scholarly journals Three-Dimensional Culture Decreases the Angiogenic Ability of Mouse Macrophages

2021 ◽  
Vol 12 ◽  
Author(s):  
Haoxin Shi ◽  
Dong Li ◽  
Qing Shi ◽  
Zhenxia Han ◽  
Yuwei Tan ◽  
...  
Keyword(s):  
Three Dimensional ◽  
3D Culture ◽  
Chorioallantoic Membrane ◽  
Tube Formation ◽  
Angiogenic Factors ◽  
Future Research ◽  
Chick Embryo Chorioallantoic Membrane ◽  
Whole Transcriptome Sequencing ◽  
3D Culture System ◽  
Almost All

Macrophages play important roles in angiogenesis; however, previous studies on macrophage angiogenesis have focused on traditional 2D cultures. In this study, we established a 3D culture system for macrophages using collagen microcarriers and assessed the effect of 3D culture on their angiogenic capabilities. Macrophages grown in 3D culture displayed a significantly different morphology and arrangement under electron microscopy compared to those grown in 2D culture. Tube formation assays and chick embryo chorioallantoic membrane assays further revealed that 3D-cultured macrophages were less angiogenic than those in 2D culture. Whole-transcriptome sequencing showed that nearly 40% of genes were significantly differently expressed, including nine important angiogenic factors of which seven had been downregulated. In addition, the expression of almost all genes related to two important angiogenic pathways was decreased in 3D-cultured macrophages, including the two key angiogenic factors, VEGFA and ANG2. Together, the findings of our study improve our understanding of angiogenesis and 3D macrophage culture in tissues, and provide new avenues and methods for future research on macrophages.

Novel three-dimensional long-term bone marrow culture system using polymer particles with grafted epoxy-polymer-chains supports the proliferation and differentiation of hematopoietic stem cells

2011 ◽  
Vol 236 (11) ◽  
pp. 1342-1350 ◽  
Author(s):  
Yukio Hirabayashi ◽  
Yoshihiro Hatta ◽  
Jin Takeuchi ◽  
Isao Tsuboi ◽  
Tomonori Harada ◽  
...  
Keyword(s):  
Stromal Cells ◽  
Culture System ◽  
3D Structure ◽  
Three Dimensional ◽  
Hematopoietic Cells ◽  
3D Culture ◽  
Polymer Chains ◽  
Polymer Particles ◽  
Hematopoietic Stem ◽  
3D Culture System

Hematopoiesis occurs in the bone marrow, where primitive hematopoietic cells proliferate and differentiate in close association with a three-dimensional (3D) hematopoietic microenvironment composed of stromal cells. We examined the hematopoietic supportive ability of stromal cells in a 3D culture system using polymer particles with grafted epoxy polymer chains. Umbilical cord blood-derived CD34+ cells were co-cultivated with MS-5 stromal cells. They formed a 3D structure in the culture dish in the presence of particles, and the total numbers of cells and the numbers of hematopoietic progenitor cells, including colony-forming unit (CFU)-Mix, CFU-granulocyte-macrophage, CFU-megakaryocyte and burst-forming unit-erythroid, were measured every seven days. The hematopoietic supportive activity of the 3D culture containing polymer particles and stromal cells was superior to that of 2D culture, and allowed the expansion and maintenance of hematopoietic progenitor cells for more than 12 weeks. Various types of hematopoietic cells, including granulocytes, macrophages and megakaryocytes at different maturation stages, appeared in the 3D culture, suggesting that the CD34+ cells were able to differentiate into a range of blood cell types. Morphological examination showed that MS-5 stromal cells grew on the surface of the particles and bridged the gaps between them to form a 3D structure. Hematopoietic cells slipped into the 3D layer and proliferated within it, relying on the presence of the MS-5 cells. These results suggest that this 3D culture system using polymer particles reproduced the hematopoietic phenomenon in vitro, and might thus provide a new tool for investigating hematopoietic stem cell–stromal cell interactions.

scholarly journals Comparison of Immunosuppressive and Angiogenic Properties of Human Amnion-Derived Mesenchymal Stem Cells between 2D and 3D Culture Systems

2019 ◽  
Vol 2019 ◽  
pp. 1-16 ◽  
Author(s):  
Vitale Miceli ◽  
Mariangela Pampalone ◽  
Serena Vella ◽  
Anna Paola Carreca ◽  
Giandomenico Amico ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Three Dimensional ◽  
3D Culture ◽  
Culture Method ◽  
Paracrine Factors ◽  
Human Amnion ◽  
Tissue Repair And Regeneration ◽  
3D Culture System

The secretion of potential therapeutic factors by mesenchymal stem cells (MSCs) has aroused much interest given the benefits that it can bring in the field of regenerative medicine. Indeed, the in vitro multipotency of these cells and the secretive capacity of both angiogenic and immunomodulatory factors suggest a role in tissue repair and regeneration. However, during culture, MSCs rapidly lose the expression of key transcription factors associated with multipotency and self-renewal, as well as the ability to produce functional paracrine factors. In our study, we show that a three-dimensional (3D) culture method is effective to induce MSC spheroid formation, to maintain the multipotency and to improve the paracrine activity of a specific population of human amnion-derived MSCs (hAMSCs). The regenerative potential of both 3D culture-derived conditioned medium (3D CM) and their exosomes (EXO) was assessed against 2D culture products. In particular, tubulogenesis assays revealed increased capillary maturation in the presence of 3D CM compared with both 2D CM and 2D EXO. Furthermore, 3D CM had a greater effect on inhibition of PBMC proliferation than both 2D CM and 2D EXO. To support this data, hAMSC spheroids kept in our 3D culture system remained viable and multipotent and secreted considerable amounts of both angiogenic and immunosuppressive factors, which were detected at lower levels in 2D cultures. This work reveals the placenta as an important source of MSCs that can be used for eventual clinical applications as cell-free therapies.

Design and Fabrication of a Three-Dimensional In Vitro System for Modeling Vascular Stenosis

2017 ◽  
Vol 23 (4) ◽  
pp. 859-871 ◽  
Author(s):  
Rebecca S. Jones ◽  
Pin H. Chang ◽  
Tzlil Perahia ◽  
Katrina A. Harmon ◽  
Lorain Junor ◽  
...  
Keyword(s):  
Three Dimensional ◽  
3D Culture ◽  
Cellular Interactions ◽  
Vessel Occlusion ◽  
Casting System ◽  
Vascular Pathogenesis ◽  
3D Culture System ◽  
Adverse Remodeling ◽  
Vascular Stenosis

AbstractVascular stenosis, the abnormal narrowing of blood vessels, arises from defective developmental processes or atherosclerosis-related adult pathologies. Stenosis triggers a series of adaptive cellular responses that induces adverse remodeling, which can progress to partial or complete vessel occlusion with numerous fatal outcomes. Despite its severity, the cellular interactions and biophysical cues that regulate this pathological progression are poorly understood. Here, we report the design and fabrication of a three-dimensional (3D) in vitro system to model vascular stenosis so that specific cellular interactions and responses to hemodynamic stimuli can be investigated. Tubular cellularized constructs (cytotubes) were produced, using a collagen casting system, to generate a stenotic arterial model. Fabrication methods were developed to create cytotubes containing co-cultured vascular cells, where cell viability, distribution, morphology, and contraction were examined. Fibroblasts, bone marrow primary cells, smooth muscle cells (SMCs), and endothelial cells (ECs) remained viable during culture and developed location- and time-dependent morphologies. We found cytotube contraction to depend on cellular composition, where SMC-EC co-cultures adopted intermediate contractile phenotypes between SMC- and EC-only cytotubes. Our fabrication approach and the resulting artery model can serve as an in vitro 3D culture system to investigate vascular pathogenesis and promote the tissue engineering field.

scholarly journals Epigallocatechin-3-Gallate Suppresses Vasculogenic Mimicry through Inhibiting the Twist/VE-Cadherin/AKT Pathway in Human Prostate Cancer PC-3 Cells

2020 ◽  
Vol 21 (2) ◽  
pp. 439 ◽  
Author(s):  
Changhwan Yeo ◽  
Deok-Soo Han ◽  
Hyo-Jeong Lee ◽  
Eun-Ok Lee
Keyword(s):  
Prostate Cancer ◽  
Cell Viability ◽  
Vasculogenic Mimicry ◽  
Three Dimensional ◽  
3D Culture ◽  
Tube Formation ◽  
Human Prostate ◽  
Akt Pathway ◽  
Human Prostate Cancer ◽  
Western Blots

Vasculogenic mimicry (VM) is the alternative process of forming vessel-like networks by aggressive tumor cells, and it has an important role in tumor survival, growth, and metastasis. Epigallocatechin-3-gallate (EGCG) is well known to have diverse bioactivities including anti-cancer effects. However, the efficacy of EGCG on VM is elusive. In this study, we explored whether and how EGCG affects VM in human prostate cancer (PCa) PC-3 cells. Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Invasive and VM formation abilities were assessed by an invasion assay and a three-dimensional (3D) culture VM tube formation assay, respectively. Western blots were carried out. An immunofluorescence assay was performed to detect nuclear twist expression. EGCG effectively inhibited the invasive ability, as well as tubular channel formation, without affecting cell viability. EGCG significantly downregulated the expression of vascular endothelial cadherin (VE-cadherin) and its transcription factor, twist, N-cadherin, vimentin, phosphor-AKT, and AKT, but not phospho-erythropoietin-producing hepatocellular receptor A2 (EphA2) and EphA2. In addition, EGCG diminished the nuclear localization of twist. Treatment with SC79, an AKT activator, effectively rescued EGCG-inhibited VM formation. These results demonstrated for the first time that EGCG causes marked suppression of VM through inhibiting the twist/VE-cadherin/AKT pathway in human PCa PC-3 cells.

scholarly journals Induced Osteogenesis in Plants Decellularized Scaffolds

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jennifer Lee ◽  
Hyerin Jung ◽  
Narae Park ◽  
Sung-Hwan Park ◽  
Ji Hyeon Ju
Keyword(s):  
Three Dimensional ◽  
3D Culture ◽  
Specific Staining ◽  
3D Culture System ◽  
Calcified Tissue ◽  
Decellularized Scaffolds ◽  
Cellular Markers ◽  
Rat Calvarial Defect

AbstractA three-dimensional (3D) culture system that closely replicates the in vivo microenvironment of calcifying osteoid is essential for in vitro cultivation of bone-like material. In this regard, the 3D cellulose constructs of plants may well serve as scaffolds to promote growth and differentiation of osteoblasts in culture. Our aim in this study was to generate bone-like tissue by seeding pluripotent stem cells (hiPSCs), stimulated to differentiate as osteoblasts in culture, onto the decellularised scaffolds of various plants. We then assessed expression levels of pertinent cellular markers and degrees of calcium-specific staining to gauge technical success. Apple scaffolding bearing regular pores of 300 μm seemed to provide the best construct. The bone-like tissue thus generated was implantable in a rat calvarial defect model where if helped form calcified tissue. Depending on the regularity and sizing of scaffold pores, this approach readily facilitates production of mineralized bone.

scholarly journals Assembly of FN-silk with laminin-521 to integrate hPSCs into a three-dimensional culture for neural differentiation

2020 ◽  
Vol 8 (9) ◽  
pp. 2514-2525
Author(s):  
Carolina Åstrand ◽  
Veronique Chotteau ◽  
Anna Falk ◽  
My Hedhammar
Keyword(s):  
Culture System ◽  
Neural Differentiation ◽  
Spider Silk ◽  
Three Dimensional ◽  
3D Culture ◽  
Silk Protein ◽  
Spider Silk Protein ◽  
3D Culture System ◽  
Three Dimensional Culture

The functionalized recombinant spider silk protein FN-silk can self-assemble into a 3D microfiber network. When combined with recombinant laminin521 it provides a 3D culture system suitable for expansion of hPSCs and following neural differentiation.

scholarly journals The Three-Dimensional Culture System with Matrigel and Neurotrophic Factors Preserves the Structure and Function of Spiral Ganglion NeuronIn Vitro

2016 ◽  
Vol 2016 ◽  
pp. 1-15 ◽  
Author(s):  
Gaoying Sun ◽  
Wenwen Liu ◽  
Zhaomin Fan ◽  
Daogong Zhang ◽  
Yuechen Han ◽  
...  
Keyword(s):  
Growth Cone ◽  
Neurotrophic Factors ◽  
Culture System ◽  
Spiral Ganglion ◽  
Three Dimensional ◽  
3D Culture ◽  
Explant Culture ◽  
Structure And Function ◽  
3D Culture System ◽  
And Function

Whole organ culture of the spiral ganglion region is a resourceful model system facilitating manipulation and analysis of live sprial ganglion neurons (SGNs). Three-dimensional (3D) cultures have been demonstrated to have many biomedical applications, but the effect of 3D culture in maintaining the SGNs structure and function in explant culture remains uninvestigated. In this study, we used the matrigel to encapsulate the spiral ganglion region isolated from neonatal mice. First, we optimized the matrigel concentration for the 3D culture system and found the 3D culture system protected the SGNs against apoptosis, preserved the structure of spiral ganglion region, and promoted the sprouting and outgrowth of SGNs neurites. Next, we found the 3D culture system promoted growth cone growth as evidenced by a higher average number and a longer average length of filopodia and a larger growth cone area. 3D culture system also significantly elevated the synapse density of SGNs. Last, we found that the 3D culture system combined with neurotrophic factors had accumulated effects in promoting the neurites outgrowth compared with 3D culture or NFs treatment only groups. Together, we conclude that the 3D culture system preserves the structure and function of SGN in explant culture.

scholarly journals Efficient Generation of Neural Stem Cells from Embryonic Stem Cells Using a Three-Dimensional Differentiation System

2021 ◽  
Vol 22 (15) ◽  
pp. 8322
Author(s):  
Sang-Hoon Yoon ◽  
Mi-Rae Bae ◽  
Hyeonwoo La ◽  
Hyuk Song ◽  
Kwonho Hong ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Neural Stem Cells ◽  
Three Dimensional ◽  
Embryonic Stem ◽  
3D Culture ◽  
Cell Types ◽  
Neural Lineage ◽  
Lineage Differentiation ◽  
3D Culture System

Mouse embryonic stem cells (ESCs) are useful tools for studying early embryonic development and tissue formation in mammals. Since neural lineage differentiation is a major subject of organogenesis, the development of efficient techniques to induce neural stem cells (NSCs) from pluripotent stem cells must be preceded. However, the currently available NSC differentiation methods are complicated and time consuming. This study aimed to propose an efficient method for the derivation of NSCs from mouse ESCs; early neural lineage commitment was achieved using a three-dimensional (3D) culture system, followed by a two-dimensional (2D) NSC derivation. To select early neural lineage cell types during differentiation, Sox1-GFP transgenic ESCs were used. They were differentiated into early neural lineage, forming spherical aggregates, which were subsequently picked for the establishment of 2D NSCs. The latter showed a morphology similar to that of brain-derived NSCs and expressed NSC markers, Musashi, Nestin, N-cadherin, and Sox2. Moreover, the NSCs could differentiate into three subtypes of neural lineages, neurons, astrocytes, and oligodendrocytes. The results together suggested that ESCs could efficiently differentiate into tripotent NSCs through specification in 3D culture (for approximately 10 days) followed by 2D culture (for seven days).

scholarly journals Strain-triggered mechanical feedback in self-organizing optic-cup morphogenesis

2018 ◽  
Vol 4 (11) ◽  
pp. eaau1354 ◽  
Author(s):  
S. Okuda ◽  
N. Takata ◽  
Y. Hasegawa ◽  
M. Kawada ◽  
Y. Inoue ◽  
...  
Keyword(s):  
Three Dimensional ◽  
3D Culture ◽  
Quantitative Prediction ◽  
Complementary Approach ◽  
Optic Cup ◽  
3D Culture System ◽  
Underlying Mechanisms ◽  
Tissue Deformations ◽  
Robust Regulation ◽  
Self Organizing

Organogenesis is a self-organizing process of multiple cells in three-dimensional (3D) space, where macroscopic tissue deformations are robustly regulated by multicellular autonomy. It is clear that this robust regulation requires cells to sense and modulate 3D tissue formation across different scales, but its underlying mechanisms are still unclear. To address this question, we developed a versatile computational model of 3D multicellular dynamics at single-cell resolution and combined it with the 3D culture system of pluripotent stem cell–derived optic-cup organoid. The complementary approach enabled quantitative prediction of morphogenesis and its corresponding verification and elucidated that the macroscopic 3D tissue deformation is fed back to individual cellular force generations via mechanosensing. We hereby conclude that mechanical force plays a key role as a feedback regulator to establish the robustness of organogenesis.

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