Coculture Assays for Endothelial Cells-Mural Cells Interactions

Sánchez-Palencia, Diana M.; Bigger-Allen, Alex; Saint‐Geniez, Magali; Arboleda‐Velásquez, Joseph F.; D’Amore, Patricia A.

doi:10.1007/978-1-4939-3999-2_4

Cited by 11 publications

(12 citation statements)

References 10 publications

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“…The findings were similar with N3ICD (V1662) antibody and Notch-3 (N3ICD) antibody (C-terminus STAN) (Supplementary Figure 1E, http://onlin elibr ary.wiley.com/doi/10.1002/art.41748/ abstract). (20). RASFs were transiently transfected with siNotch-1, siNotch-3, or scrambled control (Figure 2A).…”

Section: Resultsmentioning

confidence: 99%

“…Angiogenesis is closely related to the pathologic development of RA. To further explore whether hypoxia‐induced angiogenesis is dependent on Notch signaling, indirect coculturing of endothelial cells (human umbilical vein endothelial cells [HUVECs]) and RASFs was performed in Transwell plates (20). RASFs were transiently transfected with siNotch‐1, siNotch‐3, or scrambled control (Figure 2A).…”

Section: Resultsmentioning

confidence: 99%

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Notch‐1 and Notch‐3 Mediate Hypoxia‐Induced Activation of Synovial Fibroblasts in Rheumatoid Arthritis

Cheng

Wang

et al. 2021

Arthritis & Rheumatology

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Objective To investigate the molecular mechanism of hypoxia‐induced rheumatoid arthritis synovial fibroblast (RASF) activation via Notch‐1 and Notch‐3 signaling, and to evaluate its potential as a therapeutic target. Methods Expression of Notch‐1 intracellular domain (N1ICD), N3ICD, and hypoxia‐inducible factor 1α (HIF‐1α) was assessed by immunhistology in synovial tissue from patients with RA. RASFs were cultured under hypoxic conditions and normoxic conditions with or without small interfering RNAs (siRNAs), and N1ICD and N3ICD were overexpressed under normoxic conditions. Rats with collagen‐induced arthritis (CIA) were administered LY411575 (inhibitor of N1ICD and N3ICD) for 15 days and 28 days, and its therapeutic efficacy was assessed by histologic and radiologic evaluation of the rat synovial tissue, and by analysis of inflammatory cytokine production in the serum of rats. Results N1ICD, N3ICD, and HIF‐1α were expressed abundantly in the synovial tissue of RA patients. HIF‐1α was shown to directly regulate the expression of Notch‐1 and Notch‐3 genes under hypoxic conditions. Moreover, hypoxia‐induced N1ICD and N3ICD expression in RASFs was blocked by HIF‐1α siRNA. Notch‐1 siRNA and Notch‐3 siRNA inhibited hypoxia‐induced RASF invasion and angiogenesis in vitro, whereas overexpression of N1ICD and N3ICD promoted these processes. In addition, Notch‐1 was shown to regulate RASF migration and epithelial–mesenchymal transition under hypoxic conditions, whereas Notch‐3 was shown to regulate the processes of anti‐apoptosis and autophagy. Furthermore, in vivo studies in rats with CIA showed that the N1ICD and N3ICD inhibitor LY411575 had a therapeutic effect in terms of ameliorating the symptoms and severity of the disease. Conclusion This study identified a functional link between HIF‐1α, Notch‐1, and Notch‐3 signaling in regulating activation of RASFs and the processes involved in the pathogenesis of RA.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Notch‐1 and Notch‐3 Mediate Hypoxia‐Induced Activation of Synovial Fibroblasts in Rheumatoid Arthritis

Cheng

Wang

et al. 2021

Arthritis & Rheumatology

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“…ECs and pericytes can be co-cultured to study their interactions in vitro. D’Amore et al [ 92 , 93 ] used primary ECs and pericytes from the bovine retina to elucidate their roles in vessel development and function, as well as the regulatory mechanisms. We have co-cultured primary ECs and pericytes from the human retina to investigate the regulation of HRECs-HRPCs interaction by P l GF signaling (not yet published).…”

Section: Experimental Models To Study Pericyte-endothelial Interamentioning

confidence: 99%

Pericyte-Endothelial Interactions in the Retinal Microvasculature

2020

IJMS

114

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Retinal microvasculature is crucial for the visual function of the neural retina. Pericytes and endothelial cells (ECs) are the two main cellular constituents in the retinal microvessels. Formation, maturation, and stabilization of the micro-vasculatures require pericyte-endothelial interactions, which are perturbed in many retinal vascular disorders, such as retinopathy of prematurity, retinal vein occlusion, and diabetic retinopathy. Understanding the cellular and molecular mechanisms of pericyte-endothelial interaction and perturbation can facilitate the design of therapeutic intervention for the prevention and treatment of retinal vascular disorders. Pericyte-endothelial interactions are indispensable for the integrity and functionality of retinal neurovascular unit (NVU), including vascular cells, retinal neurons, and glial cells. The essential autocrine and paracrine signaling pathways, such as Vascular endothelial growth factor (VEGF), Platelet-derived growth factor subunit B (PDGFB), Notch, Angipointein, Norrin, and Transforming growth factor-beta (TGF-β), have been well characterized for the regulation of pericyte-endothelial interactions in the neo-vessel formation processes (vasculogenesis and angiogenesis) during embryonic development. They also play a vital role in stabilizing and remodeling mature vasculature under pathological conditions. Awry signals, aberrant metabolisms, and pathological conditions, such as oxidative stress and inflammation, can disrupt the communication between pericytes and endothelial cells, thereby resulting in the breakdown of the blood-retinal barrier (BRB) and other microangiopathies. The emerging evidence supports extracellular exosomes’ roles in the (mis)communications between the two cell types. This review summarizes the essential knowledge and updates about new advancements in pericyte-EC interaction and communication, emphasizing the retinal microvasculature.

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“…52 In the standard use of transwell inserts to model the BRB, retinal endothelial cells for the iBRB or retinal pigmented epithelial cells for the oBRB were seeded on the upper compartment of the transwell to generate monoculture devices or integrated as coor tri-culture devices where the other cell types are seeded on the opposite side of the insert and/or at the bottom of the well. [53][54][55] The inclusion of multiple cell types enhanced physiological relevance by enabling crosstalk between neighboring cell populations. 54 Further, investigating cellcell communication enabled mechanistic studies of developmental, functional, and pathological processes of the retina, as support cells are known to influence BRB permeability and endothelial cell functions.…”

Section: Conventional In Vitro Models Of the Brbmentioning

confidence: 99%

“…55 Most BRB in vitro models have been established using primary cells isolated from animal sources or from human patient samples to increase model relevance to human diseases. [53][54][55]59 Human immortalized cell lines, such as the RPE cell line ARPE-19, have been developed in an attempt to improve robustness and availability. 61,62 Recently, induced pluripotent stem cells (iPSCs), which possess selfrenewal capability and potential to differentiate into various lineages, have emerged as a source to generate retinal cells 63,64 and have been used to generate vascular progenitors and endothelial cells that express TJ and endothelial cell markers.…”

Section: Conventional In Vitro Models Of the Brbmentioning

confidence: 99%

Organ-On-A-Chip Technologies for Advanced Blood–Retinal Barrier Models

Ragelle

Gonçalves

Kustermann

et al. 2020

Journal of Ocular Pharmacology and Therapeutics

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The blood-retinal barrier (BRB) protects the retina by maintaining an adequate microenvironment for neuronal function. Alterations of the junctional complex of the BRB and consequent BRB breakdown in disease contribute to a loss of neuronal signaling and vision loss. As new therapeutics are being developed to prevent or restore barrier function, it is critical to implement physiologically relevant in vitro models that recapitulate the important features of barrier biology to improve disease modeling, target validation, and toxicity assessment. New directions in organ-on-a-chip technology are enabling more sophisticated 3-dimensional models with flow, multicellularity, and control over microenvironmental properties. By capturing additional biological complexity, organs-on-chip can help approach actual tissue organization and function and offer additional tools to model and study disease compared with traditional 2-dimensional cell culture. This review describes the current state of barrier biology and barrier function in ocular diseases, describes recent advances in organ-on-a-chip design for modeling the BRB, and discusses the potential of such models for ophthalmic drug discovery and development.

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Coculture Assays for Endothelial Cells-Mural Cells Interactions

Cited by 11 publications

References 10 publications

Notch‐1 and Notch‐3 Mediate Hypoxia‐Induced Activation of Synovial Fibroblasts in Rheumatoid Arthritis

Notch‐1 and Notch‐3 Mediate Hypoxia‐Induced Activation of Synovial Fibroblasts in Rheumatoid Arthritis

Pericyte-Endothelial Interactions in the Retinal Microvasculature

Organ-On-A-Chip Technologies for Advanced Blood–Retinal Barrier Models

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