Matrix Hydrogel Research Articles

Synergistic influence of tetraethyl orthosilicate crosslinker on mixed matrix hydrogels

Hydrogel is a 3D framework of hydrophilic polymeric material that quickly absorbs and retains a huge amount of water (or other fluid) and offers versatile functionality. A series of unique carboxymethyl cellulose/Xanthan gum/polyvinyl alcohol (CXP) blended hydrogels, containing both the natural and synthetic polymers, were prepared by following the blending and casting approach. The polymers were incorporated through chemical crosslinking by tetraethyl orthosilicate (TEOS). The fabricated hydrogels showed all required features: non-toxicity, biocompatibility, and improved mechanical strength. The addition and variation in TEOS (crosslinker) significantly impacted the key characteristics of CXP hydrogel. The scanning electron microscopy (SEM) images showed the porous structure and indicated that the pore's size and intensity were reduced with the surge in TEOS content. Fourier transform infrared spectroscopy (FTIR) results confirmed the successful incorporation of various polymeric strands through crosslinking by TEOS. The thermogravimetric analysis (TGA) highlighted the greater stability of all the hydrogels over high temperatures. The crosslinked hydrogel displayed higher thermal resilience than the uncross-linked one. The differential thermal analysis (DTA) also confirmed that the addition of TEOS content drastically enhanced the thermal endurance of crosslinked hydrogels in comparison with the neat hydrogel. All the specimens exhibited good swelling ability in distilled water during the swelling studies. This study also reflected that the addition of crosslinker in a limited amount (50 µL) has significantly enhanced the swelling but further increase in concentration hindered the water uptake. The swelling response of blends towards pH revealed low swelling of films in acidic and basic pH, but maximal swelling in neutral media. This unique pH response of hydrogels at neutral pH along with the biocompatibility made them suitable for injectable managed drug carrier.

Myocardial matrix hydrogel acts as a reactive oxygen species scavenger and supports a proliferative microenvironment for cardiomyocytes

As the native regenerative potential of adult cardiac tissue is limited post-injury, stimulating endogenous repair mechanisms in the mammalian myocardium is a potential goal of regenerative medicine therapeutics. Injection of myocardial matrix hydrogels into the heart post-myocardial infarction (MI) has demonstrated increased cardiac muscle and promotion of pathways associated with cardiac development, suggesting potential promotion of cardiomyocyte turnover. In this study, the myocardial matrix hydrogel was shown to have native capability as an effective reactive oxygen species scavenger and protect against oxidative stress induced cell cycle inhibition in vitro. Encapsulation of cardiomyocytes demonstrated an enhanced turnover in in vitro studies, and in vivo assessments of myocardial matrix hydrogel treatment post-MI showed increased thymidine analog uptake in cardiomyocyte nuclei compared to saline controls. Overall, this study provides evidence that properties of the myocardial matrix material provide a microenvironment mitigating oxidative damage and supportive of cardiomyocytes undergoing DNA synthesis, toward possible DNA repair or cell cycle activation. Statement of significanceLoss of adult mammalian cardiomyocyte turnover is influenced by shifts in oxidative damage, which represents a potential mechanism for improving restoration of cardiac muscle after myocardial infarction (MI). Injection of a myocardial matrix hydrogel into the heart post-MI previously demonstrated increased cardiac muscle and promotion of pathways associated with cardiac development, suggesting potential in promoting proliferation of cardiomyocytes. In this study, the myocardial matrix hydrogel was shown to protect cells from oxidative stress and increase proliferation in vitro. In a rat MI model, greater presence of tissue free thiol content spared from oxidative damage, lesser mitochondrial superoxide content, and increased thymidine analog uptake in cardiomyocytes was found in matrix injected animals compared to saline controls. Overall, this study provides evidence that properties of the myocardial matrix material provide a microenvironment supportive of cardiomyocytes undergoing DNA synthesis, toward possible DNA repair or cell cycle activation.

Aerobic exercise-induced circulating extracellular vesicle combined decellularized dermal matrix hydrogel facilitates diabetic wound healing by promoting angiogenesis

Background: Insufficient blood supply results in unsatisfactory wound healing, especially for challenging wound repair such as diabetic wound defects. Regular exercise training brings a lot of benefits to cardiovascular fitness and metabolic health including attenuation of T2DM progression. Circulating extracellular vesicles (EVs) are postulated to carry a variety of signals involved in tissue crosstalk by their modified cargoes, representing novel mechanisms for the effects of exercise. Prominently, both acute and chronic aerobic exercise training can promote the release of exercise-induced cytokines and enhance the angiogenic function of circulating angiogenic cell–derived EVs. Methods: We investigated the possible angiogenesis potential of aerobic exercise-induced circulating EVs (EXE-EVs) on diabetic wound healing. Circulating EVs were isolated from the plasma of rats subjected to 4 weeks of moderate aerobic exercise or sedentariness 24 h after the last training session. The therapeutic effect of circulating EVs was evaluated in vitro by proliferation, migration, and tube formation assays of human umbilical vein endothelial cells (HUVECs), as well as in vivo by quantification of angiogenesis and cutaneous wound healing in diabetic rats. Results: The number of circulating EVs did not change significantly in exercised rats 24 h post-exercise in comparison with the sedentary rats. Nevertheless, EXE-EVs showed remarkable pro-angiogenic effect by augmenting proliferation, migration, and tube formation of HUVECs. Furthermore, the findings of animal experiments revealed that the EXE-EVs delivered by decellularized dermal matrix hydrogel (DDMH) could significantly promote the repair of skin defects through stimulating the regeneration of vascularized skin. Discussion: The present study is the first attempt to demonstrate that aerobic exercise-induced circulating EVs could be utilized as a cell-free therapy to activate angiogenesis and promote diabetic wound healing. Our findings suggest that EXE-EVs may stand for a potential strategy for diabetic soft tissue wound repair.

Differentiation of neonate mouse spermatogonia on two-dimensional and three-dimensional culture systems supplemented with d-Serine and Dizocilpine (MK-801)

N-methyl-d-aspartate (NMDA) modulates the spermatogenesis process through stimulating the steroid hormone biosynthesis. The aim of this study was to evaluate the effects of NMDA receptors agonists (d-Serine) and antagonists (MK801) on spermatogonia differentiation on decellularization testicular matrix (DTM) hydrogel scaffold. Four treatment groups were planned: 2D + D-Serine, 3D + D-Serine, 2D + MK801, and 3D + MK801. Results showed that cell viability was significantly decreased after 48 h in the 3D + D-Serine group and after 24 and 48 h in the 3D + MK801 group compared to the controls. The spermatogonia proliferation after two, four, and eight weeks was significantly increased in the 3D + D-Serine culture, while it was significantly reduced in the 2D + MK801 and 3D + MK801 groups after four and eight weeks. Real-time PCR results demonstrated that pre-meiotic gene (Plzf) expression was significantly increased only in the 3D + D-Serine culture compared to the control groups after four weeks of culture. The meiotic gene (Sycp3) expression was significantly increased in the 2D + D-Serine and 3D + D-Serine compared to the 2D controls after four and eight weeks. The post-meiotic gene (Tnp1) level in the 3D + D-Serine was significantly higher than the other groups. Flow-cytometry results indicated that the protein expression of Plzf (after four and eight weeks), Sycp3 (after eight weeks), and Tnp1 (after eight weeks) in the d-Serine-treated groups was significantly increased compared with the 2D control groups. There were not any significant changes in the gene expression of spermatogenic-related markers in MK801 culture media. However, a significant decrease in the protein levels of Plzf after eight weeks and Sycp3 after four and eight weeks was observed. In conclusion, the addition of NMDARs agonists (d-Serine) could be used to regulate the differentiation of spermatogonia in the 3D culture system.

Injectable extracellular matrix hydrogels contribute to native cell infiltration in a rat partial nephrectomy model.

Decellularized extracellular matrix (dECM) hydrogels have cytocompatibility, and are currently being investigated for application in soft tissues as a material that promotes native cell infiltration and tissue reconstruction. A dECM hydrogel has broad potential for application in organs with complex structures or various tissue injury models. In this study, we investigated the practical application of a dECM hydrogel by injecting a kidney-derived dECM hydrogel into a rat partial nephrectomy model. The prepared dECM hydrogel was adjustable in viscosity to allow holding at the excision site, and after gelation, had an elastic modulus similar to that of kidney tissue. In addition, the migration of renal epithelial cells and vascular endothelial cells embedded in dECM hydrogels was observed in vitro. Four weeks after injection of the dECM hydrogel to the partial excision site of the kidneys, infiltration of renal tubular constituent cells and native cells with high proliferative activity, as well as angiogenesis, were observed inside the injected areas. This study is the first to show that dECM hydrogels can be applied to the kidney, one of the most complex structural organs and that they can function as a scaffold to induce angiogenesis and infiltration of organ-specific renal tubular constituent cells, providing fundamental insights for further application of dECM hydrogels.

3D Bioprinting of Multi-Material Decellularized Liver Matrix Hydrogel at Physiological Temperatures.

Bioprinting is an acclaimed technique that allows the scaling of 3D architectures in an organized pattern but suffers from a scarcity of appropriate bioinks. Decellularized extracellular matrix (dECM) from xenogeneic species has garnered support as a biomaterial to promote tissue-specific regeneration and repair. The prospect of developing dECM-based 3D artificial tissue is impeded by its inherent low mechanical properties. In recent years, 3D bioprinting of dECM-based bioinks modified with additional scaffolds has advanced the development of load-bearing constructs. However, previous attempts using dECM were limited to low-temperature bioprinting, which is not favorable for a longer print duration with cells. Here, we report the development of a multi-material decellularized liver matrix (dLM) bioink reinforced with gelatin and polyethylene glycol to improve rheology, extrudability, and mechanical stability. This shear-thinning bioink facilitated extrusion-based bioprinting at 37 °C with HepG2 cells into a 3D grid structure with a further enhancement for long-term applications by enzymatic crosslinking with mushroom tyrosinase. The heavily crosslinked structure showed a 16-fold increase in viscosity (2.73 Pa s−1) and a 32-fold increase in storage modulus from the non-crosslinked dLM while retaining high cell viability (85–93%) and liver-specific functions. Our results show that the cytocompatible crosslinking of dLM bioink at physiological temperatures has promising applications for extended 3D-printing procedures.

A photocrosslinking antibacterial decellularized matrix hydrogel with nanofiber for cutaneous wound healing

ddECMMA is the methacrylating product of decellularized dermal extracellular matrix with biological signals and capable of photocrosslinking. Thiolated chitosan (TCS) is an effective antibacterial component. PCLPBA is a kind of plasma-treated polycaprolactone nanofiber dispersions (PCLP) that regulates macrophage polarization and promotes angiogenesis. In this study, we obtained ddECMMA via methacrylation reaction. TCS was prepared by reaction between chitosan and thioglycolic acid. PCLPBA was fabricated via reaction between PCLP and 3-buten-1-amine. TCS and PCLPBA were mixed in ddECMMA solution and photocrosslinked to form DTP4 hydrogel. The hydrogel showed rapid gelation, good mechanical strength, antibacterial and antioxidant properties. When it was cocultured with NIH 3T3 cells, the cells showed good morphology and proliferation rate. After applying it to the full-thickness cutaneous wound, wounds almost healed in 2 weeks via re-epithelialization and neovascularization with negligible scar tissue. The results indicate that DTP4 hydrogel is a promising candidate for clinic skin wound healing.

Characterization of decellularized left and right ventricular myocardial matrix hydrogels and their effects on cardiac progenitor cells

Congenital heart defects are the leading cause of right heart failure in pediatric patients. Implantation of c-kit+ cardiac-derived progenitor cells (CPCs) is being clinically evaluated to treat the failing right ventricle (RV), but faces limitations due to reduced transplant cell survival, low engraftment rates, and low retention. These limitations have been exacerbated due to the nature of cell delivery (narrow needles) and the non-optimal recipient microenvironment (reactive oxygen species (ROS)). Extracellular matrix (ECM) hydrogels derived from porcine left ventricular (LV) myocardium have emerged as a potential therapy to treat the ischemic LV and have shown promise as a vehicle to deliver cells to injured myocardium. However, no studies have evaluated the combination of an injectable biomaterial, such as an ECM hydrogel, in combination with cell therapy for treating RV failure. In this study we characterized LV and RV myocardial matrix (MM) hydrogels and performed in vitro evaluations of their potential to enhance CPC delivery, including resistance to forces experienced during injection and exposure to ROS, as well as their potential to enhance angiogenic paracrine signaling. While physical properties of the two hydrogels are similar, the decellularized LV and RV have distinct protein signatures. Both materials were equally effective in protecting CPCs against needle forces and ROS. CPCs encapsulated in either the LV MM or RV MM exhibited similar enhanced potential for angiogenic paracrine signaling when compared to CPCs in collagen. The RV MM without cells, however, likewise improved tube formation, suggesting it should also be evaluated as a potential standalone treatment.

Adventitia-derived extracellular matrix hydrogel enhances contractility of human vasa vasorum-derived pericytes via α2 β1 integrin and TGFβ receptor.

Pericytes are essential components of small blood vessels and are found in human aortic vasa vasorum. Prior work uncovered lower vasa vasorum density and decreased levels of pro-angiogenic growth factors in adventitial specimens of human ascending thoracic aortic aneurysm. We hypothesized that adventitial extracellular matrix (ECM) from normal aorta promotes pericyte function by increasing pericyte contractile function through mechanisms deficient in ECM derived from aneurysmal aortic adventitia. ECM biomaterials were prepared as lyophilized particulates from decellularized adventitial specimens of human and porcine aorta. Immortalized human aortic adventitia-derived pericytes were cultured within Type I collagen gels in the presence or absence of human or porcine adventitial ECMs. Cell contractility index was quantified by measuring the gel area immediately following gelation and after 48 h of culture. Normal human and porcine adventitial ECM increased contractility of pericytes when compared with pericytes cultured in absence of adventitial ECM. In contrast, aneurysm-derived human adventitial ECM failed to promote pericyte contractility. Pharmacological inhibition of TGFβR1 and antibody blockade of α2 β1 integrin independently decreased porcine adventitial ECM-induced pericyte contractility. By increasing pericyte contractility, adventitial ECM may improve microvascular function and thus represents a candidate biomaterial for less invasive and preventative treatment of human ascending aortic disease.

P-248 Fallopian tube extracellular matrix hydrogels for in vitro embryo culture in rabbit models: A post-natal study

Abstract Study question Can tissue-specific extracellular matrix (ECM) hydrogels derived from decellularized (DC) fallopian tubes have phenotypic consequences in the offspring when used for in vitro embryo culture? Summary answer Tissue-specific ECM hydrogels from DC rabbit fallopian tubes influence assisted reproductive technologies (ART) outcomes, resulting in offspring with a higher body weight. What is known already The in vitro culture of embryos fails to mimic the physiological dynamism occurring in the reproductive tract. In response to this sub-optimal environment, mammalian embryo exhibits a high degree of developmental plasticity, leading to specific phenotypic deviations at birth and beyond (e.g. low weight). Recently, it has been demonstrated that using in vivo derived culture complements, these changes can be ameliorated. The ECM is one of the elements shown to be important in retaining structural and biological signals from the native source tissue. The current bioengineering techniques could provide hydrogels mimicking the native environment and offering an optimal embryo development. Study design, size, duration This study was approved by an ethics committee (2018/VSC/PEA/0116). Rabbit oviducts (n = 24) were collected, fragmented, DC and solubilized to create hydrogels. After physicochemical characterization, the tissue-specific ECM hydrogels were used for in vitro culture of pronuclear embryos during 48 hours (n = 146), which were subsequently transferred to recipient females, along with embryos cultured under control conditions (n = 138). After delivery, the rabbits were weighed during 20 weeks and blood tests were performed at weeks 8 and 20. Participants/materials, setting, methods To create hydrogels, we employed a decellularization protocol based on SDS detergent. After characterizing and validating decellularization (Francés-Herrero et al. 2021), ECM hydrogels were used to generate coatings (overnight at 37ºC) on culture plates. Pronuclear embryos were cultured until morula stage and then transferred by laparoscopy to obtain the offspring. The animals were weighed and their blood was analyzed by hemogram. T-test was used for the statistical analysis after normality verification (p < 0.05 statistically significant). Main results and the role of chance After 48 hours of embryo culture, no differences in development and morphology were detected between the coating and control conditions (Francés-Herrero et al. 2021). With a total of 284 embryos transferred, the implantation and live birth rates were slightly higher in the coating group than in the control group (31.26 ± 21.44% vs 26.53 ± 24.8% and 19.96 ± 12.5% vs 18.68 ± 24.68%, respectively), but the differences were not statistically significant. During the first 20 weeks of life, the animals from the coating group consistently showed higher weights than the control group. Although this difference was not significant at birth (75.15 ± 15.48 g vs 61.88 ± 17.79 g, respectively), it was significant at subsequent measurements (1117.73 ± 172.54 g vs 942 ± 105.51 g at week 5, 2513 ± 352.83 g vs 2256.25 ± 168.52 g at week 10, 3475.56 ± 297.91 g vs 3257.5 ± 189.94 g; at week 15 and 4151.11 ± 269.28 g vs 3808.75 ± 293.08 g at week 20; p < 0.05). The animals of both experimental groups showed normal blood parameters (white blood cells, % lymphocytes, % monocytes, % granulocytes, red blood cells and hematocrit) at 8 and 20 weeks of life. Limitations, reasons for caution This study has been carried out in an animal model and, although the offspring generated is healthy, the scope of this technique is still unknown. So, the next step would be to study the embryonic reprogramming or media metabolomic profile under ECM in vitro culture. Wider implications of the findings These findings show how culture media supplementation with fallopian tube ECM allows improving in vitro conditions and decreases postnatal phenotypic deviations after ART. This study could initiate a new embryo culture techniques era to guarantee that ART are utilized in the most efficient and safest possible practice. Trial registration number Not applicable

Abstract 1880: High throughput screening of 30 PDX cell lines in a 3D ECM hydrogel platform, incorporating tumor, stroma and immune components to demonstrate simultaneous investigation of multiple anti-tumor modalities

Abstract High throughput screening offers tangible benefits towards rapidly testing various permutations of novel or existing therapeutic agents. In particular, tumor panels that cover a range of histotypes and molecular subtypes have been previously developed, such as the NCI-60, however they utilize cell lines and, in some cases, a 2D cell culture format, which limit their translatability to preclinical and clinical trials. Moreover, the biological complexity of the tumor microenvironment (TME) has revealed a need for more translatable 3D in vitro tumor models that reflect the in vivo physiological outcome to therapies, particularly with the explosion of immunotherapy programs in drug discovery which target the immune compartment of the TME. Here, we describe for the first time a 3D in vitro PDX panel comprising 30 distinct PDX models in coculture with fibroblasts and PBMCs in engineered extracellular matrix hydrogels that display distinct similarities to the three compartments of the TME - tumor, stroma, and immune cells. Histotypes in the panel include, amongst others, Non-small cell lung, colorectal, breast, pancreatic, gastric, melanoma, and renal cancers. The panel is constructed in a high throughput 96-well format and rapidly assays tumor growth delay and other endpoints such as apoptosis in a dose-dependent manner across various drug modalities such as small molecules, biologics and cell therapy. The panel has been tested against targeted therapy (Docetaxel, Cisplatin) and immunomodulatory agents (Pembrolizumab, Atezolizumab) and the results correlate to the corresponding PDX in vivo data. Interestingly, cisplatin treatment displayed an average size reduction of >30% for melanoma but a varied response in NSCLC tumors. And, several NSCLC responded more favorably to checkpoint inhibitors, however, the lung epidermoid PDX demonstrated no significant response. Moreover, subsequent cytokine analysis and immunofluorescence staining of several models revealed protein signatures of cancer-associated fibroblasts and CD3+ sequestration in the tumor stroma in some 3D models, suggesting the fibroblasts’ critical role in regulating the immune response. In short, the 30-PDX Panel described here represents a large step forward towards achieving translatable efficacy data at the earliest stages of drug discovery where little is known about the mechanism of action for a particular therapeutic agent or combination of agents. Citation Format: Kolin C. Hribar, Bin Xue, Christopher Harrod, Timothy Jensen, Julia Schüler. High throughput screening of 30 PDX cell lines in a 3D ECM hydrogel platform, incorporating tumor, stroma and immune components to demonstrate simultaneous investigation of multiple anti-tumor modalities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1880.

Biocompatibility-on-a-chip: Characterization and evaluation of decellularized tendon extracellular matrix (tdECM) hydrogel for 3D stem cell culture in a microfluidic device

Researchers have always tried expensive in vitro tests to show the 3D usability of dECM. The use of tissue-specific hydrogels in a microfluidic device is rarely studied. In this study, we have used ECM obtained from goat digital flexor tendons by decellularization technique. The tdECM was characterized for its structural properties using Scanning Electron Microscopy (SEM). Collagen, dsDNA, GAGs, and protein contents were quantified using spectrophotometric assays. The cell viability and proliferation of human umbilical cord-derived mesenchymal stem cells (hUMSCs) encapsulated in the tdECM hydrogel inside the microfluidic device were checked using Calcein-AM/PI. The FTIR data showed prominent peaks of the amide group, indicating the presence of collagen. The SEM data showed intact fiber morphology after the decellularization process. There was a 95 % reduction in double-stranded DNA (dsDNA) content, proving the effectiveness of the decellularization technique. There was no significant difference in the collagen content of tdECM and the GAGs were also in the acceptable range compared to the native tissue. Over 90 % cell viability in hUMSCs was observed qualitatively and quantitatively in vitro and inside a microfluidic device. In conclusion, we characterized the tdECM hydrogel and demonstrated its compatibility with the microfluidic device.

Peripheral Nerve Decellularization for In Vitro Extracellular Matrix Hydrogel Use: A Comparative Study.

The rise of tissue-engineered biomaterials has introduced more clinically translatable models of disease, including three-dimensional (3D) decellularized extracellular matrix (dECM) hydrogels. Specifically, decellularized nerve hydrogels have been utilized to model peripheral nerve injuries and disorders in vitro; however, there lacks standardization in decellularization methods. Here, rat sciatic nerves of varying preparations were decellularized using previously established methods: sodium deoxycholate (SD)-based, 3-((3-cholamidopropyl)dimethylammonio)-1-propanesulfonate (CHAPS)-based, and apoptosis-mediated. These nerves were characterized for cellular debris removal, ECM retention, and low cytotoxicity with cultured Schwann cells. The best preparations of each decellularization method were digested into dECM hydrogels, and rheological characterization, gelation kinetics, and confocal reflectance imaging of collagen fibril assembly were performed. It was determined that the SD-based method with nerve epineurial removal best maintained the overall ECM composition and mechanical properties of physiological peripheral nerves while efficiently stripping the scaffolds of tissue-specific cells and debris. This method was then utilized as a culture platform for quiescent Schwann cells and cancer-nerve crosstalk. Hydrogel-embedded Schwann cells were found to have high viability and act in a more physiologically relevant manner than those cultured in monolayers, and the hydrogel platform allowed for the activation of Schwann cells following treatment with cancer secreted factors. These findings establish a standard for peripheral nerve decellularization for usage as a dECM hydrogel testbed for in vitro peripheral nerve disease modeling and may facilitate the development of treatments for peripheral nerve disease and injury.

The Influence of New Bioactive Materials on Pulp–Dentin Complex Regeneration in the Assessment of Cone Bone Computed Tomography (CBCT) and Computed Micro-Tomography (Micro-CT) from a Present and Future Perspective—A Systematic Review

The present paper is the first article providing a systematic literature review on the visualization of tertiary dentin influenced by modern bioactive materials in CBCT and micro-CT. Six database searches of studies on tertiary dentin visualization using CBCT produced 622 records in total, and the search of the studies on tertiary dentin using micro-CT produced 502 records in total. The results were thoroughly selected considering the inclusion criteria, and five research papers using CBCT and nine research papers using micro-CT for visualization of tertiary dentin were eventually qualified for the analysis. All the non-randomized and randomized studies presented good and high levels of quality evidence, respectively. Among the bioactive materials used, the most frequently analysed were: MTA, Biodentine dentin matrix hydrogel, Pro Root MTA, and EndoSequence root repair material. The highest thickness of the tertiary dentin was achieved after the use of MTA material in both imaging techniques. The remaining parameters had different results, taking into account the CBCT and micro-CT analysis. The possibilities of the qualitative and quantitative assessment of the particular parameters of tertiary dentin using CBCT and micro-CT techniques were presented and analysed. CBCT and micro-CT analyses can be useful in the assessment of tertiary dentin formed beneath the bioactive material applied during vital pulp treatment. The research argues that the presented results differ depending on the material applied to the pulp, the study duration (4–6 weeks), difference in teeth, species (rats, human), as well as the applied technique and differences in computer software used for the analysis.

Bioprinted hASC-laden collagen/HA constructs with meringue-like macro/micropores.

Extrusion‐based bioprinting is one of the most effective methods for fabricating cell‐laden mesh structures. However, insufficient cellular activities within the printed cylindrical cell‐matrix blocks, inducing low cell‐to‐cell interactions due to the disturbance of the matrix hydrogel, remain to be addressed. Hence, various sacrificial materials or void‐forming methods have been used; however, most of them cannot solve the problem completely or require complicated fabricating procedures. Herein, we suggest a bioprinted cell‐laden collagen/hydroxyapatite (HA) construct comprising meringue‐like porous cell‐laden structures to enhance osteogenic activity. A porous bioink is generated using a culinary process, i.e., the whipping method, and the whipping conditions, such as the material concentration, time, and speed, are selected appropriately. The constructs fabricated using the meringue‐like bioink with MG63 cells and human adipose stem cells exhibit outstanding metabolic and osteogenic activities owing to the synergistic effects of the efficient cell‐to‐cell interactions and HA stimulation released from the porous structure. The in vitro cellular responses indicate that the meringue‐like collagen bioink for achieving an extremely porous cell‐laden construct can be a highly promising cell‐laden material for various tissue regeneration applications.

Coagulation/anticoagulation-regulable and tough extracellular matrix hydrogels

Many active proteins and growth factors exist in extracellular matrix (ECM), regulating important physiological processes in the body. The conversion of ECM into hydrogels will significantly expand their application scope. Given the complexity of the environment in vivo, fabrication of ECM hydrogels with regulable coagulation/anticoagulation is very important yet challenging. Herein, we utilize phosphate buffer (PBS) as a concentration switch to realize the coagulation/anticoagulation transformation of ECM hydrogels. Benefiting from their regulable characteristic, these ECM hydrogels are respectively converted into hemostatic sponges/powders and tough anticoagulant films. The tensile strength of the film reached 39.23 ± 3.21 MPa. Especially, blood flow signals were still detected after two weeks transplantation of ECM films onto rat carotid arteries (diameter < 1 mm) as a vascular patch, suggesting the potential application of ECM anticoagulant films in small-caliber vascular grafts. This work provided a facile and sustainable strategy for developing ECM hydrogels with regulable coagulant/anticoagulant properties.

Bioactive Decellularized Extracellular Matrix Hydrogel Microspheres Fabricated Using a Temperature-Controlling Microfluidic System.

Hydrogel microspheres have drawn great attention as functional three-dimensional (3D) microcarriers for cell attachment and growth, which have shown great potential in cell-based therapies and biomedical research. Hydrogels derived from a decellularized extracellular matrix (dECM) retain the intrinsic physical and biological cues from the native tissues, which often exhibit high bioactivity and tissue-specificity in promoting tissue regeneration. Herein, a novel two-stage temperature-controlling microfluidic system was developed which enabled production of pristine dECM hydrogel microspheres in a high-throughput manner. Porcine decellularized peripheral nerve matrix (pDNM) was used as the model raw dECM material for continuous generation of pDNM microgels without additional supporting materials or chemical crosslinking. The sizes of the microspheres were well-controlled by tuning the feed ratios of water/oil phases into the microfluidic device. The resulting pDNM microspheres (pDNM-MSs) were relatively stable, which maintained a spherical shape and a nanofibrous ultrastructure for at least 14 days. Schwann cells and PC12 cells preseeded on the pDNM-MSs not only showed excellent viability and an adhesive property, but also promoted cell extension compared to the commercially available gelatin microspheres. Moreover, primary neural stem/progenitor cells attached well to the pDNM-MSs, which further facilitated their proliferation. The successfully fabricated dECM hydrogel microspheres provided a highly bioactive microenvironment for 3D cell culture and functionalization, which showed promising potential in versatile biomedical applications.

Tissue extracellular matrix hydrogels as alternatives to Matrigel for culturing gastrointestinal organoids

Matrigel, a mouse tumor extracellular matrix protein mixture, is an indispensable component of most organoid tissue culture. However, it has limited the utility of organoids for drug development and regenerative medicine due to its tumor-derived origin, batch-to-batch variation, high cost, and safety issues. Here, we demonstrate that gastrointestinal tissue-derived extracellular matrix hydrogels are suitable substitutes for Matrigel in gastrointestinal organoid culture. We found that the development and function of gastric or intestinal organoids grown in tissue extracellular matrix hydrogels are comparable or often superior to those in Matrigel. In addition, gastrointestinal extracellular matrix hydrogels enabled long-term subculture and transplantation of organoids by providing gastrointestinal tissue-mimetic microenvironments. Tissue-specific and age-related extracellular matrix profiles that affect organoid development were also elucidated through proteomic analysis. Together, our results suggest that extracellular matrix hydrogels derived from decellularized gastrointestinal tissues are effective alternatives to the current gold standard, Matrigel, and produce organoids suitable for gastrointestinal disease modeling, drug development, and tissue regeneration.

Future Challenges and Opportunities of Extracellular Matrix Hydrogels in Female Reproductive Medicine.

Bioengineering and reproductive medicine have progressed shoulder to shoulder for several decades. A key point of overlap is the development and clinical translation of technologies to support reproductive health, e.g., scaffold-free constructs, polymeric scaffolds, bioprinting or microfluidics, and hydrogels. Hydrogels are the focus of intense study, and those that are derived from the extracellular matrix (ECM) of reproductive tissues and organs are emerging as promising new players given their results in pre-clinical models. This literature review addresses the recent advances in the use of organ-specific ECM hydrogels in reproductive medicine, considering the entire female reproductive tract. We discuss in-depth papers describing the development of ECM hydrogels, their use in in vitro models, and their in vivo application in preclinical studies. We also summarize the functions of hydrogels, including as grafts, carriers for cell transplantation, or drug depots, and present the potential and possible scope for use of ECM hydrogels in the near future based on recent scientific advances.

Kidney Decellularized Extracellular Matrix Enhanced the Vascularization and Maturation of Human Kidney Organoids.

Kidney organoids derived from human pluripotent stem cells (hPSCs) have extensive potential for disease modelling and regenerative medicine. However, the limited vascularization and immaturity of kidney organoids have been still remained to overcome. Extracellular matrix (ECM) can provide mechanical support and a biochemical microenvironment for cell growth and differentiation. Here in vitro methods using a kidney decellularized extracellular matrix (dECM) hydrogel to culture hPSC‐derived kidney organoids, which have extensive vascular network and their own endothelial cells, are reported. Single‐cell transcriptomics reveal that the vascularized kidney organoids cultured using the kidney dECM have more mature patterns of glomerular development and higher similarity to human kidney than those cultured without the kidney dECM. Differentiation of α‐galactosidase A (GLA)‐knock‐out hPSCs generated using CRISPR/Cas9 into kidney organoids by the culture method using kidney dECM efficiently recapitulate Fabry nephropathy with vasculopathy. Transplantation of kidney organoids with kidney dECM into kidney of mouse accelerates the recruitment of endothelial cells from the host mouse kidney and maintains vascular integrity with the more organized slit diaphragm‐like structures than those without kidney dECM. The kidney dECM methodology for inducing extensive vascularization and maturation of kidney organoids can be applied to studies for kidney development, disease modeling, and regenerative medicine.