Cell Delivery Vehicle Research Articles

Nanowires and Electrical Stimulation Synergistically Improve Functions of hiPSC Cardiac Spheroids.

The advancement of human induced pluripotent stem-cell-derived cardiomyocyte (hiPSC-CM) technology has shown promising potential to provide a patient-specific, regenerative cell therapy strategy to treat cardiovascular disease. Despite the progress, the unspecific, underdeveloped phenotype of hiPSC-CMs has shown arrhythmogenic risk and limited functional improvements after transplantation. To address this, tissue engineering strategies have utilized both exogenous and endogenous stimuli to accelerate the development of hiPSC-CMs. Exogenous electrical stimulation provides a biomimetic pacemaker-like stimuli that has been shown to advance the electrical properties of tissue engineered cardiac constructs. Recently, we demonstrated that the incorporation of electrically conductive silicon nanowires to hiPSC cardiac spheroids led to advanced structural and functional development of hiPSC-CMs by improving the endogenous electrical microenvironment. Here, we reasoned that the enhanced endogenous electrical microenvironment of nanowired hiPSC cardiac spheroids would synergize with exogenous electrical stimulation to further advance the functional development of nanowired hiPSC cardiac spheroids. For the first time, we report that the combination of nanowires and electrical stimulation enhanced cell-cell junction formation, improved development of contractile machinery, and led to a significant decrease in the spontaneous beat rate of hiPSC cardiac spheroids. The advancements made here address critical challenges for the use of hiPSC-CMs in cardiac developmental and translational research and provide an advanced cell delivery vehicle for the next generation of cardiac repair.

A novel gelatin-based micro-cavitary hydrogel for potential application in delivery of anchorage dependent cells: A study with vasculogenesis model

Hydrogels have been widely regarded as promising tissue engineering scaffolds and cell delivery vehicles, however, their inherent submicron- or nano-scale polymer networks severely inhibit the settlement of anchorage dependent cells (ADCs). Here, using endothelial progenitor outgrowth cells (EPOCs) as the typical ADCs, a gelatin-based micro-cavitary gel (namely Gel-MCG) is developed with gelatin-methacrylate and gelatin microspheres as precursor and porogens, respectively, to promote cellular focal adhesion and functions. The introduction of micro-cavitary structures within the Gel-MCG improves its physical properties as well as creates numerous gel-microcavity interfaces within gel-based matrices. Compared with conventional gelatin gel (Gel-G) scaffold, the Gel-MCG provides more suitable microenvironments for EPOCs' attachment, spreading, and proliferation, and then which leads to enhanced endothelial differentiation and vascularization as demonstrated by higher expressions of endothelial markers. The Gel-MCG system shows great potential as vehicle for the delivery of ADCs in tissue engineering.

In-vitro Seeding of Bladder Smooth Muscle Cells onto Bladder Acellular Matrix as Cell Delivery Vehicle in Tissue Engineering

In-vitro Seeding of Bladder Smooth Muscle Cells onto Bladder Acellular Matrix as Cell Delivery Vehicle in Tissue Engineering

Development of size-customized hepatocarcinoma spheroids as a potential drug testing platform using a sacrificial gelatin microsphere system

Sacrificial gelatin microspheres can be developed as a cell delivery vehicle for non-anchorage dependent cells – its incorporation into a macroscopic scaffold system not only allows the cells to be cultured in suspension within cavities left behind by the sacrificial material, it also allows scaffold-free tissue development to be confined within the cavities. In this study, dense and highly viable hepatocarcinoma spheroids were developed by means of encapsulation in sacrificial gelatin microspheres produced via a simple water-in-oil emulsion technique. By initial selection of microsphere size and distribution, spheroid size can be controlled for various applications such as uniform tumor spheroids as a reproducible three-dimensional drug screening and testing platform that better mimics the in vivo nature of tumors (instead of conventional monolayer culture), as this study has suggested as a proof-of-concept with chemotherapy drug Doxorubicin.

A novel platelet lysate hydrogel for endothelial cell and mesenchymal stem cell-directed neovascularization

Mesenchymal stem cells (MSC) hold promise in promoting vascular regeneration of ischemic tissue in conditions like critical limb ischemia of the leg. However, this approach has been limited in part by poor cell retention and survival after delivery. New biomaterials offer an opportunity to localize cells to the desired tissue after delivery, but also to improve cell survival after delivery. Here we characterize the mechanical and microstructural properties of a novel hydrogel composed of pooled human platelet lysate (PL) and test its ability to promote MSC angiogenic activity using clinically relevant in vitro and in vivo models. This PL hydrogel had comparable storage and loss modulus and behaved as a viscoelastic solid similar to fibrin hydrogels despite having 1/4–1/10th the fibrin content of standard fibrin gels. Additionally, PL hydrogels enabled sustained release of endogenous PDGF-BB for up to 20days and were resistant to protease degradation. PL hydrogel stimulated pro-angiogenic activity by promoting human MSC growth and invasion in a 3D environment, and enhancing endothelial cell sprouting alone and in co-culture with MSCs. When delivered in vivo, the combination of PL and human MSCs improved local tissue perfusion after 8days compared to controls when assessed with laser Doppler perfusion imaging in a murine model of hind limb ischemia. These results support the use of a PL hydrogel as a scaffold for MSC delivery to promote vascular regeneration. Statement of SignificanceInnovative strategies for improved retention and viability of mesenchymal stem cells (MSCs) are needed for cellular therapies. Human platelet lysate is a potent serum supplement that improves the expansion of MSCs. Here we characterize our novel PL hydrogel’s desirable structural and biologic properties for human MSCs and endothelial cells. PL hydrogel can localize cells for retention in the desired tissue, improves cell viability, and augments MSCs’ angiogenic activity. As a result of these unique traits, PL hydrogel is ideally suited to serve as a cell delivery vehicle for MSCs injected into ischemic tissues to promote vascular regeneration, as demonstrated here in a murine model of hindlimb ischemia.

Evaluation of the donor cell contribution in rhBMP-2 mediated bone formation with chitosan thermogels using fluorescent protein reporter mice.

Our current understanding regarding the contribution of donor cells in growth factor and cell based tissue regeneration strategies is limited. The present study attempts to utilize fluorescent protein reporter mice [Col3.6Topaz (enhanced yellow fluorescent protein, EYFP) as host and Col3.6Cyan (enhanced cyan fluorescent protein, ECFP) as donor] to determine donor cell contribution in bone regeneration using a bilateral calvarial defect model. Thermogelling chitosan hydrogels (Chi-AHP) were used as bone marrow stromal cell (BMSC) delivery vehicle in the presence and absence of recombinant human bone morphogenetic protein-2 (rhBMP-2). Co-delivery of rhBMP-2 and donor BMSCs led to a significant increase in bone formation indicating the potential of using the combination approach for improved regeneration. On a cellular level, presence of rhBMP-2 resulted in an increased host cell derived osteoblast infiltration at 8 weeks. However, the new mineralized tissue presented two distinct morphological features based on its cellular origin. Regenerated tissue associated with ECFP positive donor cells showed a woven bone-like structure with diffuse alizarin complexone (AC) label and minimal tartrate resistant acid phosphatase (TRAP) activity, indicating the presence of immature early osteoblast cells depositing mineralized tissue without progressing to a mature lamellar bone. Host derived bone showed sharp mineralization line (AC), strong alkaline phosphatase (ALP) and TRAP activity, indicating the presence of actively mineralizing and remodeling, mature lamellar bone matrix. The study demonstrated the remarkable potential of transgenic reporters to improve our understanding of donor cell contribution during bone formation.

Injectable surgical sealant for controlled cell delivery and fusion of cartilage-to-cartilage interfaces

Event Abstract Back to Event Injectable surgical sealant for controlled cell delivery and fusion of cartilage-to-cartilage interfaces Ashkan H. Tehrani1, 2, Sumrita Bhat1, 2, Neil A. Duncan2, 3 and Arindom Sen1, 2 1 University of Calgary, Pharmaceutical Production Research Facility, Department of Chemical and Petroleum Engineering, Canada 2 University of Calgary, McCaig Institute for Bone and Joint Health, Canada 3 University of Calgary, Department of Civil Engineering, Schulich School of Engineering, Canada Introduction: Since the development of the first generation of cell-based therapies, autologous chondrocyte implantation (ACI) and mesenchymal stem cell transplantation have been used in clinical trials for joint repair[1]. However, a major challenge is the difficulty of the surgical procedure to deliver cells to the site of injury. In ACI, for example, surgeons utilize a periosteal flap to secure the cell-implant, and a high cell density to make up for hypertrophy[2]. During the culture, dedifferentiation and loss of phenotype constitute an added complexity. In this study, we introduce a novel injectable system that (i) serves as a safe vehicle for controlled cell delivery, (ii) possesses a degree of adhesiveness to confine cell to the defect site at the time of implantation, and (iii) exhibits a tunable mechanical environment to drive the commitment of the cells toward the desired lineage. This system must (i) be injectable, (ii) adhere to the chondral interface, and (iii) sustain cell release over time without compromising the level of repair. Materials and Methods: The proposed hydrogel system consisted of a crosslinked and a non-crosslinked domain. The proportion of the former impacts the mechanical environment in which the cells reside, the injectability of the system and the degree of adhesiveness, while the latter determines cell release rate. A 3% alginate/polyethylene glycol solution (1:3) was prepared in PBS at 60°C overnight followed by internal gelation during vortex with 1% calcium solution at room temperature. Porcine chondrocytes were mixed at a density of 1×106 cell/mL with the gel, and injected in a 3 mm circular defect model in a porcine patella in vitro. External gelation took place in a bath of 5% calcium solution immediately after the injection. Samples were incubated at 37°C with PPRF-MSC6 medium[3], in 5% CO2 until the completion of the delivery phase. Results and Discussion: The results showed that the gel demonstrated a viscous flow behavior at the time of injection to effectively penetrate throughout the cylindrical defect models and fully embed the implantation site, then demonstrated solid-like properties in situ post external gelation with adequate mechanical stability to not flow from the defect vicinity (Fig 1). Also, the gel was able to completely fill the focal defect model, fuse with the surrounding cartilage tissue (Fig 2a), and seal the implantation site flush to the articular surface (Fig 2). There was no need for additional fixation such as patches since the sealant demonstrated ample resistance to peeling off from the superficial layer. Figure 3 shows how the cells were released at a controlled rate due to the spontaneous discharging of the non-crosslinked domain which resulted in the concomitant breakdown of the matrix by day 7. As the release rate increased with time, the gel strength decreased and cell migration accelerated (Fig 3). Conclusion: This research introduced an injectable sealant, which also served as a cell delivery vehicle and an adhesive to treat chondral defects of different size and shape. By design, it represented a multifunctional mimicry of the chondrocytes niche in cartilage since it encompassed cells in a three-dimensional matrix, under a hypoxic condition and with a relatively similar mechanical environment to that of native cartilage. From a surgical viewpoint, the system can be cured in situ within seconds and demonstrated a reliable and a cost-effective additional step that can benefit various cell-based therapies. Alberta Innovates: Health Solutions; Natural Sciences and Engineering Research Council of Canada (NSERC)

Enzymatic control of chitosan gelation for cell delivery vehicle

Event Abstract Back to Event Enzymatic control of chitosan gelation for cell delivery vehicle Carla Tim1, Cintia C. Martignago2, Lia M. Neves3, Nivaldo A. Parizotto2 and Ana Claudia Rennó1 1 Federal University of São Paulo, Bioscience, Brazil 2 Federal University of São Carlos, Physiotherapy, Brazil 3 Federal University of São Carlos, Physiologic, Brazil Osteoarthritis (OA) is a common chronic disease among older people, which leads to a loss of articular cartilage. Due to the very limited cartilage regenerative capacity and consequently the limited efficacy of the standard treatments, the investigation of strategic innovative approaches to prevent the development of the clinical condition of OA is of great interest. Thus, development in strategies for cartilage repair have increasingly focused on cell-assisted repair, proposing to use autologous chondrocytes or other cell types to regenerate articular cartilage in situ. One promising cell delivery vehicle is chitosan hydrogels, mainly due to its several advantages including easy handling, high cell seeding efficiency and good formability to irregular defects in a minimally invasive manner. Chitosan hydrogels can be formed homogeneously by neutralization of chitosan solutions using ammonia produced in situ from enzymatic hydrolysis of urea. Therefore, this study aimed to investigate the possibility to control chitosan gelation using enzymatic hydrolysis of urea for the delivery of chodrocytes. Chitosan (Sigma–Aldrich) was dissolved at a concentration of 2.5 w/v% in 0.100M HCl. Urease (Sigma–Aldrich) was dissolved in phosphate buffered saline and Urea (Sigma–Aldrich) was dissolved in MilliQ water. To prepare 1.5 w/v% chitosan hydrogels, 1mL of chitosan was mixed with 0.67mL urease. Subsequently, 10 ul of the urea solution was added. The vials were inverted every 30 sec and the gelation time was recorded as the one at which the solution did not flow in 5 different conditions, i.e., urease concentration of 30 U/mL and urea concentrations of 7.5, 8.5, 9.5,10.5 and11.5 M. The speed at which the pH changed under different gel compositions was investigated and recorded with the use of a pHmeter electrode placed in a freshly prepared mixture of starting solutions immediately after addition of the urea. The results revealed that the gelation time of the chitosan hydrogels can be precisely controlled by varying the urea concentrations as shown in table 1. The pH during gelation of the chitosan hydrogels is displayed, which confirmed that the pH increased from initial values of about 5.7–6.8 to final values of 6.9–7.3 within 5–28 min after addition of urea. Final pH values were controlled by the concentration of urea as main source for alkaline ammonia groups, but did not depend on the concentration of urease, which allows for independent control over final pH and gelation kinetics by varying the concentrations of urea and urease, respectively. Thus, it is possible to set the desired gelation time and final pH of the gel for various applications. The results demonstrated that physically crosslinked chitosan hydrogels for biomedical purpose can be easily fabricated via enzymatic control, and such control over the gelation of chitosan hydrogels could be an option for the delivery of chondrocytes via an injectable system. We would like to acknowledge the contributions of Brazilian funding agency Fapesp (FAPESP project#2014/13702-6) for the financial support of this research.

Coaxial electrospray for cell encapsulation and delivery in cell-based tissue engineering

Event Abstract Back to Event Coaxial electrospray for cell encapsulation and delivery in cell-based tissue engineering Yu Zhou1*, Qilong Zhao1* and Min Wang1 1 The University of Hong Kong, Mechanical Engineering, Hong Kong, SAR China Introduction: For human body tissue regeneration, different approaches, including scaffold-based[1], growth factor-based[2] and cell-based tissue engineering[3], are explored. In cell-based tissue engineering, developing appropriate vehicles for cell encapsulation and delivery is highly important. The process of cell encapsulation must be cell-friendly for achieving high cell viability. Moreover, for microspherical delivery vehicles, the diameter of cell-encapsulated microspheres is a critical factor. The diameter should not be too small or too large. Degradation properties of polymers for microspheres are another important factor for cell delivery. Coaxial electrospray can produce core-shell structured microspheres and hence was investigated in this study for the encapsulation and delivery of cells. Materials and Methods: To determine suitable electrospray parameters, solid, cell-free microspheres were made first by electrospray using polymer solutions of sodium alginate (SA), poly(vinyl alcohol), gelatin and mixed solutions (1:1) of two polymers. Then, cell-free microspheres with core-shell structure were produced by coaxial electrospray where a polymer solution was fed into outer tube and PBS was fed into inner tube of the coaxial nozzle. After process optimization, PBS was replaced by aqueous cell suspensions of human umbilical vein endothelial cells (HUVECs) in coaxial electrospray for making cell-encapsulated microspheres. The morphology and structure of cell-free and cell-containing microspheres were studied. The viability of HUVECs was studied using live and dead assay. Triggered cell release was also investigated. Results and Discussion: Electrospray experiments showed that relatively large diameter (~5 micron) and uniform microspheres could be produced using mixed polymer solution of SA and gelatin. Therefore, coaxial cell electrospray was conducted using HUVEC cell suspensions and SA/gelatin polymer solution or SA solution alone, and a post-spray cross-linking treatment was added for the polymer shell. After further explorations, electrospray parameters (applied voltage, feeding rate ratio between the outer tube and inner tube, etc.) were fixed for fabricating cell-encapsulated microspheres. HUVECs with low cell density (5×106 cells/mL) and high cell density (1×107 cells/mL) were used for encapsulation. Fig.1 shows cell-encapsulated microspheres made by coaxial electrospray and post-spray crosslinking using different cell densities. Microspheres with regular spherical shape could be produced for both low cell density and high cell density. A higher cell loading efficiency was attained for microspheres using a higher cell density. Triggered cell release was achieved by treating cell-encapsulated microspheres with a 0.05M sodium citrate solution. The polymer shell of microspheres was dissolved gradually and after 10 minutes, polymer shell disappeared and cells were totally released (Fig.2). Conclusion: Coaxial electrospray proved to be a useful and adaptable technique for making cell delivery vehicles. With process optimization, sufficient cells could be encapsulated in core-shell structured microspheres via coaxial electrospray. Cells contained in the core of microspheres were well protected and showed high viability after the electrospray process. Triggered release could be realized for delivering live cells, enabling cell-based therapies. This work was supported by Hong Kong Research Grants Council through a GRF Grant (HKU 717713E).

Controlled nitric oxide release for tissue repair and regeneration

The discovery that nitric oxide (NO) plays crucial roles in mammalian systems has spurred considerable interest in its translational application. However, the major problem with its application in clinical settings is the high reactivity of NO, which makes it difficult to supply NO with spatiotemporal accuracy while maintaining its bioactivity. To deliver NO at a specified rate to a preferred location for a precise period of time is highly demanded. The significant technological advancements in controlled release make it possible to control NO release at the target site with a sustained release rate and an optimum concentration, and then provide a longer therapeutic duration of NO. Moreover, biomaterials designed for regenerative applications provide carriers with support structures for controlled NO releasing. Meanwhile, engineered matrices with controlled NO release have been used as vehicles for stem cell delivery to enhance cell engraftment and further to improve therapeutic effects of transplanted cells. In this review, we will provide an overview of controlled NO release materials, as well as the mechanisms of their treatment effects in tissue-repairing processes. Furthermore, the application of on-demand NO-releasing materials as carriers for stem cell transplantation and the NO-based therapeutic applications in translational medicine will be discussed.

Porous decellularized extracellular matrix microcarriers for tissue-specific cell expansion and delivery

Event Abstract Back to Event Porous decellularized extracellular matrix microcarriers for tissue-specific cell expansion and delivery Claire C. Yu1, Anna A. Kornmuller2 and Lauren Flynn3 1 Queen's University, Chemical Engineering, Canada 2 Western University, Chemical and Biochemical Engineering, Canada 3 Western University, Chemical and Biochemical Engineering & Anatomy and Cell Biology, Canada Introduction: The use of decellularized tissues in regenerative medicine is a promising approach due to the preservation of complex constituents and architecture of the native extracellular matrix (ECM). The tissue-specific ECM is crucial in regulating cellular behaviour and harnessing this innate function may be advantageous in strategies to direct cell proliferation, differentiation, and migration[1]. In this work, decellularized adipose tissue, dermis, or myocardium was used to fabricate non-chemically crosslinked porous microcarriers as cell expansion and injectable cell delivery vehicles for tissue-specific soft connective tissue regeneration. Materials and Methods: Decellularized human adipose tissue (DAT), porcine dermis (DDT), and porcine left ventricular myocardium (DLV) were prepared following established protocols (REB# CHEM-002-07)[2]-[4]. The tissue was enzymatically digested with α-amylase and homogenized in 0.2 M acetic acid to generate an ECM suspension. The microcarriers were fabricated by electrospraying the suspension (15, 25, and 35 mg/mL) into liquid nitrogen, followed by lyophilisation[2]. The size distribution was assessed, and the ECM composition and microarchitecture were characterized using IHC (collagen I, collagen IV, laminin, and fibronectin) and SEM. To assess the potential of the ECM-derived microcarriers as a tissue-specific cell culture and delivery platform, human adipose-derived stem cells (ASCs) were seeded onto the DAT microcarriers and dynamically cultured within a CELLSPIN system. Cell attachment, proliferation, and infiltration were monitored using Live/Dead® staining, Guava ViaCount Assay, and DAPI staining over 1 month in comparison to commercial Cultispher-S microcarriers, and the ASC immunophenotype was confirmed by flow cytometry. Viability of the ASCs seeded on the DAT microcarriers was also assessed following injection through a needle and after cryo-preservation. Results and Discussion: The DAT, DDT, and DLV microcarriers were structurally robust in culture over 1 month without the need for chemical crosslinking. SEM revealed a porous spherical morphology. IHC confirmed the preservation of collagen I and IV, fibronectin, and laminin in the DDT microcarriers, while only collagen I and IV were detected in the DAT and DLV microcarriers. Culture studies revealed enhanced ASC attachment and proliferation on the DAT microcarriers as compared to Cultispher-S microcarriers, with cell infiltration observed at 3 and 4 weeks. The injectability and ability to freeze the cell-seeded microcarriers may be advantageous in the translation of the technology as a minimally-invasive cell delivery platform for soft connective tissue regeneration. Conclusion: ECM-derived microcarriers hold promise as a clinically translatable strategy to regenerate a wide range of soft tissues and may be applied to advance current knowledge on how the tissue-specific ECM microenvironment influences cell behaviour. Funding for this study was provided by the Natural Sciences and Engineering Research Council (NSERC) of Canada and the Canadian Institutes of Health Research (CIHR). The authors would like to thank Dr. B.G. Amsden for access to his high-voltage DC apparatus and Dr. J.F. Watkins and Mrs. K. Martin for their clinical collaborations.

Integrin specificity as a novel strategy for enhancing transplanted stem cell survival and tissue repair in vivo

Event Abstract Back to Event Integrin specificity as a novel strategy for enhancing transplanted stem cell survival and tissue repair in vivo Amy Clark1, 2* and Andres J. Garcia1, 2 1 Georgia Institute of Technology, Woodruff School of Mechanical Engineering, United States 2 Georgia Institute of Technology, Petit Institute for Bioengineering and Bioscience, United States Introduction: Despite the promising clinical results for the use of human mesenchymal stem cells (hMSC) in musculoskeletal defect treatment, inadequate control over cell survival, engraftment and fate limits the success of this cell-based therapy. Integrin-mediated cell adhesion plays a central role in tissue formation, maintenance, and repair by providing anchorage forces and triggering signals that regulate cell function. We hypothesize biomaterials presenting integrin-specific adhesive motifs will direct hMSC signaling and specification. The objective of this project is to engineer bioartificial hydrogels presenting integrin-specific ligands as biomimetic cell delivery vehicles for enhanced in vivo engraftment and function – an innovative strategy as it focuses on engineering specificity to integrin receptors to promote survival and cell-based repair without the use of exogenous growth factors. Methods: Early passage hMSC (<4) were transduced to constitutively express firefly luciferase (hMSCFLuc). 4-arm PEG-maleimide was functionalized with one of three ligands: α2β1-binding collagen I-mimetic GFOGER, αvβ3-binding RGD, or non-adhesive RDG peptides and cross-linked with a protease-degradable peptide. 2.5 mm defects were created in the radii of 8-9 week old male NOD scid gamma mice and treated with cell-laden hydrogels containing 15k hMSC. Bioluminescence was monitored using an In Vivo Imaging System (PerkinElmer). New bone formation was evaluated using an in vivo microcomputer tomography scanner (Scanco Medical). Gene expression was evaluated after 1 week in vivo. Total RNA was extracted from cells in the defect space and standardized across all samples. 96 total genes were screened using Fluidigm gene chip technology. Results: Bioluminescence of transplanted hMSCFLuc was monitored after transplantation (Fig 1A). The significantly higher bioluminescent signal in GFOGER/hMSCFLuc-treated defects suggests GFOGER hydrogels support transplanted cell survival and proliferation in vivo whereas the bioluminescent signals for hMSCFLuc delivered in a RGD or RDG hydrogel remained low. GFOGER/hMSC-treated defects exhibited significantly higher bone volume at weeks 4 and 8 compared to RGD and RDG with no statistical difference between groups receiving hMSCFLuc and hMSC (Fig 1B). Differences in gene expression for approximately 60 genes were observed after 1 week in vivo, with statistically higher expression of vascularization and inflammation genes in GFOGER/hMSC treated defects (Fig 2). Conclusions: GFOGER-functionalized PEG hydrogels support enhanced bone formation and hMSC viability in vivo compared to RGD and RDG, possibly via enhanced vascularization These studies are ongoing for immunohistochemical, cytokine, and defect vascularization analyses. This work highlights integrin-specificity as an important consideration in the design of cell delivery vehicles for engraftment and tissue repair. NIH grants R01 AR062368 and R01 AR062920

Integrating Biomaterials and Stem Cells for Neural Regeneration.

The central nervous system has a limited capacity to regenerate, and thus, traumatic injuries or diseases often have devastating consequences. Therefore, there is a distinct need to develop alternative treatments that can achieve functional recovery without side effects currently observed with some pharmacological treatments. Combining biomaterials with pluripotent stem cells (PSCs), either embryonic or induced, has the potential to revolutionize the treatment of neurodegenerative diseases and traumatic injuries. Biomaterials can mimic the extracellular matrix and present a myriad of relevant biochemical cues through rational design or further functionalization. Biomaterials such as nanofibers and hydrogels, including self-assembling peptide (SAP) hydrogels can provide a superior cell culture environment. When these materials are then combined with PSCs, more accurate drug screening and disease modeling could be developed, and the generation of large number of cells with the appropriate phenotype can be achieved, for subsequent use in vitro. Biomaterials have also been shown to support endogenous cell growth after implantation, and, in particular, hydrogels and SAPs have effectively acted as cell delivery vehicles, increasing cell survival after transplantation. Few studies are yet to fully exploit the combination of PSCs and innovative biomaterials; however, initial studies with neural stem cells, for example, are promising, and, hence, such a combination for use in vitro and in vivo is an exciting new direction for the field of neural regeneration.

Data on bone marrow stem cells delivery using porous polymer scaffold

Low bioavailability and/or survival at the injury site of transplanted stem cells necessitate its delivery using a biocompatible, biodegradable cell delivery vehicle. In this dataset, we report the application of a porous biocompatible, biodegradable polymer network that successfully delivers bone marrow stem cells (BMSCs) at the wound site of a murine excisional splint wound model. In this data article, we are providing the additional data of the reference article “Porous polymer scaffold for on-site delivery of stem cells – protects from oxidative stress and potentiates wound tissue repair” (Ramasatyaveni et al., 2016) [1]. This data consists of the characterization of bone marrow stem cells (BMSCs) showing the pluripotency and stem cell-specific surface markers. Image analysis of the cellular penetration into PEG–PU polymer network and the mechanism via enzymatic activation of MMP-2 and MMP-13 are reported. In addition, we provide a comparison of various routes of transplantation-mediated BMSCs engraftment in the murine model using bone marrow transplantation chimeras. Furthermore, we included in this dataset the engraftment of BMSCs expressing Sca-1+Lin−CD133+CD90.2+ in post-surgery day 10.

Injectable and Thermosensitive Soluble Extracellular Matrix and Methylcellulose Hydrogels for Stem Cell Delivery in Skin Wounds.

Extracellular matrix (ECM) provides structural support and biochemical cues for tissue development and regeneration. Here we report a thermosensitive hydrogel composed of soluble ECM (sECM) and methylcellulose (MC) for injectable stem cell delivery. The sECM was prepared by denaturing solid ECM extracted from human adipose tissue and then blended with a MC solution. At low temperatures, the sECM-MC solution displayed a viscous solution state in which the loss modulus (G″) was predominant over the storage modulus (G'). With increasing temperature, G' increased dramatically and eventually exceeded G″ around 34 °C, characteristic of the transition from a liquid-like state to an elastic gel-like state. After a single injection of the stem cell-embedded hydrogel in full thickness cutaneous wound, the wound healed rapidly through re-epithelialization and neovascularization with minimum scar formation. The overall results suggest that in-situ-forming sECM-MC hydrogels are a promising injectable vehicle for stem cell delivery and tissue regeneration.

An In Vitro Investigation of Platelet-Rich Plasma-Gel as a Cell and Growth Factor Delivery Vehicle for Tissue Engineering.

Platelet-rich plasma (PRP) has been used for different applications in human and veterinary medicine. Many studies have shown promising therapeutic effects of PRP; however, there are still many controversies regarding its composition, properties, and clinical efficacy. The aim of this study was to evaluate the influence of different platelet concentrations on the rheological properties and growth factor (GF) release profile of PRP-gels. In addition, the viability of incorporated bone marrow-derived human mesenchymal stem cells (MSCs) was investigated. PRP (containing 1000 × 10(3), 2000 × 10(3), and 10,000 × 10(3) platelets/μL) was prepared from human platelet concentrates. Platelet activation and gelification were achieved by addition of human thrombin. Viscoelastic properties of PRP-gels were evaluated by rheological studies. The release of GFs and inflammatory proteins was measured using a membrane-based protein array and enzyme-linked immunosorbent assay. MSC viability and proliferation in PRP-gels were assessed over 7 days by cell viability staining. Cell proliferation was examined using DNA quantification. Regardless of the platelet content, all tested PRP-gels showed effective cross-linking. A positive correlation between protein release and the platelet concentration was observed at all time points. Among the detected proteins, the chemokine CCL5 was the most abundant. The greatest release appeared within the first 4 h after gelification. MSCs could be successfully cultured in PRP-gels over 7 days, with the highest cell viability and DNA content found in PRP-gels with 1000 × 10(3) platelets/μL. The results of this study suggest that PRP-gels represent a suitable carrier for both cell and GF delivery for tissue engineering. Notably, a platelet concentration of 1000 × 10(3) platelets/μL appeared to provide the most favorable environment for MSCs. Thus, the platelet concentration is an important consideration for the clinical application of PRP-gels.

Poly(ethylene glycol) hydrogels with cell cleavable groups for autonomous cell delivery

Cell-responsive hydrogels hold tremendous potential as cell delivery devices in regenerative medicine. In this study, we developed a hydrogel-based cell delivery vehicle, in which the encapsulated cell cargo control its own release from the vehicle in a protease-independent manner. Specifically, we have synthesized a modified poly(ethylene glycol) (PEG) hydrogel that undergoes degradation responding to cell-secreted molecules by incorporating disulfide moieties onto the backbone of the hydrogel precursor. Our results show the disulfide-modified PEG hydrogels disintegrate seamlessly into solution in presence of cells without any external stimuli. The rate of hydrogel degradation, which ranges from hours to months, is found to be dependent upon the type of encapsulated cells, cell number, and fraction of disulfide moieties present in the hydrogel backbone. The differentiation potential of human mesenchymal stem cells released from the hydrogels is maintained in vitro. The in vivo analysis of these cell-laden hydrogels, through a dorsal window chamber and intramuscular implantation, demonstrated autonomous release of cells to the host environment. The hydrogel-mediated implantation of cells resulted in higher cell retention within the host tissue when compared to that without a biomaterial support. Biomaterials that function as a shield to protect cell cargos and assist their delivery in response to signals from the encapsulated cells could have a wide utility in cell transplantation and could improve the therapeutic outcomes of cell-based therapies.

Development of recombinant collagen-peptide-based vehicles for delivery of adipose-derived stromal cells.

Stem cell therapy is a promising approach for repair, remodeling and even regenerate tissue of otherwise irreparable damage, such as after myocardial infarction (aMI). A severe limitation of cardiac stem cell therapy is the generally poor retention of administered cells in the target tissue. In tissue repair the main mode of action of adipose tissue-derived stem cells (ADSC) is the production of various growth factors, cytokines, anti-inflammatory and anti-apoptotic factors that together augment repair, remodeling, and regeneration. In this experiment, we used recombinant collagen peptide (RCP) with additional integrin-binding motives and different crosslinkers. Formulated as 50-100 μm microspheres with bound ADSC, we hypothesized that this would improve ADSC retention and function. Crosslinking was performed with chemical crosslinkers (EDC and HMDIC) at high and low concentrations or by thermal treatment (DHT). ADSC adhesion, proliferation, apoptosis/necrosis, and gene expressions in two-dimensional and three-dimensional were analyzed. In addition, the effect of ADSC conditioned medium (ADSC-CM) on proapoptotic/sprouting HUVEC was examined. Our results show that all materials support cell adhesion in short time point, however, EDC-High crosslinker induced ADSC apoptosis/necrosis. Gene expression results revealed lower expression of proinflammatory genes in chemical crosslinked materials, despite EDC-High the proinflammatory genes expressions were similar or higher than TCPS. In addition, cultured ADSC on DHT crosslinked RCP showed a proinflammatory phenotype compared to TCPS. Sprouting assay results confirmed the protective effect of ADSC-CM derived from TCPS and HMDIC-High crosslinked RCP proapoptotic HUVEC. We conclude that ADSC adhere to the materials and maintain their therapeutic profile.

A Neuroprotective Sericin Hydrogel As an Effective Neuronal Cell Carrier for the Repair of Ischemic Stroke.

Ischemic stroke causes extensive cellular loss that impairs brain functions, resulting in severe disabilities. No effective treatments are currently available for brain tissue regeneration. The need to develop effective therapeutic approaches for treating stroke is compelling. A tissue engineering approach employing a hydrogel carrying both cells and neurotrophic cytokines to damaged regions is an encouraging alternative for neuronal repair. However, this approach is often challenged by low in vivo cell survival rate, and low encapsulation efficiency and loss of cytokines. To address these limitations, we propose to develop a biomaterial that can form a matrix capable of improving in vivo survival of transplanted cells and reducing in vivo loss of cytokines. Here, we report that using sericin, a natural protein from silk, we have fabricated a genipin-cross-linked sericin hydrogel (GSH) with porous structure and mild swelling ratio. The GSH supports the effective attachment and growth of neurons in vitro. Strikingly, our data reveal that sericin protein is intrinsically neurotrophic and neuroprotective, promoting axon extension and branching as well as preventing primary neurons from hypoxia-induced cell death. Notably, these functions are inherited by the GSH's degradation products, which might spare a need of incorporating costly cytokines. We further demonstrate that this neurotrophic effect is dependent on the Lkb1-Nuak1 pathway, while the neuroprotective effect is realized through regulating the Bcl-2/Bax protein ratio. Importantly, when transplanted in vivo, the GSH gives a high cell survival rate and allows the cells to continuously proliferate. Together, this work unmasks the neurotrophic and neuroprotective functions for sericin and provides strong evidence justifying the GSH's suitability as a potential neuronal cell delivery vehicle for ischemic stroke repair.

Porous polymer scaffold for on-site delivery of stem cells – Protects from oxidative stress and potentiates wound tissue repair

Wound healing by cell transplantation techniques often suffer setbacks due to oxidative stress encountered at injury sites. A porous polyethyleneglycol–polyurethane (PEG–PU) scaffold that facilitates cell delivery and boosts tissue repair was developed through semi-interpenetrating polymer network approach. The key physico-chemical properties assessed confirms these polymeric matrices are highly thermostable, barostable, degrade at an acidic pH (5.8), biodegradable, cytocompatible and possess excellent porosity. Mechanism of cellular penetration into porous polymer networks was evident by a ≥6 – fold increase in gene expression of MMP-13 and MMP-2 via activation of Akt and Erk. H2O2-induced apoptosis of mouse bone marrow stem cells (BMSCs) was abrogated in presence of polymer networks indicating a protective effect from oxidative stress. Transplantation of BMSC + PEG–PU at murine excisional splint wound site depicted significant increase in fibroblast proliferation, collagen deposition, anti-oxidant enzyme activities of catalase, SOD and GPx. Furthermore it significantly decreased expression of pro-inflammatory cytokines (IL-1β, TNF-α, IL-8, etc) with a concomitant increase in anti-inflammatory cytokines (IL-10, IL-13) at an early healing period of day 7. Finally, immunostaining revealed an enhanced engraftment and vascularity indicating an accelerated wound tissue closure. This pre-clinical study demonstrates the proof-of-concept and further necessitates their clinical evaluation as potential cell delivery vehicle scaffolds.