Mechanical and physicochemical behavior of a 3D hydrogel scaffold during cell growth and proliferation

Rivero, Rebeca Edith; Capella, Virginia; Liaudat, Ana Cecilia; Bosch, Pablo; Barbero, César Alberto; Rodríguez, Nancy; Rivarola, Claudia R.

doi:10.1039/c9ra08162c

Cited by 42 publications

(25 citation statements)

References 46 publications

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“…However, the resazurin-based assays are dependent on the cell metabolic state, and it is known that, in various conditions, cell metabolism can change, thus over-or under-estimation of the overall cell population might occur [18]. Assessment of the entire cellular DNA content is becoming a reliable method for cell proliferation analysis [28][29][30]. The development of a 3D compatible DNA concentration-based method was the goal of this study.…”

Section: Discussionmentioning

confidence: 99%

“…The other strategy is based on the quantification of cellular DNA and can measure cell number at the population level or even at the individual cell level [28]. These methods are generally based on stoichiometric fluorophore binding to DNA, meaning that cell number can be evaluated using fluorescent microscopy, PCR quantification, or fluorescence measurements [28][29][30]. Only several fluorophores are usually used for cell number evaluation in 2D and 3D-PicoGreen, CyQuant, and Hoechst 33258.…”

Section: Introductionmentioning

confidence: 99%

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DNA-DAPI Interaction-Based Method for Cell Proliferation Rate Evaluation in 3D Structures

Šimoliūnas

Kantakevičius

Kalvaitytė

et al. 2021

CIMB

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Effective cell number monitoring throughout the three-dimensional (3D) scaffold is a key factor in tissue engineering. There are many methods developed to evaluate cell number in 2D environments; however, they often encounter limitations in 3D. Therefore, there is a demand for reliable methods to measure cell proliferation in 3D surroundings. Here, we report a novel technique for the DNA content-based evaluation of cell proliferation using DNA-binding dye DAPI. We demonstrated the method’s compatibility with four different cell cultures: cancer lines MCF-7 and MH-22a, embryonic fibroblast cell line Swiss 3T3, and primary mesenchymal stem cell culture isolated from rat’s incisors. The DAPI based method was able to successfully evaluate cell proliferation in 2D, 2.5D, and 3D environments. Even though the proposed method does not discriminate between viable and dead cells, it might give a convenient snapshot of the cell number at a given time point. This should help to more reliably evaluate various processes proceeding in 2.5D and 3D cultures.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

DNA-DAPI Interaction-Based Method for Cell Proliferation Rate Evaluation in 3D Structures

Šimoliūnas

Kantakevičius

Kalvaitytė

et al. 2021

CIMB

View full text Add to dashboard Cite

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“…This is consistent with our previous studies, which showed that encapsulated cell growth was proportional to the rate of scaffold swelling/degradation [ 36 , 37 ]. Furthermore, hydrogel scaffold degradation via hydrolysis over time has been shown to lead to changes in the mechanical properties of the scaffolds [ 52 ]. As such, the mechanical strength and dimensions of the scaffolds play important roles in both the restriction of colony size and the rate of cell proliferation.…”

Section: Discussionmentioning

confidence: 99%

Transcriptomic Analysis of Naïve Human Embryonic Stem Cells Cultured in Three-Dimensional PEG Scaffolds

et al. 2020

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Naïve human embryonic stem cells (ESCs) are characterized by improved viability, proliferation, and differentiation capacity in comparison to traditionally derived primed human ESCs. However, currently used two-dimensional (2-D) cell culture techniques fail to mimic the three-dimensional (3-D) in vivo microenvironment, altering morphological and molecular characteristics of ESCs. Here, we describe the use of 3-D self-assembling scaffolds that support growth and maintenance of the naïve state characteristics of ESC line, Elf1. Scaffolds were formed via a Michael addition reaction upon the combination of two 8-arm polyethylene glycol (PEG) polymers functionalized with thiol (PEG-8-SH) and acrylate (PEG-8-Acr) end groups. 3-D scaffold environment maintained the naïve state and supported the long-term growth of ESCs. RNA-sequencing demonstrated significant changes in gene expression profiles between 2-D and 3-D grown cells. Gene ontology analysis revealed upregulation of biological processes involved in the regulation of transcription and translation, extracellular matrix organization, and chromatin remodeling in 3-D grown cells. 3-D culture conditions also induced upregulation of genes associated with Wnt and focal adhesion signaling, while p53 signaling pathway associated genes were downregulated. Our findings, for the first time, provide insight into the possible mechanisms of self-renewal of naïve ESCs stimulated by the transduction of mechanical signals from the 3-D microenvironment.

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“…[ 17 ] Basic in vitro studies have shed light on mammalian cell responses (cellular adhesion, proliferation, morphogenesis, and differentiation) when growing on surfaces or into structures of biocompatible hydrogels. [ 18,19 ] Studies conducted in our laboratory show that synthetic hydrogels based on poly(N‐isopropylacrylamide) (PNIPAM) and copolymers are non‐cytotoxic, [ 20 ] feature that make them ideal candidates to work with biological samples. Adhesion and proliferation of bovine fetal fibroblasts on hydrogel surfaces [ 19–21 ] and into 3D hydrogel structures, [ 18 ] bioethanol production by encapsulated Saccharomyces cerevisiae, [ 22 ] cytotoxicity and bioadhesive properties of murine preadipose cells (3T3‐L1), human embryonic kidney cells (HEK293), and human lung carcinoma‐derived cells (A549), [ 23 ] are some of the biologic systems used to characterize the physicochemical interactions between cells and polymeric materials based on PNIPAM and copolymers.…”

Section: Introductionmentioning

confidence: 99%

Interaction Between Hyaluronic Acid Semi‐Interpenetrated Hydrogel with Bull Spermatozoa: Studies of Sperm Attachment–Release and Sperm Quality

Blois

Liaudat

Capella

et al. 2021

Adv Materials Inter

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The study of living systems in contact with biocompatible synthetic hydrogels is developing. The authors’ group is the first to propose use of polymeric materials to accomplish sperm selection process by attachment of bull sperm to hydrogel surfaces, this being a crucial step for successful assisted reproductive techniques (ARTs). Surfaces based on poly(N‐isopropylacrylamide) hydrogels (PNIPAM) and ionic and neutral copolymers are synthetized. Their physicochemical properties in aqueous and culture medium are related to the percentage of attachment and subsequent release of bull spermatozoa from the hydrogel surface. High attaching capacity (30–45%) of sperm on cationic and neutral surfaces is mainly observed, however, sperm detachment from the hydrogel using traditional induction processes (i.e., sperm capacitation) is not significant. Therefore, PNIPAM‐co‐20%‐N‐Tris(hydroxymethyl)methyl acrylamide (PNIPAM‐HMA) hydrogel surface is semi‐interpenetrated with hyaluronic acid (HA). In this case, 50% of spermatozoa are attached to PNIPAM‐HMA‐HA hydrogel and after treatment with hyaluronidase, 47% of them are released. Collected sperm show acceptable characteristics of progressive motility (70% with score 4 for vigor), high viability, and cytoplasmic membrane integrity. Noteworthy, the study of the interaction of hydrogel surface/spermatozoa can be extended to human and other mammalian species, in order to provide advanced alternatives for gamete selection for ARTs.

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Mechanical and physicochemical behavior of a 3D hydrogel scaffold during cell growth and proliferation

Abstract: 3D cell scaffold based on macroporous PNIPAM is cytocompatible and preserves the cell viability for more than 75 culture days.

Cited by 42 publications

References 46 publications

DNA-DAPI Interaction-Based Method for Cell Proliferation Rate Evaluation in 3D Structures

DNA-DAPI Interaction-Based Method for Cell Proliferation Rate Evaluation in 3D Structures

Transcriptomic Analysis of Naïve Human Embryonic Stem Cells Cultured in Three-Dimensional PEG Scaffolds

Interaction Between Hyaluronic Acid Semi‐Interpenetrated Hydrogel with Bull Spermatozoa: Studies of Sperm Attachment–Release and Sperm Quality

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