Organoid > Volume 3; 2023 > Article
Jo: Natural biopolymer-based hydrogels as designer matrices for organoid cultures

Abstract

Matrigel, a mouse sarcoma-derived extract, is considered the gold standard for organoid cultures. However, it has several drawbacks, including inconsistent and ill-defined composition, varying quality between batches, and potential cancer-related health risks. These factors highlight the need to develop chemically defined alternatives to Matrigel. Natural biopolymers derived from living organisms have emerged as promising substitutes capable of creating chemically defined extracellular matrix (ECM)-mimicking materials to support organoids in a 3-dimensional (3D) environment. This article provides an overview of natural biopolymeric hydrogel-based bioengineering approaches for constructing 3D matrices resembling artificial ECM for organoid cultures. It discusses the latest developments in utilizing natural biopolymers to direct the growth, differentiation, and maturation of organoids, along with their translational applications in the fields of bioengineering and biomedicine. Additionally, the article offers perspectives on multidisciplinary research on natural biopolymer-based hydrogels for more practical applications as next-generation matrices for organoid cultures.

Introduction

Organoids are 3-dimensional (3D) mini-organs formed by the self-organization of stem cells, progenitor cells, or tissue fragments, in the presence of biophysical and biochemical signals that simulate the corresponding organ’s in vivo milieu [1,2]. Their tissue-specific structural and functional characteristics, along with their multicellular complexity, provide a potent platform for advancing organ developmental research, disease modeling, drug screening, and tissue engineering [3-5]. Most organoid culture systems have relied heavily on Matrigel, a basement membrane extract produced from Engelbreth-Holm-Swarm mouse sarcoma [6]. Although Matrigel provides a highly functional matrix for cell proliferation and differentiation due to its remarkable stem cell niche signaling properties and tissue-like mechanical properties [7], its heterogeneity, batch-to-batch variation, and ill-defined composition lead to uncontrollable microenvironments and thus poor reproducibility of organoids [8]. Furthermore, the mouse tumor origin of Matrigel limits its use for therapeutic transplantation in vivo due to potential risks of immunogenicity and carcinogenicity [9]. A chemically defined extracellular matrix (ECM)-mimicking material that would allow the precise modulation of the physical and biochemical properties of cellular microenvironments and thus guarantees more consistent generation of organoids is urgently needed to achieve downstream translational applications such as drug screening, tissue engineering, and personalized medicine.
Natural biopolymers, which are biomacromolecules sourced from living organisms such as bacteria, plants, and animals, have become increasingly popular in the creation of hydrogels that mimic the ECM. These hydrogels provide three-dimensional support for organoids. This popularity is due to the molecular structures of natural biopolymers, which closely resemble the ECM, as well as their superior biocompatibility and biodegradability [10-13]. The high content of functional groups (for example, hydroxyl, amino, and carboxylic acid groups) in natural biopolymers allows for the easy impartation of desired bioactivities to cellular microenvironments [14]. This is particularly beneficial when customizing the microenvironmental cues of an artificial 3D matrix. In addition, the abundance of biopolymers in the natural world and their ability to be produced in a cost-effective manner make them highly attractive for organoid cultures. These cultures can be utilized in various industrial fields, ranging from biomedicine (including regenerative therapy, biobanks, and personalized medicine), to foods, pharmaceuticals, and cosmetics.
In this study, we present an overview of cutting-edge natural biopolymer-based hydrogels as a bioengineered matrix for organoid culture. First, we describe the important biochemical properties of natural biopolymers for the creation of hydrogels (Table 1). Next, we present recent advances in organoid culturing in (1) polysaccharide- and (2) protein-based hydrogels. Finally, we examine the existing problems with these natural biopolymeric hydrogels for artificial ECM engineering, as well as future views and prospects.

Preparation of natural biopolymer-based hydrogels

Ethics statement: This study was a literature review of previously published studies and was therefore exempt from institutional review board approval.
Natural biopolymer-based hydrogels have been evaluated as 3D matrices for cell culture due to their ability to deliver superior nutrition to cells, protect cells and delicate medications, and have inherent biocompatibility [15]. The abundance of hydrophilic groups in the backbone of these hydrogels contributes to the matrices’ high water absorption capacity, which is beneficial for cell growth [16]. The formation of complex structures, along with the enhancement of mechanical properties and stability in physiological environments, is achieved through inter/intramolecular crosslinking within the biopolymeric chain. This crosslinking responds to changes in environmental conditions such as pH and temperature, or the introduction of a crosslinking agent like a chemical crosslinker [17-21]. However, physical or chemical crosslinking can lead to a reduction in the availability of functional groups in biopolymers and poorer degradability [22]. Therefore, an effective crosslinking method is crucial for producing hydrogels intended for successful organoid applications (Fig. 1).
Biopolymers can self-assemble into aggregates through reversible physical interactions such as hydrogen bonding, electrostatic interactions, hydrophobic interactions, or host-guest interactions [23-27]. This process allows the creation of hydrogels under mild conditions without the need for crosslinking agents, which is beneficial for encapsulating cells or biomolecules, as it reduces the risk of unwanted interactions with bioactive agents [28]. Physically crosslinked biopolymeric hydrogels are often sensitive to changes in environmental factors such as pH, temperature, or ionic strength, allowing for variations in hydrogel matrix mechanics [29]. The reversible nature of physically crosslinked hydrogels allows for dynamic behaviors such as shear-thinning or self-healing, as well as native ECM-like physicochemical properties, which offer bioengineering options for enhanced organoid formation [30].
Covalently crosslinked biopolymeric hydrogels, which are produced by covalent bonds between polymer chains (e.g., thiol-ene click chemistry and Schiff’s base reaction), have improved mechanical properties in vivo due to their relatively robust gel networks compared to those formed by physical interactions [31]. The rapid gelation period of these hydrogels, typically less than 10 minutes, is achieved through strong covalent bonding under physiologically moderate conditions. This has led to a particular interest in enzymatic crosslinking and photo-crosslinking for the in situ formation of hydrogel matrices [32,33]. Covalently crosslinked hydrogels, in general, act as linearly elastic materials, and organoid morphogenesis may be physically impeded owing to the poor dissipation of significant compressive pressures during colony expansion [34,35]. To circumvent the irreversible nature of covalent bonding, which causes the progressive softening of covalently crosslinked hydrogels, many techniques for enzymatic degradation [36-38], photo-responsive degradation [39], and passive hydrolysis [34,40] have been proposed.

Polysaccharide-based hydrogels

Polysaccharides are polymeric carbohydrates that consist of multiple monosaccharide units covalently linked by glycosidic bonds [41,42]. Many polysaccharides possess ionizable functional groups, such as amines (-NH3+) in chitosan and carboxylates (-COO-) in hyaluronic acid (HA), and the distribution of those functional groups determines their charge density in different pH environments [43]. The physicochemical properties of polysaccharides can be modified through physical, chemical, and enzymatic processes [44]. Hydrogels based on polysaccharides have recently gained attention as platforms for culturing and/or delivering organoids (Table 2) due to their high water-retaining capacity, biocompatibility, and biodegradability [45-59].

1. Alginate-based hydrogels

Alginate is a linear polysaccharide composed of negatively charged 1,4-linked β-D-mannuronic acid (M-block) and α-L-guluronic acid (G-block) units (Fig. 2A). It is obtained from brown algae through alkali treatment [47,60]. This polysaccharide, approved by the Food and Drug Administration, has garnered significant attention for cell encapsulation techniques due to its low toxicity and ease of manipulation [60,61]. Alginate offers several advantages for organoid culture, including cost-effectiveness, the ability to modulate physical and biochemical properties [62,63], and its viscoelastic nature [64].
The addition of multivalent cations, with Ca2+ being the most commonly used, enables the rapid gelation of alginate through ionic crosslinking under mild conditions (Fig. 2B) [10,65]. Calcium-crosslinked alginate hydrogels (Ca-alginate hydrogels) have been explored for growing mouse small intestinal stem cell-derived intestinal organoids [48]. However, the colony formation efficiency of mouse small intestinal stem cells within the Ca-alginate hydrogel was significantly lower compared to the counterpart grown in Matrigel (Fig. 2C and 2D). Conversely, another study has proposed that Ca-alginate hydrogel can support the growth and development of human intestinal organoids both in vitro and in vivo, despite its lack of cell adhesive properties (Fig. 2E and 2F) [47]. By culturing human pluripotent stem cell (PSC)-derived hindgut spheroids within a Ca-alginate hydrogel with an appropriate level of stiffness for approximately 30 days, the researchers were able to generate intestinal organoids that closely resembled Matrigel-grown organoids. Moreover, the resulting Ca-alginate hydrogel-grown intestinal organoids exhibited engraftment and maturation after transplantation in vivo to a similar extent as Matrigel-grown organoids, suggesting the potential applicability of Ca-alginate hydrogel as an alternative matrix to Matrigel for culturing intestinal organoids.
Alginate’s inertness and biostability enable cell-specific spatiotemporal imaging and tracking of cells trapped inside an alginate hydrogel for lengthy periods of time [66]. In general, embedding organoids in 3D hydrogels reduces nutrient delivery during conventional static culture because the gel matrix functions as an additional barrier to solute diffusion, making the culture of organoids with high metabolic activity, such as islets, highly unstable for maintaining long-term cell viability and function [66]. However, the continuous dynamic culture of human and rodent pancreatic islets within a 3D alginate hydrogel gelled by BaCl2 solution allowed for the elucidation of complex islet physiological and pathophysiological processes via optical assessment and functional assays using a microphysiological system [49].
Alginate beads were crosslinked using BaCl2 and functionalized with type I collagen, in order to build a platform for disease modeling and medication development for lung illnesses such as idiopathic pulmonary fibrosis [50]. Culture of fetal lung fibroblasts with functionalized alginate beads resulted in the production of cohesive organoids as a consequence of cellular adhesion to the bead surface and subsequent cellular growth and contraction, enabling the formation of self-assembled human lung tissue encompassing numerous cell types.
Alginate modification enables cation-free crosslinking of hydrogels, which may alter their mechanical characteristics and enhance bioactivity [47,67]. The covalent modification of alginate with norbornene (NB-alginate) results in a UV-crosslinkable hydrogel through thiol-ene chemistry, allowing the creation of an ECM-like environment that allows the unhindered passage of most nutrients in lower concentrations of hydrogel [51]. When compared to a standard culture technique on the air-liquid interface, simple encapsulation of kidney organoids within the NB-alginate hydrogel resulted in lower expression of aberrant type 1a1 collagen, with no alterations in organoid structural shape. This synthetic microenvironment, which replicates the in vivo circumstances of the growing kidney, has shown the potential for producing organoids for therapeutic applications.
Furthermore, the potential and relevance of modifying hydrogel characteristics to regulate kidney organoids have been demonstrated using oxidized alginate hydrogels created by imine-type dynamic covalent crosslinking [52]. Kidney organoids were encapsulated in three different stiffness levels (0.1-20 kPa) of hydrogels and two soft hydrogels (0.1 kPa) with adjustable stress relaxation, following induced PSC (iPSC) differentiation (7 days) and air-liquid interface culture (14 days). Kidney organoids grown in soft, rapidly relaxing hydrogels showed a higher degree of maturity in terms of renal structure formation and the expression of epithelial-mesenchymal transition markers, compared to those grown in stiffer or slow-relaxing hydrogels.

2. Hyaluronic acid-based hydrogels

The negatively charged polysaccharide HA is composed of D-glucuronic acid and N-acetyl-D-glucosamine. Because of its capacity to bind with transmembrane receptors (e.g., CD44, CD54, and CD168), HA, a non-sulfated glycosaminoglycan (GAG) abundantly found in the ECM, is implicated in different signaling cascades that impact cell attachment, migration, proliferation, and morphogenesis [68]. HA may be used to create organoid microenvironments because of its high bioactivity; however, its poor degradation rates and unfavorable mechanical properties make it difficult to apply alone in hard tissue [69]. Hybrid hydrogels formed by incorporating HA into enzymatically crosslinked poly(ethylene glycol) (PEG) matrices demonstrated the ability to maintain, expand, or differentiate human bone marrow-derived stromal cells and human hematopoietic stem cells in vitro, eventually generating bone marrow organoids [53]. Furthermore, another HA/chitosan hybrid hydrogel has been reported to promote cerebral organoid development by iPSC culture in Essential 8 (E8) media without the inclusion of neural induction components [54]. Within 10 to 14 days of culture, iPSCs encapsulated in the HA-chitosan hydrogel showed morphological features of cerebral organoids and growth up to 3 mm in the greatest dimension at day 28, while exhibiting specific behaviors of early corticogenesis (e.g., neural rosette and neural tube-like structures).

3. Heparin-based hydrogels

Heparin is a negatively charged polysaccharide composed of L-iduronic acid and D-glucosamine repeats [70]. This highly sulfated GAG molecule has been shown to regulate cell signaling by sequestering heparin-binding domain-associated growth factors [71], as well as blocking coagulation and thrombosis [72]. Heparin’s capacity to bind and stabilize numerous growth factors and proteins, in particular, makes it useful as a building element of 3D matrices for organoid cultures [73,74]. The use of a matrix metalloproteinase-cleavable peptide linker to crosslink heparin hydrogel with 4-armed PEG was shown to stimulate the morphogenesis of proximal tubule epithelial cells (HK-2) into physiologically sized tubule structures [55]. The resultant tubules demonstrated the form and function of the in vivo renal proximal tubule and responded to nephrotoxins, demonstrating their potential for disease modeling and medication toxicity research. This heparin hydrogel was also used to cultivate multicellular polarized mammary epithelial organoids [56]. Human mammary epithelial cells immersed in heparin-based hydrogel demonstrated laminin secretion and organization into basement membrane-like assemblies, enhancing integrin signaling and encouraging the creation of polarized acini.

4. Cellulose-based hydrogels

Cellulose is a plant-derived structural polysaccharide that is separated into nanofibers with diameters ranging from 2 to 60 nm [75,76]. The oxidation mediated by 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) enables the conversion of cellulose strands’ main alcohol groups into negatively charged carboxyl groups (Fig. 3A) [77]. When compared to Matrigel, the resulting cellulose nanofibrils (CNFs) form hydrogels with remarkable mechanical properties, such as rapid self-healing and shear-thinning behavior, supporting the differentiation of liver organoids, while exhibiting comparable or even superior levels of hepatic gene expression, hepatocyte function and organoid polarization [57]. The functionalization of oxidized CNFs with a fibronectin-derived cell adhesion moiety, RGD peptide, has shown promise in improving cellular contact between organoids and the cellulose backbone (Fig. 3B and 3C) [58]. Ca2+-mediated ionic crosslinking between CNF carboxyl groups created a milieu conductive to the development and budding of tiny intestine organoids. Furthermore, TEMPO-periodate oxidation has been shown to allow the incorporation of a greater number of carboxyl groups into the cellulose molecule [78], resulting in the functionalization of CNFs with more RGD peptides than CNFs treated alone with TEMPO-mediated oxidation [59]. Mg2+-generated RGD-grafted CNF-based hydrogel-based cationic crosslinking aided in the development of intestinal organoids, while also enabling their long-term culture following passage.

Protein-based hydrogels

Proteins are biomacromolecules composed of numerous amino acids linked by peptide bonds [79,80]. They serve as the essential building blocks for highly structured systems that underpin life's critical functions [10]. The sequence of amino acids in a polypeptide chain dictates the protein's physicochemical properties, such as molecular weight, shape, and hydrophobicity, as well as its biological characteristics [43,79,80]. The process of proteins unfolding and refolding mechanically results in inherent viscoelasticity, akin to that of the ECM [81]. The limitless design possibilities and diverse functions of proteins, coupled with their excellent biocompatibility and biodegradability [82], make them particularly attractive for creating hydrogel matrices for organoid cultures (Table 3) [48,83-97].

1. Collagen-based hydrogels

Collagen is the most prevalent ECM protein, providing mechanical support to vertebrate connective tissues and promoting cell proliferation, migration, and differentiation [83,98,99]. The main structure of collagen is a triple-stranded helix stabilized by intra- and inter-chain hydrogen bonding, which is responsible for collagen’s thermo-responsive activity [100,101]. Due to its low antigenicity and high mechanical strength, collagen is often proposed for the construction of fibrous matrices in organoid culture [67,84,85,102]. In particular, the biomimetic properties of collagen make its hydrogel amenable to cell adhesion without modification and capable of presenting a native viscoelastic environment for resident cells [98].
Collagen-based hydrogels have been shown to facilitate the growth of mouse and human organoids for the gastrointestinal system, including the small intestine, colon, and stomach, in non-toxic, favorable environments for both organoids and normal tissue [86]. These collagen hydrogels exhibit thermo-responsive behavior, transforming into a solution at lower temperatures of 4 to 8 °C and a gel when heated to 37 °C. When compared to Matrigel-grown organoids, the organoids produced in collagen hydrogels had identical shapes, specific markers, and proliferation rates. Notably, in an ethylenediaminetetraacetic acid-colitis animal model, the transplantation of mouse colon organoids with the collagen hydrogel resulted in effective engraftment in injured tissue, showing the usefulness of regenerative medicine in vivo [86]. After in vivo engraftment, mouse and human intestinal organoids co-cultured with intestinal subepithelial myofibroblasts in collagen hydrogel recapitulated an autonomous experimental stem cell niche [85,87]. Furthermore, centimeter-long macroscopic units of intestinal epithelium were generated by embedding intestinal organoids in a floating collagen hydrogel [84]. Proliferating organoids aligned and fused to form a hollow structure of epithelial tubes containing all intestine-specific cell types, including Lgr5+ stem cells. The combination of collagen and CNFs has been investigated as a way to produce hybrid hydrogels with improved mechanical properties and bioactive effects, allowing the embedded crypts to undergo epithelial budding while maintaining cell viability and metabolic activity and expressing tissue-specific cell markers [88].
Human basal cells placed in floating collagen hydrogels grew mammary gland organoids, demonstrating the role of mechanical signals in controlling ductal branch elongation [89]. This process involved the cells migrating back and 4th within the surrounding collagen network, generating tension, promoting branch outgrowth, and causing plastic deformation of the matrix. The identified equilibrium of mechanical tension has been suggested as a promising area for future research into branching morphogenesis during organogenesis.

2. Gelatin-based hydrogels

Gelatin is a fibrous protein generated from the denaturation or hydrolysis of native collagen that is more water-soluble and less immunogenic than collagen [103-105]. It forms gels at approximately 30 °C by transitioning gelatin chains from disordered random coils to ordered helices through hydrogen bonding [106]. This process results in an intermediate biological complexity matrix following in situ gelation [101]. In addition, several advantageous properties of gelatin, such as biocompatibility, biodegradability, and the capability to promote cell adhesion and proliferation, make it highly attractive for organoid applications [103,105]. However, the poor mechanical properties and short degradation times, especially under physiological conditions, of native gelatin necessitate crosslinking for practical applications [107]. The covalent crosslinking of a gelatin-based hydrogel with 8-arm PEG was achieved through an enzymatic reaction with coagulation factor XIII (FXIII) [90]. This hybrid hydrogel demonstrated the ability to support cell differentiation and matrix secretion for liver organoid creation, with the potential to further stimulate tissue development by fine-tuning hydrogels and covalently immobilizing essential proteins.

3. Fibrin-based hydrogels

Fibrin is a blood protein formed by the activation of the serine protease thrombin during the coagulation process [108]. It possesses an abundance of naturally occurring RGD adhesion domains, which makes it useful as a substrate for cell proliferation and improved ECM deposition [108,109]. However, like other natural biopolymeric materials, fibrin has poor mechanical properties, necessitating a crosslinking approach [98]. FXIII-mediated enzymatic crosslinking of fibrin-based hydrogels has been used to enable the development of mouse intestinal organoids, as well as human intestinal, pancreatic, and liver organoids [48]. Notably, the combination of internal pressure and increased cell contractility inside the hydrogel enabled the formation of budding intestine organoids. Furthermore, laminin supplementation supported the long-term development of all tested epithelial organoids, demonstrating its effectiveness as a specific alternative to Matrigel.
An oil-free droplet microfluidic method was designed to fabricate hydrogel capsules with a fibrin hydrogel core and an alginate-chitosan composite shell in a single step (Fig. 4A) [91]. The produced hydrogel capsules with the prescribed compositions demonstrated good homogeneity and stability, as well as outstanding biocompatibility and high-throughput productivity. Human iPSC-derived hepatic cells self-organized into liver organoids with consistent sizes, composed of hepatocyte- and cholangiocyte-like cells in the core hydrogel generated by the enzymatic reaction of fibrinogen and thrombin (Fig. 4B). The produced liver organoids demonstrated the preservation of liver-specific activities, such as urea production and albumin secretion, indicating a successful recapitulation of the fundamental properties of the human liver (Fig. 4C and 4D).

4. Recombinant protein-based hydrogels

Over the past two decades, recombinant protein-based hydrogels have advanced rapidly, thanks to considerable improvements in recombinant DNA technology and protein engineering [110-113]. In general, recombinant protein-based hydrogels demonstrate superior mechanical properties and consistency between batches when compared to their natural protein-based counterparts [110,114-116]. Genetic engineering of the amino acid sequence allows for precise control over the structural and functional aspects of protein building blocks, such as folding structure, chain length, and stereochemistry [117]. Specifically, the integration of signaling sequences into recombinant proteins provides the building blocks for artificial microenvironments that mimic the ECM [118-120].
Elastin-like proteins (ELPs) are recombinantly produced protein polymers consisting of conserved repeating units, as observed in tropoelastin’s hydrophobic domains (Fig. 5A) [121]. The pentapeptide Val-Pro-Gly-Val-Gly (VPGVG) is the most common repeating motif with lower critical solution temperature phase behavior [121,122]. Most ELPs are composed of the Val-Pro-Gly-X-Gly (VPGXG) pentapeptide repeat, where different physicochemical features may be accurately programmed depending on which amino acid is present in the guest residue “X” [123]. ELPs offer a modulable design due to their inherent biocompatibility, biodegradability, stimuli-responsiveness, and viscoelastic properties, making them potential biomaterials for 3D cell cultures, including ECM mimetics [123]. A genetic union of an ELP-based structural backbone with a fibronectin-derived, cell-adhesive, extended RGD sequence resulted in the development of a recombinantly designed ECM (Fig. 5B) [92]. Engineered ECM-based hydrogels were created by chemically crosslinking lysine residues with tetrakis(hydroxymethyl) phosphonium chloride and provided a microenvironment suitable for the formation and growth of mouse intestinal organoids by providing cell adhesive biochemical cues and elastomeric biomechanical cues. A recombinant triblock protein (PEP) made up of two leucine zippers (P) separated by an ELP (E) was also used to create a 3D matrix for organoid development (Fig. 5C) [93]. The PEP protein can self-associate into hydrogels thanks to its coiled-coil helix domains. Furthermore, combining PEP with cell-binding ECM motifs derived from fibronectin or laminin alpha 3, which are key components in pancreatic endocrine activities, resulted in the formation of pancreatic organoids composed of primary endocrine and endocrine progenitor cells.
Spider silk, due to its exceptional biocompatibility and unique mechanical properties, can be utilized to construct 3D matrices for organoid culturing [124]. Recombinant spider silk, which is inspired by its natural counterpart, has been suggested as a consistent and highly reproducible scaffold material for 3D cell culturing [125]. Microfibers of recombinant spider silk protein undergo thermal transition polymerization, resulting in sturdy, elastic, and biocompatible matrices that facilitate the self-assembly of human PSCs into cerebral organoids [94,95]. When human PSCs are introduced into these silk microfiber networks, they stimulate neuroectoderm development and organoid maturation in relation to neuronal functioning [95].

5. Peptide-based hydrogels

Peptides, which are short chains of naturally occurring amino acids (usually 2 to 50 residues) connected by peptide bonds, can also be used to combine the beneficial qualities of natural and synthetic matrices for organoid cultures [126]. The simplicity of peptides’ structure allows for a more predictable design of hydrogels based on their sequences, compared to recombinant proteins, despite their relatively lower mechanical properties [127]. Among these, self-assembling peptides have been engineered to spontaneously form fibrillar structures in aqueous solutions, leading to physical gelation with architecture and characteristics similar to native ECM [127,128]. When human iPSCs were implanted in self-assembling peptide hydrogels, they successfully generated kidney organoids with complex architectures comparable to those in Matrigel [96]. Moreover, chemically crosslinked peptide hydrogels composed of specific ECM protein-mimicking fragments, such as collagen-like peptide (CLP) and CLP combined with RGD peptide (CLP-RGD), showed improved neural cell differentiation. This resulted in the rapid development of self-assembled cerebellar organoids [97]. Primary cerebellar cells spontaneously organized into tissue-like clusters capable of producing action potentials within the elastomechanical environment of ECM-mimetic matrices of CLP-based hydrogels.

Summary and perspectives

In this review, we highlight recent advances in natural hydrogels composed of polysaccharides or proteins, specifically for the production of organoids. These hydrogels have significant biological and biomedical applications, including developmental studies, disease modeling, drug screening, and regenerative medicine. Natural biopolymer-based hydrogels, due to their inherent benefits such as high activity (for instance, a wealth of cell recognition motifs), superior biocompatibility, and excellent degradability, have proven to be ideal for organoid culture. This is because they can mimic the microenvironmental features of the ECM. The molecular behaviors of natural biopolymers can be altered through physical or chemical crosslinking, chemical modification, or by combining them with other materials. This results in 3D matrices with mechanical and biochemical properties that encourage organoid growth, proliferation, or differentiation. The use of such natural biopolymeric hydrogels holds significant potential, especially for transplantation treatments. This is because they can reduce health risks associated with synthetic polymers, such as unwanted immunogenicity and in vivo toxicity [11].
However, significant material considerations need to be addressed for biopolymeric hydrogels to replace traditional Matrigel and be used as next-generation matrices in organoid technology: (1) Due to the lot-to-lot variability of biopolymers, consistently controlling the size and cellular composition of biopolymeric hydrogel-based organoids is a significant problem [129-134]. More comprehensive efforts to standardize the structural and functional features of biopolymeric hydrogels are required to make bioengineered matrices more effective and predictable. (2) To accurately replicate the dynamic process of organoid formation, the matrix material must be engineered to modify the microenvironment and surrounding stromal matrix in a spatiotemporally controlled manner. Natural biopolymers can change their physical and biochemical characteristics in response to stimuli commonly found in biological environments, such as light, temperature, and pH [10,135]. By controlling biomaterial parameters such as structural geometry, mechanical properties, and cell-binding ligands, this inherent stimulus-responsive behavior can be advantageously applied to the organoid system to better mimic the dynamic changes in extracellular microenvironmental inputs derived from the evolution of biological properties. (3) Integrative solutions combining biopolymeric hydrogel-based organoids with droplet microfluidics are highly desired as a future approach in organoid engineering. This approach would provide a programmable 3D scaffold for synthesizing organoids in a high-throughput manner. Recent studies suggest that microfluidic technologies, which allow for precise control of organoids and dynamic physical conditions, could be used to produce large quantities of highly uniform organoids in biopolymeric microdroplet hydrogels [136]. While this method holds great potential, especially for high-throughput drug candidate screening [137,138], it is still in the early stages of development. (4) Through 3D methods such as bioprinting, biopolymeric hydrogel-based organoids could be used as a building block to assemble larger, more complicated structures resembling genuine organs [139-142]. The design of biopolymeric hydrogels should allow for the embedding of organoids while incorporating their unique mechanical and biochemical properties to achieve the desired bulk properties. This would allow for the inclusion of key tissue compartments of native organs, such as the immune system and vascularization.
By addressing the aforementioned challenges through the integration of multiple disciplines, such as stem cell biology and materials engineering, natural biopolymer-based hydrogels could become an emerging framework to achieve cellular diversity, maturation, and full functionality of organoids with greater controllability and fidelity, facilitating an eventual shift away from the use of Matrigel and opening up a large design space to transform the field of numerous downstream translational applications.

NOTES

Conflict of interest

No potential conflict of interest relevant to this article was reported.

Funding

This research was supported by a Korean Fund for Regenerative Medicine (KFRM) grant funded by the Korean government (the Ministry of Science and ICT, the Ministry of Health & Welfare) (grant number: 23A0105L1) and a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI) funded by the Ministry of Health & Welfare, Korea (grant number: HI22C1754).

Data availability

Please contact the corresponding author for data availability.

Fig. 1.
Crosslinking strategies for generating natural biopolymer-based hydrogels.
organoid-2023-3-e13f1.jpg
Fig. 2.
Alginate-based hydrogels for intestinal organoid culture. (A) Chemical structure of alginate. (B) Schematic representation of Ca2+-based ionic crosslinking of alginate. Reproduced from Jo and Lee. Small 2020;16:e1903736, with permission from John Wiley and Sons [10]. (C) Colony formation efficiency of mouse small intestinal stem cells in various hydrogel backbones with or without Matrigel supplementation [48]. (D) Bright-field images of the cultures after 3 days of culture in different hydrogels and Matrigel. Scale bars: 200 μm. Reproduced and slightly modified from Broguiere et al. Adv Mater 2018;30:e1801621, with permission from John Wiley and Sons [48]. (E) Hematoxylin and eosin staining of intestinal organoids cultured in Ca-alginate hydrogel and Matrigel for 28 days. Dashed lines outline the epithelium. (F) Frequency of mature cell type differentiation in Ca-alginate hydrogel and Matrigel. Reproduced and slightly modified from Capeling et al. Stem Cell Rep 2019;12:381-94, with permission from Cell Press [47].
organoid-2023-3-e13f2.jpg
Fig. 3.
Cellulose-based hydrogels for culture of intestinal organoids. (A) Schematic description of TEMPO-mediated oxidation of cellulose. (B) Scheme of small intestinal organoids cultured in an oxidized cellulose nanofiber (CNF)-based hydrogel [58]. (C) Intestinal organoids cultured in oxidized CNF hydrogels. Cystic organoids are generated only upon the addition of glycine (GLY). The growth of organoids is sustained in the hydrogel of CNFs functionalized with RGD peptide. Scale bars: 100 μm. Reproduced from Curvello et al. Adv Sci (Weinh) 2020;8:2002135, with permission from John Wiley and Sons [58].
organoid-2023-3-e13f3.jpg
Fig. 4.
Fibrin-based composite hydrogel capsules for culture of liver organoids. (A) Schematic description of the oil-free droplet microfluidic system to fabricate composite hydrogel capsules with a fibrin hydrogel core and alginate-chitosan composite shell. (B) Immunohistochemical staining images of hepatocyte markers (ALB and CYP3A4) and cholangiocyte markers (CK7 and CK19) in liver organoids after 7 days of encapsulation in capsules. Scale bars: 50 μm. (C) Albumin secretion and (D) urea synthesis in liver organoids after encapsulation of 2, 4, 6 and 8 days. Reproduced from Wang et al. Biomater Sci 2020;8:5476-88, with permission from the Royal Society of Chemistry [91].
organoid-2023-3-e13f4.jpg
Fig. 5.
Molecular structures of elastin-like proteins (ELPs). (A) Schematic representation of amino acid sequence domain arrangement in tropoelastin. Yellow rectangles indicate hydrophobic domains. Reproduced from Acosta et al. Adv Funct Mater 2020;30:1909050, with permission from John Wiley and Sons [123]. (B) Scheme of an engineered extracellular matrix (eECM) polypeptide chain with an extended, cell-adhesive RGD domain. Reproduced from DiMarco et al. Biomater Sci 2015;3:1376-85 [92]. (C) Scheme of PEP-FN and PEP-LAMA3 proteins containing leucine zipper (green), ELPs (yellow), and the cell binding peptide (blue). Reproduced from Kozlowski et al. Front Bioeng Biotechnol 2023;11:1144209, according to the Creative Commons license [93].
organoid-2023-3-e13f5.jpg
Table 1.
Summary of the general advantages and limitations of natural biopolymers for organoid cultures
Category Biopolymer Advantages Limitations
Polysaccharide Alginate Low toxicity Absence of a cell adhesion motif
Ease of manipulation Limited long-term stability
Low cost
Ease of gelation
Hyaluronic acid High bioactivity Poor degradation rate
Biological relevance Unfavorable mechanical properties
Chemical tunability
Heparin Affinity to various growth factors Supply and safety problems regarding animal sources
Cellulose Remarkable mechanical properties Limited application in the form of nanocellulose fibers
Protein Collagen Structural and mechanical properties reminiscent of native tissues Low stiffness
Amenable to cell adhesion without modification Limited long-term stability
Batch-to-batch variability
Gelatin Low immunogenicity Poor mechanical properties
Amenable to cell adhesion without modification Short degradation time
Fibrin Abundant cell adhesion domain Poor mechanical properties
Recombinant protein Lot-to-lot repeatability Unpredictable properties from protein folding
Excellent design flexibility
Peptide Excellent design flexibility Poor mechanical properties
Highly predictable properties according to sequence
Table 2.
Polysaccharide-based hydrogels for organoid engineering
Biopolymer Fabrication method Organoid type Origin Cell source Features of hydrogel Ref.
Alginate Ionic crosslinking Intestine Mouse ASC Provides space for organoid growth [48]
Human PSC Provides space for organoid growth [47]
Pancreas Human ASC Supports dynamic culture of organoids in a microphysiological system [49]
Rat ASC [49]
Lung Human ASC Allows a bead template for the alveolar sac [50]
Norbornene-alginate Photocrosslinking Kidney Human iPSC Tunable mechanical properties without Ca2+ ions [51]
Oxidized alginate Covalent crosslinking Kidney Human iPSC Allows dynamic reshuffling of the crosslinks in cell culture conditions [52]
Hyaluronic acid Enzymatic crosslinking Bone marrow Human HSPC or BMSC Tunable physical and biological properties [53]
Polyelectrolytic complexation Cerebral Human iPSC Provides space for organoid growth [54]
Heparin Peptide linker Kidney Human ASC Modulates growth factor release [55]
Maintains the polarization of proximal tubule cells
Mammary Human ASC Precisely controllable biochemical properties [56]
Cellulose TEMPO-mediated oxidation Liver Human ASC Exhibits rapid self-healing and shear-thinning behavior [57]
Ionic crosslinking Intestine Mouse ASC Very low-cost but performant for organoid growth [58,59]
Tunable and compatible with ECM-components

Ref., reference, ASC, adult stem cell; PSC, pluripotent stem cell; iPSC, induced pluripotent stem cell; HSPC, hematopoietic stem and progenitor cell; BMSC, bone marrow stromal cell; ECM, extracellular matrix.

Table 3.
Protein-based hydrogels for organoid engineering
Biopolymer Fabrication method Organoid type Origin Cell source Features of hydrogel Ref.
Collagen Thermal crosslinking Intestine Mouse ASC Easily tunable physical properties [83,84,87,88]
Human ASC [85]
Colon Mouse ASC Easily tunable physical properties [86]
Human ASC [86]
Stomach Mouse ASC Easily tunable physical properties [86]
Mammary gland Human ASC Exhibits mechanical plasticity [89]
Gelatin Enzymatic crosslinking Liver Human ASC Tunable physical and biological properties [90]
Fibrin Enzymatic crosslinking Intestine Mouse ASC Provides physical support and RGD adhesion domains [48]
Human ASC [48]
Pancreas Human ASC Provides physical support and RGD adhesion domains [48]
Liver Human ASC Provides physical support and RGD adhesion domains [48]
iPSC Provides a controllable and stable environment for organoid generation [91]
Recombinant protein Chemical crosslinking Intestine Mouse ASC Tunable physical and biological properties [92]
Self-assembly Pancreas Mouse ASC Tunable physical and biological properties [93]
Thermal crosslinking Cerebral Human PSC Easy to functionalize with bioactive molecules [94,95]
Reduces inter-organoid variability
Peptide Chemical crosslinking Cerebral Rat PSC Tunable physical and biological properties [97]
Self-assembly Kidney Human iPSC Tunable physical and biological properties [96]

Ref., reference, ASC, adult stem cell; PSC, pluripotent stem cell; iPSC, induced pluripotent stem cell.

References

1. Hofer M, Lutolf MP. Engineering organoids. Nat Rev Mater 2021;6:402-20.
crossref pmid pmc pdf
2. Garreta E, Kamm RD, Chuva de Sousa Lopes SM, Lancaster MA, Weiss R, Trepat X, et al. Rethinking organoid technology through bioengineering. Nat Mater 2021;20:145-55.
crossref pmid pdf
3. Kim MB, Hwangbo S, Jang S, Jo YK. Bioengineered co-culture of organoids to recapitulate host-microbe interactions. Mater Today Bio 2022;16:100345.
crossref pmid pmc
4. Kaur S, Kaur I, Rawal P, Tripathi DM, Vasudevan A. Non-matrigel scaffolds for organoid cultures. Cancer Lett 2021;504:58-66.
crossref pmid
5. Kozlowski MT, Crook CJ, Ku HT. Towards organoid culture without Matrigel. Commun Biol 2021;4:1387.
crossref pmid pmc pdf
6. Hughes CS, Postovit LM, Lajoie GA. Matrigel: a complex protein mixture required for optimal growth of cell culture. Proteomics 2010;10:1886-90.
crossref pmid pdf
7. Rossi G, Manfrin A, Lutolf MP. Progress and potential in organoid research. Nat Rev Genet 2018;19:671-87.
crossref pmid pdf
8. Czerwinski M, Spence JR. Hacking the matrix. Cell Stem Cell 2017;20:9-10.
crossref pmid pmc
9. Schneeberger K, Spee B, Costa P, Sachs N, Clevers H, Malda J. Converging biofabrication and organoid technologies: the next frontier in hepatic and intestinal tissue engineering? Biofabrication 2017;9:013001.
crossref pmid pmc pdf
10. Jo YK, Lee D. Biopolymer microparticles prepared by microfluidics for biomedical applications. Small 2020;16:e1903736.
crossref pmid pdf
11. Stoppel WL, Ghezzi CE, McNamara SL, Black LD, Kaplan DL. Clinical applications of naturally derived biopolymer-based scaffolds for regenerative medicine. Ann Biomed Eng 2015;43:657-80.
crossref pmid pdf
12. Patel SN, Ishahak M, Chaimov D, Velraj A, LaShoto D, Hagan DW, et al. Organoid microphysiological system preserves pancreatic islet function within 3D matrix. Sci Adv 2021;7:eaba5515.
crossref pmid pmc
13. Bao Z, Xian C, Yuan Q, Liu G, Wu J. Natural polymer-based hydrogels with enhanced mechanical performances: preparation, structure, and property. Adv Healthc Mater 2019;8:e1900670.
crossref pmid pdf
14. Li Y, Maciel D, Rodrigues J, Shi X, Tomás H. Biodegradable polymer nanogels for drug/nucleic acid delivery. Chem Rev 2015;115:8564-608.
crossref pmid
15. Hoffman AS. Hydrogels for biomedical applications. Adv Drug Deliv Rev 2002;54:3-12.
crossref pmid
16. Sánchez-Cid P, Jiménez-Rosado M, Romero A, Pérez-Puyana V. Novel trends in hydrogel development for biomedical applications: a review. Polymers (Basel) 2022;14:3023.
crossref pmid pmc
17. Xiong R, Grant AM, Ma R, Zhang S, Tsukruk VV. Naturally-derived biopolymer nanocomposites: interfacial design, properties and emerging applications. Mater Sci Eng R 2018;125:1-41.
crossref
18. McClements DJ. Designing biopolymer microgels to encapsulate, protect and deliver bioactive components: physicochemical aspects. Adv Colloid Interface Sci 2017;240:31-59.
crossref pmid
19. Lin S, Cao C, Wang Q, Gonzalez M, Dolbow JE, Zhao X. Design of stiff, tough and stretchy hydrogel composites via nanoscale hybrid crosslinking and macroscale fiber reinforcement. Soft Matter 2014;10:7519-27.
crossref pmid
20. Li J, Illeperuma WR, Suo Z, Vlassak JJ. Hybrid hydrogels with extremely high stiffness and toughness. ACS Macro Lett 2014;3:520-3.
crossref pmid
21. Zhao X. Multi-scale multi-mechanism design of tough hydrogels: building dissipation into stretchy networks. Soft Matter 2014;10:672-87.
crossref pmid pmc
22. Reddy N, Reddy R, Jiang Q. Crosslinking biopolymers for biomedical applications. Trends Biotechnol 2015;33:362-9.
crossref pmid
23. Zhang X, Malhotra S, Molina M, Haag R. Micro- and nanogels with labile crosslinks: from synthesis to biomedical applications. Chem Soc Rev 2015;44:1948-73.
crossref pmid
24. Cao J, Cai Y, Yu L, Zhou J. Dual physically crosslinked hydrogels based on the synergistic effects of electrostatic and dipole-dipole interactions. J Mater Chem B 2019;7:676-83.
crossref pmid
25. Fredrick R, Podder A, Viswanathan A, Bhuniya S. Synthesis and characterization of polysaccharide hydrogel based on hydrophobic interactions. J Appl Polym Sci 2019;136:47665.
crossref pdf
26. Peng G, Wang J, Yang F, Zhang S, Hou J, Xing W, et al. In situ formation of biodegradable dextran-based hydrogel via Michael addition. J Appl Polym Sci 2013;127:577-84.
crossref
27. Dragan ES, Dinu MV. Polysaccharides constructed hydrogels as vehicles for proteins and peptides. A review. Carbohydr Polym 2019;225:115210.
crossref pmid
28. Teixeira LS, Feijen J, van Blitterswijk CA, Dijkstra PJ, Karperien M. Enzyme-catalyzed crosslinkable hydrogels: emerging strategies for tissue engineering. Biomaterials 2012;33:1281-90.
crossref pmid
29. Li X, Su X. Multifunctional smart hydrogels: potential in tissue engineering and cancer therapy. J Mater Chem B 2018;6:4714-30.
crossref pmid
30. Blondel D, Lutolf MP. Bioinspired hydrogels for 3D organoid culture. Chimia (Aarau) 2019;73:81-5.
crossref pmid pdf
31. Hu W, Wang Z, Xiao Y, Zhang S, Wang J. Advances in crosslinking strategies of biomedical hydrogels. Biomater Sci 2019;7:843-55.
crossref pmid
32. Yao H, Wang J, Mi S. Photo processing for biomedical hydrogels design and functionality: a review. Polymers (Basel) 2017;10:11.
crossref pmid pmc
33. Sperinde JJ, Griffith LG. Synthesis and characterization of enzymatically-cross-linked poly(ethylene glycol) hydrogels. Macromolecules 1997;30:5255-64.
crossref
34. Gjorevski N, Sachs N, Manfrin A, Giger S, Bragina ME, Ordóñez-Morán P, et al. Designer matrices for intestinal stem cell and organoid culture. Nature 2016;539:560-4.
crossref pmid pdf
35. Cantini M, Donnelly H, Dalby MJ, Salmeron-Sanchez M. The plot thickens: the emerging role of matrix viscosity in cell mechanotransduction. Adv Healthc Mater 2020;9:e1901259.
crossref pmid pdf
36. Lutolf MP, Hubbell JA. Synthesis and physicochemical characterization of end-linked poly(ethylene glycol)-co-peptide hydrogels formed by Michael-type addition. Biomacromolecules 2003;4:713-22.
crossref pmid
37. Lutolf MP, Hubbell JA. Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering. Nat Biotechnol 2005;23:47-55.
crossref pmid pdf
38. Lutolf MP, Lauer-Fields JL, Schmoekel HG, Metters AT, Weber FE, Fields GB, et al. Synthetic matrix metalloproteinase-sensitive hydrogels for the conduction of tissue regeneration: engineering cell-invasion characteristics. Proc Natl Acad Sci U S A 2003;100:5413-8.
crossref pmid pmc
39. Kloxin AM, Kasko AM, Salinas CN, Anseth KS. Photodegradable hydrogels for dynamic tuning of physical and chemical properties. Science 2009;324:59-63.
crossref pmid pmc
40. Gjorevski N, Lutolf MP. Synthesis and characterization of well-defined hydrogel matrices and their application to intestinal stem cell and organoid culture. Nat Protoc 2017;12:2263-74.
crossref pmid pdf
41. Lee KY, Jeong L, Kang YO, Lee SJ, Park WH. Electrospinning of polysaccharides for regenerative medicine. Adv Drug Deliv Rev 2009;61:1020-32.
crossref pmid
42. Lovegrove A, Edwards CH, De Noni I, Patel H, El SN, Grassby T, et al. Role of polysaccharides in food, digestion, and health. Crit Rev Food Sci Nutr 2017;57:237-53.
crossref pmid pdf
43. Jones OG, McClements DJ. Functional biopolymer particles: design, fabrication, and applications. Compr Rev Food Sci Food Saf 2010;9:374-97.
crossref pmid
44. Oh JK, Lee DI, Park JM. Biopolymer-based microgels/nanogels for drug delivery applications. Prog Polym Sci 2009;34:1261-82.
crossref
45. Okeyoshi K, Joshi G, Okajima MK, Kaneko T. Formation of polysaccharide membranes by splitting of evaporative air-LC interface. Adv Mater Interfaces 2018;5:1701219.
crossref pdf
46. Jankolovits J, Gazit OM, Nigra MM, Bohling J, Roper JA, Katz A. Single-pot encapsulation of oxide particles within a polysaccharide multilayer nanocoating. Adv Mater Interfaces 2015;2:1400465.
crossref
47. Capeling MM, Czerwinski M, Huang S, Tsai YH, Wu A, Nagy MS, et al. Nonadhesive alginate hydrogels support growth of pluripotent stem cell-derived intestinal organoids. Stem Cell Rep 2019;12:381-94.
crossref
48. Broguiere N, Isenmann L, Hirt C, Ringel T, Placzek S, Cavalli E, et al. Growth of epithelial organoids in a defined hydrogel. Adv Mater 2018;30:e1801621.
crossref pmid pdf
49. Patel SN, Ishahak M, Chaimov D, Velraj A, LaShoto D, Hagan DW, et al. Organoid microphysiological system preserves pancreatic islet function within 3D matrix. Sci Adv 2021;7:eaba5515.
crossref pmid pmc
50. Wilkinson DC, Alva-Ornelas JA, Sucre JM, Vijayaraj P, Durra A, Richardson W, et al. Development of a three-dimensional bioengineering technology to generate lung tissue for personalized disease modeling. Stem Cells Transl Med 2017;6:622-33.
crossref pmid pdf
51. Geuens T, Ruiter FA, Schumacher A, Morgan FL, Rademakers T, Wiersma LE, et al. Thiol-ene cross-linked alginate hydrogel encapsulation modulates the extracellular matrix of kidney organoids by reducing abnormal type 1a1 collagen deposition. Biomaterials 2021;275:120976.
crossref pmid
52. Ruiter FA, Morgan FL, Roumans N, Schumacher A, Slaats GG, Moroni L, et al. Soft, dynamic hydrogel confinement improves kidney organoid lumen morphology and reduces epithelial-mesenchymal transition in culture. Adv Sci (Weinh) 2022;9:e2200543.
crossref pmid
53. Vallmajo-Martin Q, Broguiere N, Millan C, Zenobi-Wong M, Ehrbar M. PEG/HA hybrid hydrogels for biologically and mechanically tailorable bone marrow organoids. Adv Funct Mater 2020;30:1910282.
crossref pdf
54. Lindborg BA, Brekke JH, Vegoe AL, Ulrich CB, Haider KT, Subramaniam S, et al. Rapid induction of cerebral organoids from human induced pluripotent stem cells using a chemically defined hydrogel and defined cell culture medium. Stem Cells Transl Med 2016;5:970-9.
crossref pmid pmc pdf
55. Weber HM, Tsurkan MV, Magno V, Freudenberg U, Werner C. Heparin-based hydrogels induce human renal tubulogenesis in vitro. Acta Biomater 2017;57:59-69.
crossref pmid
56. Nowak M, Freudenberg U, Tsurkan MV, Werner C, Levental KR. Modular GAG-matrices to promote mammary epithelial morphogenesis in vitro. Biomaterials 2017;112:20-30.
crossref pmid
57. Krüger M, Oosterhoff LA, van Wolferen ME, Schiele SA, Walther A, Geijsen N, et al. Cellulose nanofibril hydrogel promotes hepatic differentiation of human liver organoids. Adv Healthc Mater 2020;9:e1901658.
pmid
58. Curvello R, Kerr G, Micati DJ, Chan WH, Raghuwanshi VS, Rosenbluh J, et al. Engineered plant-based nanocellulose hydrogel for small intestinal organoid growth. Adv Sci (Weinh) 2020;8:2002135.
crossref pmid pmc pdf
59. Curvello R, Garnier G. Cationic cross-linked nanocellulose-based matrices for the growth and recovery of intestinal organoids. Biomacromolecules 2021;22:701-9.
crossref pmid
60. Sun J, Tan H. Alginate-based biomaterials for regenerative medicine applications. Materials (Basel) 2013;6:1285-309.
crossref pmid pmc
61. Agarwal T, Kabiraj P, Narayana GH, Kulanthaivel S, Kasiviswanathan U, Pal K, et al. Alginate bead based hexagonal close packed 3D implant for bone tissue engineering. ACS Appl Mater Interfaces 2016;8:32132-45.
crossref pmid
62. Lee KY, Mooney DJ. Alginate: properties and biomedical applications. Prog Polym Sci 2012;37:106-26.
crossref pmid pmc
63. Samorezov JE, Morlock CM, Alsberg E. Dual ionic and photo-crosslinked alginate hydrogels for micropatterned spatial control of material properties and cell behavior. Bioconjug Chem 2015;26:1339-47.
crossref pmid pmc
64. Webber RE, Shull KR. Strain dependence of the viscoelastic properties of alginate hydrogels. Macromolecules 2004;37:6153-60.
crossref
65. Kulanthaivel S, Rathnam V S S, Agarwal T, Pradhan S, Pal K, Giri S, et al. Gum tragacanth-alginate beads as proangiogenic-osteogenic cell encapsulation systems for bone tissue engineering. J Mater Chem B 2017;5:4177-89.
crossref pmid
66. Papas KK, De Leon H, Suszynski TM, Johnson RC. Oxygenation strategies for encapsulated islet and beta cell transplants. Adv Drug Deliv Rev 2019;139:139-56.
crossref pmid
67. Capeling M, Huang S, Mulero-Russe A, Cieza R, Tsai YH, Garcia A, et al. Generation of small intestinal organoids for experimental intestinal physiology. Methods Cell Biol 2020;159:143-74.
crossref pmid
68. Oh EJ, Park K, Kim KS, Kim J, Yang JA, Kong JH, et al. Target specific and long-acting delivery of protein, peptide, and nucleotide therapeutics using hyaluronic acid derivatives. J Control Release 2010;141:2-12.
crossref pmid
69. Wang M, Deng Z, Guo Y, Xu P. Designing functional hyaluronic acid-based hydrogels for cartilage tissue engineering. Mater Today Bio 2022;17:100495.
crossref pmid pmc
70. Anderson E, Pierre-Louis WS, Wong CJ, Lary JW, Cole JL. Heparin activates PKR by inducing dimerization. J Mol Biol 2011;413:973-84.
crossref pmid pmc
71. Belair DG, Le NN, Murphy WL. Design of growth factor sequestering biomaterials. Chem Commun (Camb) 2014;50:15651-68.
crossref pmid pmc
72. Liang Y, Kiick KL. Heparin-functionalized polymeric biomaterials in tissue engineering and drug delivery applications. Acta Biomater 2014;10:1588-600.
crossref pmid
73. Capila I, Linhardt RJ. Heparin-protein interactions. Angew Chem Int Ed Engl 2002;41:391-412.
crossref pmid
74. Freudenberg U, Liang Y, Kiick KL, Werner C. Glycosaminoglycan-based biohybrid hydrogels: a sweet and smart choice for multifunctional biomaterials. Adv Mater 2016;28:8861-91.
crossref pmid pmc pdf
75. Salas C, Nypelö T, Rodriguez-Abreu C, Carrillo C, Rojas OJ. Nanocellulose properties and applications in colloids and interfaces. Curr Opin Colloid Interface Sci 2014;19:383-96.
crossref
76. Curvello R, Raghuwanshi VS, Garnier G. Engineering nanocellulose hydrogels for biomedical applications. Adv Colloid Interface Sci 2019;267:47-61.
crossref pmid
77. Saito T, Kimura S, Nishiyama Y, Isogai A. Cellulose nanofibers prepared by TEMPO-mediated oxidation of native cellulose. Biomacromolecules 2007;8:2485-91.
crossref pmid
78. Mendoza DJ, Browne C, Raghuwanshi VS, Simon GP, Garnier G. One-shot TEMPO-periodate oxidation of native cellulose. Carbohydr Polym 2019;226:115292.
crossref pmid
79. Ninan N, Muthiah M, Park IK, Wong TW, Thomas S, Grohens Y. Natural polymer/inorganic material based hybrid scaffolds for skin wound healing. Polym Rev 2015;55:453-90.
crossref
80. Gupta P, Nayak KK. Characteristics of protein-based biopolymer and its application. Polym Eng Sci 2015;55:485-98.
crossref
81. Huerta-López C, Alegre-Cebollada J. Protein hydrogels: the Swiss army knife for enhanced mechanical and bioactive properties of biomaterials. Nanomaterials (Basel) 2021;11:1656.
crossref pmid pmc
82. Xu X, Xu Z, Yang X, He Y, Lin R. Construction and characterization of a pure protein hydrogel for drug delivery application. Int J Biol Macromol 2017;95:294-8.
crossref pmid
83. Jee JH, Lee DH, Ko J, Hahn S, Jeong SY, Kim HK, et al. Development of collagen-based 3D matrix for gastrointestinal tract-derived organoid culture. Stem Cells Int 2019;2019:8472712.
crossref pmid pmc pdf
84. Sachs N, Tsukamoto Y, Kujala P, Peters PJ, Clevers H. Intestinal epithelial organoids fuse to form self-organizing tubes in floating collagen gels. Development 2017;144:1107-12.
crossref pmid pdf
85. Jabaji Z, Brinkley GJ, Khalil HA, Sears CM, Lei NY, Lewis M, et al. Type I collagen as an extracellular matrix for the in vitro growth of human small intestinal epithelium. PLoS One 2014;9:e107814.
crossref pmid pmc
86. Jee JH, Lee DH, Ko J, Hahn S, Jeong SY, Kim HK, et al. Development of collagen-based 3D matrix for gastrointestinal tract-derived organoid culture. Stem Cells Int 2019;2019:8472712.
crossref pmid pmc pdf
87. Jabaji Z, Sears CM, Brinkley GJ, Lei NY, Joshi VS, Wang J, et al. Use of collagen gel as an alternative extracellular matrix for the in vitro and in vivo growth of murine small intestinal epithelium. Tissue Eng Part C Methods 2013;19:961-9.
crossref pmid pmc
88. Curvello R, Alves D, Abud HE, Garnier G. A thermo-responsive collagen-nanocellulose hydrogel for the growth of intestinal organoids. Mater Sci Eng C Mater Biol Appl 2021;124:112051.
crossref pmid
89. Buchmann B, Engelbrecht LK, Fernandez P, Hutterer FP, Raich MK, Scheel CH, et al. Mechanical plasticity of collagen directs branch elongation in human mammary gland organoids. Nat Commun 2021;12:2759.
crossref pmid pmc pdf
90. Klotz BJ, Oosterhoff LA, Utomo L, Lim KS, Vallmajo-Martin Q, Clevers H, et al. A versatile biosynthetic hydrogel platform for engineering of tissue analogues. Adv Healthc Mater 2019;8:e1900979.
crossref pmid pmc pdf
91. Wang Y, Liu H, Zhang M, Wang H, Chen W, Qin J. One-step synthesis of composite hydrogel capsules to support liver organoid generation from hiPSCs. Biomater Sci 2020;8:5476-88.
crossref pmid
92. DiMarco RL, Dewi RE, Bernal G, Kuo C, Heilshorn SC. Protein-engineered scaffolds for in vitro 3D culture of primary adult intestinal organoids. Biomater Sci 2015;3:1376-85.
crossref pmid pmc
93. Kozlowski MT, Zook HN, Chigumba DN, Johnstone CP, Caldera LF, Shih HP, et al. A matrigel-free method for culture of pancreatic endocrine-like cells in defined protein-based hydrogels. Front Bioeng Biotechnol 2023;11:1144209.
crossref pmid pmc
94. Fiorenzano A, Sozzi E, Birtele M, Kajtez J, Giacomoni J, Nilsson F, et al. Single-cell transcriptomics captures features of human midbrain development and dopamine neuron diversity in brain organoids. Nat Commun 2021;12:7302.
crossref pmid pmc pdf
95. Sozzi E, Kajtez J, Bruzelius A, Wesseler MF, Nilsson F, Birtele M, et al. Silk scaffolding drives self-assembly of functional and mature human brain organoids. Front Cell Dev Biol 2022;10:1023279.
crossref pmid pmc
96. Treacy NJ, Clerkin S, Davis JL, Kennedy C, Miller AF, Saiani A, et al. Growth and differentiation of human induced pluripotent stem cell (hiPSC)-derived kidney organoids using fully synthetic peptide hydrogels. Bioact Mater 2023;21:142-56.
crossref pmid
97. Balion Z, Cėpla V, Svirskiene N, Svirskis G, Druceikaitė K, Inokaitis H, et al. Cerebellar cells self-assemble into functional organoids on synthetic, chemically crosslinked ECM-mimicking peptide hydrogels. Biomolecules 2020;10:754.
crossref pmid pmc
98. Caliari SR, Burdick JA. A practical guide to hydrogels for cell culture. Nat Methods 2016;13:405-14.
crossref pmid pmc pdf
99. Echave MC, Saenz del Burgo L, Pedraz JL, Orive G. Gelatin as biomaterial for tissue engineering. Curr Pharm Des 2017;23:3567-84.
crossref pmid
100. Zhang Y, Olsen K, Grossi A, Otte J. Effect of pretreatment on enzymatic hydrolysis of bovine collagen and formation of ACE-inhibitory peptides. Food Chem 2013;141:2343-54.
crossref pmid
101. Agarwal T, Celikkin N, Costantini M, Maiti TK, Makvandi P. Recent advances in chemically defined and tunable hydrogel platforms for organoid culture. Bio-Des Manuf 2021;4:641-74.
crossref pdf
102. Lee CH, Singla A, Lee Y. Biomedical applications of collagen. Int J Pharm 2001;221:1-22.
crossref pmid
103. Lee KY, Mooney DJ. Hydrogels for tissue engineering. Chem Rev 2001;101:1869-79.
crossref pmid
104. Yuan L, Li B, Yang J, Ni Y, Teng Y, Guo L, et al. Effects of composition and mechanical property of injectable collagen I/II composite hydrogels on chondrocyte behaviors. Tissue Eng Part A 2016;22:899-906.
crossref pmid
105. Truong VX, Hun ML, Li F, Chidgey AP, Forsythe JS. In situ-forming click-crosslinked gelatin based hydrogels for 3D culture of thymic epithelial cells. Biomater Sci 2016;4:1123-31.
crossref pmid
106. Parker NG, Povey MJ. Ultrasonic study of the gelation of gelatin: phase diagram, hysteresis and kinetics. Food Hydrocolloids 2012;26:99-107.
crossref
107. Guizzardi R, Vaghi L, Marelli M, Natalello A, Andreosso I, Papagni A, et al. Gelatin-based hydrogels through homobifunctional triazolinediones targeting tyrosine residues. Molecules 2019;24:589.
crossref pmid pmc
108. Catoira MC, Fusaro L, Di Francesco D, Ramella M, Boccafoschi F. Overview of natural hydrogels for regenerative medicine applications. J Mater Sci Mater Med 2019;30:115.
crossref pmid pmc pdf
109. Seifu DG, Purnama A, Mequanint K, Mantovani D. Small-diameter vascular tissue engineering. Nat Rev Cardiol 2013;10:410-21.
crossref pmid pdf
110. Li H, Kong N, Laver B, Liu J. Hydrogels constructed from engineered proteins. Small 2016;12:973-87.
crossref pmid
111. Petka WA, Harden JL, McGrath KP, Wirtz D, Tirrell DA. Reversible hydrogels from self-assembling artificial proteins. Science 1998;281:389-92.
crossref pmid
112. Wang C, Stewart RJ, Kopecek J. Hybrid hydrogels assembled from synthetic polymers and coiled-coil protein domains. Nature 1999;397:417-20.
crossref pmid pdf
113. Li Y, Xue B, Cao Y. 100th anniversary of macromolecular science viewpoint: synthetic protein hydrogels. ACS Macro Lett 2020;9:512-24.
crossref pmid
114. Gomes S, Leonor IB, Mano JF, Reis RL, Kaplan DL. Natural and genetically engineered proteins for tissue engineering. Prog Polym Sci 2012;37:1-17.
crossref pmid pmc
115. Jeong Y, Jo YK, Kim MS, Joo KI, Cha HJ. Tunicate-inspired photoactivatable proteinic nanobombs for tumor-adhesive multimodal therapy. Adv Healthc Mater 2021;10:e2101212.
crossref pmid pdf
116. Choi HS, Jo YK, Ahn GN, Kim DP, Joo KI, Cha HJ. Magnetically guidable proteinaceous adhesive microbots for targeted locoregional therapeutics delivery in the highly dynamic environment of the esophagus. Adv Funct Mater 2021;31:2104602.
crossref pdf
117. Chow D, Nunalee ML, Lim DW, Simnick AJ, Chilkoti A. Peptide-based biopolymers in biomedicine and biotechnology. Mater Sci Eng R Rep 2008;62:125-55.
crossref pmid pmc
118. Li H, Kong N, Laver B, Liu J. Hydrogels constructed from engineered proteins. Small 2016;12:973-87.
crossref pmid
119. Jo YK, Choi BH, Zhou C, Jun SH, Cha HJ. Cell recognitive bioadhesive-based osteogenic barrier coating with localized delivery of bone morphogenetic protein-2 for accelerated guided bone regeneration. Bioeng Transl Med 2023;8:e10493.
pmid pmc
120. Choi BH, Jo YK, Zhou C, Jang HS, Ahn JS, Jun SH, et al. Sticky bone-specific artificial extracellular matrix for stem cell-mediated rapid craniofacial bone therapy. Appl Mater Today 2020;18:100531.
crossref
121. Urry DW. Characterization of soluble peptides of elastin by physical techniques. Methods Enzymol 1982;82 Pt A:673-716.
pmid
122. Tamura T, Yamaoka T, Kunugi S, Panitch A, Tirrell DA. Effects of temperature and pressure on the aggregation properties of an engineered elastin model polypeptide in aqueous solution. Biomacromolecules 2000;1:552-5.
crossref pmid
123. Acosta S, Quintanilla-Sierra L, Mbundi L, Reboto V, Rodríguez-Cabello JC. Elastin-like recombinamers: deconstructing and recapitulating the functionality of extracellular matrix proteins using recombinant protein polymers. Adv Funct Mater 2020;30:1909050.
crossref pdf
124. Rising A, Widhe M, Johansson J, Hedhammar M. Spider silk proteins: recent advances in recombinant production, structure-function relationships and biomedical applications. Cell Mol Life Sci 2011;68:169-84.
crossref pmid pdf
125. Johansson U, Widhe M, Shalaly ND, Arregui IL, Nilebäck L, Tasiopoulos CP, et al. Assembly of functionalized silk together with cells to obtain proliferative 3D cultures integrated in a network of ECM-like microfibers. Sci Rep 2019;9:6291.
crossref pmid pmc pdf
126. Slyke DV. Physiology of the amino acids. Nature 1942;149:342-5.
crossref pdf
127. Bakhtiary N, Ghalandari B, Ghorbani F, Varma SN, Liu C. Advances in peptide-based hydrogel for tissue engineering. Polymers (Basel) 2023;15:1068.
crossref pmid pmc
128. Wychowaniec JK, Smith AM, Ligorio C, Mykhaylyk OO, Miller AF, Saiani A. Role of sheet-edge interactions in β-sheet self-assembling peptide hydrogels. Biomacromolecules 2020;21:2285-97.
crossref pmid pmc pdf
129. Vishakha SK, Kishor DB, Sudha SR. Natural polymers: a comprehensive review. Int J Res Pharm Biomed Sci 2012;3:1597-613.
130. Ye S, Boeter JW, Mihajlovic M, van Steenbeek FG, van Wolferen ME, Oosterhoff LA, et al. A chemically defined hydrogel for human liver organoid culture. Adv Funct Mater 2020;30:2000893.
crossref pmid pmc pdf
131. Eiraku M, Takata N, Ishibashi H, Kawada M, Sakakura E, Okuda S, et al. Self-organizing optic-cup morphogenesis in three-dimensional culture. Nature 2011;472:51-6.
crossref pmid pdf
132. Xu R, Zhou X, Wang S, Trinkle C. Tumor organoid models in precision medicine and investigating cancer-stromal interactions. Pharmacol Ther 2021;218:107668.
crossref pmid
133. Zhao H, Chen Y, Shao L, Xie M, Nie J, Qiu J, et al. Airflow-assisted 3D bioprinting of human heterogeneous microspheroidal organoids with microfluidic nozzle. Small 2018;14:e1802630.
crossref pmid pdf
134. Li Y, Kumacheva E. Hydrogel microenvironments for cancer spheroid growth and drug screening. Sci Adv 2018;4:eaas8998.
crossref pmid pmc
135. Wei M, Gao Y, Li X, Serpe MJ. Stimuli-responsive polymers and their applications. Polym Chem 2017;8:127-43.
crossref
136. Brandenberg N, Hoehnel S, Kuttler F, Homicsko K, Ceroni C, Ringel T, et al. High-throughput automated organoid culture via stem-cell aggregation in microcavity arrays. Nat Biomed Eng 2020;4:863-74.
crossref pmid pdf
137. Jin Y, Kim J, Lee JS, Min S, Kim S, Ahn DH, et al. Vascularized liver organoids generated using induced hepatic tissue and dynamic liver-specific microenvironment as a drug testing platform. Adv Funct Mater 2018;28:1801954.
crossref pdf
138. Schuster B, Junkin M, Kashaf SS, Romero-Calvo I, Kirby K, Matthews J, et al. Automated microfluidic platform for dynamic and combinatorial drug screening of tumor organoids. Nat Commun 2020;11:5271.
crossref pmid pmc pdf
139. Moroni L, Burdick JA, Highley C, Lee SJ, Morimoto Y, Takeuchi S, et al. Biofabrication strategies for 3D in vitro models and regenerative medicine. Nat Rev Mater 2018;3:21-37.
crossref pmid pmc pdf
140. Workman MJ, Mahe MM, Trisno S, Poling HM, Watson CL, Sundaram N, et al. Engineered human pluripotent-stem-cell-derived intestinal tissues with a functional enteric nervous system. Nat Med 2017;23:49-59.
crossref pmid pdf
141. Xiang Y, Tanaka Y, Cakir B, Patterson B, Kim KY, Sun P, et al. hESC-derived thalamic organoids form reciprocal projections when fused with cortical organoids. Cell Stem Cell 2019;24:487-97.
crossref pmid pmc
142. Du Y, Lo E, Ali S, Khademhosseini A. Directed assembly of cell-laden microgels for fabrication of 3D tissue constructs. Proc Natl Acad Sci U S A 2008;105:9522-7.
crossref pmid pmc
TOOLS
METRICS Graph View
  • 0 Crossref
  •  0 Scopus
  • 1,321 View
  • 33 Download
ORCID iDs

Yun Kee Jo
https://orcid.org/0000-0003-4240-7821

Related articles


ABOUT
BROWSE ARTICLES
EDITORIAL POLICY
FOR CONTRIBUTORS
Editorial Office
Room 319, Hall 1 of Chonbuk National University Dental College, 20, Geonji-ro, Deokjin-gu, Jeonju 54907, Korea
Tel: +82-63-270-4024    E-mail: editor@j-organoid.org                

Copyright © 2024 by The Organoid Society.

Developed in M2PI

Close layer
prev next