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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

The effect of PMSNs-siCCR2 on inflammatory infiltration post-AMI (n=12). (a), Flow cytometry reveals a different pattern of CD11b+Ly6Chigh in the circulation and the spleen of AMI mice one day after treatment with PMSNs-siCCR2. Significantly reduced counts of the cells were shown in the siCCR2 group. (b), Flow cytometry revealed reduced CD11b-positive cells in the infarct zones of AMI mice one day after treatment with PMSNs-siCCR2. The expression of CCR2 in the infarction was further determined by Western-blot (A1~3 in the PMSNs-siCCR2 group, B1~3 in the control). (c), FACS analysis of the CCR2 protein levels on the spleen CD11b+Ly6Chigh monocytes of the AMI mice confirmed the decreased levels of CCR2 in the PMSN-siCCR2 group. (d), Levels of the inflammatory cytokines IL-1, IL-6, TNF-α and IFN-γ in the serum of AMI mice after treatment with PMSNs-siCCR2 were determined using immunosorbent assays (ELISAs), *p<0.05 vs. the corresponding control group.
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Figure 4: The effect of PMSNs-siCCR2 on inflammatory infiltration post-AMI (n=12). (a), Flow cytometry reveals a different pattern of CD11b+Ly6Chigh in the circulation and the spleen of AMI mice one day after treatment with PMSNs-siCCR2. Significantly reduced counts of the cells were shown in the siCCR2 group. (b), Flow cytometry revealed reduced CD11b-positive cells in the infarct zones of AMI mice one day after treatment with PMSNs-siCCR2. The expression of CCR2 in the infarction was further determined by Western-blot (A1~3 in the PMSNs-siCCR2 group, B1~3 in the control). (c), FACS analysis of the CCR2 protein levels on the spleen CD11b+Ly6Chigh monocytes of the AMI mice confirmed the decreased levels of CCR2 in the PMSN-siCCR2 group. (d), Levels of the inflammatory cytokines IL-1, IL-6, TNF-α and IFN-γ in the serum of AMI mice after treatment with PMSNs-siCCR2 were determined using immunosorbent assays (ELISAs), *p<0.05 vs. the corresponding control group.

Mentions: Ly6Chigh monocytes are potent inflammatory mediators and are believed to be the dominant source of inflammation after AMI; therefore, we tested the deployment of Ly6Chigh monocytes after AMI and PMSN-siCCR2 administration. Flow cytometric analysis showed a significant reduction in the number of inflammatory Ly6Chigh monocytes (43.62%±11.32% vs. 67.95%±17.04%, p<0.001) in the circulation one day after AMI in mice treated with PMSNs-siCCR2-PEI compared with the control group. The number of cells present in the spleen also changed significantly between the groups (29.78%±4.38% vs. 20.06%± 4.25%, p<0.001) (Fig. 4a). CD11b-positive monocytes in the infarcts were further examined at the same time, and the results revealed that PMSNs-siCCR2-PEI could significantly reduce the levels of the cells compared with the groups not given siCCR2 (49.3%±17.34% vs. 61.32%± 22.43%, p<0.001) (Fig. 4b). These findings were further confirmed by the decreased levels of CCR2 protein expression on spleen monocytes, as examined by flow cytometry (8.04%±2.07% vs. 20.02%±4.55%, p<0.001) and Western blot (1.78±0.47 vs. 2.71±0.69, p<0.001) (Fig. 4c), and the decreased expression of inflammatory cytokines (pg/ml), including IL-1 (157.49±62.36 vs. 233.37±45.78, p<0.001), IL-6 (207.88±73.3 vs. 298.62±75.5, p<0.001), TNF-α (105.74±33.87 vs. 161.09±50.24, p<0.001) and IFN-γ (131.07±40.34 vs. 197.98±49.77, p<0.001), in the serum, as detected by ELISA kits (Fig. 4d).


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)

The effect of PMSNs-siCCR2 on inflammatory infiltration post-AMI (n=12). (a), Flow cytometry reveals a different pattern of CD11b+Ly6Chigh in the circulation and the spleen of AMI mice one day after treatment with PMSNs-siCCR2. Significantly reduced counts of the cells were shown in the siCCR2 group. (b), Flow cytometry revealed reduced CD11b-positive cells in the infarct zones of AMI mice one day after treatment with PMSNs-siCCR2. The expression of CCR2 in the infarction was further determined by Western-blot (A1~3 in the PMSNs-siCCR2 group, B1~3 in the control). (c), FACS analysis of the CCR2 protein levels on the spleen CD11b+Ly6Chigh monocytes of the AMI mice confirmed the decreased levels of CCR2 in the PMSN-siCCR2 group. (d), Levels of the inflammatory cytokines IL-1, IL-6, TNF-α and IFN-γ in the serum of AMI mice after treatment with PMSNs-siCCR2 were determined using immunosorbent assays (ELISAs), *p<0.05 vs. the corresponding control group.
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Related In: Results  -  Collection

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Figure 4: The effect of PMSNs-siCCR2 on inflammatory infiltration post-AMI (n=12). (a), Flow cytometry reveals a different pattern of CD11b+Ly6Chigh in the circulation and the spleen of AMI mice one day after treatment with PMSNs-siCCR2. Significantly reduced counts of the cells were shown in the siCCR2 group. (b), Flow cytometry revealed reduced CD11b-positive cells in the infarct zones of AMI mice one day after treatment with PMSNs-siCCR2. The expression of CCR2 in the infarction was further determined by Western-blot (A1~3 in the PMSNs-siCCR2 group, B1~3 in the control). (c), FACS analysis of the CCR2 protein levels on the spleen CD11b+Ly6Chigh monocytes of the AMI mice confirmed the decreased levels of CCR2 in the PMSN-siCCR2 group. (d), Levels of the inflammatory cytokines IL-1, IL-6, TNF-α and IFN-γ in the serum of AMI mice after treatment with PMSNs-siCCR2 were determined using immunosorbent assays (ELISAs), *p<0.05 vs. the corresponding control group.
Mentions: Ly6Chigh monocytes are potent inflammatory mediators and are believed to be the dominant source of inflammation after AMI; therefore, we tested the deployment of Ly6Chigh monocytes after AMI and PMSN-siCCR2 administration. Flow cytometric analysis showed a significant reduction in the number of inflammatory Ly6Chigh monocytes (43.62%±11.32% vs. 67.95%±17.04%, p<0.001) in the circulation one day after AMI in mice treated with PMSNs-siCCR2-PEI compared with the control group. The number of cells present in the spleen also changed significantly between the groups (29.78%±4.38% vs. 20.06%± 4.25%, p<0.001) (Fig. 4a). CD11b-positive monocytes in the infarcts were further examined at the same time, and the results revealed that PMSNs-siCCR2-PEI could significantly reduce the levels of the cells compared with the groups not given siCCR2 (49.3%±17.34% vs. 61.32%± 22.43%, p<0.001) (Fig. 4b). These findings were further confirmed by the decreased levels of CCR2 protein expression on spleen monocytes, as examined by flow cytometry (8.04%±2.07% vs. 20.02%±4.55%, p<0.001) and Western blot (1.78±0.47 vs. 2.71±0.69, p<0.001) (Fig. 4c), and the decreased expression of inflammatory cytokines (pg/ml), including IL-1 (157.49±62.36 vs. 233.37±45.78, p<0.001), IL-6 (207.88±73.3 vs. 298.62±75.5, p<0.001), TNF-α (105.74±33.87 vs. 161.09±50.24, p<0.001) and IFN-γ (131.07±40.34 vs. 197.98±49.77, p<0.001), in the serum, as detected by ELISA kits (Fig. 4d).

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