scholarly journals Mesenchymal-like glioma cells are enriched in the gelatin methacrylate hydrogels

2021 ◽  
Author(s):  
Nameeta Shah ◽  
Pavan M. Hallur ◽  
Raksha A. Ganesh ◽  
Pranali Sonpatki ◽  
Divya Naik ◽  
...  
Keyword(s):  
Therapeutic Response ◽  
Immune Cell ◽  
Three Dimensional ◽  
Glioma Cells ◽  
Cellular Phenotype ◽  
3D Scaffold ◽  
Promising Alternative ◽  
Cells Cultured

AbstractGlioblastoma is the most lethal primary malignant brain tumor in adults. Simplified two-dimensional (2D) cell culture and neurospheres in vitro models fail to recapitulate the complexity of the tumor microenvironment, limiting its ability to predict therapeutic response. Three-dimensional (3D) scaffold-based models have emerged as a promising alternative for addressing these concerns. One such 3D system is gelatin methacrylate (GelMA) hydrogels, which can be used for modeling the glioblastoma microenvironment. We characterized the phenotype of patient-derived glioma cells cultured in GelMA hydrogels (3D-GMH) for their tumorigenic properties using invasion and chemoresponse assays. In addition, we used integrated single-cell and spatial transcriptome analysis to compare cells cultured in 3D-GMH to cells in vivo. Finally, we assessed tumor-immune cell interactions with a macrophage infiltration assay and a cytokine array. We show that cells cultured in 3D-GMH develop a mesenchymal-like cellular phenotype found in perivascular and hypoxic regions present in the core of the tumor, and recruit macrophages by secreting cytokines in contrast to the cells grown as neurospheres that match the phenotype of cells of the infiltrative edge of the tumor.

scholarly journals Gelatin methacrylate hydrogels culture model for glioblastoma cells enriches for mesenchymal-like state and models interactions with immune cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Nameeta Shah ◽  
Pavan M. Hallur ◽  
Raksha A. Ganesh ◽  
Pranali Sonpatki ◽  
Divya Naik ◽  
...  
Keyword(s):  
Immune Cell ◽  
Three Dimensional ◽  
Glioma Cells ◽  
Glioblastoma Cells ◽  
Treatment Resistant ◽  
3D Scaffold ◽  
Promising Alternative ◽  
Cells Cultured

AbstractGlioblastoma is the most lethal primary malignant brain tumor in adults. Simplified two-dimensional (2D) cell culture and neurospheres in vitro models fail to recapitulate the complexity of the tumor microenvironment, limiting its ability to predict therapeutic response. Three-dimensional (3D) scaffold-based models have emerged as a promising alternative for addressing these concerns. One such 3D system is gelatin methacrylate (GelMA) hydrogels, and we aimed to understand the suitability of using this system to mimic treatment-resistant glioblastoma cells that reside in specific niches. We characterized the phenotype of patient-derived glioma cells cultured in GelMA hydrogels (3D-GMH) for their tumorigenic properties using invasion and chemoresponse assays. In addition, we used integrated single-cell and spatial transcriptome analysis to compare cells cultured in 3D-GMH to neoplastic cells in vivo. Finally, we assessed tumor-immune cell interactions with a macrophage infiltration assay and a cytokine array. We show that the 3D-GMH system enriches treatment-resistant mesenchymal cells that are not represented in neurosphere cultures. Cells cultured in 3D-GMH resemble a mesenchymal-like cellular phenotype found in perivascular and hypoxic regions and recruit macrophages by secreting cytokines, a hallmark of the mesenchymal phenotype. Our 3D-GMH model effectively mimics the phenotype of glioma cells that are found in the perivascular and hypoxic niches of the glioblastoma core in situ, in contrast to the neurosphere cultures that enrich cells of the infiltrative edge of the tumor. This contrast highlights the need for due diligence in selecting an appropriate model when designing a study‘s objectives.

scholarly journals Preparation of Gelatin and Gelatin/Hyaluronic Acid Cryogel Scaffolds for the 3D Culture of Mesothelial Cells and Mesothelium Tissue Regeneration

2019 ◽  
Vol 20 (18) ◽  
pp. 4527 ◽  
Author(s):  
Hao-Hsi Kao ◽  
Chang-Yi Kuo ◽  
Kuo-Su Chen ◽  
Jyh-Ping Chen
Keyword(s):  
Hyaluronic Acid ◽  
Three Dimensional ◽  
3D Culture ◽  
Mesothelial Cells ◽  
In Vivo Studies ◽  
3D Scaffold ◽  
Scaffold Degradation ◽  
Cells Cultured

Mesothelial cells are specific epithelial cells that are lined in the serosal cavity and internal organs. Nonetheless, few studies have explored the possibility to culture mesothelial cells in a three-dimensional (3D) scaffold for tissue engineering applications. Towards this end, we fabricated macroporous scaffolds from gelatin and gelatin/hyaluronic acid (HA) by cryogelation, and elucidated the influence of HA on cryogel properties and the cellular phenotype of mesothelial cells cultured within the 3D scaffolds. The incorporation of HA was found not to significantly change the pore size, porosity, water uptake kinetics, and swelling ratios of the cryogel scaffolds, but led to a faster scaffold degradation in the collagenase solution. Adding 5% HA in the composite cryogels also decreased the ultimate compressive stress (strain) and toughness of the scaffold, but enhanced the elastic modulus. From the in vitro cell culture, rat mesothelial cells showed quantitative cell viability in gelatin (G) and gelatin/HA (GH) cryogels. Nonetheless, mesothelial cells cultured in GH cryogels showed a change in the cell morphology and cytoskeleton arrangement, reduced cell proliferation rate, and downregulation of the mesothelium specific maker gene expression. The production of key mesothelium proteins E-cadherin and calretinin were also reduced in the GH cryogels. Choosing the best G cryogels for in vivo studies, the cell/cryogel construct was used for the transplantation of allograft mesothelial cells for mesothelium reconstruction in rats. A mesothelium layer similar to the native mesothelium tissue could be obtained 21 days post-implantation, based on hematoxylin and eosin (H&E) and immunohistochemical staining.

scholarly journals The Revolutionary Roads to Study Cell–Cell Interactions in 3D In Vitro Pancreatic Cancer Models

Cancers ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 930
Author(s):  
Donatella Delle Cave ◽  
Riccardo Rizzo ◽  
Bruno Sainz ◽  
Giuseppe Gigli ◽  
Loretta L. del Mercato ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Therapeutic Response ◽  
Three Dimensional ◽  
Cellular Models ◽  
Common Cancer ◽  
Cancer Models ◽  
Anti Cancer ◽  
Tumor Environment

Pancreatic cancer, the fourth most common cancer worldwide, shows a highly unsuccessful therapeutic response. In the last 10 years, neither important advancements nor new therapeutic strategies have significantly impacted patient survival, highlighting the need to pursue new avenues for drug development discovery and design. Advanced cellular models, resembling as much as possible the original in vivo tumor environment, may be more successful in predicting the efficacy of future anti-cancer candidates in clinical trials. In this review, we discuss novel bioengineered platforms for anticancer drug discovery in pancreatic cancer, from traditional two-dimensional models to innovative three-dimensional ones.

scholarly journals A Three-Dimensional Co-Culture Model for Rheumatoid Arthritis Pannus Tissue

2021 ◽  
Vol 9 ◽  
Author(s):  
Jietao Lin ◽  
Antonia RuJia Sun ◽  
Jian Li ◽  
Tianying Yuan ◽  
Wenxiang Cheng ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Vascular Endothelial Cells ◽  
Three Dimensional ◽  
Synovial Fibroblasts ◽  
Vascular Endothelial ◽  
3D Scaffold ◽  
Culture Model ◽  
Pannus Tissue

Three-dimensional (3D) co-culture models have closer physiological cell composition and behavior than traditional 2D culture. They exhibit pharmacological effects like in vivo responses, and therefore serve as a high-throughput drug screening model to evaluate drug efficacy and safety in vitro. In this study, we created a 3D co-culture environment to mimic pathological characteristics of rheumatoid arthritis (RA) pannus tissue. 3D scaffold was constructed by bioprinting technology with synovial fibroblasts (MH7A), vascular endothelial cells (EA.hy 926) and gelatin/alginate hydrogels. Cell viability was observed during 7-day culture and the proliferation rate of co-culture cells showed a stable increase stage. Cell-cell interactions were evaluated in the 3D printed scaffold and we found that spheroid size increased with time. TNF-α stimulated MH7A and EA.hy 926 in 3D pannus model showed higher vascular endothelial growth factor (VEGF) and angiopoietin (ANG) protein expression over time. For drug validation, methotrexate (MTX) was used to examine inhibition effects of angiogenesis in 3D pannus co-culture model. In conclusion, this 3D co-culture pannus model with biological characteristics may help the development of anti-RA drug research.

scholarly journals Multiscale modelling of drug transport and metabolism in liver spheroids

2020 ◽  
Vol 10 (2) ◽  
pp. 20190041 ◽  
Author(s):  
Joseph A. Leedale ◽  
Jonathan A. Kyffin ◽  
Amy L. Harding ◽  
Helen E. Colley ◽  
Craig Murdoch ◽  
...  
Keyword(s):  
Drug Transport ◽  
Temporal Dynamics ◽  
Three Dimensional ◽  
Culture Conditions ◽  
Isolated Cells ◽  
Modelling Framework ◽  
The Impact ◽  
Cells Cultured

In early preclinical drug development, potential candidates are tested in the laboratory using isolated cells. These in vitro experiments traditionally involve cells cultured in a two-dimensional monolayer environment. However, cells cultured in three-dimensional spheroid systems have been shown to more closely resemble the functionality and morphology of cells in vivo . While the increasing usage of hepatic spheroid cultures allows for more relevant experimentation in a more realistic biological environment, the underlying physical processes of drug transport, uptake and metabolism contributing to the spatial distribution of drugs in these spheroids remain poorly understood. The development of a multiscale mathematical modelling framework describing the spatio-temporal dynamics of drugs in multicellular environments enables mechanistic insight into the behaviour of these systems. Here, our analysis of cell membrane permeation and porosity throughout the spheroid reveals the impact of these properties on drug penetration, with maximal disparity between zonal metabolism rates occurring for drugs of intermediate lipophilicity. Our research shows how mathematical models can be used to simulate the activity and transport of drugs in hepatic spheroids and in principle any organoid, with the ultimate aim of better informing experimentalists on how to regulate dosing and culture conditions to more effectively optimize drug delivery.

scholarly journals Ultrasound-triggered on Demand Lidocaine Release Relieves Postoperative Pain

2021 ◽  
Author(s):  
Xiaohong Chen ◽  
Jianfeng Zhang ◽  
Yan Yu ◽  
Haoran Wang ◽  
Genshan Ma ◽  
...  
Keyword(s):  
Postoperative Pain ◽  
Three Dimensional ◽  
Systemic Effect ◽  
Tissue Reaction ◽  
Control Group ◽  
3D Scaffold ◽  
Nerve Blocks ◽  
On Demand

Abstract BackgroundSafe and noninvasive on-demand relief is a crucial and effective treatment for postoperative pain because it considers variable timing and intensity of anesthetics. Ultrasound modulation is a promising technique for this treatment because it allows convenient timed and noninvasive controlled drug release.MethodsWe created an ultrasound-triggered lidocaine (Lido) release platform using an amino acid hydrogel functioning as three dimensional (3D) scaffold material (Lido-PPIX@ER hydrogel). Optimal preparation conditions and ultrasound-triggered parameters were evaluated. In the postoperative pain SD rat model, the Lido-PPIX@ER hydrogel or free lidocaine was administered by subcutaneous injection immediately after making the paw incision. Mechanical hypersensitivity was assessed using calibrated von Frey filaments after an individualized (highly variable) ultrasound-triggered process. The safety of the treatment was also evaluated.ResultsThe Lido-PPIX@ER hydrogel allows control of the timing, intensity and duration of lidocaine (Lido) to relieve postoperative pain. The hydrogel releases Lido due to the elevated reactive oxygen species (ROS) levels generated by PPIX under ultrasound triggering. The optimal ultrasound parameter (0.3 W/cm2, 30 s) was chosen for in vitro and in vivo studies. The Lido-PPIX@ER hydrogel (with lidocaine, 5.6 mg/mL) under individualized ultrasound triggering (every 2 h in the first 12 h, every 4 h for the next 36 h, and then every 6 h until 72 h postsurgery) released lidocaine and provided effective analgesia for more than 72 h. Additionally, the withdrawal threshold was higher than that in the control group at all time points measured. The hydrogel showed repeatable and adjustable ultrasound-triggered nerve blocks in vivo, the duration of which depended on the extent and intensity of insonation. On histopathology, no systemic effect or tissue reaction was observed in the ultrasound-triggered Lido-PPIX@ER hydrogel-treated group.ConclusionsThe Lido-PPIX@ER hydrogel with individualized (highly variable) ultrasound triggering is a convenient and effective method that offers timed and spatiotemporally controlled Lido release to manage postoperative pain.

scholarly journals Experimental and numerical investigations of fluid flow in bioreactors for optimized in vitro stem cell loading in xenografts

2018 ◽  
Vol 4 (1) ◽  
pp. 423-427
Author(s):  
Robert Ott ◽  
Carolin Wüstenhagen ◽  
Michael Stiehm ◽  
Klaus-Peter Schmitz ◽  
Stefan Siewert ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cfd Simulation ◽  
Healing Process ◽  
Three Dimensional ◽  
Flow Conditions ◽  
Micro Computed Tomography ◽  
3D Scaffold ◽  
Numerical Investigations

AbstractIn tissue engineering and regenerative medicine mesenchymal stem cells (MSC) are widely used to replace and restore the function of dysfunctional or missing tissue. Recent studies have shown significant enhancements of the in vivo healing process following dentofacial bone augmentation procedures employing stem cell-loaded xenografts. We conducted experimental and numerical investigations in perfusion flow bioreactor-xenograft-systems to identify flow conditions as well as bioreactor design features that allow for homogeneous MSC-distribution in Geistlich Bio- Oss Block xenografts. Pressure gradient - velocity characteristics and flow distributions were investigated experimentally and numerically for two bioreactor designs at steady-state flow conditions with Reynolds numbers (Re) ranging from 0.01 ≤ Re ≤ 0.32. Distilled water at 20°C with a dynamic viscosity of 1.002 mPa∙s and a density of 998 kg/m3 was used. The geometry of the xenograft utilized in three-dimensional computational fluid dynamics (CFD) simulation was obtained by means of micro-computed tomography (μCT) at an isotropic spatial resolution of 9.5 μm. The permeability values calculated from the experimental data are in good accordance with the numerical results. The investigations showed that the increase of the inflow- and outflow-area diameter, as well as the decrease of the volumetric flow rate, result in a decreasing heterogeneity of the flow distribution within the xenograft. The calculated wall shear stress rates in the three-dimensional (3D) scaffold range from 1∙10-12Pa ≤ τ ≤ 0.2 Pa. Experimentally validated CFD simulations introduced in this study provide an applicable tool to assess optimal flow conditions for homogeneous MSC distribution in bioreactor-xenograft-systems.

scholarly journals Biofilm-stimulated epithelium modulates the inflammatory responses in co-cultured immune cells

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jason L. Brown ◽  
William Johnston ◽  
Chris Delaney ◽  
Ranjith Rajendran ◽  
John Butcher ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Immune Cells ◽  
Three Dimensional ◽  
Epithelial Tissue ◽  
Interface Model ◽  
Inflammatory Status ◽  
Gingival Epithelium ◽  
Cells Cultured

Abstract The gingival epithelium is a physical and immunological barrier to the microbiota of the oral cavity, which interact through soluble mediators with the immune cells that patrol the tissue at the gingival epithelium. We sought to develop a three-dimensional gingivae-biofilm interface model using a commercially available gingival epithelium to study the tissue inflammatory response to oral biofilms associated with “health”, “gingivitis” and “periodontitis”. These biofilms were developed by sequential addition of microorganisms to mimic the formation of supra- and sub-gingival plaque in vivo. Secondly, to mimic the interactions between gingival epithelium and immune cells in vivo, we integrated peripheral blood mononuclear cells and CD14+ monocytes into our three-dimensional model and were able to assess the inflammatory response in the immune cells cultured with and without gingival epithelium. We describe a differential inflammatory response in immune cells cultured with epithelial tissue, and more so following incubation with epithelium stimulated by “gingivitis-associated” biofilm. These results suggest that gingival epithelium-derived soluble mediators may control the inflammatory status of immune cells in vitro, and therefore targeting of the epithelial response may offer novel therapies. This multi-cellular interface model, both of microbial and host origin, offers a robust in vitro platform to investigate host-pathogens at the epithelial surface.

scholarly journals Aligned 3D porous polyurethane scaffolds for biological anisotropic tissue regeneration

2019 ◽  
Author(s):  
Weiwei Lin ◽  
Wanling Lan ◽  
Yingke Wu ◽  
Daiguo Zhao ◽  
Yanchao Wang ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Tissue Regeneration ◽  
Three Dimensional ◽  
Vertical Direction ◽  
Waterborne Polyurethane ◽  
3D Scaffold ◽  
Pure Material ◽  
Muscle Nerve

Abstract A green fabrication process (organic solvent-free) of artificial scaffolds is required in tissue engineering field. In this work, a series of aligned three-dimensional (3D) scaffolds are made from biodegradable waterborne polyurethane (PU) emulsion via directional freeze–drying method to ensure no organic byproducts. After optimizing the concentration of polymer in the emulsion and investigating different freezing temperatures, an aligned PUs scaffold (PU14) generated from 14 wt% polymer content and processed at −196°C was selected based on the desired oriented porous structure (pore size of 32.5 ± 9.3 μm, porosity of 92%) and balanced mechanical properties both in the horizontal direction (strength of 41.3 kPa, modulus of 72.3 kPa) and in the vertical direction (strength of 45.5 kPa, modulus of 139.3 kPa). The response of L929 cells and the regeneration of muscle tissue demonstrated that such pure material-based aligned 3D scaffold can facilitate the development of orientated cells and anisotropic tissue regeneration both in vitro and in vivo. Thus, these pure material-based scaffolds with ordered architecture have great potentials in tissue engineering for biological anisotropic tissue regeneration, such as muscle, nerve, spinal cord and so on.

Constitutive IFN signaling to modulate the immunogenicity of glioma cells.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. e13528-e13528
Author(s):  
Patrick Roth ◽  
Sara Nagy ◽  
Caroline Happold ◽  
Hannah Schneider ◽  
Michael Weller ◽  
...  
Keyword(s):  
Cellular Proliferation ◽  
Cell Function ◽  
Immune Cell ◽  
Acute Infection ◽  
Constitutive Expression ◽  
Glioma Cells ◽  
Type I ◽  
Type I Ifn

e13528 Background: Interferons (IFN) are cytokines that might be constitutively expressed also in the absence of acute infection and thereby regulate various physiological processes such as cellular proliferation and differentiation, cell viability or immune cell function. Alterations in IFN signaling have been observed in numerous malignancies, but their role in glioblastoma, particularly in glioma-initiating cells, has not been investigated in detail. Methods: We characterized constitutive type I IFN signaling and its functional impact in glioma cells using expression analyses, RNA interference-mediated silencing of various genes involved in the IFN signaling pathway and immune cell lysis assays. Results: We found constitutive expression of pSTAT1 and Myxovirus A (MxA), a classical IFN-response marker, in the absence of exogenous IFN-β in vitro and higher MxA expression in gliomas than in normal tissue in vivo, indicating that IFN signaling is constitutively active in these tumors. Silencing of the type I IFN receptor, IFNAR1/2, or its ligands, IFN-α or IFN-β, decreased MxA levels, providing evidence for the existence of an autocrine type I IFN signaling loop. On a functional level, we observed reduced PD-L1, MHC class I and class II expression and increased susceptibility to immune cell-mediated lysis upon disruption of IFN signaling, suggesting that constitutive IFN signaling in gliomas contributes to the immune evasion of glioma cells. Conclusions: Our findings point to an important role of constitutive IFN signaling in glioma cells in the regulation of anti-tumor immune responses.

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