Digital Image Analysis Applied to Tumor Cell Proliferation, Aggressiveness, and Migration-Related Protein Synthesis in Neuroblastoma 3D Models

Monferrer, Ezequiel; Sanegre, Sabina; Martín-Vañó, Susana; García-Lizarribar, Andrea; Burgos‐Panadero, Rebeca; López-Carrasco, Amparo; Navarro, Samuel; Samitier, J.; Noguera, Rosa

doi:10.3390/ijms21228676

Cited by 10 publications

(19 citation statements)

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“…Based on higher cluster and VN Frontiers in Cell and Developmental Biology frontiersin.org integrated densities, together with increased degradation of the stiff hydrogels, we infer that 3 wt% VN/PEG conditions better recreate NB cell behavior. However, cell adaptive response depends on stiffness, time and the genetic background of the cell line, as previously suggested in the literature (López-Carrasco et al, 2020;Monferrer, Sanegre, et al, 2020), so further genetic and drug-testing studies on long-cultured models, using various MYCN amplified and/or ALK mutated NB cell lines, should be performed to confirm which model better recapitulates high-risk NB behavior. Finally, VN synthesis even in VN rich ECM conditions strengthen the relevance of testing VN targeted preclinical therapies in NB.…”

Section: Discussionmentioning

confidence: 85%

“…In particular, our previous results highlighted VN as a relevant glycoprotein related to NB patients with poor prognosis (Burgos-Panadero, Vicente-Munuera et al, 2020). Our previous results with a clinical cohort and preclinical models (orthotopic xenograft VN knock-out (KO) mice and 3D bioprinted hydrogels with different stiffness) have also established that the interaction of VN, its ligands (e.g., αv integrins), and genomic intratumor heterogeneity in MYCN-amplified NB cell line are related to increased ECM stiffness (Burgos-Panadero, López-Carrasco et al, 2020;Monferrer, Sanegre, et al, 2020;Vicente-Munuera et al, 2020). Furthermore, we performed preclinical therapeutic studies on NB monolayer cell cultures, targeting VN function blockade by employing cilengitide (αv integrin antagonist) and combination therapy with etoposide-loaded (cytotoxin used in high-risk NB treatment) lipid nanoparticles (Burgos-Panadero et al, 2021).…”

Section: Introductionmentioning

confidence: 97%

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Vitronectin-based hydrogels recapitulate neuroblastoma growth conditions

Monferrer

Dobre

Trujillo

et al. 2022

Front. Cell Dev. Biol.

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The tumor microenvironment plays an important role in cancer development and the use of 3D in vitro systems that decouple different elements of this microenvironment is critical for the study of cancer progression. In neuroblastoma (NB), vitronectin (VN), an extracellular matrix protein, has been linked to poor prognosis and appears as a promising therapeutic target. Here, we developed hydrogels that incorporate VN into 3D polyethylene glycol (PEG) hydrogel networks to recapitulate the native NB microenvironment. The stiffness of the VN/PEG hydrogels was modulated to be comparable to the in vivo values reported for NB tissue samples. We used SK-N-BE (2) NB cells to demonstrate that PEGylated VN promotes cell adhesion as the native protein does. Furthermore, the PEGylation of VN allows its crosslinking into the hydrogel network, providing VN retention within the hydrogels that support viable cells in 3D. Confocal imaging and ELISA assays indicate that cells secrete VN also in the hydrogels and continue to reorganize their 3D environment. Overall, the 3D VN-based PEG hydrogels recapitulate the complexity of the native tumor extracellular matrix, showing that VN-cell interaction plays a key role in NB aggressiveness, and that VN could potentially be targeted in preclinical drug studies performed on the presented hydrogels.

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Section: Discussionmentioning

confidence: 85%

Section: Introductionmentioning

confidence: 97%

Vitronectin-based hydrogels recapitulate neuroblastoma growth conditions

Monferrer

Dobre

Trujillo

et al. 2022

Front. Cell Dev. Biol.

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“…In addition, cells in vivo may not only receive signals on one side but also receive signals from all directions (Baker and Chen, 2012). Therefore, cell migration is considered to be different in dimensionality (Rebelo et al, 2018;Young et al, 2018;Monferrer et al, 2020). The scratch assay is typically used to measure basic cell migration parameters such as speed, persistence and polarity.…”

Section: Introductionmentioning

confidence: 99%

Controlled Fabrication of Bioactive Microtubes for Screening Anti-Tongue Squamous Cell Migration Drugs

Tang

Shen

et al. 2022

Front. Chem.

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The treatment of tongue squamous cell carcinoma (TSCC) faces challenges because TSCC has an aggressive biological behavior and manifests usually as widespread metastatic disease. Therefore, it is particularly important to screen out and develop drugs that inhibit tumor invasion and metastasis. Two-dimensional (2D) cell culture has been used as in vitro models to study cellular biological behavior, but growing evidence now shows that the 2D systems can result in cell bioactivities that deviate appreciably the in vivo response. It is urgent to develop a novel 3D cell migration model in vitro to simulate the tumor microenvironment as much as possible and screen out effective anti-migration drugs. Sodium alginate, has a widely used cell encapsulation material, as significant advantages. We have designed a microfluidic device to fabricate a hollow alginate hydrogel microtube model. Based on the difference in liquid flow rate, TSCC cells (Cal27) were able to be evenly distributed in the hollow microtubes, which was confirmed though fluorescence microscope and laser scanning confocal microscope (LSCM). Our microfluidic device was cheap, and commercially available and could be assembled in a modular way, which are composed of a coaxial needle, silicone hose, and syringes. It was proved that the cells grow well in artificial microtubes with extracellular matrix (ECM) proteins by LSCM and flow cytometry. Periodic motility conferred a different motor state to the cells in the microtubes, more closely resembling the environment in vivo. The quantitative analysis of tumor cell migration could be achieved simply by determining the position of the cell in the microtube cross-section. We verified the anti-migration effects of three NSAIDs drugs (aspirin, indomethacin, and nimesulide) with artificial microtubes, obtaining the same results as conventional migration experiments. The results showed that among the three NSAIDs, nimesulide showed great anti-migration potential against TSCC cells. Our method holds great potential for application in the more efficient screening of anti-migration tumor drugs.

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“…In this regard, our group has previously described a common aggressive pattern of rigid ECM in HR-NB [14], rich in cross-linked collagen III fibres, poor in glycosaminoglycans, supporting sinusoidal vascular structures (blood and lymphatic) and with a high amount of territorial vitronectin (located in cytoplasmic compartments and a thin layer around tumour cells) [15][16][17][18]. In a recent study we observed the dramatic impact of ECM stiffness on ITH and clonal evolution of a MNA NB cell line, in which dominant clones were selected when cultured in stiff 3D-bioprinted hydrogels and in xenograft tumours; these changes were not found in an ALK mutated NB cell-line [3,19]. A continuous interplay therefore exists between TME and genetics, tumour cell proliferation, aggressiveness and migration [20].…”

Section: Introductionmentioning

confidence: 99%

Intra-Tumour Genetic Heterogeneity and Prognosis in High-Risk Neuroblastoma

López-Carrasco

Berbegall

Martín-Vañó

et al. 2021

Cancers

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Spatial ITH is defined by genomic and biological variations within a tumour acquired by tumour cell evolution under diverse microenvironments, and its role in NB patient prognosis is understudied. In this work, we applied pangenomic techniques to detect chromosomal aberrations in at least two different areas of each tumour and/or in simultaneously obtained solid and liquid biopsies, detecting ITH in the genomic profile of almost 40% of HR-NB. ITH was better detected when comparing one or more tumour pieces and liquid biopsy (50%) than between different tumour pieces (21%). Interestingly, we found that patients with ITH analysed by pangenomic techniques had a significantly better survival rate that those with non-heterogeneous tumours, especially in cases without MYCN amplification. Moreover, all patients in the studied cohort with high ITH (defined as 50% or more genomic aberration differences between areas of a tumour or simultaneously obtained samples) survived after 48 months. These results clearly support analysing at least two solid tumour areas (separately or mixed) and liquid samples to provide more accurate genomic diagnosis, prognosis and therapy options in HR-NB.

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Digital Image Analysis Applied to Tumor Cell Proliferation, Aggressiveness, and Migration-Related Protein Synthesis in Neuroblastoma 3D Models

Cited by 10 publications

References 49 publications

Vitronectin-based hydrogels recapitulate neuroblastoma growth conditions

Vitronectin-based hydrogels recapitulate neuroblastoma growth conditions

Controlled Fabrication of Bioactive Microtubes for Screening Anti-Tongue Squamous Cell Migration Drugs

Intra-Tumour Genetic Heterogeneity and Prognosis in High-Risk Neuroblastoma

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