- Email: [email protected]
The mTOR inhibitor rapamycin inhibits proadhesive activity of arterial intimal smooth muscle cells
S102
Vascular Surgery I
The mTOR inhibitor rapamycin inhibits proadhesive activity of arterial intimal smooth muscle cells J Daniel Day MD, Jeffery L Walding MD, Benjamin Christie MD, Jing Kang MD, Martin L Dalton MD, Zhongbiao Wang MD Department of Surgery, Mercer University School of Medicine and the Medical, Macon, GA INTRODUCTION: Balloon catheter injury causes de-endothelialization and neointimal hyperplasia. Intimal smooth muscle cells (I-SMCs), forming a pseudoendothelium in the endotheliumdenuded vessels, continue to express high levels of adhesive molecules P-selectin and monocyte chemotactic protein-1 (MCP-1) and to recruit mononuclear cells, thus perpetuating the inflammatory response. Since rapamycin, an inhibitor of mammalian target of rapamycin (mTOR), can attenuate inflammation after vascular intervention, we investigated whether rapamycin would affect the proadhesive activity of I-SMCs, which are distinctly different from normal medial SMCs (M-SMCs). METHODS: I-SMCs and M-SMCs were cultured from ballooninjured rat arteries and from unmanipulated arteries. The cells were incubated in medium with or without rapamycin (10 ng/ml) for 24 hours. Phosphorylated mTOR, MCP-1 and P-selectin were detected with ELISA. Proadhesive activity of the cells was assessed by quantitating adhesion of Jurkat T-cells to SMCs. RESULTS: Higher levels of phosphorylated mTOR, P-selectin and MCP-1 were found in I-SMCs than in M-SMCs. Consistent with levels of the adhesion molecules, more Jurkat T-cells adhered to I-SMCs than to M-SMC. Treatment of I-SMCs with rapamycin reduced constitutive mTOR phosphorylation, expression of P-selectin and MCP-1, and adhesion of Jurkat T-cells. Furthermore, antibody to P-selectin inhibited the adhesion whereas antibody to MCP-1 had little effect. CONCLUSIONS: I-SMCs are a highly proadhesive phenotype. Rapamycin can inhibit expression of P-slectin and MCP-1, and adhesion of mononuclear cells to I-SMC through reducing expression of P-selectin. It is suggested that rapamycin might reduce inflammation response after vascular intervention through inhibiting the proadhesive activity of I-SMCs.
J Am Coll Surg
METHODS: Microarray experiments utilizing rabbit VG samples suggested extremely high IL-1beta mRNA transcription induction early (24h; confirmed by real time PCR) compared to thousands of other genes. IL-1beta protein localized to the developing neointima. Using IL-1RI⫺/⫺ (B6129 background) and wild-type (IL1RI⫹/⫹) mice, we investigated VG morphological changes 4 weeks after isograft from donor inferior vena cava to recipient interposition carotid artery (cuff anastomotic technique). RESULTS: No significant differences were observed on the traditional endpoints of neointimal area and calculated luminal area, yet clear histologic differentials between the VG walls existed. On crosssection, the mean thickness of the vascular wall outside the internal elastic laminae of IL-1RI⫺/⫺ mice was 44% smaller (59.1⫾28.6 m [n⫽9] vs 104.7⫾37.7 m [n⫽10]; p⬍0.01) than IL-1RI⫹/⫹ mice. The neointimal/circumferential thickness ratios tended to be higher in the IL-1RI⫺/⫺ animals (1.1⫾0.8 vs 0.5⫾0.2; p⫽0.06). CONCLUSIONS: The early VG wall exhibits high IL-1beta expression. When lacking IL-1 signaling, the VG wall adapts differently compared to controls (with a decreased outer wall thickness), directly emphasizing the relevance of IL-1 signaling in VG adaptations.
In vivo evaluation of a magnetic stent based drug delivery system Zachary G Forbes PhD, Frederick R Stoddard MD, Shami Jagtap MD, David Hansberry BS, Benjamin B Yellen PhD, Kenneth A Barbee PhD, Gennady Friedman PhD, Ari D Brooks MD Drexel University College of Medicine, Philadelphia, PA INTRODUCTION: Biological polymers have been used for controlled drug release. Such vehicles often lack a mechanism for site specific delivery. We are developing a drug delivery system using magnetic stents to capture magnetically susceptible drug carriers loaded with therapeutic agents. This study is the first demonstration of this principle in-vivo.
Peng Yu MD, Zhihua Jiang MD, PhD, Ming Tao MD, Scott A Berceli MD, PhD, C Keith Ozaki MD University of Florida College of Medicine and Malcom Randall VAMC, Gainesville, FL
METHODS: Magnetic stents were surgically implanted into the common Iliac arteries of Sprague Dawley rats. Intra-aortic injections of 2.5mg of Technetium-99m labeled, 350nm diameter poly(propyleneglycol)-coated magnetic particles were administered during exposure to a 0.1T magnetic field. Particles circulated for 5 minutes under the magnetic field, followed by sacrifice and organ/ stent harvest. Labeled particle activity was used to determine capture efficiency and biodistribution of uncaptured particles. Stent efficiency was calculated as number of particles captured per 10,000 particles injected (ppt) and biodistribution as particles per gram of tissue per 10,000 particles injected (ppgpt).
INTRODUCTION: Vein grafts (VG) fail due to wall maladaptations to surgical injury and dramatic hemodynamic perturbations. Interleukin-1 (IL-1) has emerged as an important mediator of hemodynamically induced arterial vascular lesions. We therefore hypothesized that IL-1 drives early VG wall adaptations. Specifically, we explored the temporal expression of IL-1beta after VG arterialization, and utilized mice lacking functional IL-1 Type I receptor (IL1RI⫺/⫺) to delineate the impact of deficient IL-1 signaling on early VG wall adaptations.
RESULTS: Experiments comprising 15 rats demonstrated considerably higher capture of particles to magnetizable stents within a field when compared to non-magnetic stents without a field (3.05⫹/ ⫺1.08ppt vs. 0.34ppt). field when compared to non-magnetic stents without a field (3.05⫹/⫺1.08ppt vs. 0.34ppt). Particle distribution peaked in the liver (374.9⫹/⫺78.1ppgpt) and spleen (159.9⫹/ ⫺67.9ppgpt) with lower particle counts in lungs (88.9⫹/ ⫺59.0ppgpt), blood (50.4⫹/⫺18.2ppgpt), kidney (32.1⫹/ ⫺11.5ppgpt) and heart (12.8⫹/⫺14.1ppgpt).
IL-1 signaling in the early vein graft—Impact of IL-1 type I receptor deficiency
Vascular Surgery I
The mTOR inhibitor rapamycin inhibits proadhesive activity of arterial intimal smooth muscle cells J Daniel Day MD, Jeffery L Walding MD, Benjamin Christie MD, Jing Kang MD, Martin L Dalton MD, Zhongbiao Wang MD Department of Surgery, Mercer University School of Medicine and the Medical, Macon, GA INTRODUCTION: Balloon catheter injury causes de-endothelialization and neointimal hyperplasia. Intimal smooth muscle cells (I-SMCs), forming a pseudoendothelium in the endotheliumdenuded vessels, continue to express high levels of adhesive molecules P-selectin and monocyte chemotactic protein-1 (MCP-1) and to recruit mononuclear cells, thus perpetuating the inflammatory response. Since rapamycin, an inhibitor of mammalian target of rapamycin (mTOR), can attenuate inflammation after vascular intervention, we investigated whether rapamycin would affect the proadhesive activity of I-SMCs, which are distinctly different from normal medial SMCs (M-SMCs). METHODS: I-SMCs and M-SMCs were cultured from ballooninjured rat arteries and from unmanipulated arteries. The cells were incubated in medium with or without rapamycin (10 ng/ml) for 24 hours. Phosphorylated mTOR, MCP-1 and P-selectin were detected with ELISA. Proadhesive activity of the cells was assessed by quantitating adhesion of Jurkat T-cells to SMCs. RESULTS: Higher levels of phosphorylated mTOR, P-selectin and MCP-1 were found in I-SMCs than in M-SMCs. Consistent with levels of the adhesion molecules, more Jurkat T-cells adhered to I-SMCs than to M-SMC. Treatment of I-SMCs with rapamycin reduced constitutive mTOR phosphorylation, expression of P-selectin and MCP-1, and adhesion of Jurkat T-cells. Furthermore, antibody to P-selectin inhibited the adhesion whereas antibody to MCP-1 had little effect. CONCLUSIONS: I-SMCs are a highly proadhesive phenotype. Rapamycin can inhibit expression of P-slectin and MCP-1, and adhesion of mononuclear cells to I-SMC through reducing expression of P-selectin. It is suggested that rapamycin might reduce inflammation response after vascular intervention through inhibiting the proadhesive activity of I-SMCs.
J Am Coll Surg
METHODS: Microarray experiments utilizing rabbit VG samples suggested extremely high IL-1beta mRNA transcription induction early (24h; confirmed by real time PCR) compared to thousands of other genes. IL-1beta protein localized to the developing neointima. Using IL-1RI⫺/⫺ (B6129 background) and wild-type (IL1RI⫹/⫹) mice, we investigated VG morphological changes 4 weeks after isograft from donor inferior vena cava to recipient interposition carotid artery (cuff anastomotic technique). RESULTS: No significant differences were observed on the traditional endpoints of neointimal area and calculated luminal area, yet clear histologic differentials between the VG walls existed. On crosssection, the mean thickness of the vascular wall outside the internal elastic laminae of IL-1RI⫺/⫺ mice was 44% smaller (59.1⫾28.6 m [n⫽9] vs 104.7⫾37.7 m [n⫽10]; p⬍0.01) than IL-1RI⫹/⫹ mice. The neointimal/circumferential thickness ratios tended to be higher in the IL-1RI⫺/⫺ animals (1.1⫾0.8 vs 0.5⫾0.2; p⫽0.06). CONCLUSIONS: The early VG wall exhibits high IL-1beta expression. When lacking IL-1 signaling, the VG wall adapts differently compared to controls (with a decreased outer wall thickness), directly emphasizing the relevance of IL-1 signaling in VG adaptations.
In vivo evaluation of a magnetic stent based drug delivery system Zachary G Forbes PhD, Frederick R Stoddard MD, Shami Jagtap MD, David Hansberry BS, Benjamin B Yellen PhD, Kenneth A Barbee PhD, Gennady Friedman PhD, Ari D Brooks MD Drexel University College of Medicine, Philadelphia, PA INTRODUCTION: Biological polymers have been used for controlled drug release. Such vehicles often lack a mechanism for site specific delivery. We are developing a drug delivery system using magnetic stents to capture magnetically susceptible drug carriers loaded with therapeutic agents. This study is the first demonstration of this principle in-vivo.
Peng Yu MD, Zhihua Jiang MD, PhD, Ming Tao MD, Scott A Berceli MD, PhD, C Keith Ozaki MD University of Florida College of Medicine and Malcom Randall VAMC, Gainesville, FL
METHODS: Magnetic stents were surgically implanted into the common Iliac arteries of Sprague Dawley rats. Intra-aortic injections of 2.5mg of Technetium-99m labeled, 350nm diameter poly(propyleneglycol)-coated magnetic particles were administered during exposure to a 0.1T magnetic field. Particles circulated for 5 minutes under the magnetic field, followed by sacrifice and organ/ stent harvest. Labeled particle activity was used to determine capture efficiency and biodistribution of uncaptured particles. Stent efficiency was calculated as number of particles captured per 10,000 particles injected (ppt) and biodistribution as particles per gram of tissue per 10,000 particles injected (ppgpt).
INTRODUCTION: Vein grafts (VG) fail due to wall maladaptations to surgical injury and dramatic hemodynamic perturbations. Interleukin-1 (IL-1) has emerged as an important mediator of hemodynamically induced arterial vascular lesions. We therefore hypothesized that IL-1 drives early VG wall adaptations. Specifically, we explored the temporal expression of IL-1beta after VG arterialization, and utilized mice lacking functional IL-1 Type I receptor (IL1RI⫺/⫺) to delineate the impact of deficient IL-1 signaling on early VG wall adaptations.
RESULTS: Experiments comprising 15 rats demonstrated considerably higher capture of particles to magnetizable stents within a field when compared to non-magnetic stents without a field (3.05⫹/ ⫺1.08ppt vs. 0.34ppt). field when compared to non-magnetic stents without a field (3.05⫹/⫺1.08ppt vs. 0.34ppt). Particle distribution peaked in the liver (374.9⫹/⫺78.1ppgpt) and spleen (159.9⫹/ ⫺67.9ppgpt) with lower particle counts in lungs (88.9⫹/ ⫺59.0ppgpt), blood (50.4⫹/⫺18.2ppgpt), kidney (32.1⫹/ ⫺11.5ppgpt) and heart (12.8⫹/⫺14.1ppgpt).
IL-1 signaling in the early vein graft—Impact of IL-1 type I receptor deficiency