Regenerative Therapy Applications Research Articles

Exosomes derived from Polygonum multiflorum-treated human dental pulp stem cells (hDPSCs): New approach in regenerative medicine

Extracellular vesicles (EVs) are currently being extensively utilized in clinical trials. Our previous study demonstrated that the conditioned medium of human dental pulp stem cells (hDPSCs) treated by 2,3,5,4′-tetrahydroxystilbene-2-O-β-D-glucoside (THSG) exhibits biological activity and anti-inflammatory responses, which are believed to be regulated by EVs. To date, no study has thoroughly investigated the potential of the exosomes released by THSG-treated hDPSCs. In this study, we utilized the conditioned medium (CM) of hDPSCs with/without THSG stimulation to compare the differences in exosome extraction using ultracentrifugation (UC), tangential flow filtration (TFF), and TFF combined with size-exclusion chromatography (TFF + SEC) methods, aiming to establish an effective purification method. Subsequently, we analyzed the contents of the exosomes using proteomic strategy. The results showed that TFF + SEC was more effective in reducing protein contamination compared to CM and TFF alone. TFF effectively concentrated large-scale CM and maintained exosome particle sizes consistently around 85–100 nm. Flow cytometry analysis revealed that the presence of THSG increased exosome secretion from hDPSCs, concurrently leading to the identification of 219 unique proteins through proteomic analysis, including BMP-1, VEGF, and members of the MMP family. Compared to non-lyophilized exosomes, lyophilized exosomes showed no change in protein activity, indicating that lyophilization was an effective preservation method. Collectively, these findings present a combined extraction strategy for dental stem cell exosomes and reveal the high purity and proteome composition of THSG-exosomes, thereby highlighting their potential applications in regenerative therapy.

Growth Factor Delivery Using a Collagen Membrane for Bone Tissue Regeneration.

The use of biomaterials and bioactive agents has shown promise in bone defect repair, leading to the development of strategies for bone regeneration. Various artificial membranes, especially collagen membranes (CMs) that are widely used for periodontal therapy and provide an extracellular matrix-simulating environment, play a significant role in promoting bone regeneration. In addition, numerous growth factors (GFs) have been used as clinical applications in regenerative therapy. However, it has been established that the unregulated administration of these factors may not work to their full regenerative potential and could also trigger unfavorable side effects. The utilization of these factors in clinical settings is still restricted due to the lack of effective delivery systems and biomaterial carriers. Hence, considering the efficiency of bone regeneration, both spaces maintained using CMs and GFs can synergistically create successful outcomes in bone tissue engineering. Therefore, recent studies have demonstrated a significant interest in the potential of combining CMs and GFs to effectively promote bone repair. This approach holds great promise and has become a focal point in our research. The purpose of this review is to highlight the role of CMs containing GFs in the regeneration of bone tissue, and to discuss their use in preclinical animal models of regeneration. Additionally, the review addresses potential concerns and suggests future research directions for growth factor therapy in the field of regenerative science.

Plant-Derived Exosome-like Nanoparticles for Biomedical Applications and Regenerative Therapy.

Plant-derived exosome-like nanoparticles (PDENs) comprise various bioactive biomolecules. As an alternative cell-free therapeutic approach, they have the potential to deliver nano-bioactive compounds to the human body, and thus lead to various anti-inflammatory, antioxidant, and anti-tumor benefits. Moreover, it is known that Indonesia is one of the herbal centers of the world, with an abundance of unexplored sources of PDENs. This encouraged further research in biomedical science to develop natural richness in plants as a source for human welfare. This study aims to verify the potential of PDENs for biomedical purposes, especially for regenerative therapy applications, by collecting and analyzing data from the latest relevant research and developments.

A review of regenerative therapy for spinal cord injury using human iPS cells

Spinal cord injury (SCI) has been considered to cause sudden, irreversible loss of function in patients. However, developments in stem cell biology and regenerative medicine are changing this conventional notion. Here we reviewed the overview of regenerative medicine of SCI.As a consequence of the establishment of human induced pluripotent stem cells (hiPSCs), hiPSC-based therapies for SCI, such as neural stem/progenitor cell (NS/PC) transplantation, have emerged as promising therapeutic modalities. Using several animal models, hiPSC-NS/PC transplantation into subacute injured spinal cords has been repeatedly demonstrated to improve locomotor function.Some biological mechanisms underlying this improvement have been proposed. In particular, combined with advanced neuroscience techniques such as designer receptors exclusively activated by designer drugs (DREADDs), neuronal relay theory, in which the transplanted cell-derived neurons reconstruct disrupted neuronal circuits, was proven to be involved histologically, pharmaceutically, electrophysiologically, and via in vivo bioimaging.Based on these findings, hiPSC-NS/PC transplantation for subacute SCI was moved ahead to a clinical study on human patients. At the same time, the search for effective treatments for chronic SCI is proceeding gradually, combining hiPSC-NS/PC transplantation with other treatment modalities such as rehabilitation, pharmaceutical interventions, or optimal scaffolds. In addition to NS/PCs, oligodendrocyte precursor cells (OPCs) are also a promising cell source for transplantation, as demyelinated axons affected by SCI can be repaired by OPCs. Therapies with OPCs derived from hiPSCs are still in preclinical studies but have shown favorable outcomes in animal models.In the future, several therapeutic options may be available according to the pathological conditions and the time period of SCI. Moreover, the application of regenerative therapy for the spinal cord could be broadened to degenerative disorders, such as spinal canal stenosis.Summary sentence: A historical review of human induced pluripotent stem cell (hiPSC) based cell transplantation therapy for spinal cord injury (SCI), in particular about footsteps of hiPSC-derived neural stem/progenitor cell transplantation, recent clinical study, and its future perspective.

Otolog Trombosit Konsantrelerinin İmmunolojik ve Antimikrobiyal Etkileri

Antibiotic resistance has remarkable potential in human beings and veterinary medicine. However, to prevent the clinical reflection of this resistance from reaching the feared dimensions, there is a requirement for antimicrobial treatment options supported and improved with new molecular biocursors at the preclinical point. Platelet-rich plasma (PRP) and fibrin (PRF) are biomaterial products that recently used to increase the anti-infective defense system by platelet growth factors to support postoperative wound healing, bone regeneration, graft stabilization, biofilm inhibition, catheter hygiene, and hemostasis. Recently, research has been carried out on antibacterial, antifungal, and prevention of clinical biofilm formation. Autologous platelet concentrates are autogenous and do not cause any immunological reaction or infection. Therefore, the choice and application of regenerative therapies are being favored due to their nominal invasive procedures. In particular, PRP and PRF are of interest because of their influence to stimulate and speed up the injury area healing process. Cytokines and growth factors involved in the formation of PRP are played an important role in the recovery process. This article aims to evaluate the antibacterial, antifungal and antibiofilm properties of PRP and PRF in the field of microbiology. In addition, the act of growth factors in the process of healing and their use in regenerative treatments were also evaluated.

PD34-02 CHEMOKINE THERAPY FOR ANAL SPHINCTER INJURY: DEVELOPMENT OF A RAT MODEL OF CHRONIC INJURY

You have accessJournal of UrologyCME1 May 2022PD34-02 CHEMOKINE THERAPY FOR ANAL SPHINCTER INJURY: DEVELOPMENT OF A RAT MODEL OF CHRONIC INJURY Amr El Haraki, Shannon Lankford, Wencheng Li, Catherine Matthews, Koudy Williams, and Gopal Badlani Amr El HarakiAmr El Haraki More articles by this author , Shannon LankfordShannon Lankford More articles by this author , Wencheng LiWencheng Li More articles by this author , Catherine MatthewsCatherine Matthews More articles by this author , Koudy WilliamsKoudy Williams More articles by this author , and Gopal BadlaniGopal Badlani More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002585.02AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Stromal cell-derived factor 1 (CXCL12) is a potential alternative to stem cell augmentation therapy for management of fecal incontinence. The objective of this study was to develop a rat model of anal sphincter injury and determine the predictability of using anal manometry in the rat model to collaborate histologic findings. METHODS: Adult female rats were divided into three groups: Group A was a control group that did not undergo surgery; group B had an anal sphincterotomy with primary surgical repair and group C had an anal sphincterotomy with primary surgical repair and intra-sphincteric injection of 200 ng of CXCL12 at 6 weeks post-injury. All rats underwent anal manometry measurements at baseline and at 6 weeks and 12 weeks post-injury. In each rat, the three strongest contractions were chosen for measurement and mean values of the following: total duration (D), time to peak (Tmax), and difference between maximal and minimal pressure value (P∆) were calculated and compared between groups. Mason-Trichrome staining of the anal sphincters was performed and histologic analysis was obtained. RESULTS: 16 rats, 5 in Group A, 5 in Group B and 6 in Group C were investigated. At baseline and at 6 weeks, there were no statistically significant differences between D, Tmax and P∆ of Groups A, B and C. At 12-week manometry, the total duration of contractions on anal manometry was significantly less in Group C compared to Groups A and B (3.65, 5.5, 5.3 p<0.01) as was time to peak of contraction at 12 weeks (1.6,2.1,3.1 p<0.01); however, group C had a significantly higher P∆ at 12 weeks compared to Groups A and B (2.25,1.4, 0.34 p<0.01). On histologic analysis, there were no statistically significant differences in the ratio of muscle to collagen at the site of injury between the different groups, however, muscle fibers were significantly smaller in group C and less per bundle than groups A and B. CONCLUSIONS: Delayed injection of CXCL12 seemed to enhance the magnitude of anal sphincter contractions on anorectal manometry in a rat model of anal sphincter injury. The mechanism of increased anal sphincter contraction magnitude was not explained by histologic differences in explanted specimens. A deficit in the rat model, the rat's intrinsic self-healing ability and difficulty to accurately measure direct sphincter activity, likely contributed to this discrepancy. A better animal model will be needed in future experiments of fecal incontinence to assess the application of regenerative therapy. Source of Funding: Wake Forest Institute of Regenerative Medicine © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e559 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Amr El Haraki More articles by this author Shannon Lankford More articles by this author Wencheng Li More articles by this author Catherine Matthews More articles by this author Koudy Williams More articles by this author Gopal Badlani More articles by this author Expand All Advertisement PDF DownloadLoading ...

Effects of concentrated growth factor on cells and tissues and applications in regenerative therapy

Effects of concentrated growth factor on cells and tissues and applications in regenerative therapy

Isolation and Culture of Primary Oral Keratinocytes from the Adult Mouse Palate.

For years, most studies involving keratinocytes have been conducted using human and mouse skin epidermal keratinocytes. Recently, oral keratinocytes have attracted attention because of their unique function and characteristics. They maintain the homeostasis of the oral epithelium and serve as resources for applications in regenerative therapies. However, in vitro studies that use oral primary keratinocytes from adult mice have been limited due to the lack of an efficient and well-established culture protocol. Here, oral primary keratinocytes were isolated from the palate tissues of adult mice and cultured in a commercial low-calcium medium supplemented with a chelexed-serum. Under these conditions, keratinocytes were maintained in a proliferative or stem cell-like state, and their differentiation was inhibited even after increased passages. Marker expression analysis showed that the cultured oral keratinocytes expressed the basal cell markers p63, K14, and α6-integrin and were negative for the differentiation marker K13 and the fibroblast marker PDGFRα. This method produced viable and culturable cells suitable for downstream applications in the study of oral epithelial stem cell functions in vitro.

Isolation and Culture of Primary Oral Keratinocytes from the Adult Mouse Palate

For years, most studies involving keratinocytes have been conducted using human and mouse skin epidermal keratinocytes. Recently, oral keratinocytes have attracted attention because of their unique function and characteristics. They maintain the homeostasis of the oral epithelium and serve as resources for applications in regenerative therapies. However, in vitro studies that use oral primary keratinocytes from adult mice have been limited due to the lack of an efficient and well-established culture protocol. Here, oral primary keratinocytes were isolated from the palate tissues of adult mice and cultured in a commercial low-calcium medium supplemented with a chelexed-serum. Under these conditions, keratinocytes were maintained in a proliferative or stem cell-like state, and their differentiation was inhibited even after increased passages. Marker expression analysis showed that the cultured oral keratinocytes expressed the basal cell markers p63, K14, and α6-integrin and were negative for the differentiation marker K13 and the fibroblast marker PDGFRα. This method produced viable and culturable cells suitable for downstream applications in the study of oral epithelial stem cell functions in vitro.

Comparison of stem cells from human exfoliated deciduous posterior teeth with varying levels of root resorption.

Stem cells from human exfoliated deciduous teeth (SHED) are regarded as an attractive cell source for tissue regeneration. However, the effect of different levels of root resorption on the characteristics of SHED remains less understood. Thus, the tooth source that is most suitable for the isolation of SHEDs needs to be determined. To compare cellular and biological characteristics of stem cells from human exfoliated deciduous posterior teeth with varying levels of root resorption. The pulp was obtained from the deciduous posterior teeth depending on the level of root resorption, and isolated SHEDs were grouped as follows: Teeth with 0 to 1/3rd root resorption as SHEDs (G1) and 1/3rd to 2/3rd root resorption as SHEDs (G2). Teeth were also collected from >2/3rd root resorption status, but failed to establish primary culture of SHED as the availability of pulp tissue was too less. Later, isolated SHEDs were compared on their morphology, viability, growth kinetics, colony-forming ability, expression of cell surface markers and in vitro differentiation into osteocytes and adipocytes. No major differences were observed in terms of cellular morphology, viability, proliferation rate, colony-forming ability, cell surface markers expression, and mesenchymal lineage differentiation of SHEDs isolated from posterior teeth with 0 to 1/3rd and 1/3rd to 2/3rd root resorption. However, SHED from teeth with 0 to 1/3rd root resorption (G1) displayed relatively higher proliferation capacity and expression of selected markers. Collectively, SHEDs (G1) and SHEDs (G2) showed comparable cellular and biological characteristics that enable their possible applications in regenerative therapies.

Effect of Ascorbic Acid on Differentiation, Secretome and Stemness of Stem Cells from Human Exfoliated Deciduous Tooth (SHEDs)

Stem cells from human exfoliated deciduous teeth (SHEDs) are considered a type of mesenchymal stem cells (MSCs) because of their unique origin from the neural crest. SHEDs can self-renewal and multi-lineage differentiation with the ability to differentiate into odontoblasts, osteoblast, chondrocytes, neuronal cells, hepatocytes, adipocytes, etc. They are emerging as an ideal source of MSCs because of their easy availability and extraordinary cell number. Ascorbic acid, or vitamin C, has many cell-based applications, such as bone regeneration, osteoblastic differentiation, or extracellular matrix production. It also impacts stem cell plasticity and the ability to sustain pluripotent activity. In this study, we evaluate the effects of ascorbic acid on stemness, paracrine secretion, and differentiation into osteoblast, chondrocytes, and adipocytes. SHEDs displayed enhanced multifaceted activity, which may have applications in regenerative therapy.

Corneal Ulcer in Dogs and Cats: Novel Clinical Application of Regenerative Therapy Using Subconjunctival Injection of Autologous Platelet-Rich Plasma.

Background: Corneal ulcer could be a major source of distress in small animals, with many contributing agents. In recent years, few studies evaluated the efficacy of platelet-rich plasma (PRP) in healing corneal ulcers.Aim: This study aimed to assess the ability of subconjunctival injection of autologous PRP in the treatment of corneal ulcers in dogs and cats as well as estimate the expression of matrix metalloproteinase (MMP)-2, MMP-9, and oxidative stress biomarkers in these patients.Methods: A total number of 28 animals (16 cats and 12 dogs) were enrolled in this study. Each animal was subjected to clinical, neurologic, and ophthalmic examinations where the type of ulcer was documented. Tear samples were collected for evaluation of oxidative biomarkers and MMPs; conjunctival swabs were taken to identify the involved organism. PRP was prepared from each animal and given as subconjunctival injection; numbers of injections were done according to case response. Clinical follow-up was done and documented for each case.Results: In cat patients, female and Persian cats were most affected; unilateral and superficial ulcers were most recorded. In male dogs, unilateral, and superficial ulcers were most recorded. FHV-1 was most identified in cats, while Staphylococcus aureus was most identified in dogs. Numbers of injections needed to achieve healing were recorded, with 50% of dogs needing two injections with 1-week intervals and 50% of cats needed three injections with 1-week intervals. Alterations in both oxidative biomarkers and MMPs were recorded in affected animals.Conclusion: The use of autologous PRP as a subconjunctival injection in treating corneal ulcers in dogs and cats is effective. The number of injections is the case and corneal ulcer type-dependent.Clinical Significance: Autologous PRP as a subconjunctival injection in treating corneal ulcer is a relatively cheap, safe method and can be done in the clinical setting.

Prospects for 3D bioprinting of organoids

Three-dimensional (3D) organoids derived from pluripotent or adult tissue stem cells seem to possess excellent potential for studying development and disease mechanisms alongside having a myriad of applications in regenerative therapies. However, lack of precise architectures and large-scale tissue sizes are some of the key limitations of current organoid technologies. 3D bioprinting of organoids has recently emerged to address some of these impediments. In this review, we discuss 3D bioprinting with respect to the use of bioinks and bioprinting methods and highlight recent studies that have shown success in bioprinting of stem cells and organoids. We also summarize the use of several vascularization strategies for the bioprinted organoids, that are critical for a complex tissue organization. To fully realize the translational applications of organoids in disease modeling and regenerative medicine, these areas in 3D bioprinting need to be appropriately harnessed and channelized.

Subchondral Bone Remodeling: A Therapeutic Target for Osteoarthritis.

There is emerging awareness that subchondral bone remodeling plays an important role in the development of osteoarthritis (OA). This review presents recent investigations on the cellular and molecular mechanism of subchondral bone remodeling, and summarizes the current interventions and potential therapeutic targets related to OA subchondral bone remodeling. The first part of this review covers key cells and molecular mediators involved in subchondral bone remodeling (osteoclasts, osteoblasts, osteocytes, bone extracellular matrix, vascularization, nerve innervation, and related signaling pathways). The second part of this review describes candidate treatments for OA subchondral bone remodeling, including the use of bone-acting reagents and the application of regenerative therapies. Currently available clinical OA therapies and known responses in subchondral bone remodeling are summarized as a basis for the investigation of potential therapeutic mediators.

Overcoming Current Dilemma in Cartilage Regeneration: Will Direct Conversion Provide a Breakthrough?

Direct reprogramming/direct conversion/transdifferentiation is a process that induces conversion between completely different matured (differentiated) cells in higher organisms. Unlike the process of reprogramming of differentiated cells into induced pluripotent stem cells (iPSCs) and re-differentiation into the desired cell types, differentiated cells undergo the conversion into another type of differentiated cells without going through the iPSCs state. Osteoarthritis (OA) is the most common type of arthritis that causes a significant deterioration in patients' quality of life. The high prevalence of OA as well as the current lack of disease-modifying drugs has led to a rise in regenerative strategy for OA treatment. Regenerative therapy in OA started with the concept of engraftment of the administered cells within the cartilage lesion and differentiation to chondrocytes after the engraftment. However, recent studies show that cells, particularly when injected in suspension, rapidly undergo apoptosis after exerting a transient paracrine effect. In this perspective review, the general overview and current status of direct conversion are introduced along with the conceptual strategy and future directions for possible application of regenerative therapy using stem cells in OA. In vivo direct conversion may open a new stage of regenerative medicine for OA treatment. Recent advances in in vivo gene transfer and smart biomaterials can bring the concept into reality in near future. Direct conversion can be a new type of treatment technology that has the potential to overcome the limitations of current cell therapy.

Synthetic tissue engineering: Programming multicellular self-organization by designing customized cell-cell communication.

Cells communicate with each other to organize multicellular collective systems and assemble complex, elaborate tissue structures by themselves during development. Despite intensive biological studies, what kind of cell-cell communication can sufficiently drive self-organization of specific tissue architectures remain unclear. Thanks to recent advances on genetic engineering technologies, synthetic biologists start to build customized cell-cell communication with user-defined signal input and gene expression output to program multicellular behaviors using mammalian systems. This review article introduces how we can design and engineer customized cell-cell communication to program synthetic self-organizing multicellular structures. Creating tissue formation processes with synthetic genetic programs will help understanding of fundamental principles of how genetic programs drive tissue self-organization and provide new capabilities on tissue engineering for cell-based regenerative therapy applications.

Baboon induced pluripotent stem cell generation by piggyBac transposition of reprogramming factors.

Clinical application of regenerative therapies using embryonic or induced pluripotent stem cells is within reach. Progress made during recent years has encouraged researchers to address remaining open questions in order to finally translate experimental cell replacement therapies into application in patients. To achieve this, studies in translationally relevant animal models are required to make the final step to the clinic. In this context, the baboon (Papio anubis) may represent a valuable nonhuman primate (NHP) model to test cell replacement therapies because of its close evolutionary relationship to humans and its large body size. In this study, we describe the reprogramming of adult baboon skin fibroblasts using the piggyBac transposon system. Via transposon-mediated overexpression of six reprogramming factors, we generated five baboon induced pluripotent stem cell (iPSC) lines. The iPSC lines were characterized with respect to alkaline phosphatase activity, pluripotency factor expression analysis, teratoma formation potential, and karyotype. Furthermore, after initial cocultivation with mouse embryonic fibroblasts, we were able to adapt iPSC lines to feeder-free conditions. In conclusion, we established a robust and efficient protocol for iPSC generation from adult baboon fibroblasts.

An Optimized System for Effective Derivation of Three-Dimensional Retinal Tissue via Wnt Signaling Regulation.

Effective derivation of three-dimensional (3D) retinal tissue from human-induced pluripotent stem cells (hiPSCs) could provide models for drug screening and facilitate patient-specific retinal cell replacement therapy. However, some hiPSC lines cannot undergo 3D self-organization and show inadequate differentiation efficiency to meet clinical demand. In this study, we developed an optimized system for derivation of 3D retinal tissue. We found that the Wnt signaling pathway antagonist Dickkopf-related protein 1 (DKK-1) rescued the inability of differentiated retinal progenitors to self-organize. By evaluating DKK-1 expression and supplying DKK-1 if necessary, retinal organoids were differentiated from six hiPSC lines, which were reprogramed from three common initiating cell types. Retinal tissues derived from the optimized system were well organized and capable of surviving for further maturation. Thus, using this system, we generated retinal tissues from various hiPSC lines with high efficiency. This novel system has many potential applications in regenerative therapy and precision medicine. Stem Cells 2018;36:1709-1722.

Current Strategies to Generate Human Mesenchymal Stem Cells In Vitro.

Mesenchymal stem cells (MSCs) are heterogeneous multipotent stem cells that are involved in the development of mesenchyme-derived evolving structures and organs during ontogeny. In the adult organism, reservoirs of MSCs can be found in almost all tissues where MSCs contribute to the maintenance of organ integrity. The use of these different MSCs for cell-based therapies has been extensively studied over the past years, which highlights the use of MSCs as a promising option for the treatment of various diseases including autoimmune and cardiovascular disorders. However, the proportion of MSCs contained in primary isolates of adult tissue biopsies is rather low and, thus, vigorous ex vivo expansion is needed especially for therapies that may require extensive and repetitive cell substitution. Therefore, more easily and accessible sources of MSCs are needed. This review summarizes the current knowledge of the different strategies to generate human MSCs in vitro as an alternative method for their applications in regenerative therapy.

Epigallocatechin Gallate-Modified Gelatin Sponges Treated by Vacuum Heating as a Novel Scaffold for Bone Tissue Engineering.

Chemical modification of gelatin using epigallocatechin gallate (EGCG) promotes bone formation in vivo. However, further improvements are required to increase the mechanical strength and bone-forming ability of fabricated EGCG-modified gelatin sponges (EGCG-GS) for practical applications in regenerative therapy. In the present study, we investigated whether vacuum heating-induced dehydrothermal cross-linking of EGCG-GS enhances bone formation in critical-sized rat calvarial defects. The bone-forming ability of vacuum-heated EGCG-GS (vhEGCG-GS) and other sponges was evaluated by micro-computed tomography and histological staining. The degradation of sponges was assessed using protein assays, and cell morphology and proliferation were verified by scanning electron microscopy and immunostaining using osteoblastic UMR106 cells in vitro. Four weeks after the implantation of sponges, greater bone formation was detected for vhEGCG-GS than for EGCG-GS or vacuum-heated gelatin sponges (dehydrothermal cross-linked sponges without EGCG). In vitro experiments revealed that the relatively low degradability of vhEGCG-GS supports cell attachment, proliferation, and cell–cell communication on the matrix. These findings suggest that vacuum heating enhanced the bone forming ability of EGCG-GS, possibly via the dehydrothermal cross-linking of EGCG-GS, which provides a scaffold for cells, and by maintaining the pharmacological effect of EGCG.