Neovascularization and angiogenesis in the brain are important physiological processes for normal brain development and repair/regeneration following insults. Integrins are cell surface adhesion receptors mediating im...Neovascularization and angiogenesis in the brain are important physiological processes for normal brain development and repair/regeneration following insults. Integrins are cell surface adhesion receptors mediating important function of cells such as survival, growth and development during tissue organization, differentiation and organogenesis. In this study, we used an integrin-binding array platform to identify the important types of integrins and their binding peptides that facilitate adhesion, growth, development, and vascular-like network formation of rat primary brain microvascular endothelial cells. Brain microvascular endothelial cells were isolated from rat brain on post-natal day 7. Cells were cultured in a custom-designed integrin array system containing short synthetic peptides binding to 16 types of integrins commonly expressed on cells in vertebrates. After 7 days of culture, the brain microvascular endothelial cells were processed for immunostaining with markers for endothelial cells including von Willibrand factor and platelet endothelial cell adhesion molecule. 5-Bromo-2′-dexoyuridine was added to the culture at 48 hours prior to fixation to assess cell proliferation. Among 16 integrins tested, we found that α5β1, αvβ5 and αvβ8 greatly promoted proliferation of endothelial cells in culture. To investigate the effect of integrin-binding peptides in promoting neovascularization and angiogenesis, the binding peptides to the above three types of integrins were immobilized to our custom-designed hydrogel in three-dimensional(3 D) culture of brain microvascular endothelial cells with the addition of vascular endothelial growth factor. Following a 7-day 3 D culture, the culture was fixed and processed for double labeling of phalloidin with von Willibrand factor or platelet endothelial cell adhesion molecule and assessed under confocal microscopy. In the 3 D culture in hydrogels conjugated with the integrin-binding peptide, brain microvascular endothelial cells formed interconnected vascular-like network with clearly discernable lumens, which is reminiscent of brain microvascular network in vivo. With the novel integrin-binding array system, we identified the specific types of integrins on brain microvascular endothelial cells that mediate cell adhesion and growth followed by functionalizing a 3 D hydrogel culture system using the binding peptides that specifically bind to the identified integrins, leading to robust growth and lumenized microvascular-like network formation of brain microvascular endothelial cells in 3 D culture. This technology can be used for in vitro and in vivo vascularization of transplants or brain lesions to promote brain tissue regeneration following neurological insults.展开更多
Biomaterial bridging provides physical substrates to guide axonal growth across the lesion.To achieve efficient directional guidance,combinatory strategies using permissive matrix,cells and trophic factors are necessa...Biomaterial bridging provides physical substrates to guide axonal growth across the lesion.To achieve efficient directional guidance,combinatory strategies using permissive matrix,cells and trophic factors are necessary.In the present study,we evaluated permissive effect of poly(acrylonitrile-co-vinyl chloride)guidance channels filled by different densities of laminin-precoated unidirectional polypropylene filaments combined with Schwann cells,and glial cell line-derived neurotrophic factor for axonal regeneration through a T10 hemisected spinal cord gap in adult rats.We found that channels with filaments significantly reduced the lesion cavity,astrocytic gliosis,and inflammatory responses at the graft-host boundaries.The laminin coated low density filament provided the most favorable directional guidance for axonal regeneration which was enhanced by co-grafting of Schwann cells and glial cell line-derived neurotrophic factor.These results demonstrate that the combinatorial strategy of filament-filled guiding scaffold,adhesive molecular laminin,Schwann cells,and glial cell line-derived neurotrophic factor,provides optimal topographical cues in stimulating directional axonal regeneration following spinal cord injury.This study was approved by Indiana University Institutional Animal Care and Use Committees(IACUC#:11011)on October 29,2015.展开更多
Vulnerable atherosclerotic plaques are responsible for most cardiovascular diseases(CVDs).Folate receptor(FR)positive activated macrophages were thought to be a prominent component in the development of vulnerable pla...Vulnerable atherosclerotic plaques are responsible for most cardiovascular diseases(CVDs).Folate receptor(FR)positive activated macrophages were thought to be a prominent component in the development of vulnerable plaque.The objective of this study is to develop folate conjugated two-dimensional(2D)Pd@Au nanomaterials(Pd@Au-PEG-FA)for targeted multimodal imaging of the FRs in advanced atherosclerotic plaques.Pharmacokinetic and imaging studies(single photon emission computed tomography(SPECT),computed tomography(CT)and photoacoustic(PA)imaging)were performed to confirm the prolonged blood half-life and enrichment of radioactivity in atherosclerotic plaques.Strong signals were detected in vivo with SPECT,CT and PA imaging in heavy atherosclerotic plaques,which were significantly higher than those of the normal aortas after injection of Pd@Au-PEG-FA.Blocking studies with preinjection of excess FA could effectively reduce the targeting ability of Pd@Au-PEG-FA in atherosclerotic plaques,further demonstrating the specific binding of Pd@Au-PEG-FA for plaque lesions.Histopathological characterization revealed that the signal of probe was in accordance with the high-risk plaques.In summary,the Pd@Au-PEG-FA has favorable pharmacokinetic properties and provides a valuable approach for detecting high-risk plaques in the presence of FRs in atherosclerotic plaques.展开更多
基金supported by NIH grant RO1 NS093985 (to DS, NZ, XW) and RO1 NS101955 (to DS)the VCU Microscopy Facility,supported,in part,by funding from NIH-NCI Cancer Center Support Grant P30 CA016059。
文摘Neovascularization and angiogenesis in the brain are important physiological processes for normal brain development and repair/regeneration following insults. Integrins are cell surface adhesion receptors mediating important function of cells such as survival, growth and development during tissue organization, differentiation and organogenesis. In this study, we used an integrin-binding array platform to identify the important types of integrins and their binding peptides that facilitate adhesion, growth, development, and vascular-like network formation of rat primary brain microvascular endothelial cells. Brain microvascular endothelial cells were isolated from rat brain on post-natal day 7. Cells were cultured in a custom-designed integrin array system containing short synthetic peptides binding to 16 types of integrins commonly expressed on cells in vertebrates. After 7 days of culture, the brain microvascular endothelial cells were processed for immunostaining with markers for endothelial cells including von Willibrand factor and platelet endothelial cell adhesion molecule. 5-Bromo-2′-dexoyuridine was added to the culture at 48 hours prior to fixation to assess cell proliferation. Among 16 integrins tested, we found that α5β1, αvβ5 and αvβ8 greatly promoted proliferation of endothelial cells in culture. To investigate the effect of integrin-binding peptides in promoting neovascularization and angiogenesis, the binding peptides to the above three types of integrins were immobilized to our custom-designed hydrogel in three-dimensional(3 D) culture of brain microvascular endothelial cells with the addition of vascular endothelial growth factor. Following a 7-day 3 D culture, the culture was fixed and processed for double labeling of phalloidin with von Willibrand factor or platelet endothelial cell adhesion molecule and assessed under confocal microscopy. In the 3 D culture in hydrogels conjugated with the integrin-binding peptide, brain microvascular endothelial cells formed interconnected vascular-like network with clearly discernable lumens, which is reminiscent of brain microvascular network in vivo. With the novel integrin-binding array system, we identified the specific types of integrins on brain microvascular endothelial cells that mediate cell adhesion and growth followed by functionalizing a 3 D hydrogel culture system using the binding peptides that specifically bind to the identified integrins, leading to robust growth and lumenized microvascular-like network formation of brain microvascular endothelial cells in 3 D culture. This technology can be used for in vitro and in vivo vascularization of transplants or brain lesions to promote brain tissue regeneration following neurological insults.
基金Research in the Xu laboratory is supported by NIH 1R01100531,1R01 NS103481Merit Review Award I01 BX002356,I01 BX003705,I01 RX002687 from the U.S.Department of Veterans AffairsMari Hulman George Endowment Funds.
文摘Biomaterial bridging provides physical substrates to guide axonal growth across the lesion.To achieve efficient directional guidance,combinatory strategies using permissive matrix,cells and trophic factors are necessary.In the present study,we evaluated permissive effect of poly(acrylonitrile-co-vinyl chloride)guidance channels filled by different densities of laminin-precoated unidirectional polypropylene filaments combined with Schwann cells,and glial cell line-derived neurotrophic factor for axonal regeneration through a T10 hemisected spinal cord gap in adult rats.We found that channels with filaments significantly reduced the lesion cavity,astrocytic gliosis,and inflammatory responses at the graft-host boundaries.The laminin coated low density filament provided the most favorable directional guidance for axonal regeneration which was enhanced by co-grafting of Schwann cells and glial cell line-derived neurotrophic factor.These results demonstrate that the combinatorial strategy of filament-filled guiding scaffold,adhesive molecular laminin,Schwann cells,and glial cell line-derived neurotrophic factor,provides optimal topographical cues in stimulating directional axonal regeneration following spinal cord injury.This study was approved by Indiana University Institutional Animal Care and Use Committees(IACUC#:11011)on October 29,2015.
基金supported by the National Postdoctoral Program for Innovative Talents(No.BX201700142)Postdoctoral Science Foundation of China(No.2018M630732)+3 种基金the National Natural Science Foundation of China(Nos.81901805,21906135,81471707,21705037,and 91539126)Hunan Provincial Natural Science Foundation of China(No.2018JJ3092)Chinese Academy of Medical Sciences Innovation Fund for Medical Sciences(CIFMS 2016-I2M-1-009)Drug Innovation Major Project(2018ZX09711001-003-011).
文摘Vulnerable atherosclerotic plaques are responsible for most cardiovascular diseases(CVDs).Folate receptor(FR)positive activated macrophages were thought to be a prominent component in the development of vulnerable plaque.The objective of this study is to develop folate conjugated two-dimensional(2D)Pd@Au nanomaterials(Pd@Au-PEG-FA)for targeted multimodal imaging of the FRs in advanced atherosclerotic plaques.Pharmacokinetic and imaging studies(single photon emission computed tomography(SPECT),computed tomography(CT)and photoacoustic(PA)imaging)were performed to confirm the prolonged blood half-life and enrichment of radioactivity in atherosclerotic plaques.Strong signals were detected in vivo with SPECT,CT and PA imaging in heavy atherosclerotic plaques,which were significantly higher than those of the normal aortas after injection of Pd@Au-PEG-FA.Blocking studies with preinjection of excess FA could effectively reduce the targeting ability of Pd@Au-PEG-FA in atherosclerotic plaques,further demonstrating the specific binding of Pd@Au-PEG-FA for plaque lesions.Histopathological characterization revealed that the signal of probe was in accordance with the high-risk plaques.In summary,the Pd@Au-PEG-FA has favorable pharmacokinetic properties and provides a valuable approach for detecting high-risk plaques in the presence of FRs in atherosclerotic plaques.
