Regenerative Medicine Applications Research Articles

Decellularization and characterization of camel pericardium as a new scaffold for tissue engineering and regenerative medicine.

Valvular heart diseases (VHDs) have become prevalent in populations due to aging. Application of different biomaterials for cardiac valve regeneration and repair holds a great promise for treatment of VHD. Aortic valve replacement using tissue-engineered xenografts is a considered approach, and the pericardium of different species such as porcine and bovine has been studied over the last few years. It has been suggested that the animal origin can affect the outcomes of replacement. So, herein, we at first decellularized and characterized the camel pericardium (dCP), then characterized dCP with H&E staining, in vitro and in vivo biocompatibility and mechanical tests and compared it with decellularized bovine pericardium (dBP), to describe the potency of dCP as a new xenograft and bio scaffold. The histological assays indicated less decluttering and extracellular matrix damage in dCP after decellularization compared to the dBP also dCP had higher Young Modulus (105.11), and yield stress (1.57 ± 0.45). We observed more blood vessels and also less inflammatory cells in the dCP sections after implantation. In conclusion, the results of this study showed that the dCP has good capabilities not only for use in VHD treatment but also for other applications in tissue engineering and regenerative medicine.

Emerging Trends and Future Prospects in Microfluidic Systems for Prevention and Diagnosis of Infection Pathogens

AbstractInfectious diseases caused by bacteria, viruses, fungi, and parasites pose a significant societal challenge. In response, scientists are developing advanced technology to enhance the prevention, diagnosis, and treatment of such diseases. One such promising technology is microfluidic systems, which are utilized in organ‐on‐a‐chip systems to replicate the microenvironments of organs. These systems have potential applications in drug screening, disease modeling, and personalized medicine. This review provides an overview of recent advances in organ‐on‐a‐chip platforms and their potential for preventing and diagnosing various infections. After discussing traditional techniques employed in studying infectious diseases, the role of microfluidic platforms in detecting infections is delved in. It is expound on advanced microfluidic‐based strategies for infection diagnosis, such as the polymerase chain reaction‐based microfluidic devices, enzyme linked immunosorbent assay‐based microfluidic devices, hierarchical nanofluidic molecular enrichment systemand µWestern blotting‐based microfluidic devices, and smartphone‐based microfluidic devices. Additionally, future research challenges and perspectives are discussed on microfluidic systems in biomedical and regenerative medicine applications. Consequently, microfluidic platforms have the potential to serve as fundamental frameworks for understanding infectious diseases, thereby leading to personalized regenerative medicine. hierarchical nanofluidic molecular enrichment system

ATPSpin: A Single Microfluidic Platform that Produces Diversified ATPS-Alginate Microfibers.

Microfluidic spinning is emerging as a useful technique in the fabrication of alginate fibers, enabling applications in drug screening, disease modeling, and disease diagnostics. In this paper, by capitalizing on the benefits of aqueous two-phase systems (ATPS) to produce diverse alginate fiber forms, we introduce an ATPS-Spinning platform (ATPSpin). This ATPS-enabled method efficiently circumvents the rapid clogging challenges inherent to traditional fiber production techniques by regulating the interaction between alginate and cross-linking agents like Ba2+ ions. By varying system parameters under the guidance of a regime map, our system produces several fiber forms─solid, hollow, and droplet-filled─consistently and reproducibly from a single device. We demonstrate that the resulting alginate fibers possess distinct features, including biocompatibility. We also encapsulate HEK293 cells in the microfibers as a proof-of-concept that this versatile microfluidic fiber generation platform may have utility in tissue engineering and regenerative medicine applications.

3D-printed short nanofibers/hydrogel-based vaginal films as a novel system for the delivery of anti-HIV microbicide drugs

Acquired immunodeficiency syndrome (AIDS) is a global epidemic that affects the lives of millions of young women, particularly those who live in poverty-stricken regions and engage in unsafe sexual behaviors, forcing the development of unique pre-exposure anti-HIV prophylaxis tools. Using a combination of antiretroviral medications with various antiviral mechanisms significantly improves the prognosis for HIV patients. The integration of 3D printing technology, short nanofiber technology, and hydrogel into a single system known as “3D-printed nanocomposites” is an emerging approach that is gaining traction in tissue engineering, regenerative medicine, and biomedical applications. Based on these foundations, the current study aims to develop, for the first time, a 3D-printed short nanofiber/film for the vaginal delivery of ritonavir and efavirenz in a fixed-dose combination. To identify an optimal 3D-printable short nanofibers/hydrogel formulation, multiple formulations, and fragmentation methods were investigated. Following that, the generated vaginal films were evaluated using several physicochemical characterizations, mechanical properties measurements, and in vitro release studies. The findings revealed that only cryocutting is effective for polycaprolactone nanofiber fragmentation. The 3D-printed short nanofibers-loaded films had an ultimate tensile strength of 6.75 ± 1.27 MPa and elongation at break of 262.67 ± 44.32 MPa, suggesting their reliability for handling and vaginal application. Furthermore, the in vitro release study demonstrated that rapid EFZ release occurred during the first 45 min, whereas RIT release lasted more than 72 h, indicating that the complex drug release profile was met. In summary, the study emphasizes that 3D printing of short nanofiber-based films is likely to be an effective platform for localized vaginal drug delivery, particularly anti-HIV therapies.

Utilizing stem cell-secreted molecules as a versatile toolbox for skin regenerative medicine

Stem cells are recognized as an important target and tool in regenerative engineering. In this study, we explored the feasibility of engineering amniotic fluid-derived mesenchymal stem cell-secreted molecules (afMSC-SMs) as a versatile bioactive material for skin regenerative medicine applications in a time- and cost-efficient and straightforward manner. afMSC-SMs, obtained in powder form through ethanol precipitation, effectively contributed to preserving the self-renewal capacity and differentiation potential of primary human keratinocytes (pKCs) in a xeno-free environment, offering a potential alternative to traditional culture methods for their long-term in vitro expansion, and allowed them to reconstitute a fully stratified epithelium sheet on human dermal fibroblasts. Furthermore, we demonstrated the flexibility of afMSC-SMs in wound healing and hair regrowth through injectable hydrogel and nanogel-mediated transdermal delivery systems, respectively, expanding the pool of regenerative applications. This cell-free approach may offer several potential advantages, including streamlined manufacturing processes, scalability, controlled formulation, longer shelf lives, and mitigation of risks associated with living cell transplantation. Accordingly, afMSC-SMs could serve as a promising therapeutic toolbox for advancing cell-free regenerative medicine, simplifying their broad applicability in various clinical settings.

Chromatin Condensation Delays Senescence in Human Mesenchymal Stem Cells by Safeguarding Nuclear Damages during In Vitro Expansion

Human mesenchymal stem cells (hMSCs) are multipotent cells that differentiate into adipocytes, chondrocytes, and osteoblasts. Owing to their differentiation potential, hMSCs are among the cells most frequently used for therapeutic applications in tissue engineering and regenerative medicine. However, the number of cells obtained through isolation alone is insufficient for hMSC-based therapies and basic research, which necessitates in vitro expansion. Conventionally, this is often performed on rigid surfaces such as tissue culture plates (TCPs). However, during in vitro expansion, hMSCs lose their proliferative ability and multilineage differentiation potential, rendering them unsuitable for clinical use. Although multiple approaches have been attempted to maintain hMSC stemness during prolonged expansion, finding a suitable culture system remains an unmet need. Recently, a few research groups have shown that hMSCs maintain their stemness over long passages when cultured on soft substrates. In addition, it has been shown that hMSCs cultured on soft substrates have more condensed chromatin and lower levels of histone acetylation compared to those cultured on stiff substrates. Furthermore, it has also been shown that condensing/decondensing chromatin by deacetylation/acetylation can delay replicative senescence in hMSCs during long-term expansion on TCPs. However, the mechanism by which chromatin condensation/decondensation influences nuclear morphology and DNA damage, which are strongly related to the onset of senescence, remains unknown. To answer this question, we cultured hMSCs for long duration in the presence of epigenetic modifiers, histone acetyltransferase inhibitor (HATi), which promotes chromatin condensation by preventing histone acetylation, and histone deacetylase inhibitor (HDACi), which promotes chromatin decondensation, and investigated their effects on various nuclear markers related to senescence. We found that consistent acetylation causes severe nuclear abnormalities, whereas chromatin condensation by deacetylation helps to safeguard the nucleus from damage caused by in vitro expansion.

Decellularized porcine vena cava grafts are fully repopulated after orthotopic implantation

Tissue-engineered organs, based on native extracellular matrix (ECM) scaffolds, could be a game changer in regenerative medicine applications. Decellularization technology provides such scaffolds with organ-typic ECM composition and architecture. Despite limitations such as the requirement of huge cell numbers and finding the optimal route of entry, recellularized scaffolds provide alternative grafts for transplantation. In this study we assessed whether decellularized scaffolds, when implanted, are repopulated from the adjacent tissue. Since the vasculature plays an important role in tissue functionality, our main focus was to evaluate in situ repopulation of decellularized veins in a pig model. For this, porcine inferior vena cava grafts were decellularized and orthotopically implanted in recipient pigs (n = 12). To evaluate possible immune responses to the scaffolds and to assess potential thrombus formation, cellular allogeneic vena cava grafts were transplanted in control pigs (n = 8). Within 28 days after implantation, the decellularized veins were fully recellularized with endothelial cells and smooth muscle cells. Quantitative histological analysis showed a comparable amount of smooth muscle actin in the repopulated decellularized grafts similar to the native IVC. Lymphocyte infiltrates representing signs of graft rejection were not detected in the pigs, as opposed to the control group that received the allogeneic grafts. The decellularized grafts provoked a higher incidence of thrombosis in comparison with allogeneic grafts (33.3 vs. 12.5%). With this study, we show efficient in situ repopulation of decellularized vein grafts. These findings are insightful and promising to further explore the use of decellularized tissue without the need for full pre-transplant recellularization.Graphical abstract

Cryo-storage of porcine hides at the industrial scale for tissue engineering and regenerative medicine application

BACKGROUND: The industrial scale cryo-storage of raw tissue materials requires a robust, low-cost and easy-to-operate method that can facilitate the down-stream process. OBJECTIVE: The study was aimed to develop the multifunctional protective solutions (MPS) for transportation at ambient conditions and also subsequent cryo-storage below −20°C of raw porcine hides for tissue engineering and regenerative medicine. MATERIALS AND METHODS: Protective solutions with antimicrobial activity and proteinase-inhibiting activity were developed and tested for its efficacy in preserving the extracellular matrix of porcine dermis from microbial spoilage, proteolytic degradation, freeze damage and excessive dehydration during shipping and cryo-storage. The MPSs contained phosphate-buffered saline with ethylene diamine tetra acetic acid (EDTA) added as chelator and proteinase inhibitor, as well as glycerol or maltodextrin (M180) as cryoprotectants. RESULTS: MPSs prepared with EDTA and glycerol or M180 had significant antimicrobial activity and proteinase-inhibiting activity during the period of shipping and handling. Glycerol and M180 prevented eutectic salt precipitation and excessive freeze dehydration upon cryo-storage of porcine hides. Without glycerol or M180, hides could be freeze-dehydrated to the low hydration at ˜0.4 g/g dw, and formed irreversible plications after freezing. A critical hydration (0.8˜0.9 g/g dw) was observed for the extracellular matrix of porcine dermis, and dehydration to a lower level could impose enormous stress and potential damage. The soaking of porcine hides in MPSs decreased water content as glycerol and M180 entered into dermis. Upon equilibration, the glycerol content in the tissue was about 94% of the incubating glycerol solution, but the M180 content in the tissue was only about 50% of the incubating M180 solution, indicating that M180 did not get into the entire aqueous domain within dermis. MPSs reduced ice formation and increased the unfrozen water content of porcine raw hides upon cryo-storage. CONCLUSION: MPSs prepared with EDTA and glycerol or M180 have antimicrobial activity and proteinase-inhibiting activity, which can be used for transportation and cryo-storage of raw hides at the industrial scale. Glycerol at 7.5% w/v and M180 at 20% w/v were sufficient to prevent freeze damage and excessive freeze dehydration.

Amplifying endogenous stem cell migration for in situ bone tissue formation: Substance P analog and BMP mimetic peptide-loaded click-crosslinked hyaluronic acid hydrogel

Endogenous stem cell-driven in situ bone tissue formation has recently garnered increasing attention. Therefore, our study sought to refine methods to enhance the migration and subsequent osteogenic differentiation of these cells. Our innovative approach involves using an injectable hydrogel that combines click cross-linking sites and a BMP-2 mimetic peptide (BP) with hyaluronic acid (HA). This injectable formulation, hereinafter referred to as SPa + Cx-HA-BP, incorporates a substance P analog peptide (SPa) with Cx-HA-BP, proving versatile for in vitro and in vivo applications without cytotoxicity. The controlled release of SPa creates a gradient that guides endogenous stem cells towards the Cx-HA scaffold from specific tissue niches. Both Cx-HA and SPa+Cx-HA induced minimal changes in the expression of genes associated with osteogenic differentiation. In contrast, these genes were robustly induced by both SPa + Cx-HA+BP and SPa + Cx-HA-BP, in which BP was respectively integrated via physical and chemical methods. Remarkably, chemically incorporating BP (Cx-HA-BP) resulted in 4–9 times higher osteogenic gene expression than physically mixed BP in Cx-HA+BP. This study validates the role of SPa role in guiding endogenous stem cells toward the hydrogel and underscores the substantial impact of sustained BP presence within the hydrogel. Collectively, our findings offer valuable insights for the development of innovative strategies to promote endogenous stem cell-based tissue regeneration. The developed hydrogel effectively guides stem cells from their natural locations and facilitates sustained osteogenic differentiation, thus holding great promise for applications in regenerative medicine.

Human mesenchymal stromal cells-laden crosslinked hyaluronic acid-alginate bioink for 3D bioprinting applications in tissue engineering.

Three-dimensional (3D) bioprinting is considered one of the most advanced tools to build up materials for tissue engineering. The aim of this work was the design, development and characterization of a bioink composed of human mesenchymal stromal cells (hMSC) for extrusion through nozzles to create these 3D structures that might potentially be apply to replace the function of damaged natural tissue. In this study, we focused on the advantages and the wide potential of biocompatible biomaterials, such as hyaluronic acid and alginate for the inclusion of hMSC. The bioink was characterized for its physical (pH, osmolality, degradation, swelling, porosity, surface electrical properties, conductivity, and surface structure), mechanical (rheology and printability) and biological (viability and proliferation) properties. The developed bioink showed high porosity and high swelling capacity, while the degradation rate was dependent on the temperature. The bioink also showed negative electrical surface and appropriate rheological properties required for bioprinting. Moreover, stress-stability studies did not show any sign of physical instability. The developed bioink provided an excellent environment for the promotion of the viability and growth of hMSC cells. Our work reports the first-time study of the effect of storage temperature on the cell viability of bioinks, besides showing that our bioink promoted a high cell viability after being extruded by the bioprinter. These results support the suggestion that the developed hMSC-composed bioink fulfills all the requirements for tissue engineering and can be proposed as a biological tool with potential applications in regenerative medicine and tissue engineering.

Self-Assembling Triple-Helix Recombinant Collagen Hydrogel Enriched with Tyrosine.

The self-assembly of collagen within the human body creates a complex 3D fibrous network, providing structural integrity and mechanical strength to connective tissues. Recombinant collagen plays a pivotal role in the realm of biomimetic natural collagen. However, almost all of the reported recombinant collagens lack the capability of self-assembly, severely hindering their application in tissue engineering and regenerative medicine. Herein, we have for the first time constructed a series of self-assembling tyrosine-rich triple helix recombinant collagens, mimicking the structure and functionality of natural collagen. The recombinant collagen consists of a central triple-helical domain characterized by the (Gly-Xaa-Yaa)n sequence, along with N-terminal and C-terminal domains featuring the GYY sequence. The introduction of GYY has a negligible impact on the stability of the triple-helical structure of recombinant collagen while simultaneously promoting its self-assembly into fibers. In the presence of [Ru(bpy)3]Cl2 and APS as catalysts, tyrosine residues in the recombinant collagen undergo covalent cross-linking, resulting in a hydrogel with exceptional mechanical properties. The recombinant collagen hydrogel exhibits outstanding biocompatibility and bioactivity, significantly enhancing the proliferation, adhesion, migration, and differentiation of HFF-1 cells. This innovative self-assembled triple-helix recombinant collagen demonstrates significant potential in the fields of tissue engineering and medical materials.

Exploring Present and Future Directions in Nano-Enhanced Optoelectronic Neuromodulation.

ConspectusElectrical neuromodulation has achieved significant translational advancements, including the development of deep brain stimulators for managing neural disorders and vagus nerve stimulators for seizure treatment. Optoelectronics, in contrast to wired electrical systems, offers the leadless feature that guides multisite and high spatiotemporal neural system targeting, ensuring high specificity and precision in translational therapies known as "photoelectroceuticals". This Account provides a concise overview of developments in novel optoelectronic nanomaterials that are engineered through innovative molecular, chemical, and nanostructure designs to facilitate neural interfacing with high efficiency and minimally invasive implantation.This Account outlines the progress made both within our laboratory and across the broader scientific community, with particular attention to implications in materials innovation strategies, studying bioelectrical activation with spatiotemporal methods, and applications in regenerative medicine. In materials innovation, we highlight a nongenetic, biocompatible, and minimally invasive approach for neuromodulation that spans various length scales, from single neurons to nerve tissues using nanosized particles and monolithic membranes. Furthermore, our discussion exposes the critical unresolved questions in the field, including mechanisms of interaction at the nanobio interface, the precision of cellular or tissue targeting, and integration into existing neural networks with high spatiotemporal modulation. In addition, we present the challenges and pressing needs for long-term stability and biocompatibility, scalability for clinical applications, and the development of noninvasive monitoring and control systems.In addressing the existing challenges in the field of nanobio interfaces, particularly for neural applications, we envisage promising strategic directions that could significantly advance this burgeoning domain. This involves a deeper theoretical understanding of nanobiointerfaces, where simulations and experimental validations on how nanomaterials interact spatiotemporally with biological systems are crucial. The development of more durable materials is vital for prolonged applications in dynamic neural interfaces, and the ability to manipulate neural activity with high specificity and spatial resolution, paves the way for targeting individual neurons or specific neural circuits. Additionally, integrating these interfaces with advanced control systems, possibly leveraging artificial intelligence and machine learning algorithms and programming dynamically responsive materials designs, could significantly ease the implementation of stimulation and recording. These innovations hold the potential to introduce novel treatment modalities for a wide range of neurological and systemic disorders.

Comprehensive characterisation and cryopreservation optimisation of buffalo (Bubalus bubalis) adipose tissue-derived mesenchymal stem cells

Over half of the world's buffalo (Bubalus bubalis) inhabit India, and buffaloes frequently encounter health challenges that resist conventional treatments, prompting the exploration of alternative therapeutic strategies. One promising approach is stem cell therapy, particularly multipotent mesenchymal/stromal stem cells (MSCs). These cells have shown significant efficacy in addressing various diseases in livestock that exhibit resistance to conventional therapies. Adipose tissue-derived MSCs (ADSCs) have garnered attention due to their accessibility and robust expansion potential. The current study comprehensively characterises buffalo ADSCs (bADSCs), confirming their identity as MSCs capable of differentiating into diverse cell lineages—the identified characteristics position bADSCs as promising candidates for applications in regenerative medicine, applicable in veterinary contexts. Notably, the study established that a cryoprotective solution comprising 10 % dimethyl sulfoxide and 90 % fetal bovine serum is optimal for preserving bADSCs. This cryoprotective solution maintains vital parameters, including viability, apoptosis, senescence, cell adherence, adherent cell viability, metabolic and clonogenic efficiency, and the activity of reactive oxygen species and trilineage differentiation potential following thawing. These findings lay the foundation for developing a cryo-banking system for bADSCs. Subsequent research efforts are focused on exploring the therapeutic potential of bADSCs in specific disease models and clinical settings. The outcomes of such investigations may pave the way for innovative and effective treatments, further enhancing our understanding of the regenerative capabilities of bADSCs.

Development of porous hydroxyapatite/PVA/gelatin/alginate hybrid flexible scaffolds with improved mechanical properties for bone tissue engineering

Porous scaffolds based on gelatin and hydroxyapatite particles (HAp) are promising for applications in bone tissue engineering and regenerative medicine. However, they are not ideal for stress and shock protection due to low compressive strength, brittleness, and poor toughness. Herein, we developed porous hybrid scaffolds by combining multiple components into a single bone scaffold, including hydroxyapatite nanoparticles (HAp NPs), alginate, polyvinyl alcohol (PVA), and gelatin with different gelatin/PVA composition ratios. HAp NPs were synthesized in situ in PVA solution and scaffolds were fabricated using a freeze-drying method. The results showed good physicochemical properties of the scaffolds: formation of pure HAp NPs phase, high porosity, large pore sizes, large swelling capacity depending on varying gelatin/PVA ratios, as well as a long-term degradation rate up to 28 days. The porous scaffolds exhibited compressive strength close to the cancellous bone with stress-strain behavior exhibiting three-stage flexible behavior indicating improved fracture resistance with an energy absorbing capability up to 1.9 MJ/m3. The scaffolds have a yield strength of 70–403 kPa, a compressive strength at 65 % strain of 0.96–1.80 MPa, a nonporous elastic modulus of 10.44–12.40 GPa, and a densification strain of approximately 0.92 %. This work develops three-dimensional (3D) porous hybrid bone scaffolds with mechanical strength within the minimum compressive strength of cancellous bone and mechanical energy absorption capacity favor for cancellous bone repair, bone tissue engineering, and regenerative medicine applications.

Computational Studies Using COMSOL Multiphysics on Dielectrophoretic Characterization and Separation of Mesenchymal Stem Cells Undergoing Tenogenic Differentiation

Stem cells have unique self-renewal and differentiation capacities, which are advantageous for regenerative medicine and tissue engineering applications. Stem cells can recreate cells needed to repair injured tissues and organs in the body, for example, by regenerating connective tissues, which provide support, protection, and a structural framework for various organs and other tissues. However, a significant limitation is the susceptibility of tendons to injury with long-term loss of function. Mesenchymal Stem Cell (MSC) therapies are promising for healing tendon injuries and tears but controlling stem cell differentiation and generating homogeneous tenogenic stem cell populations remains challenging. Therefore, separating differentiating and non-differentiating MSCs to collect homogenous cells is critical for further tissue repair after transplantation. To substantiate our hypothesis that differentiating and non-differentiating stem cells have unique dielectric properties, we focused on characterizing-obtaining dielectric signatures for differentiating tenogenic and non-tenogenic MSCs on the third day of differentiation using dielectrophoresis (DEP) focused on crossover frequencies [1, 2]. The estimated values for membrane capacitance, conductivity, and permittivity after 3-day treatment of undifferentiated cells differentiating into tenogenic MSCs are 2.46±0.1 pF, 0.82±0.01 S/m, and 1.97±0.05, respectively, and cytoplasm conductivity 0.82±0.02 S/m. The treated stem cells undergo differentiation, but the untreated cells don’t. The results showed a significant difference between the 3-day treatment and no-treatment group. The estimated properties from the dielectrophoretic characterization studies are crucial in designing a DEP-based enrichment microdevice to collect homogeneous differentiated stem cell populations for tendon repair. Using particle tracing, creeping flow (transport of diluted species model), and electric current physics in COMSOL Multiphysics simulation software, we designed a microdevice that achieves 100% separation of untreated and treated mesenchymal stem cells undergoing differentiation towards tendon. Applying dielectrophoresis to the microdevice architecture demonstrated high selectivity and yield, processing one million treated and untreated stem cells in 6.4 and 27.8 hours, respectively.

Application of pulsed electric field technology to skin engineering.

Tissue engineering encompasses a range of techniques that direct the growth of cells into a living tissue construct for regenerative medicine applications, disease models, drug discovery, and safety testing. These techniques have been implemented to alleviate the clinical burdens of impaired healing of skin, bone, and other tissues. Construct development requires the integration of tissue-specific cells and/or an extracellular matrix-mimicking biomaterial for structural support. Production of such constructs is generally expensive and environmentally costly, thus eco-sustainable approaches should be explored. Pulsed electric field (PEF) technology is a nonthermal physical processing method commonly used in food production and biomedical applications. In this review, the key principles of PEF and the application of PEF technology for skin engineering will be discussed, with an emphasis on how PEF can be applied to skin cells to modify their behaviour, and to biomaterials to assist in their isolation or sterilisation, or to modify their physical properties. The findings indicate that the success of PEF in tissue engineering will be reliant on systematic evaluation of key parameters, such as electric field strength, and their impact on different skin cell and biomaterial types. Linking tangible input parameters to biological responses critical to healing will assist with the development of PEF as a sustainable tool for skin repair and other tissue engineering applications.

The Combination of Bioactive Herbal Compounds with Biomaterials for Regenerative Medicine.

Regenerative medicine aims to restore the function of diseased or damaged tissues and organs by cell therapy, gene therapy, and tissue engineering, along with the adjunctive application of bioactive molecules. Traditional bioactive molecules, such as growth factors and cytokines, have shown great potential in the regulation of cellular and tissue behavior, but have the disadvantages of limited source, high cost, short half-life, and side effects. In recent years, herbal compounds extracted from natural plants/herbs have gained increasing attention. This is not only because herbal compounds are easily obtained, inexpensive, mostly safe, and reliable, but also owing to their excellent effects, including anti-inflammatory, antibacterial, antioxidative, proangiogenic behavior and ability to promote stem cell differentiation. Such effects also play important roles in the processes related to tissue regeneration. Furthermore, the moieties of the herbal compounds can form physical or chemical bonds with the scaffolds, which contributes to improved mechanical strength and stability of the scaffolds. Thus, the incorporation of herbal compounds as bioactive molecules in biomaterials is a promising direction for future regenerative medicine applications. Herein, an overview on the use of bioactive herbal compounds combined with different biomaterial scaffolds for regenerative medicine application is presented. We first introduce the classification, structures, and properties of different herbal bioactive components and then provide a comprehensive survey on the use of bioactive herbal compounds to engineer scaffolds for tissue repair/regeneration of skin, cartilage, bone, neural, and heart tissues. Finally, we highlight the challenges and prospects for the future development of herbal scaffolds toward clinical translation. Overall, it is believed that the combination of bioactive herbal compounds with biomaterials could be a promising perspective for the next generation of regenerative medicine.

Comprehensive assessment of goat adiposetissue-derived mesenchymal stem cells cultured in different media.

Mesenchymal stem cells (MSCs) have demonstrated potential in treating livestock diseases that are unresponsive to conventional therapies. MSCs derived from goats, a valuable model for studying orthopaedic disorders in humans, offer insights into bone formation and regeneration. Adipose tissue-derived MSCs (ADSCs) are easily accessible and have a high capacity for expansion. Although the choice of culture media significantly influences the biological properties of MSCs, the optimal media for goat ADSCs (gADSCs) remains unclear. This study aimed to assess the effects of four commonly used culture media on gADSCs' culture characteristics, stem cell-specific immunophenotype, and differentiation. Results showed that MEM, DMEM/F12, and DMEM-LG were superior in maintaining cell morphology and culture parameters of gADSCs, such as cell adherence, metabolic activity, colony-forming potential, and population doubling. Conversely, DMEM-HG exhibited poor performance across all evaluated parameters. The gADSCs cultured in DMEM/F12 showed enhanced early proliferation and lower apoptosis. The cell surface marker distribution exhibited superior characteristics in gADSCs cultured in MEM and DMEM/F12. In contrast, the distribution was inferior in gADSCs cultured in DMEM-LG. DMEM/F12 and DMEM-LG culture media demonstrated a significantly higher potential for chondrogenic differentiation and DMEM-LG for osteogenic differentiation. In conclusion, DMEM/F12 is a suitable culture medium for propagating gADSCs as it effectively maintains cell morphology, growth parameters, proliferation and lower apoptosis while exhibiting desirable expression patterns of MSC-specific markers. These findings contribute to optimising culture conditions for gADSCs, enhancing their potential applications in disease treatment and regenerative medicine.

Chitosan-PNIPAM Thermogel Associated with Hydrogel Microspheres as a Smart Formulation for MSC Injection.

Regenerative medicine based on cell therapy has emerged as a promising approach for the treatment of various medical conditions. However, the success of cell therapy heavily relies on the development of suitable injectable hydrogels that can encapsulate cells and provide a conducive environment for their survival, proliferation, and tissue regeneration. Herein, we address the medical need for cyto- and biocompatible injectable hydrogels by reporting on the synthesis of a hydrogel-forming thermosensitive copolymer. The copolymer was synthesized by grafting poly(N-isopropylacrylamide-co-carboxymethyl acrylate) (PNIPAM-COOH) onto chitosan through amide coupling. This chemical modification resulted in the formation of hydrogels that exhibit a sol-gel transition with an onset at approximately 27 °C, making them ideal for use in injectable applications. The hydrogels supported the survival and proliferation of cells for several days, which is critical for cell encapsulation. Furthermore, the study evaluates the addition of collagen/chitosan hybrid microspheres to support the adhesion of mesenchymal stem cells within the hydrogels. Altogether, these results demonstrate the potential of the PNIPAM-chitosan thermogel for cell encapsulation and its possible applications in regenerative medicine.

Synergistic large segmental bone repair by 3D printed bionic scaffolds and engineered ADSC nanovesicles: Towards an optimized regenerative microenvironment

Achieving sufficient bone regeneration in large segmental defects is challenging, with the structure of bone repair scaffolds and their loaded bioactive substances crucial for modulating the local osteogenic microenvironment. This study utilized digital laser processing (DLP)-based 3D printing technology to successfully fabricate high-precision methacryloylated polycaprolactone (PCLMA) bionic bone scaffold structures. Adipose-derived stem cell-engineered nanovesicles (ADSC-ENs) were uniformly and stably modified onto the bionic scaffold surface using a perfusion device, constructing a conducive microenvironment for tissue regeneration and long bone defect repair through the scaffold's structural design and the vesicles' biological functions. Scanning electron microscopy (SEM) examination of the scaffold surface confirmed the efficient loading of ADSC-ENs. The material group loaded with vesicles (PCLMA-BAS-ENs) demonstrated good cell compatibility and osteogenic potential when analyzed for the adhesion and osteogenesis of primary rabbit bone marrow mesenchymal stem cells (BMSCs) on the material surface. Tested in a 15 mm critical rabbit radial defect model, the PCLMA-BAS-ENs scaffold facilitated near-complete bone defect repair after 12 weeks. Immunofluorescence and proteomic results indicated that the PCLMA-BAS-ENs scaffold significantly improved the osteogenic microenvironment at the defect site in vivo, promoted angiogenesis, and enhanced the polarization of macrophages towards M2 phenotype, and facilitated the recruitment of BMSCs. Thus, the PCLMA-BAS-ENs scaffold was proven to significantly promote the repair of large segmental bone defects. Overall, this strategy of combining engineered vesicles with highly biomimetic scaffolds to promote large-segment bone tissue regeneration holds great potential in orthopedic and other regenerative medicine applications.