation. (f), Infarct size at baseline and at 21 days after MSC transplantation, Arrowheads (blue) indicate the diastolic thickness of the LV posterior walls (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) over time in the two groups. *p<0.05 vs. the corresponding 1-day-post-AMI group; #p<0.05 vs. the control group at 21 days after cell transplantation.
Mentions: A current limitation of MSC therapy is the impact of the inflammatory microenvironment on cell behavior 25. We next tested the therapeutic potential of MSCs in the context of PMSN-siCCR2 treatment. After infarction, the hearts were harvested and embedded in paraffin, and the sections were incubated with azide-conjugated Alexa Fluor 488. We then observed a significant difference in the number of EdU-positive cells (Fig. 5a) in the infarction (13±3/mm2 vs. 4±1/mm2, p<0.001) between the two groups three days after MSC transplantation. In addition, increased levels of SDF-1 (0.047±0.023 vs. 0.025±0.014, p<0.001) and Ang-1 (1.05±0.51 vs. 0.54±0.29, p<0.001) mRNA were identified in the infarct zone of the siCCR2 group (Fig. 5b).

Bottom Line: The therapeutic effects of PMSNs-siCCR2 for MSC transplantation were determined at the mRNA, protein and functional levels.In addition, significant amelioration of left ventricular (LV) remodeling (thickness of the LV posterior walls) (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) was also observed at the same time compared with the control group.These results suggest that PMSNs-siCCR2 could potentially be used to develop an anti-inflammatory therapy for post-AMI MSC transplantation.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Cardiology, ZhongDa Hospital affiliated with Southeast University, China; ; 2. Department of Cardiology, the Second Hospital affiliated with Southeast University, China;

ABSTRACT

Background: Despite the benefits of mesenchymal stromal cell (MSC) transplantation in cardiac tissue, detailed in vivo observations have shown that MSCs only survive for a brief period after transplantation due to harsh microenvironmental conditions, including ischemia, inflammation and anoikis, in the infarcted myocardium. Thus, new strategies are needed to enhance MSC survival and inhibit cardiac remodeling. Studies have now demonstrated that chemokine [C-C motif] ligand 2 (CCL2) and its cognate receptor C-C chemokine receptor 2 (CCR2) promote excessive Ly6C(high) inflammatory monocyte infiltration at the infarct in response to ischemic myocardial injury. Therefore, decreasing the activities of these monocytes immediately after acute myocardial infarction (AMI) could be beneficial for AMI patients.

Objectives: This study tested the hypothesis that therapeutic siRNA-loaded photoluminescent mesoporous silicon nanoparticles (PMSNs) targeting CCR2 expression in Ly6C(high) inflammatory monocytes decrease the accumulation of these cells in the infarct, improve the efficacy of MSC transplantation and attenuate myocardial remodeling.

Methods: PMSNs carrying therapeutic siCCR2 were first synthesized without the inclusion of fluorescent materials or dyes. After AMI BALB/c mice were established, 10(5) 5-ethynyl-2'- deoxyuridine (EdU)-labeled MSCs suspended in 100 µl of phosphate buffered saline (PBS) were injected into the border zone of the infarct of each mouse. PMSNs-siCCR2 (25 µg/g) were also intravenously injected via the tail vein immediately following AMI induction. Control mice were injected with an equal amount of PMSNs without siCCR2. PMSNs-siCCR2 were examined in vivo using near-infrared imaging technology. The therapeutic effects of PMSNs-siCCR2 for MSC transplantation were determined at the mRNA, protein and functional levels.

Results: PMSNs-siCCR2 circulated freely in vivo and were cleared in a relatively short period of time (t(½)=37 min) with no evidence of toxicity. The therapeutic PMSNs-siCCR2 showed higher levels of cellular accumulation in Ly6C(high) monocytes in the spleen and more efficient degradation of CCR2 compared with the control (8.04%±2.17% vs. 20.02%±4.55%, p<0.001). Subsequently, the PMSNs-siCCR2 decreased the accumulation of CD11b-positive monocytes at the infarct (49.3%±17.34% vs. 61.32%±22.43%, p<0.001) on day 1. Increased survival of transplanted MSCs (13±3/mm(2) vs. 4±1/mm(2), p<0.001) and significantly decreased TdT-mediated dUTP nick end labeling (TUNEL)(+) cardiac myocytes (17.44%±6.26% vs. 39.49%±13.28%, p<0.001) were then identified in the infarct zone three days after AMI induction in the PMSNs-siCCR2 group. Three weeks after MSC injection, significant increases were observed in the vascular density (235.5±39.6/mm(2) vs. 147.4±20.3/mm(2), p<0.001) and the cardiac myosin-positive area (21.7%±8.4% vs. 13.2%±4.4%, p<0.001) of the infarct border zone. In addition, significant amelioration of left ventricular (LV) remodeling (thickness of the LV posterior walls) (0.84±0.11 mm vs. 0.61±0.08 mm, p<0.001) was also observed at the same time compared with the control group.

Conclusions: PMSNs-siCCR2-mediated CCR2 gene silencing in Ly6C(high) monocytes improved the effectiveness of MSC transplantation and selectively ameliorated myocardial remodeling after AMI. These results suggest that PMSNs-siCCR2 could potentially be used to develop an anti-inflammatory therapy for post-AMI MSC transplantation.

No MeSH data available.


Related in: MedlinePlus