Bioprinting stem cells: building physiological tissues one cell at a time

Bioprinting aims to direct the spatial arrangement in three dimensions of cells, biomaterials, and growth factors. The biofabrication of clinically relevant constructs for the repair or modeling of either diseased or damaged tissues is rapidly advancing, resulting in the ability to three-dimensional (3D) print biomimetic platforms which imitate a large number of tissues in the human body. Primary tissue-specific cells are typically isolated from patients and used for the fabrication of 3D models for drug screening or tissue repair purposes. However, the lack of resilience of these platforms, due to the difficulties in harnessing, processing, and implanting patient-specific cells can limit regeneration ability. The printing of stem cells obviates these hurdles, producing functional in vitro models or implantable constructs. Advancements in biomaterial science are helping the development of inks suitable for the encapsulation and the printing of stem cells, promoting their functional growth and differentiation. This review specifically aims to investigate the most recent studies exploring innovative and functional approaches for the printing of 3D constructs to model disease or repair damaged tissues. Key concepts in tissue physiology are highlighted, reporting stem cell applications in biofabrication. Bioprinting technologies and biomaterial inks are listed and analyzed, including recent advancements in biomaterial design for bioprinting applications, commenting on the influence of biomaterial inks on the encapsulated stem cells. Ultimately, most recent successful efforts and clinical potentials for the manufacturing of functional physiological tissue substitutes are reported here, with a major focus on specific tissues, such as vasculature, heart, lung and airways, liver, bone and muscle.

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Physical confinement signals regulate the organization of stem cells in three dimensions

Journal of The Royal Society Interface ◽

10.1098/rsif.2016.0613 ◽

2016 ◽

Vol 13 (123) ◽

pp. 20160613 ◽

Cited By ~ 7

Author(s):

Sebastian V. Hadjiantoniou ◽

David Sean ◽

Maxime Ignacio ◽

Michel Godin ◽

Gary W. Slater ◽

...

Keyword(s):

Stem Cells ◽

Inner Cell Mass ◽

Cell Mass ◽

Three Dimensional ◽

Embryonic Stem ◽

Cell Interactions ◽

Three Dimensions ◽

Cell Substrate ◽

Cell Cell

During embryogenesis, the spherical inner cell mass (ICM) proliferates in the confined environment of a blastocyst. Embryonic stem cells (ESCs) are derived from the ICM, and mimicking embryogenesis in vitro , mouse ESCs (mESCs) are often cultured in hanging droplets. This promotes the formation of a spheroid as the cells sediment and aggregate owing to increased physical confinement and cell–cell interactions. In contrast, mESCs form two-dimensional monolayers on flat substrates and it remains unclear if the difference in organization is owing to a lack of physical confinement or increased cell–substrate versus cell–cell interactions. Employing microfabricated substrates, we demonstrate that a single geometric degree of physical confinement on a surface can also initiate spherogenesis. Experiment and computation reveal that a balance between cell–cell and cell–substrate interactions finely controls the morphology and organization of mESC aggregates. Physical confinement is thus an important regulatory cue in the three-dimensional organization and morphogenesis of developing cells.

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Living the heart in three dimensions: applications of 3D printing in CHD

Cardiology in the Young ◽

10.1017/s1047951119000398 ◽

2019 ◽

Vol 29 (06) ◽

pp. 733-743 ◽

Cited By ~ 2

Author(s):

Mari Nieves Velasco Forte ◽

Tarique Hussain ◽

Arno Roest ◽

Gorka Gomez ◽

Monique Jongbloed ◽

...

Keyword(s):

3D Printing ◽

Heart Surgery ◽

Wide Spectrum ◽

Three Dimensional ◽

Structural Heart Disease ◽

Medical Personnel ◽

Three Dimensions ◽

Patient Specific ◽

3D Printed

AbstractAdvances in biomedical engineering have led to three-dimensional (3D)-printed models being used for a broad range of different applications. Teaching medical personnel, communicating with patients and relatives, planning complex heart surgery, or designing new techniques for repair of CHD via cardiac catheterisation are now options available using patient-specific 3D-printed models. The management of CHD can be challenging owing to the wide spectrum of morphological conditions and the differences between patients. Direct visualisation and manipulation of the patients’ individual anatomy has opened new horizons in personalised treatment, providing the possibility of performing the whole procedure in vitro beforehand, thus anticipating complications and possible outcomes. In this review, we discuss the workflow to implement 3D printing in clinical practice, the imaging modalities used for anatomical segmentation, the applications of this emerging technique in patients with structural heart disease, and its limitations and future directions.

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Designing Hydrogel-Based Bone-On-Chips for Personalized Medicine

Applied Sciences ◽

10.3390/app11104495 ◽

2021 ◽

Vol 11 (10) ◽

pp. 4495

Author(s):

Gabriele Nasello ◽

Mar Cóndor ◽

Ted Vaughan ◽

Jessica Schiavi

Keyword(s):

Three Dimensional ◽

In Vitro Models ◽

Patient Specific ◽

Specific Situation ◽

Culture Chamber ◽

Definition Of ◽

And Function ◽

Tissue Matrix ◽

Hematopoietic Niche

The recent development of bone-on-chips (BOCs) holds the main advantage of requiring a low quantity of cells and material, compared to traditional In Vitro models. By incorporating hydrogels within BOCs, the culture system moved to a three dimensional culture environment for cells which is more representative of bone tissue matrix and function. The fundamental components of hydrogel-based BOCs, namely the cellular sources, the hydrogel and the culture chamber, have been tuned to mimic the hematopoietic niche in the bone aspirate marrow, cancer bone metastasis and osteo/chondrogenic differentiation. In this review, we examine the entire process of developing hydrogel-based BOCs to model In Vitro a patient specific situation. First, we provide bone biological understanding for BOCs design and then how hydrogel structural and mechanical properties can be tuned to meet those requirements. This is followed by a review on hydrogel-based BOCs, developed in the last 10 years, in terms of culture chamber design, hydrogel and cell source used. Finally, we provide guidelines for the definition of personalized pathological and physiological bone microenvironments. This review covers the information on bone, hydrogel and BOC that are required to develop personalized therapies for bone disease, by recreating clinically relevant scenarii in miniaturized devices.

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Application of three-dimensional neuronal cell cultures in the studies of mechanisms of neurodegenerative diseases

Postępy Higieny i Medycyny Doświadczalnej ◽

10.5604/01.3001.0010.3832 ◽

2017 ◽

Vol 71 (1) ◽

pp. 0-0 ◽

Cited By ~ 2

Author(s):

Anna Słońska ◽

Joanna Cymerys

Keyword(s):

Neurodegenerative Diseases ◽

Cell Cultures ◽

Three Dimensional ◽

Neuronal Cell ◽

3D Models ◽

In Vitro Models ◽

Gene And Protein Expression ◽

Or Gene

In vitro models utilizing cells in planar two-dimensional (2D) cultures do not reflect the in vivo environment and are increasingly replaced by three-dimensional (3D) cultures. Fundamental differences between 2D and 3D cell cultures systems include cell attach, spread and grow, their morphology, proliferation, differentiation or gene and protein expression. For that reason 3D models have been proven to be invaluable tools of study for the various fields of science, such as drug discovery, cancer research, differentiation studies or neuroscience. In the present review, we discuss 3D neural in vitro models that might provide important insides about the mechanisms of pathogenesis of neurodegenerative diseases.

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Human Dental Pulp Stem Cells: recent findings and current research

Medical Journal of Cell Biology ◽

10.2478/acb-2019-0016 ◽

2019 ◽

Vol 7 (3) ◽

pp. 119-124

Author(s):

Krzysztof Janowicz ◽

Paul Mozdziak ◽

Artur Bryja ◽

Bartosz Kempisty ◽

Marta Dyszkiewicz-Konwińska

Keyword(s):

Stem Cells ◽

Dental Pulp ◽

Three Dimensional ◽

Clinical Problem ◽

Dental Pulp Stem Cells ◽

Patient Specific ◽

Culture Methods ◽

Regenerative Dentistry ◽

Promising Source

AbstractPrevalence of neurodegenerative diseases, most of which are life threatening and incurable, is an increasing clinical problem. To date, studies have demonstrated a superior proliferation rate of dental pulp stem cells (DPSCs) compared to other mesenchymal stem cells in vitro. DPSCs has recently been recognized as a novel treatment strategy for neurodegenerative disease, due to their advanced potential for neurogenic differentiation. The oral cavity has been described as a promising source of dental pulp stem cells. DPSCs are widely used in regenerative dentistry holding alternative capacity for osteogenic differentiation and therefore new promises for tissue and whole tooth regeneration. Dental stem cell banking offers a plentiful source of stem cells representing great potential for cell reprogramming and thus cell therapy. Recently, the association of pulp stem cells with three – dimensional scaffold templates allows for building up naturally derived implants. This review introduces to unique properties of DPSCs and biological factors influencing mineralization, proliferation and differentiation of pulp stem cells. Latest research studies are compared in terms of effectiveness and limitations of techniques for the isolation of pulp stem cells, including the enzymatic digestion and the explant culture methods. Moreover, a short overview of most recent findings and clinical application of DPSCs is proffered including progress of current research and limitations still to be addressed in the nearest future. Finally, the article presents new advances in the area of regenerative dentistry and regenerative medicine, including three dimensional printing and three dimensional analysis, emerged to deepen studies under procedures to replace the non patient specific artificial implants.Running title: DPSCs - review

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Comparative Analysis of Cell–Cell Contact Abundance in Ovarian Carcinoma Cells Cultured in Two- and Three-Dimensional In Vitro Models

Biology ◽

10.3390/biology9120446 ◽

2020 ◽

Vol 9 (12) ◽

pp. 446

Author(s):

Olga M. Kutova ◽

Ludmila M. Sencha ◽

Anton D. Pospelov ◽

Olga E. Dobrynina ◽

Anna A. Brilkina ◽

...

Keyword(s):

Tumor Microenvironment ◽

Connexin 43 ◽

Three Dimensional ◽

3D Models ◽

In Vitro Models ◽

Cell Contact ◽

Tumor Resistance ◽

Ovarian Adenocarcinoma ◽

Junction Protein

Tumor resistance to therapy is associated with the 3D organization and peculiarities of the tumor microenvironment, of which intercellular adhesion is a key participant. In this work, the abundance of contact proteins was compared in SKOV-3 and SKOV-3.ip human ovarian adenocarcinoma cell lines, cultivated in monolayers, tumor spheroids and collagen hydrogels. Three-dimensional models were characterized by extremely low expression of basic molecules of adherens junctions E-cadherin and demonstrated a simultaneous decrease in desmosomal protein desmoglein-2, gap junction protein connexin-43 and tight junction proteins occludin and ZO-1. The reduction in the level of contact proteins was most pronounced in collagen hydrogel, accompanied by significantly increased resistance to treatment with doxorubicin and targeted anticancer toxin DARPin-LoPE. Thus, we suggest that 3D models of ovarian cancer, especially matrix-based models, tend to recapitulate tumor microenvironment and treatment responsiveness to a greater extent than monolayer culture, so they can be used as a highly relevant platform for drug efficiency evaluation.

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Adipose-Derived Stem Cells Primed with Paclitaxel Inhibit Ovarian Cancer Spheroid Growth and Overcome Paclitaxel Resistance

Pharmaceutics ◽

10.3390/pharmaceutics12050401 ◽

2020 ◽

Vol 12 (5) ◽

pp. 401 ◽

Cited By ~ 1

Author(s):

Cinzia Borghese ◽

Naike Casagrande ◽

Giuseppe Corona ◽

Donatella Aldinucci

Keyword(s):

Ovarian Cancer ◽

Stem Cells ◽

Three Dimensional ◽

In Vitro Models ◽

Adipose Derived Stem Cells ◽

Cytotoxic Effects ◽

Spheroid Growth ◽

Ic50 Values ◽

Ovca Cell

Adipose-derived stem cells (ADSCs) primed with paclitaxel (PTX) are now hypothesized to represent a potential Trojan horse to vehicle and deliver PTX into tumors. We analyzed the anticancer activity of PTX released by ADSCs primed with PTX (PTX-ADSCs) (~20 ng/mL) in a panel of ovarian cancer (OvCa) cells sensitive or resistant to PTX. We used two (2D) and three dimensional (3D) in vitro models (multicellular tumor spheroids, MCTSs, and heterospheroids) to mimic tumor growth in ascites. The coculture of OvCa cells with PTX-ADSCs inhibited cell viability in 2D models and in 3D heterospheroids (SKOV3-MCTSs plus PTX-ADSCs) and counteracted PTX-resistance in Kuramochi cells. The cytotoxic effects of free PTX and of equivalent amounts of PTX secreted in PTX-ADSC-conditioned medium (CM) were compared. PTX-ADSC-CM decreased OvCa cell proliferation, was more active than free PTX and counteracted PTX-resistance in Kuramochi cells (6.0-fold decrease in the IC50 values). Cells cultivated as 3D aggregated MCTSs were more resistant to PTX than 2D cultivation. PTX-ADSC-CM (equivalent-PTX) was more active than PTX in MCTSs and counteracted PTX-resistance in all cell lines. PTX-ADSC-CM also inhibited OvCa-MCTS dissemination on collagen-coated wells. In conclusion, PTX-ADSCs and PTX-MSCs-CM may represent a new option with which to overcome PTX-resistance in OvCa.

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Pre-Clinical In Vitro Models Used in Cancer Research: Results of a Worldwide Survey

Cancers ◽

10.3390/cancers13236033 ◽

2021 ◽

Vol 13 (23) ◽

pp. 6033

Author(s):

Sarai Martinez-Pacheco ◽

Lorraine O’Driscoll

Keyword(s):

Cancer Research ◽

Three Dimensional ◽

3D Cell Culture ◽

Cost Effective ◽

Cell Types ◽

3D Models ◽

In Vitro Models ◽

3D Cultures

To develop and subsequently get cancer researchers to use organotypic three-dimensional (3D) models that can recapitulate the complexity of human in vivo tumors in an in vitro setting, it is important to establish what in vitro model(s) researchers are currently using and the reasons why. Thus, we developed a survey on this topic, obtained ethics approval, and circulated it throughout the world. The survey was completed by 101 researchers, across all career stages, in academia, clinical or industry settings. It included 40 questions, many with multiple options. Respondents reported on their field of cancer research; type of cancers studied; use of two-dimensional (2D)/monolayer, 2.5D and/or 3D cultures; if using co-cultures, the cell types(s) they co-culture; if using 3D cultures, whether these involve culturing the cells in a particular way to generate spheroids, or if they use additional supports/scaffolds; techniques used to analyze the 2D/2.5D/3D; and their downstream applications. Most researchers (>66%) only use 2D cultures, mainly due to lack of experience and costs. Despite most cancer researchers currently not using the 3D format, >80% recognize their importance and would like to progress to using 3D models. This suggests an urgent need to standardize reliable, robust, reproducible methods for establishing cost-effective 3D cell culture models and their subsequent characterization.

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3D Breast Tumor Models for Radiobiology Applications

Cancers ◽

10.3390/cancers13225714 ◽

2021 ◽

Vol 13 (22) ◽

pp. 5714

Author(s):

Akhilandeshwari Ravichandran ◽

Julien Clegg ◽

Mark N. Adams ◽

Madison Hampson ◽

Andrew Fielding ◽

...

Keyword(s):

Breast Cancer ◽

Radiation Therapy ◽

Three Dimensional ◽

3D Models ◽

In Vitro Models ◽

Breast Cancers ◽

Malignant Breast ◽

Effects Of Radiation ◽

Models Of Disease

Breast cancer is a leading cause of cancer-associated death in women. The clinical management of breast cancers is normally carried out using a combination of chemotherapy, surgery and radiation therapy. The majority of research investigating breast cancer therapy until now has mainly utilized two-dimensional (2D) in vitro cultures or murine models of disease. However, there has been significant uptake of three-dimensional (3D) in vitro models by cancer researchers over the past decade, highlighting a complimentary model for studies of radiotherapy, especially in conjunction with chemotherapy. In this review, we underline the effects of radiation therapy on normal and malignant breast cells and tissues, and explore the emerging opportunities that pre-clinical 3D models offer in improving our understanding of this treatment modality.

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Epigallocatechin-3-gallate inhibits the growth of three-dimensional in vitro models of neuroblastoma cell SH-SY5Y

Molecular and Cellular Biochemistry ◽

10.1007/s11010-021-04154-w ◽

2021 ◽

Author(s):

Xiao Wan ◽

Wenbo Wang ◽

Zhu Liang

Keyword(s):

Endothelial Cells ◽

Vascular Endothelial Cells ◽

Umbilical Vein ◽

Neuroblastoma Cell ◽

Three Dimensional ◽

3D Models ◽

In Vitro Models ◽

Neuroblastoma Cell Line ◽

Egcg Treatment

AbstractThe aim of the study is to investigate the potential of using three-dimensional (3D) in vitro neuroblastoma models to mimic the neuroblastoma microenvironment by testing a potential therapeutic compound—the natural extract epigallocatechin-3-gallate (EGCG), and to further elucidate the roles of DYRK1A in the growth and differentiation of neuroblastoma tissue. In vitro models based on a classic neuroblastoma cell line SH-SY5Y were employed, including 3D models with extracellular matrix and co-cultured with vascular endothelial cells. Cell viability was tested using AlamarBlue and Resazurin assay. The growth and differentiation of in vitro models of SH-SY5Y were analysed based on microscopy images obtained from immunofluorescence or real-time imaging. Protein expression level was investigated using immunoblotting analysis. The two-dimensional (2D) in vitro model implies the cytotoxicity and DYRK1A inhibition effect of EGCG and shows the induction of neuronal differentiation marker TuJ1. 3D in vitro models suggest that EGCG treatment compromised the growth of SH-SY5Y multicellular 3D spheroids and the viability of SH-SY5Y cultured in 3D Matrigel matrix. In addition, co-culture of SH-SY5Y with human vascular umbilical vein endothelial cells implied the inhibitory effects by EGCG in a vascularised microenvironment. In this study, novel 3D in vitro models of neuroblastoma were established in the application of testing a potential anti-cancer candidate compound EGCG. In pursuit of the goals of the 3Rs (replacement, reduction and refinement), the usage of these 3D in vitro models has the potential to reduce and eventually replace current animal models used in neuroblastoma research. The DYRK1A inhibiting nature of EGCG, together with the facts that EGCG inhibits the growth and induces the differentiation of neuroblastoma in vitro models, suggests an oncogene role of DRYK1A.

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