Limits...
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

Biocompatibility of PMSNs-siCCR2-PEI and monocytes. (a), Determination of the cell viability of monocytes by the MTT assay after incubation with PMSNs-siCCR2-PEI at a concentration of 0.02-0.1 µg/ul; no significant difference was shown at any of the concentrations tested (p=NS). (b), Comparison of the phagocytic capacity of Ly6Chigh/low monocytes for PMSNs-siCCR2-PEI, determined by immunofluorescence staining and flow cytometry. (c), Migration assay of cultured Ly-6Chigh monocytes using CCL2 (10 ng/ml) as a chemoattractant. Cells were harvested after incubation with PMSNs-siCCR2-PEI for 48 h. (d), Analysis of CCR2 expression on Ly-6Chigh monocytes by flow cytometry (n=12).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4508497&req=5

Figure 2: Biocompatibility of PMSNs-siCCR2-PEI and monocytes. (a), Determination of the cell viability of monocytes by the MTT assay after incubation with PMSNs-siCCR2-PEI at a concentration of 0.02-0.1 µg/ul; no significant difference was shown at any of the concentrations tested (p=NS). (b), Comparison of the phagocytic capacity of Ly6Chigh/low monocytes for PMSNs-siCCR2-PEI, determined by immunofluorescence staining and flow cytometry. (c), Migration assay of cultured Ly-6Chigh monocytes using CCL2 (10 ng/ml) as a chemoattractant. Cells were harvested after incubation with PMSNs-siCCR2-PEI for 48 h. (d), Analysis of CCR2 expression on Ly-6Chigh monocytes by flow cytometry (n=12).

Mentions: We first tested the biocompatibility of PMSNs-siCCR2-PEI in vitro. CD11b-positive monocytes obtained from spleens and cultured in vitro were incubated with increasing concentrations of the particles for 48 h, and the cell viability was quantified using an MTT assay. The results showed that the delivery formulation is relatively non-toxic (p=NS) to CD11b-positive monocytes (Fig. 2a). We next analyzed the toxicity of the particles to mice in vivo. Consistent with the results obtained in vitro, the hepatic and renal function of the AMI mice treated with or without PMSNs-siCCR2-PEI were not significantly different at 24 hours (Supplementary Table 1). The phagocytic capacity of Ly6Chigh/low monocytes for the particle was subsequently determined by fluorescence microscopy and flow cytometry. As shown in Fig. 2b, Ly6Chigh monocytes exhibited greater phagocytosis of the particles compared with Ly6Clow monocytes (89.2%±17.5% vs. 26.5%±6.4%, p<0.001). After the cells phagocytized the PMSNs-siCCR2-PEI, a transwell migration assay was introduced to test the migratory response of the cells to CCL2 (10 ng/ml). We then observed a significant decrease in the migratory response of the monocytes compared with the control, as shown in Fig. 2c (13±4/mm2 vs. 35±11/mm2, p<0.001). Finally, the expression of the chemokine receptor CCR2 on the monocytes was examined. Consistent with the results of the migratory assays, a significant decrease in CCR2 expression was observed in the PMSNs-siCCR2 group (Fig. 2d) compared with the control (19.6%±8.4% vs. 37.1%±13.5%, p<0.001).


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)

Biocompatibility of PMSNs-siCCR2-PEI and monocytes. (a), Determination of the cell viability of monocytes by the MTT assay after incubation with PMSNs-siCCR2-PEI at a concentration of 0.02-0.1 µg/ul; no significant difference was shown at any of the concentrations tested (p=NS). (b), Comparison of the phagocytic capacity of Ly6Chigh/low monocytes for PMSNs-siCCR2-PEI, determined by immunofluorescence staining and flow cytometry. (c), Migration assay of cultured Ly-6Chigh monocytes using CCL2 (10 ng/ml) as a chemoattractant. Cells were harvested after incubation with PMSNs-siCCR2-PEI for 48 h. (d), Analysis of CCR2 expression on Ly-6Chigh monocytes by flow cytometry (n=12).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4508497&req=5

Figure 2: Biocompatibility of PMSNs-siCCR2-PEI and monocytes. (a), Determination of the cell viability of monocytes by the MTT assay after incubation with PMSNs-siCCR2-PEI at a concentration of 0.02-0.1 µg/ul; no significant difference was shown at any of the concentrations tested (p=NS). (b), Comparison of the phagocytic capacity of Ly6Chigh/low monocytes for PMSNs-siCCR2-PEI, determined by immunofluorescence staining and flow cytometry. (c), Migration assay of cultured Ly-6Chigh monocytes using CCL2 (10 ng/ml) as a chemoattractant. Cells were harvested after incubation with PMSNs-siCCR2-PEI for 48 h. (d), Analysis of CCR2 expression on Ly-6Chigh monocytes by flow cytometry (n=12).
Mentions: We first tested the biocompatibility of PMSNs-siCCR2-PEI in vitro. CD11b-positive monocytes obtained from spleens and cultured in vitro were incubated with increasing concentrations of the particles for 48 h, and the cell viability was quantified using an MTT assay. The results showed that the delivery formulation is relatively non-toxic (p=NS) to CD11b-positive monocytes (Fig. 2a). We next analyzed the toxicity of the particles to mice in vivo. Consistent with the results obtained in vitro, the hepatic and renal function of the AMI mice treated with or without PMSNs-siCCR2-PEI were not significantly different at 24 hours (Supplementary Table 1). The phagocytic capacity of Ly6Chigh/low monocytes for the particle was subsequently determined by fluorescence microscopy and flow cytometry. As shown in Fig. 2b, Ly6Chigh monocytes exhibited greater phagocytosis of the particles compared with Ly6Clow monocytes (89.2%±17.5% vs. 26.5%±6.4%, p<0.001). After the cells phagocytized the PMSNs-siCCR2-PEI, a transwell migration assay was introduced to test the migratory response of the cells to CCL2 (10 ng/ml). We then observed a significant decrease in the migratory response of the monocytes compared with the control, as shown in Fig. 2c (13±4/mm2 vs. 35±11/mm2, p<0.001). Finally, the expression of the chemokine receptor CCR2 on the monocytes was examined. Consistent with the results of the migratory assays, a significant decrease in CCR2 expression was observed in the PMSNs-siCCR2 group (Fig. 2d) compared with the control (19.6%±8.4% vs. 37.1%±13.5%, p<0.001).

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