scholarly journals Rapid and efficient immunomagnetic isolation of endothelial cells from human peripheral nerves

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Patrick Dömer ◽  
Janine Kayal ◽  
Ulrike Janssen-Bienhold ◽  
Bettina Kewitz ◽  
Thomas Kretschmer ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Peripheral Nerves ◽  
Peripheral Nerve Regeneration ◽  
The Novel ◽  
Isolated Cells ◽  
Nerve Grafts ◽  
Marker Proteins ◽  
Immunomagnetic Isolation ◽  
Human Nerve

AbstractEndothelial cells (ECs) have gained an increased scientific focus since they were reported to provide guidance for Schwann cells and subsequently following axons after nerve injuries. However, previous protocols for the isolation of nerve-derived ECs from human nerves are ineffective regarding time and yield. Therefore, we established a novel and efficient protocol for the isolation of ECs from human peripheral nerves by means of immunomagnetic CD31-antibody conjugated Dynabeads and assessed the purity of the isolated cells. The easy-to-follow and time-effective isolation method allows the isolation of > 95% pure ECs. The isolated ECs were shown to express highly specific EC marker proteins and revealed functional properties by formation of CD31 and VE-cadherin positive adherens junctions, as well as ZO-1 positive tight-junctions. Moreover, the formation of capillary EC-tubes was observed in-vitro. The novel protocol for the isolation of human nerve-derived ECs allows and simplifies the usage of ECs in research of the human blood-nerve-barrier and peripheral nerve regeneration. Additionally, a potential experimental application of patient-derived nerve ECs in the in-vitro vascularization of artificial nerve grafts is feasible.

scholarly journals Pluripotent Stem Cells Can Be Isolated from Human Peripheral Nerves after in vitro BMP-2 Stimulation

2019 ◽  
Author(s):  
Ren-Yi Sun ◽  
Michael H. Heggeness ◽  
Tanghong Jia ◽  
Sunaina Shrestha ◽  
Bradley Dart ◽  
...  
Keyword(s):  
Stem Cell ◽  
Transcription Factors ◽  
Peripheral Nerves ◽  
Nerve Tissue ◽  
Isolated Cells ◽  
Adult Mice ◽  
Stem Cell Medium ◽  
Natural Mechanism ◽  
Human Nerve

AbstractWe have recently identified a population of cells within the peripheral nerves of adult mice that can respond to BMP-2 exposure or physical injury to rapidly proliferate. More importantly, these cells exhibited embryonic differentiation potentials that could be induced into osteoblastic and endothelial cells in vitro. The current study examined human nerve specimens to compare and characterize the cells after BMP-2 stimulation. Fresh pieces of human nerve tissue were minced and treated with either BMP-2 (750ng/ml) or vehicle for 12 hours at 37°C, before digested in 0.2% collagenase and 0.05% trypsin-EDTA. Isolated cells were cultured in restrictive stem cell medium. Significantly more cells were obtained from the nerve pieces with BMP-2 treatment in comparison with the non-treated controls. Cell colonies were starting to form at day 3. Expressions of the 4 transcription factors Klf4, c-Myc, Sox2 and Oct4 were confirmed at both transcriptional and translational levels. The cells can be maintained in the stem cell culture medium for at least 6 weeks without changing morphologies. When the cells were switched to fibroblast growth medium, dispersed spindle-shaped cells were noted and became fibroblast activated protein-α (FAP) positive following immunocytochemistry staining. The data suggested that human peripheral nerve tissue also contain a population of cells that can respond to BMP-2 and express all four transcription factors KLF4, Sox2, cMyc, and Oct4. These cells are capable to differentiate into FAP-positive fibroblasts. It is proposed that these cells are possibly at the core of a previously unknown natural mechanism for healing injury.

Earthworm Extracts Facilitate PC12 Cell Differentiation and Promote Axonal Sprouting in Peripheral Nerve Injury

2010 ◽  
Vol 38 (03) ◽  
pp. 547-560 ◽  
Author(s):  
Chao-Tsung Chen ◽  
Jaung-Geng Lin ◽  
Tung-Wu Lu ◽  
Fuu-Jen Tsai ◽  
Chih-Yang Huang ◽  
...  
Keyword(s):  
Peripheral Nerve ◽  
Peripheral Nerves ◽  
Peripheral Nerve Regeneration ◽  
In Vitro Study ◽  
Potential Growth ◽  
Marked Enhancement ◽  
Growth Promoting ◽  
Success Percentage

The present study provides in vitro and in vivo evaluations of earthworm (Pheretima aspergilum) on peripheral nerve regeneration. In the in vitro study, we found the earthworm (EW) water extracts caused a marked enhancement of the nerve growth factor-mediated neurite outgrowth from PC12 cells as well as the expressions of growth associated protein 43 and synapsin I. In the in vivo study, silicone rubber chambers filled with EW extracts were used to bridge a 10 mm sciatic nerve defect in rats. Eight weeks after implantation, the group receiving EW extracts had a much higher success percentage of regeneration (90%) compared to the control (60%) receiving the saline. In addition, quantitative histology of the successfully regenerated nerves revealed that myelinated axons in EW group at 31.25 μg/ml was significantly more than those in the controls (p < 0.05). These results showed that EW extracts can be a potential growth-promoting factor on regenerating peripheral nerves.

scholarly journals The novel mitochondria-targeted hydrogen sulfide (H 2 S) donors AP123 and AP39 protect against hyperglycemic injury in microvascular endothelial cells in vitro

2016 ◽  
Vol 113 ◽  
pp. 186-198 ◽  
Author(s):  
Domokos Gerő ◽  
Roberta Torregrossa ◽  
Alexis Perry ◽  
Alicia Waters ◽  
Sophie Le-Trionnaire ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Hydrogen Sulfide ◽  
Microvascular Endothelial Cells ◽  
The Novel ◽  
Microvascular Endothelial

scholarly journals Biased agonism of protease-activated receptor 1 by activated protein C caused by noncanonical cleavage at Arg46

Blood ◽  
2012 ◽  
Vol 120 (26) ◽  
pp. 5237-5246 ◽  
Author(s):  
Laurent O. Mosnier ◽  
Ranjeet K. Sinha ◽  
Laurent Burnier ◽  
Eveline A. Bouwens ◽  
John H. Griffin
Keyword(s):  
Endothelial Cells ◽  
Protein C ◽  
Glycogen Synthase ◽  
Activated Protein C ◽  
The Novel ◽  
Biased Agonism ◽  
Protease Activated Receptor 1 ◽  
Protective Signaling

Abstract Activated protein C (APC) exerts endothelial cytoprotective actions that require protease-activated receptor 1 (PAR1), whereas thrombin acting via PAR1 causes endothelial disruptive, proinflammatory actions. APC's activities, but not thrombin's, require PAR1 located in caveolae. PAR1 is a biased 7-transmembrane receptor because G proteins mediate thrombin's signaling, whereas β-arrestin 2 mediates APC's signaling. Here we elucidate novel mechanisms for APC's initiation of signaling. Biochemical studies of APC's protease specificity showed that APC cleaved PAR1 sequences at both Arg41 and Arg46. That PAR1 cleavage at Arg46 can occur on cells was supported by APC's cleavage of N-terminal-SEAP-tagged R41Q-PAR1 but not R41Q/R46Q-PAR1 mutants transfected into cells and by anti-PAR1 epitope mapping of APC-treated endothelial cells. A synthetic peptide composing PAR1 residues 47-66, TR47, stimulated protective signaling in endothelial cells as reflected in Akt and glycogen synthase kinase 3β phosphorylation, Ras-related C3 botulinum toxin substrate 1 activation, and barrier stabilization effects. In mice, the TR47 peptide reduced VEGF-induced vascular leakage. These in vitro and in vivo data imply that the novel PAR1 N-terminus beginning at residue Asn47, which is generated by APC cleavage at Arg46, mediates APC's cytoprotective signaling and that this unique APC-generated N-terminal peptide tail is a novel biased agonist for PAR1.

scholarly journals Blood–Nerve Barrier (BNB) Pathology in Diabetic Peripheral Neuropathy and In Vitro Human BNB Model

2020 ◽  
Vol 22 (1) ◽  
pp. 62
Author(s):  
Yukio Takeshita ◽  
Ryota Sato ◽  
Takashi Kanda
Keyword(s):  
Endothelial Cells ◽  
Peripheral Neuropathy ◽  
Schwann Cells ◽  
Cell Lines ◽  
Diabetic Peripheral Neuropathy ◽  
Peripheral Nerves ◽  
Vascular Endothelial Cells ◽  
Vascular Endothelial ◽  
Immortalized Cell

In diabetic peripheral neuropathy (DPN), metabolic disorder by hyperglycemia progresses in peripheral nerves. In addition to the direct damage to peripheral neural axons, the homeostatic mechanism of peripheral nerves is disrupted by dysfunction of the blood–nerve barrier (BNB) and Schwann cells. The disruption of the BNB, which is a crucial factor in DPN development and exacerbation, causes axonal degeneration via various pathways. Although many reports revealed that hyperglycemia and other important factors, such as dyslipidemia-induced dysfunction of Schwann cells, contributed to DPN, the molecular mechanisms underlying BNB disruption have not been sufficiently elucidated, mainly because of the lack of in vitro studies owing to difficulties in establishing human cell lines from vascular endothelial cells and pericytes that form the BNB. We have developed, for the first time, temperature-sensitive immortalized cell lines of vascular endothelial cells and pericytes originating from the BNB of human sciatic nerves, and we have elucidated the disruption to the BNB mainly in response to advanced glycation end products in DPN. Recently, we succeeded in developing an in vitro BNB model to reflect the anatomical characteristics of the BNB using cell sheet engineering, and we established immortalized cell lines originating from the human BNB. In this article, we review the pathologic evidence of the pathology of DPN in terms of BNB disruption, and we introduce the current in vitro BNB models.

scholarly journals Recent Advances and Developments in Neural Repair and Regeneration for Hand Surgery

2012 ◽  
Vol 6 (1) ◽  
pp. 103-107 ◽  
Author(s):  
Mukai Chimutengwende-Gordon ◽  
Wasim Khan
Keyword(s):  
Gold Standard ◽  
Peripheral Nerves ◽  
Nerve Repair ◽  
Hand Surgery ◽  
Matrix Proteins ◽  
Neural Repair ◽  
Nerve Grafts ◽  
Nerve Conduits

End-to-end suture of nerves and autologous nerve grafts are the ‘gold standard’ for repair and reconstruction of peripheral nerves. However, techniques such as sutureless nerve repair with tissue glues, end-to-side nerve repair and allografts exist as alternatives. Biological and synthetic nerve conduits have had some success in early clinical studies on reconstruction of nerve defects in the hand. The effectiveness of nerve regeneration could potentially be increased by using these nerve conduits as scaffolds for delivery of Schwann cells, stem cells, neurotrophic and neurotropic factors or extracellular matrix proteins. There has been extensivein vitroandin vivoresearch conducted on these techniques. The clinical applicability and efficacy of these techniques needs to be investigated fully.

scholarly journals Human Peripheral Nerve-Derived Pluripotent Cells Can Be Stimulated by In Vitro Bone Morphogenetic Protein-2

2021 ◽  
Vol 8 (10) ◽  
pp. 132
Author(s):  
Renyi Sun ◽  
Tanghong Jia ◽  
Bradley Dart ◽  
Sunaina Shrestha ◽  
Morgan Bretches ◽  
...  
Keyword(s):  
Stem Cell ◽  
Transcription Factors ◽  
Peripheral Nerve ◽  
Bone Morphogenetic Protein ◽  
Peripheral Nerves ◽  
Bone Morphogenetic Protein 2 ◽  
Nerve Tissue ◽  
Morphogenetic Protein ◽  
Human Nerve

We have recently identified a population of cells within the peripheral nerves of adult rodent animals (mice and rats) that can respond to Bone Morphogenetic Protein-2 (BMP-2) exposure or physical injury to rapidly proliferate. More importantly, these cells exhibited embryonic differentiation potentials that could be induced into osteoblastic and endothelial cells in vitro. The current study examined human nerve specimens to compare and characterize the cells after BMP-2 stimulation. Fresh pieces of human nerve tissue were minced and treated with either BMP-2 (750 ng/mL) or a PBS vehicle for 12 h at 37 °C, before being digested in 0.2% collagenase and 0.05% trypsin-EDTA. Isolated cells were cultured in a restrictive stem cell medium. Significantly more cells were obtained from the nerve pieces with the BMP-2 treatment in comparison with the PBS vehicle controls. Cell colonies started to form at Day 3. Expressions of the four transcription factors, namely, Klf4, c-Myc, Sox2, and Oct4, were confirmed at both the transcriptional and translational levels. The cells can be maintained in the stem cell culture medium for at least 6 weeks without changing their morphology. When the cells were transferred to a fibroblast growth medium, dispersed spindle-shaped motile cells were noted and became fibroblast activated protein-α (FAP) positive with immunocytochemistry staining. The data suggest that human peripheral nerve tissue also contains a population of cells that can respond to BMP-2 and express Klf4, Sox2, cMyc, and Oct4—the four transcription factors driving cell pluripotency. These cells are able to differentiate into FAP-positive fibroblasts. In summary, in human peripheral nerves also reside a population of quiescent cells with pluripotency potential that may be the same cells as rodent nerve-derived adult stem (NEDAPS) cells. It is proposed that these cells are possibly at the core of a previously unknown natural mechanism for healing an injury.

scholarly journals The Use of Degradable Nerve Conduits for Human Nerve Repair: A Review of the Literature

2005 ◽  
Vol 2 (1) ◽  
pp. 39-43 ◽  
Author(s):  
M. F. Meek ◽  
K. Jansen ◽  
P. H. Robinson
Keyword(s):  
Peripheral Nerve ◽  
Peripheral Nerves ◽  
Nerve Repair ◽  
Experimental Studies ◽  
Review Of The Literature ◽  
Nerve Grafts ◽  
Nerve Guides ◽  
Clinical Challenge ◽  
Nerve Conduits ◽  
Human Nerve

The management of peripheral nerve injury continues to be a major clinical challenge. The most widely used technique for bridging defects in peripheral nerves is the use of autologous nerve grafts. This technique, however, has some disadvantages. Many alternative experimental techniques have thus been developed, such as degradable nerve conduits. Degradable nerve guides have been extensively studied in animal experimental studies. However, the repair of human nerves by degradable nerve conduits has been limited to only a few clinical studies. In this paper, an overview of the available international published literature on degradable nerve conduits for bridging human peripheral nerve defects is presented for literature available until 2004. Also, the philosophy on the use of nerve guides and nerve grafts is given.

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