Ultrasound-Based Scaffold-Free Core-Shell Multicellular Tumor Spheroid Formation

In cancer research and drug screening, multicellular tumor spheroids (MCTSs) are a popular model to bridge the gap between in vitro and in vivo. However, the current techniques to culture mixed co-culture MCTSs do not mimic the structural architecture and cellular spatial distribution in solid tumors. In this study we present an acoustic trapping-based core-shell MCTSs culture method using sequential seeding of the core and shell cells into microwells coated with a protein repellent coating. Scaffold-free core-shell ovarian cancer OVCAR-8 cell line MCTSs were cultured, stained, cleared and confocally imaged on-chip. Image analysis techniques were used to quantify the shell thickness (23.2 ± 1.8 µm) and shell coverage percentage (91.2 ± 2.8%). We also show that the shell thickness was evenly distributed over the MCTS cores with the exception of being slightly thinner close to the microwell bottom. This scaffold-free core-shell MCTSs formation technique and the analysis tools presented herein could be used as an internal migration assay within the MCTS or to form core-shell MCTS co-cultures to study therapy response or the interaction between tumor and stromal cells.

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Synthesis and characterization of core-shell mesoporous silica nanoparticles with various shell thickness as indomethacin carriers: In vitro and in vivo evaluation

Microporous and Mesoporous Materials ◽

10.1016/j.micromeso.2020.110043 ◽

2020 ◽

Vol 297 ◽

pp. 110043 ◽

Cited By ~ 2

Author(s):

Jia Ke ◽

Yumei Wang ◽

Lingmei Wang ◽

Baixue Yang ◽

Kaijun Gou ◽

...

Keyword(s):

Mesoporous Silica ◽

Silica Nanoparticles ◽

Shell Thickness ◽

Mesoporous Silica Nanoparticles ◽

Synthesis And Characterization ◽

Core Shell ◽

In Vivo Evaluation

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Transcriptome comparisons of in vitro intestinal epithelia grown under static and microfluidic gut-on-chip conditions with in vivo human epithelia

Scientific Reports ◽

10.1038/s41598-021-82853-6 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Kornphimol Kulthong ◽

Guido J. E. J. Hooiveld ◽

Loes Duivenvoorde ◽

Ignacio Miro Estruch ◽

Victor Marin ◽

...

Keyword(s):

Gene Expression ◽

Culture Conditions ◽

Gene Set Enrichment Analysis ◽

Shear Stresses ◽

Static Culture ◽

Dynamic Culture ◽

Intestinal Segments ◽

On Chip

AbstractGut-on-chip devices enable exposure of cells to a continuous flow of culture medium, inducing shear stresses and could thus better recapitulate the in vivo human intestinal environment in an in vitro epithelial model compared to static culture methods. We aimed to study if dynamic culture conditions affect the gene expression of Caco-2 cells cultured statically or dynamically in a gut-on-chip device and how these gene expression patterns compared to that of intestinal segments in vivo. For this we applied whole genome transcriptomics. Dynamic culture conditions led to a total of 5927 differentially expressed genes (3280 upregulated and 2647 downregulated genes) compared to static culture conditions. Gene set enrichment analysis revealed upregulated pathways associated with the immune system, signal transduction and cell growth and death, and downregulated pathways associated with drug metabolism, compound digestion and absorption under dynamic culture conditions. Comparison of the in vitro gene expression data with transcriptome profiles of human in vivo duodenum, jejunum, ileum and colon tissue samples showed similarities in gene expression profiles with intestinal segments. It is concluded that both the static and the dynamic gut-on-chip model are suitable to study human intestinal epithelial responses as an alternative for animal models.

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Astrocyte-Derived CCL2 is Associated with M1 Activation and Recruitment of Cultured Microglial Cells

Cellular Physiology and Biochemistry ◽

10.1159/000443040 ◽

2016 ◽

Vol 38 (3) ◽

pp. 859-870 ◽

Cited By ~ 27

Author(s):

Mingfeng He ◽

Hongquan Dong ◽

Yahui Huang ◽

Shunmei Lu ◽

Shu Zhang ◽

...

Keyword(s):

Central Nervous System ◽

Nervous System ◽

Microglial Activation ◽

Culture Media ◽

Microglial Cells ◽

Migration Ability ◽

Migration Assay ◽

Tnf Α

Background/Aims: Microglia are an essential player in central nervous system inflammation. Recent studies have demonstrated that the astrocytic chemokine, CCL2, is associated with microglial activation in vivo. However, CCL2-induced microglial activation has not yet been studied in vitro. The purpose of the current study was to understand the role of astrocyte-derived CCL2 in microglial activation and to elucidate the underlying mechanism(s). Methods: Primary astrocytes were pre-treated with CCL2 siRNA and stimulated with TNF-α. The culture medium (CM) was collected and added to cultures of microglia, which were incubated with and without CCR2 inhibitor. Microglial cells were analyzed by quantitative RT-PCR to determine whether they polarized to the M1 or M2 state. Microglial migratory ability was assessed by transwell migration assay. Results: TNF-α stimulated the release of CCL2 from astrocytes, even if the culture media containing TNF-α was replaced with fresh media after 3 h. CM from TNF-α-stimulated astrocytes successfully induced microglial activation, which was ascertained by increased activation of M1 and enhanced migration ability. In contrast, CM from astrocytes pretreated with CCL2 siRNA showed no effect on microglial activation, compared to controls. Additionally, microglia pre-treated with RS102895, a CCR2 inhibitor, were resistant to activation by CM from TNF-α-stimulated astrocytes. Conclusion: This study demonstrates that the CCL2/CCR2 pathway of astrocyte-induced microglial activation is associated with M1 polarization and enhanced migration ability, indicating that this pathway could be a useful target to ameliorate inflammation in the central nervous system.

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Contrasting migratory response of astrocytoma cells to tenascin mediated by different integrins

Journal of Cell Science ◽

10.1242/jcs.109.8.2161 ◽

1996 ◽

Vol 109 (8) ◽

pp. 2161-2168 ◽

Cited By ~ 5

Author(s):

A. Giese ◽

M.A. Loo ◽

S.A. Norman ◽

S. Treasurywala ◽

M.E. Berens

Keyword(s):

Cell Adhesion ◽

Matrix Protein ◽

Glioma Cells ◽

Extracellular Matrix Protein ◽

Integrin Receptors ◽

Migration Assay ◽

Dose Dependent Fashion ◽

Beta 1 Integrin

Tenascin, an extracellular matrix protein, is expressed in human gliomas in vitro and in vivo. The distribution of tenascin at the invasive edge of these tumors, even surrounding solitary invading cells, suggests a role for this protein as a regulator of glioma cell migration. We tested whether purified tenascin, passively deposited on surfaces, influenced the adhesion or migration of a human gliomaderived cell line, SF-767. Adhesion of glioma cells to tenascin increased in a dose-dependent fashion up to a coating concentration of 10 micrograms/ml. Higher coating concentrations resulted in progressively fewer cells attaching. Cell adhesion could be blocked to basal levels using anti-beta 1 integrin antibodies. In contrast, when anti-alpha v antibodies were added to the medium of cells on tenascin, cell adhesion was enhanced slightly. Using a microliter scale migration assay, we found that cell motility on tenascin was dose dependently stimulated at coating concentrations of 1 and 3 micrograms/ml, but migration was inhibited below levels of non-specific motility when tested at coating concentrations of 30 and 100 micrograms/ml. Migration on permissive concentrations of tenascin could be reversibly inhibited with anti-beta 1, while treatment with anti-alpha v antibodies increased migration rates. We conclude that SF-767 glioma cells express two separate integrin receptors that mediate contrasting adhesive and migratory responses to tenascin.

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Overview of existing in vitro BBB models: advantages and disadvantages, current state and future prospects

Complex Issues of Cardiovascular Diseases ◽

10.17802/2306-1278-2021-10-3-109-120 ◽

2021 ◽

Vol 10 (3) ◽

pp. 109-120

Author(s):

A. I. Mosiagina ◽

A. V. Morgun ◽

A. B. Salmina

Keyword(s):

Neurovascular Unit ◽

Clinical Trial Data ◽

Advantages And Disadvantages ◽

The Central Nervous System ◽

Complex Relationships ◽

On Chip ◽

Induced Pluripotent ◽

Level Of Understanding

There is growing research focusing on endothelial cells as separate units of the blood-brain barrier (BBB), and on the complex relationships between different types of cells within a neurovascular unit. To conduct this type of studies, researches use vastly different in vitro BBB models. The main objective of such models is to study the BBB permeability for different molecules, and to advance the current level of understanding the mechanisms of disease and to develop methods of targeted therapy for the central nervous system. The analysis of the existing Abstract in vitro BBB models and their advantages/disadvantages was conducted using the clinical trial data obtained in Russian/foreign countries. In this review, the authors highlight the most relevant assessment parameters and propose a unified classification of in vitro BBB models. According to the performed analysis, there is a tendency to move from 2D BBB models based on semipermeable inserts to 3D BBB spheroid and microfluidic organ-on-chip models. Moreover, the use of human induced pluripotent stem cells instead of animal primary cells will make it possible to reliably scale the results obtained in vitro to conditions in vivo.

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Berberine inhibits the proliferation of pancreatic cancer targeting pancreatic cancer stem cells

10.21203/rs.3.rs-282783/v1 ◽

2021 ◽

Author(s):

Mengmeng Liu ◽

Yue Pan ◽

Xufeng Tao ◽

Ning Li ◽

Kun Li ◽

...

Keyword(s):

Pancreatic Cancer ◽

Clinical Effectiveness ◽

The Novel ◽

Migration Assay ◽

Medicinal Value ◽

Anti Cancer ◽

Cancer Agent ◽

Pancreatic Cancer Stem Cells

Abstract BackgroundPDAC is universally acknowledged to be one of the highest mortality rate of cancer-related deaths. PCSCs, regulated by EMT, could promote the proliferation of PDAC. Berberine with high medicinal value has usually been used as an anti-cancer agent. Hence the purpose of this study is to investigate the anti-cancer effect of berberine in PDAC. MethodsMTT assay was used to verify berberine inhibiting the proliferation of PDAC. Immunofluorescence staining, stem cell sphere, wound healing and transwell migration assay were demonstrated the anti-proliferation and anti-stemness of PCSCs in vitro . PANC-02 cells were injected in C57BL/6 mice to establish the orthotopic pancreatic-cancer model in vivo . H&E and Ki67 immunohistogical staining assay were used to evaluated the effect of berberine in PDAC in vivo. q-PCR and Western blot methods were applied to detect the expression of EMT procedure.ResultsIn this study, berberine has selective anti-cancer effect in PDAC in vitro . Moreover, berberine suppressed the proliferation and stemness of PCSCs in PDAC. In vivo , berberine reduced the tumor size and decreased the expression of Ki67 in orthotopic pancreatic-cancer pancreases. In addition, berberine inhibit the EMT signaling pathway both in vitro and in vivo . ConclusionsOur study indicates that berberine inhibit the proliferation of PDAC in vivo and vitro . The mechanism of anti-cancer effect on berberine may suppress the PCSCs through inhibiting EMT procedure. Therefore, berberine may be the novel antineoplastic drug with clinical effectiveness in PDAC. Keywords: Berberine, PDAC, PCSCs, EMT, berberine

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ASAI KEMAMPUAN BAKTERI ENDOFIT DARI KACANG TANAH DALAM MENGHAMBAT PERTUMBUHAN Sclerotium sp. PADA KECAMBAH KACANG TANAH

JURNAL HAMA DAN PENYAKIT TUMBUHAN TROPIKA ◽

10.23960/j.hptt.214178-186 ◽

2020 ◽

Vol 14 (2) ◽

pp. 178-186

Author(s):

Lisa Novita Arios ◽

Dwi Suryanto . ◽

Kiki Nurtjahja . ◽

Erman Munir .

Keyword(s):

Endophytic Bacteria ◽

Culture Method ◽

Dry Weight ◽

Dual Culture ◽

Seedling Height ◽

Bacterial Isolates ◽

Peanut Seed ◽

Number Of Leaves

Assay on ability of endophytic bacteria isolated from peanut to inhibit Sclerotium sp. growth in peanut seedlings. A study on assay of ability of endophytic bacteria to inhibit Sclerotium sp. in peanut seedling has been done. The bacteria were isolated from peanut healthy plants, while Sclerotium sp. was isolated from infected peanaut plant. Antagonistic assay was conducted by dual culture method. In vivo assay of inhibiting Sclerotium sp. was conducted by dipping peanut seed in bacterial solution, and planting the seed in soil:compost (3:1) growing media. Six endophytic bacterial isolates showed to inhibit the growth of Sclerotium sp. in vitro. LN1 seemed to inhibit more of Sclerotium sp., while LN5 showed to inhibit less. Two potential isolates LN1 of gram-negative and LN2 of gram-positive using for further study showed to decrease more of dumping off. It also seemed that the isolates increased the seedling height, number of leaves, and dry weight.

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CTCF Interacts with and Recruits the Largest Subunit of RNA Polymerase II to CTCF Target Sites Genome-Wide

Molecular and Cellular Biology ◽

10.1128/mcb.01993-06 ◽

2007 ◽

Vol 27 (5) ◽

pp. 1631-1648 ◽

Cited By ~ 104

Author(s):

Igor Chernukhin ◽

Shaharum Shamsuddin ◽

Sung Yun Kang ◽

Rosita Bergström ◽

Yoo-Wook Kwon ◽

...

Keyword(s):

Rna Polymerase ◽

Rna Polymerase Ii ◽

Gene Activation ◽

Ctcf Binding ◽

Pol Ii ◽

Genome Wide ◽

Target Sites ◽

On Chip

ABSTRACT CTCF is a transcription factor with highly versatile functions ranging from gene activation and repression to the regulation of insulator function and imprinting. Although many of these functions rely on CTCF-DNA interactions, it is an emerging realization that CTCF-dependent molecular processes involve CTCF interactions with other proteins. In this study, we report the association of a subpopulation of CTCF with the RNA polymerase II (Pol II) protein complex. We identified the largest subunit of Pol II (LS Pol II) as a protein significantly colocalizing with CTCF in the nucleus and specifically interacting with CTCF in vivo and in vitro. The role of CTCF as a link between DNA and LS Pol II has been reinforced by the observation that the association of LS Pol II with CTCF target sites in vivo depends on intact CTCF binding sequences. “Serial” chromatin immunoprecipitation (ChIP) analysis revealed that both CTCF and LS Pol II were present at the β-globin insulator in proliferating HD3 cells but not in differentiated globin synthesizing HD3 cells. Further, a single wild-type CTCF target site (N-Myc-CTCF), but not the mutant site deficient for CTCF binding, was sufficient to activate the transcription from the promoterless reporter gene in stably transfected cells. Finally, a ChIP-on-ChIP hybridization assay using microarrays of a library of CTCF target sites revealed that many intergenic CTCF target sequences interacted with both CTCF and LS Pol II. We discuss the possible implications of our observations with respect to plausible mechanisms of transcriptional regulation via a CTCF-mediated direct link of LS Pol II to the DNA.

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