PMCC PMCC

Search tips
Search criteria

Advanced
Results 1-25 (146059)

Clipboard (0)
None

Related Articles

1.  Sustained Delivery of Dibutyryl Cyclic Adenosine Monophosphate to the Transected Spinal Cord Via Oligo [(Polyethylene Glycol) Fumarate] Hydrogels 
Tissue Engineering. Part A  2011;17(9-10):1287-1302.
This study describes the use of oligo [(polyethylene glycol) fumarate] (OPF) hydrogel scaffolds as vehicles for sustained delivery of dibutyryl cyclic adenosine monophosphate (dbcAMP) to the transected spinal cord. dbcAMP was encapsulated in poly(lactic-co-glycolic acid) (PLGA) microspheres, which were embedded within the scaffolds architecture. Functionality of the released dbcAMP was assessed using neurite outgrowth assays in PC12 cells and by delivery to the transected spinal cord within OPF seven channel scaffolds, which had been loaded with Schwann cells or mesenchymal stem cells (MSCs). Our results showed that encapsulation of dbcAMP in microspheres lead to prolonged release and continued functionality in vitro. These microspheres were then successfully incorporated into OPF scaffolds and implanted in the transected thoracic spinal cord. Sustained delivery of dbcAMP inhibited axonal regeneration in the presence of Schwann cells but rescued MSC-induced inhibition of axonal regeneration. dbcAMP was also shown to reduce capillary formation in the presence of MSCs, which was coupled with significant functional improvements. Our findings demonstrate the feasibility of incorporating PLGA microsphere technology for spinal cord transection studies. It represents a novel sustained delivery mechanism within the transected spinal cord and provides a platform for potential delivery of other therapeutic agents.
doi:10.1089/ten.tea.2010.0396
PMCID: PMC3079174  PMID: 21198413
2.  Repair of Osteochondral Defects with Biodegradable Hydrogel Composites Encapsulating Marrow Mesenchymal Stem Cells in a Rabbit Model 
Acta biomaterialia  2009;6(1):39-47.
This work investigated the delivery of marrow mesenchymal stem cells (MSCs), with or without the growth factor transforming growth factor-β1 (TGF-β1), from biodegradable hydrogel composites on the repair of osteochondral defects in a rabbit model. Three formulations of oligo(poly(ethylene glycol) fumarate) (OPF) hydrogel composites containing gelatin microparticles (GMPs) and MSCs were implanted in osteochondral defects, including (1) OPF/GMP hydrogel composites; (2) OPF/GMP hydrogel composites encapsulating MSCs; and (3) OPF hydrogel composites containing TGF-β1 loaded GMPs and MSCs. At 12 weeks, the quality of new tissue formed in chondral and subchondral regions of defects was evaluated based on subjective and quantitative histological analysis. OPF hydrogel composites were partially degraded and the defects were filled with newly formed tissue at 12 weeks with no sign of persistent inflammation. With the implantation of scaffolds alone, newly formed chondral tissue had an appearance of hyaline cartilage with zonal organization and intense staining for glycosaminoglycans, while in the subchondral region hypertrophic cartilage with some extent of bone formation was often observed. The addition of MSCs, especially with TGF-β1 loaded GMPs, facilitated subchondral bone formation, as evidenced by more trabecular bone appearance. However, the delivery of MSCs with or without TGF-β1 at the dosage investigated did not improve cartilage morphology. While OPF-based hydrogel composites supported osteochondral tissue generation, further investigations are necessary to elucidate the effects of MSC seeding density and differentiation stage on new tissue formation and regeneration.
doi:10.1016/j.actbio.2009.07.041
PMCID: PMC2787824  PMID: 19660580
cartilage tissue engineering; mesenchymal stem cells; hydrogel composites; osteochondral defects
3.  Synthesis of oligo(poly(ethylene glycol) fumarate) 
Nature protocols  2012;7(6):1219-1227.
This protocol describes the synthesis of oligo(poly(ethylene glycol) fumarate) (OPF) (1–35 kDa)(a polymer useful for tissue engineering applications) by a one-pot reaction of poly(ethylene glycol) (PEG) and fumaryl chloride. The procedure involves three parts: dichloromethane and PEG are first dried; the reaction step follows in which fumaryl chloride and triethylamine are added dropwise to a solution of PEG in dichloromethane; and finally the product solution is filtered to remove byproduct salt, and the OPF product is twice crystallized, washed, and dried under vacuum. The reaction is affected by PEG molecular weight and reactant molar ratio. The OPF product is cross-linked by radical polymerization by either a thermally induced or UV-induced radical initiator, and the physical properties of the OPF oligomer and resulting cross-linked hydrogel are easily tailored by varying PEG molecular weight. OPF hydrogels are injectable, polymerize in situ, and undergo biodegradation by hydrolysis of ester bonds. The expected time required to complete this protocol is 6 d.
doi:10.1038/nprot.2012.055
PMCID: PMC3513913  PMID: 22653160
oligo(poly(ethylene glycol) fumarate); OPF; polymer; tissue engineering; polymer synthesis; radical polymerization; hydrogel; PEG
4.  Injectable Biodegradable Hydrogels for Embryonic Stem Cell Transplantation: Improved Cardiac Remodeling and Function of Myocardial Infarction 
In this study, an injectable, biodegradable hydrogel composite of oligo(poly(ethylene glycol) fumarate) (OPF) was investigated as a carrier of mouse embryonic stem cells (mESCs) for the treatment of myocardial infarction (MI). 10K OPF hydrogels were used to encapsulate mESCs. The cell differentiation in vitro over 14 days was determined via immunohistochemical examination. Then, mESCs encapsulated in OPF hydrogels were injected into the left ventricular wall of a rat myocardial infarction model. Detailed histological analysis and echocardiography were used to determine the structural and functional consequences after 4 weeks of transplantation. With ascorbic acid induction, mESCs could differentiate into cardiomyocytes and other cell types in all three lineages in the OPF hydrogel. After transplantation, both the 24h-cell retention and 4-week graft size were significantly greater in the OPF + ESC group than that of the PBS + ESC group (p<0.01). Four weeks after transplantation, OPF hydrogel alone significantly reduced the infarct size and collagen deposition and improved the cardiac function. The heart function and revascularization improved significantly, while the infarct size and fibrotic area decreased significantly in the OPF + ESC group compared with that of the PBS +ESC group, OPF group and PBS group (p<0.01). All treatments had significantly reduced MMP2 and MMP9 protein levels compared to the PBS control group, and the OPF + ESC group decreased most by Western blotting. Transplanted mESCs expressed cardiovascular markers. This study suggest the potential of a method for heart regeneration involving oligo(poly(ethylene glycol) fumarate) hydrogels for stem cell encapsulation and transplantation.
doi:10.1111/j.1582-4934.2011.01409.x
PMCID: PMC3227757  PMID: 21838774
cardiac tissue engineering; injectable hydrogels; cell encapsulation; embryonic stem cell; myocardial infarction
5.  Development of Electrically Conductive Oligo(polyethylene Glycol) Fumarate-Polypyrrole Hydrogels for Nerve Regeneration 
Biomacromolecules  2010;11(11):2845-2853.
Electrically conductive hydrogel composites consisting of oligo(polyethylene glycol) fumarate (OPF) and polypyrrole (PPy) were developed for applications in nerve regeneration. OPF-PPy scaffolds were synthesized using three different anions: naphthalene-2-sulfonic acid sodium salt (NSA), dodecylbenzenesulfonic acid sodium salt (DBSA), and dioctyl sulfosuccinate sodium salt (DOSS). Scaffolds were characterized by ATR-FTIR, XPS, AFM, dynamic mechanical analysis, electrical resistivity measurements, and swelling experiments. OPF-PPy scaffolds were shown to consist of up to 25 mol% polypyrrole with a compressive modulus ranging from 265 to 323 kPa and a sheet resistance ranging from 6 to 30 × 103 Ohms/square. In vitro studies using PC12 cells showed OPF-PPy materials had no cytotoxicity and PC12 cells showed distinctly better cell attachment and an increase in the percent of neurite bearing cells on OPF-PPy materials compared to OPF. The neurite lengths of PC12 cells were significantly higher on OPF-PPyNSA and OPF-PPyDBSA. These results show that electrically conductive OPF-PPy hydrogels are promising candidates for future applications in nerve regeneration.
doi:10.1021/bm100526a
PMCID: PMC3947846  PMID: 20942380
hydrogel; electrical; conductive; nerve; tissue regeneration
6.  Oligo[poly(ethylene glycol)fumarate] Hydrogel Enhances Osteochondral Repair in Porcine Femoral Condyle Defects 
Background
Management of osteochondritis dissecans remains a challenge. Use of oligo[poly(ethylene glycol)fumarate] (OPF) hydrogel scaffold alone has been reported in osteochondral defect repair in small animal models. However, preclinical evaluation of usage of this scaffold alone as a treatment strategy is limited.
Questions/purposes
We therefore (1) determined in vitro pore size and mechanical stiffness of freeze-dried and rehydrated freeze-dried OPF hydrogels, respectively; (2) assessed in vivo gross defect filling percentage and histologic findings in defects implanted with rehydrated freeze-dried hydrogels for 2 and 4 months in a porcine model; (3) analyzed highly magnified histologic sections for different types of cartilage repair tissues, subchondral bone, and scaffold; and (4) assessed neotissue filling percentage, cartilage phenotype, and Wakitani scores.
Methods
We measured pore size of freeze-dried OPF hydrogel scaffolds and mechanical stiffness of fresh and rehydrated forms. Twenty-four osteochondral defects from 12 eight-month-old micropigs were equally divided into scaffold and control (no scaffold) groups. Gross and histologic examination, one-way ANOVA, and one-way Mann-Whitney U test were performed at 2 and 4 months postoperatively.
Results
Pore sizes ranged from 20 to 433 μm in diameter. Rehydrated freeze-dried scaffolds had mechanical stiffness of 1 MPa. The scaffold itself increased percentage of neotissue filling at both 2 and 4 months to 58% and 54%, respectively, with hyaline cartilage making up 39% of neotissue at 4 months.
Conclusions
Rehydrated freeze-dried OPF hydrogel can enhance formation of hyaline-fibrocartilaginous mixed repair tissue of osteochondral defects in a porcine model.
Clinical Relevance
Rehydrated freeze-dried OPF hydrogel alone implanted into cartilage defects is insufficient to generate a homogeneously hyaline cartilage repair tissue, but its spacer effect can be enhanced by other tissue-regenerating mediators.
doi:10.1007/s11999-012-2487-0
PMCID: PMC3586016  PMID: 22826014
7.  Effect of Swelling Ratio of Injectable Hydrogel Composites on Chondrogenic Differentiation of Encapsulated Rabbit Marrow Mesenchymal Stem Cells In Vitro 
Biomacromolecules  2009;10(3):541-546.
An injectable, biodegradable hydrogel composite of oligo(poly(ethylene glycol) fumarate) (OPF) and gelatin microparticles (MPs) has been investigated as a cell and growth factor carrier for cartilage tissue engineering applications. In this study, hydrogel composites with different swelling ratios were prepared by crosslinking OPF macromers with poly(ethylene glycol) (PEG) repeating units of varying molecular weights from 1,000 ~ 35,000. Rabbit marrow mesenchymal stem cells (MSCs) and MPs loaded with transforming growth factor-β1 (TGF-β1) were encapsulated in the hydrogel composites in order to examine the effect of the swelling ratio of the hydrogel composites on the chondrogenic differentiation of encapsulated rabbit marrow MSCs both in the presence and absence of TGF-β1. The swelling ratio of the hydrogel composites increased as the PEG molecular weight in the OPF macromers increased. Chondrocyte-specific genes were expressed at higher levels in groups containing TGF-β1-loaded MPs and varied with the swelling ratio of the hydrogel composites. OPF hydrogel composites with PEG repeating units of molecular weight 35,000 and 10,000 with TGF-β1-loaded MPs exhibited a 159 ± 95 and a 89 ± 31 fold increase in type II collagen gene expression at day 28, respectively, while OPF hydrogel composites with PEG repeating units of molecular weight 3,000 and 1,000 with TGF-β1-loaded MPs showed a 27 ± 10 and a 17 ± 7 fold increase in type II collagen gene expression, respectively, as compared to the composites with blank MPs at day 0. The results indicate that chondrogenic differentiation of encapsulated rabbit marrow MSCs within OPF hydrogel composites could be affected by their swelling ratio, thus suggesting the potential of OPF composite hydrogels as part of a novel strategy for controlling the differentiation of stem cells.
doi:10.1021/bm801197m
PMCID: PMC2765566  PMID: 19173557
injectable hydrogels; crosslinking; marrow mesenchymal stem cells; gelatin microparticles; TGF-β1; chondrogenic differentiation; cartilage tissue engineering
8.  The effects of fixed electrical charge on chondrocyte behavior 
Acta biomaterialia  2011;7(5):2080-2090.
In this study, we have compared the effects of negative and positive fixed charge on chondrocyte behavior in vitro. Electrical charges have been incorporated into oligo(poly(ethylene glycol) fumarate) (OPF) using small charged monomers such as sodium methacrylate (SMA) and (2-(methacryloyloxy) ethyl)-trimethyl ammonium chloride (MAETAC) to produce negatively and positively charged hydrogels, respectively. The hydrogel physical and electrical properties were characterized through measuring and calculating the swelling ratio and zeta potential, respectively. Our results revealed that the properties of these OPF modified hydrogels varied according to the concentration of charged monomers. Zeta potential measurements demonstrated that the electrical property of the OPF hydrogel surfaces changed due to incorporation of SMA and MAETAC and that this change in electrical property was dose-dependent. Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy was used to determine the hydrogel surface composition. To assess the effects of surface properties on chondrocyte behavior, primary chondrocytes isolated from rabbit ears were seeded as a monolayer on top of the hydrogels. We demonstrated that the cells remained viable over 7 days and began to proliferate while seeded on top of the hydrogels. Collagen type II staining was positive in all samples; however, the intensity of the stain was higher on negatively charged hydrogels. Similarly, GAG production was significantly higher on negatively charged hydrogels compared to neutral hydrogel. Reverse transcription polymerase chain reaction showed up-regulation of collagen type II and down-regulation of collagen type I on the negatively charged hydrogels. These findings indicate that charge plays an important role in establishing an appropriate environment for chondrocytes and hence in the engineering of cartilage. Thus, further investigation into charged hydrogels for cartilage tissue engineering is merited.
doi:10.1016/j.actbio.2011.01.012
PMCID: PMC3103083  PMID: 21262395
hydrogel; cartilage tissue engineering; OPF; scaffold
9.  Axon Regeneration through Scaffold into Distal Spinal Cord after Transection 
Journal of neurotrauma  2009;26(10):1759-1771.
We employed Fast Blue (FB) axonal tracing to determine the origin of regenerating axons after thoracic spinal cord transection injury in rats. Schwann cell (SC)-loaded, biodegradable, poly(lactic-co-glycolic acid) (PLGA) scaffolds were implanted after transection. Scaffolds loaded with solubilized basement membrane preparation (without SCs) were used for negative controls, and nontransected cords were positive controls. One or 2 months after injury and scaffold implantation, FB was injected 0–15 mm caudal or about 5 mm rostral to the scaffold. One week later, tissue was harvested and the scaffold and cord sectioned longitudinally (30 μm) on a cryostat. Trans-scaffold labeling of neuron cell bodies was identified with confocal microscopy in all cell-transplanted groups. Large (30–50 μm diameter) neuron cell bodies were predominantly labeled in the ventral horn region. Most labeled neurons were seen 1–10 mm rostral to the scaffold, although some neurons were also labeled in the cervical cord. Axonal growth occurred bidirectionally after cord transection, and axons regenerated up to 14 mm beyond the PLGA scaffolds and into distal cord. The extent of FB labeling was negatively correlated with distance from the injection site to the scaffold. Electron microscopy showed myelinated axons in the transverse sections of the implanted scaffold 2 months after implantation. The pattern of myelination, with extracellular collagen and basal lamina, was characteristic of SC myelination. Our results show that FB labeling is an effective way to measure the origin of regenerating axons.
doi:10.1089/neu.2008-0610
PMCID: PMC2763055  PMID: 19413501
axonal tracing; biodegradable polymers; Fast Blue; Schwann cells; spinal cord injury
10.  Axon Regeneration through Scaffold into Distal Spinal Cord after Transection 
Journal of Neurotrauma  2009;26(10):1759-1771.
Abstract
We employed Fast Blue (FB) axonal tracing to determine the origin of regenerating axons after thoracic spinal cord transection injury in rats. Schwann cell (SC)-loaded, biodegradable, poly(lactic-co-glycolic acid) (PLGA) scaffolds were implanted after transection. Scaffolds loaded with solubilized basement membrane preparation (without SCs) were used for negative controls, and nontransected cords were positive controls. One or 2 months after injury and scaffold implantation, FB was injected 0–15 mm caudal or about 5 mm rostral to the scaffold. One week later, tissue was harvested and the scaffold and cord sectioned longitudinally (30 μm) on a cryostat. Trans-scaffold labeling of neuron cell bodies was identified with confocal microscopy in all cell-transplanted groups. Large (30–50 μm diameter) neuron cell bodies were predominantly labeled in the ventral horn region. Most labeled neurons were seen 1–10 mm rostral to the scaffold, although some neurons were also labeled in the cervical cord. Axonal growth occurred bidirectionally after cord transection, and axons regenerated up to 14 mm beyond the PLGA scaffolds and into distal cord. The extent of FB labeling was negatively correlated with distance from the injection site to the scaffold. Electron microscopy showed myelinated axons in the transverse sections of the implanted scaffold 2 months after implantation. The pattern of myelination, with extracellular collagen and basal lamina, was characteristic of SC myelination. Our results show that FB labeling is an effective way to measure the origin of regenerating axons.
doi:10.1089/neu.2008.0610
PMCID: PMC2763055  PMID: 19413501
axonal tracing; biodegradable polymers; Fast Blue; Schwann cells; spinal cord injury
11.  A Stimuli-Responsive Hydrogel for Doxorubicin Delivery 
Biomaterials  2010;31(31):8051-8062.
The goal of this study was to develop a polymeric carrier for delivery of anti-tumor drugs and sustained release of these agents in order to optimize anti-tumor activity while minimizing systemic effects. We used oligo(poly(ethylene glycol) fumarate) (OPF) hydrogels modified with small negatively charged molecules, sodium methacrylate (SMA), for delivery of doxorubicin (DOX). SMA at different concentrations was incorporated into the OPF hydrogel with a photo-crosslinking method. The resulting hydrogels exhibited sensitivity to the pH and ionic strength of the surrounding environment. Our results revealed that DOX was bound to the negatively charged hydrogel through electrostatic interaction and was released in a timely fashion with an ion exchange mechanism. Release kinetics of DOX was directly correlated to the concentration of SMA in the hydrogel formulations. Anti-tumor activity of the released DOX was assessed using a human osteosarcoma cell line. Our data revealed that DOX released from the modified, charged hydrogels remained biologically active and had the capability to kill cancer cells. In contrast, control groups of unmodified OPF hydrogels with or without DOX did not exhibit any cytotoxicity. This study demonstrates the feasibility of using SMA-modified OPF hydrogels as a potential carrier for chemotherapeutic drugs for cancer treatments.
doi:10.1016/j.biomaterials.2010.06.054
PMCID: PMC2936247  PMID: 20696470
12.  Injectable biodegradable hydrogel composites for rabbit marrow mesenchymal stem cell and growth factor delivery for cartilage tissue engineering 
Biomaterials  2007;28(21):3217-3227.
We investigated the development of an injectable, biodegradable hydrogel composite of oligo(poly(ethylene glycol) fumarate) (OPF) with encapsulated rabbit marrow mesenchymal stem cells (MSCs) and gelatin microparticles (MPs) loaded with transforming growth factor-β1 (TGF-β1) for cartilage tissue engineering applications. Rabbit MSCs and TGF-β1-loaded MPs were mixed with OPF, a poly(ethylene glycol)-diacrylate crosslinker and the radical initiators ammonium persulfate and N,N,N’,N’-tetramethylethylenediamine, and then crosslinked at 37°C for 8 min to form hydrogel composites. Three studies were conducted over 14 days in order to examine the effects of: 1) the composite formulation, 2) the MSC seeding density, and 3) the TGF-β1 concentration on the chondrogenic differentiation of encapsulated rabbit MSCs. Bioassay results showed no significant difference in DNA amount between groups, however, groups with MPs had a significant increase in glycosaminoglycan content per DNA starting at day 7 as compared to controls at day 0. Chondrocyte-specific gene expression of type II collagen and aggrecan were only evident in groups containing TGF-β1-loaded MPs and varied with TGF-β1 concentration in a dose dependent manner. Specifically, type II collagen gene expression exhibited a 161 ± 49 fold increase and aggrecan gene expression a 221 ± 151 fold increase after 14 days with the highest dose of TGF-β1 (16 ng/ml). These results indicate that encapsulated rabbit MSCs remained viable over the culture period and differentiated into chondrocyte-like cells, thus suggesting the potential of OPF composite hydrogels as part of a novel strategy for localized delivery of stem cells and bioactive molecules.
doi:10.1016/j.biomaterials.2007.03.030
PMCID: PMC2964378  PMID: 17445882
Cartilage tissue engineering; marrow mesenchymal stem cells; gelatin microparticles; injectable hydrogels; TGF-β1
13.  Cyclic Tensile Culture Promotes Fibroblastic Differentiation of Marrow Stromal Cells Encapsulated in Poly(Ethylene Glycol)-Based Hydrogels 
Tissue Engineering. Part A  2010;16(11):3457-3466.
To inform future efforts in tendon/ligament tissue engineering, our laboratory has developed a well-controlled model system with the ability to alter both external tensile loading parameters and local biochemical cues to better understand marrow stromal cell differentiation in response to both stimuli concurrently. In particular, the synthetic, poly(ethylene glycol)-based hydrogel material oligo(poly(ethylene glycol) fumarate) (OPF) has been explored as a cell carrier for this system. This biomaterial can be tailored to present covalently incorporated bioactive moieties and can be loaded in our custom cyclic tensile bioreactor for up to 28 days with no loss of material integrity. Human marrow stromal cells encapsulated in these OPF hydrogels were cultured (21 days) under cyclic tensile strain (10%, 1 Hz, 3 h of strain followed by 3 h without) or at 0% strain. No difference was observed in cell number due to mechanical stimulation or across time (n = 4), with cells remaining viable (n = 4) through 21 days. Cyclic strain significantly upregulated all tendon/ligament fibroblastic genes examined (collagen I, collagen III, and tenascin-C) by day 21 (n ≥ 6), whereas genes for other pathways (osteogenic, chondrogenic, and adipogenic) did not increase. After 21 days, the presence of collagen I and tenascin-C was observed via immunostaining (n = 2). This study demonstrates the utility of this hydrogel/bioreactor system as a versatile, yet well-controlled, model environment to study marrow stromal cell differentiation toward the tendon/ligament phenotype under a variety of conditions.
doi:10.1089/ten.tea.2010.0233
PMCID: PMC2965196  PMID: 20666585
14.  Three-Dimensional Porous Biodegradable Polymeric Scaffolds Fabricated with Biodegradable Hydrogel Porogens 
We have developed a new fabrication technique to create three-dimensional (3D) porous poly(ε-caprolactone fumarate) (PCLF) scaffolds using hydrogel microparticle porogens, as an alternative to overcome certain limitations of traditional scaffold fabrication techniques such as a salt leaching method. Both natural hydrogel, gelatin, and synthetic hydrogel, poly(ethylene glycol) sebacic acid diacrylate, were used as porogens to fabricate 3D porous PCLF scaffolds. Hydrogel microparticles were prepared by a single emulsion technique with the particle size in the range of 100–500 μm after equilibrium in water. The pore size distribution, porosity, pore interconnectivity, and spatial pore heterogeneity of the 3D PCLF scaffolds were assessed using micro-computed tomography and imaging analysis. Scaffolds fabricated with the hydrogel porogens had higher porosity and pore interconnectivity as well as more homogeneous spatial pore distribution, compared to the scaffolds made from the salt leaching process. Compressive moduli of the scaffolds were also measured and showed that lower porosity yielded greater modulus of the scaffolds. Overall, the new fabrication technology using hydrogel porogens may be beneficial for certain tissue engineering applications.
doi:10.1089/ten.tec.2008.0642
PMCID: PMC2819712  PMID: 19216632
15.  Stimulation of neurite outgrowth using positively charged hydrogels 
Biomaterials  2009;30(23-24):3874-3881.
Autologous nerve grafts are currently the best option for the treatment of segmental peripheral nerve defects. However, autografts have several drawbacks including size mismatch and loss of sensation in the donor nerve’s sensory distribution. In this work, we have investigated the development of a synthetic hydrogel that contains positive charge for use as a substrate for nerve cell attachment and neurite outgrowth in culture. We have demonstrated that modification of oligo-(polyethylene glycol) fumarate (OPF) with a positively charged monomer improves primary sensory rat neuron attachment and differentiation in a dose-dependent manner. Positively charged hydrogels also supported attachment of dorsal root ganglion (DRG) explants that contain sensory neurons, Schwann cells and neuronal support cells. Furthermore, charged hydrogels were analyzed for the appearance of myelinated structures in a co-culture containing DRG neurons and Schwann cells. DRGs and Schwann cells remained viable on charged hydrogels for a time period of three weeks and neurites extended from the DRGs. Sudan black staining revealed that neurites emerging from DRGs were accompanied by migrating Schwann cells. These findings suggest that charged OPF hydrogels are capable of sustaining both primary nerve cells and the neural support cells that are critical for regeneration.
doi:10.1016/j.biomaterials.2009.04.018
PMCID: PMC2716054  PMID: 19427689
hydrogel; nerve regeneration; Schwann cells; scaffold
16.  Neural Stem Cell– and Schwann Cell–Loaded Biodegradable Polymer Scaffolds Support Axonal Regeneration in the Transected Spinal Cord 
Tissue Engineering. Part A  2009;15(7):1797-1805.
Biodegradable polymer scaffolds provide an excellent approach to quantifying critical factors necessary for restoration of function after a transection spinal cord injury. Neural stem cells (NSCs) and Schwann cells (SCs) support axonal regeneration. This study examines the compatibility of NSCs and SCs with the poly-lactic-co-glycolic acid polymer scaffold and quantitatively assesses their potential to promote regeneration after a spinal cord transection injury in rats. NSCs were cultured as neurospheres and characterized by immunostaining for nestin (NSCs), glial fibrillary acidic protein (GFAP) (astrocytes), βIII-tubulin (immature neurons), oligodendrocyte-4 (immature oligodendrocytes), and myelin oligodendrocyte (mature oligodendrocytes), while SCs were characterized by immunostaining for S-100. Rats with transection injuries received scaffold implants containing NSCs (n = 17), SCs (n = 17), and no cells (control) (n = 8). The degree of axonal regeneration was determined by counting neurofilament-stained axons through the scaffold channels 1 month after transplantation. Serial sectioning through the scaffold channels in NSC- and SC-treated groups revealed the presence of nestin, neurofilament, S-100, and βIII tubulin–positive cells. GFAP-positive cells were only seen at the spinal cord–scaffold border. There were significantly more axons in the NSC- and SC- treated groups compared to the control group. In conclusion, biodegradable scaffolds with aligned columns seeded with NSCs or SCs facilitate regeneration across the transected spinal cord. Further, these multichannel biodegradable polymer scaffolds effectively serve as platforms for quantitative analysis of axonal regeneration.
doi:10.1089/ten.tea.2008.0364
PMCID: PMC2792101  PMID: 19191513
17.  Effect of Hydrogel Porosity on Marrow Stromal Cell Phenotypic Expression 
Biomaterials  2008;29(14):2193-2202.
This study describes investigation of porous photocrosslinked oligo[(polyethylene glycol) fumarate] (OPF) hydrogels as potential matrix for osteoblastic differentiation of marrow stromal cells (MSCs). The porosity and interconnectivity of porous hydrogels were assessed using magnetic resonance microscopy (MRM) as a noninvasive investigative tool that could image the water construct inside the hydrogels at a high spatial resolution. MSCs were cultured onto the porous hydrogels and cell number was assessed using PicoGreen DNA assay. Our results showed 10% of cells initially attached to the surface of scaffolds. However, cells did not show significant proliferation over a time period of 14 days. MSCs cultured on porous hydrogels had increased alkaline phosphatase activity as well as deposition of calcium, suggesting successful differentiation and maturation to the osteoblastic phenotype. Moreover, continued expression of type I collagen and osteonectin over 14 days confirmed osteoblastic differentiation of MSCs. MRM was also applied to monitor osteogenesis of MSCs on porous hydrogels. MRM images showed porous scaffolds became consolidated with osteogenic progression of cell differentiation. These findings indicate that porous OPF scaffolds enhanced MSC differentiation leading to development of bone-like mineralized tissue.
doi:10.1016/j.biomaterials.2008.01.006
PMCID: PMC2386206  PMID: 18262642
Hydrogel; oligo[(polyethylene glycol) fumarate] (OPF); Marrow stromal cells; Magnetic resonance microscopy; Osteogenesis
18.  Relationship between Scaffold Channel Diameter and Number of Regenerating Axons in the Transected Rat Spinal Cord 
Acta biomaterialia  2009;5(7):2551-2559.
Regeneration of endogenous axons through a Schwann cell (SC)-seeded scaffold implant has been demonstrated in the transected rat spinal cord. The formation of a cellular lining in the scaffold channel may limit the degree of axonal regeneration. Spinal cords of adult rats were transected and implanted with the SC-loaded polylactic co-glycollic acid (PLGA) scaffold implants containing seven parallel-aligned channels, either 450-μm (n=19) or 660-μm in diameter (n=14). Animals were sacrificed after 1, 2, and 3 months. Immunohistochemistry for neurofilament-expression was performed. The cross-sectional area of fibrous tissue and regenerative core was calculated. We found that the 450-μm scaffolds had significantly greater axon fibers per channel at the one month (186 ± 37) and three month (78 ± 11) endpoints than the 660-μm scaffolds (90 ± 19 and 40 ± 6, respectively) (P=0.0164 & 0.0149, respectively). The difference in the area of fibrous rim between the 450-μm and 660-μm channels was most pronounced at the one month endpoint, at 28,046 μm2 ± 6,551 and 58,633 μm2 ± 7,063, respectively (P=0.0105). Our study suggests that fabricating scaffolds with smaller diameter channels promotes greater regeneration over larger diameter channels. Axonal regeneration was reduced in the larger channels due to the generation of a large fibrous rim. Optimization of this scaffold environment establishes a platform for future studies of the effects of cell types, trophic factors or pharmacological agents on the regenerative capacity of the injured spinal cord.
doi:10.1016/j.actbio.2009.03.021
PMCID: PMC2731813  PMID: 19409869
Biomedical Engineering; Tissue Development and Growth; Central Nervous System; Polymeric Scaffolds
19.  Hydrogel-mediated DNA delivery confers estrogenic response in non-responsive osteoblast cells 
Oligo (polyethylene glycol) fumarate (OPF) hydrogel has been employed in musculoskeletal tissue engineering for photo-encapsulation of chondrocytes and as a matrix for marrow stromal cells differentiation. In this study, we have studied the application of OPF hydrogel for co-encapsulation of DNA and bone cells and examined whether co-encapsulation can enhance gene transfer by maintaining the DNA within the cellular microenvironment. Our results showed that plasmid DNA encoding green fluorescence protein (GFP), co-encapsulated with bone tumor cells, was capable of transfecting the cells and the transfected tumor cells continuously expressed GFP protein over the time course of study (21 days). Furthermore, we have examined the co-encapsulation of estrogen receptor (ER) encoding plasmid DNA and human fetal osteoblast cells (hFOB) that lack endogenous ER. Our results show that the transfected cells responded to estrogen as alkaline phosphatase (ALP), and estrogen response element (ERE)-directed luciferase enzyme activities increased with estrogen-treatment. Taken together, these studies show that OPF hydrogel could be further explored for targeted gene delivery in bone and other tissues encapsulated within the hydrogels.
doi:10.1002/jbm.a.32291
PMCID: PMC2783666  PMID: 19148929
Bone tissue engineering; DNA delivery, Hydrogel; Osteoblast, Estrogen receptor
20.  In Vitro Generation of an Osteochondral Construct Using Injectable Hydrogel Composites Encapsulating Rabbit Marrow Mesenchymal Stem Cells 
Biomaterials  2009;30(14):2741-2752.
Injectable, biodegradable hydrogel composites of crosslinked oligo(poly(ethylene glycol) fumarate) (OPF) and gelatin microparticles (MPs) were utilized to fabricate a bilayered osteochondral construct consisting of a chondrogenic layer and an osteogenic layer, and to investigate the differentiation of rabbit marrow mesenchymal stem cells (MSCs) encapsulated in both layers in vitro. The results showed that MSCs in the chondrogenic layer were able to undergo chondrogenic differentiation, especially in the presence of TGF-β1-loaded MPs. In the osteogenic layer, cells maintained their osteoblastic phenotype. Although calcium deposition in the osteogenic layer was limited, cells in the osteogenic layer significantly enhanced chondrogenic differentiation of MSCs in the chondrogenic layer. The greatest effect was observed when MSCs were encapsulated with TGF-β1-loaded MPs and cultured with osteogenic cells in the bilayered constructs. Overall, this study demonstrates the fabrication of bilayered hydrogel composites that mimic the structure and function of osteochondral tissue, along with the application of these composites as cell and growth factor carriers.
doi:10.1016/j.biomaterials.2009.01.048
PMCID: PMC2877597  PMID: 19232711
mesenchymal stem cell; cartilage tissue engineering; hydrogel; cell encapsulation; drug delivery; growth factors; composite; biomimetic material
21.  The Effects of TGF-β3 and Preculture Period of Osteogenic Cells on the Chondrogenic Differentiation of Rabbit Marrow Mesenchymal Stem Cells Encapsulated in a Bilayered Hydrogel Composite 
Acta biomaterialia  2010;6(8):2920-2931.
In this work, injectable, biodegradable hydrogel composites of crosslinked oligo(poly(ethylene glycol) fumarate) (OPF) and gelatin microparticles (MPs) were utilized to fabricate a bilayered osteochondral construct. Rabbit marrow mesenchymal stem cells (MSCs) were encapsulated with transforming growth factor-β3 (TGF-β3)-loaded MPs in the chondrogenic layer and cocultured with cells of different periods of osteogenic preculture (0, 3, 6 and 12 days) in the osteogenic layer to investigate the effects of TGF-β3 delivery and coculture on the proliferation and differentiation of cells in both layers. The results showed that, in the chondrogenic layer, TGF-β3 significantly stimulated chondrogenic differentiation of MSCs. Additionally, cells of various osteogenic preculture periods in the osteogenic layer, along with TGF-β3, enhanced gene expression for MSC chondrogenic markers to different extents. In the osteogenic layer, cells maintained their alkaline phosphatase activity during the coculture; however, mineralization was delayed by the presence of TGF-β3. Overall, this study demonstrated the fabrication of bilayered hydrogel composites that mimic the structure and function of osteochondral tissue, along with the application of these composites as cell and growth factor carriers, while illustrating that encapsulated cells of different degrees of osteogenic differentiation can significantly influence the chondrogenic differentiation of cocultured progenitor cells in both the presence and absence of chondrogenic growth factors.
doi:10.1016/j.actbio.2010.02.046
PMCID: PMC2882985  PMID: 20197126
bilayered hydrogel composites; mesenchymal stem cell; cell differentiation; coculture
22.  Enzymatic mineralization of hydrogels for bone tissue engineering by incorporation of alkaline phosphatase 
Macromolecular bioscience  2012;12(8):1077-1089.
Alkaline Phosphatase (ALP), an enzyme involved in mineralization of bone, was incorporated into three hydrogel biomaterials to induce their mineralization with calcium phosphate (CaP). These were collagen type I, a mussel protein-inspired adhesive consisting of PEG substituted with catechol groups, cPEG, and the PEG-fumaric acid copolymer OPF. After incubation in calcium glycerophosphate (Ca-GP) solution, FTIR, EDS, SEM, XRD, SAED, ICP-OES and von Kossa staining confirmed CaP formation. The amount of mineral formed decreased in the order cPEG > collagen > OPF. Mineral:polymer ratio decreased in the order collagen > cPEG > OPF. Mineralization increased Young’s modulus, most profoundly for cPEG. Such enzymatically mineralized hydrogel-CaP composites could find application as bone regeneration materials.
doi:10.1002/mabi.201100501
PMCID: PMC3690824  PMID: 22648976
enzymes; biomineralization; hydrogels; biomaterials; composites
23.  Synthesis, Material Properties and Biocompatibility of a Novel Self-Crosslinkable Poly(caprolactone fumarate) as an Injectable Tissue Engineering Scaffold 
Biomacromolecules  2005;6(5):2503-2511.
A novel self-crosslinkable and biodegradable macromer poly(caprolactone fumarate) (PCLF) has been developed for guided bone regeneration. This macromer is a copolymer of fumaryl chloride, which contains double bonds for in-situ crosslinking, and poly(ε-caprolactone) that has a flexible chain to facilitate self-crosslinkability. PCLF was characterized with Fourier transform infrared (FTIR) spectroscopy, 1H and 13C nuclear magnetic resonance (NMR) spectroscopy, and gel permeation chromatography (GPC). Porous scaffolds were fabricated with sodium chloride particles as the porogen and a chemical initiation system. The PCLF scaffolds were characterized with scanning electron microscopy (SEM) and micro-computed tomography (micro-CT). The cytotoxicity and in vivo biocompatibility of PCLF were also assessed. Our results suggest that this novel copolymer, PCLF, is an injectable, self-crosslinkable, and biocompatible macromer that may be potentially used as a scaffold for tissue engineering applications.
doi:10.1021/bm050206y
PMCID: PMC2530909  PMID: 16153086
24.  Controlled release of doxorubicin from pH-responsive microgels 
Acta biomaterialia  2012;9(3):5438-5446.
Stimuli-responsive hydrogels have enormous potential in drug delivery applications. They can be used for site-specific drug delivery due to environmental variables in the body such as pH and temperature. In this study, we have developed pH-responsive microgels for the delivery of doxorubicin (DOX) in order to optimize its anti-tumor activity while minimizing its systemic toxicity. We used a copolymer of oligo(polyethylene glycol) fumarate (OPF) and sodium methacrylate (SMA) to fabricate the pH-responsive microgels. We demonstrated that the microgels were negatively charged, and the amounts of charge on the microgels were correlated with the SMA concentration in their formulation. The resulting microgels exhibited sensitivity to the pH and ionic strength of the surrounding environment. We demonstrated that DOX was efficiently loaded into the microgels and released in a controlled fashion via an ion-exchange mechanism. Our data revealed that the DOX release was influenced by the pH and ionic strength of the solution. Moreover, we designed a phenomenological mathematical model, based on a stretched exponential function, to quantitatively analyze the cumulative release of DOX. We found a linear correlation between the maximum release of DOX calculated from the model and the SMA concentration in the microgel formulation. The anti-tumor activity of the released DOX was assessed using a human chordoma cell line. Our data revealed that OPF–SMA microgels prolonged the cell killing effect of DOX.
doi:10.1016/j.actbio.2012.09.019
PMCID: PMC3970914  PMID: 23022545
pH-responsive; Doxorubicin; Microgels; Chordoma; Oligo(polyethylene glycol) fumarate
25.  Nanofibrous Patches for Spinal Cord Regeneration 
Advanced functional materials  2010;20(9):1433-1440.
The difficulty in spinal cord regeneration is related to the inhibitory factors for axon growth and the lack of appropriate axon guidance in the lesion region. Here we developed scaffolds with aligned nanofibers for nerve guidance and drug delivery in spinal cord. Blended polymers including Poly (l-lactic acid) (PLLA) and Poly (lactide-co-glycolide) (PLGA) were used to electrospin nanofibrous scaffolds with two-layer structure: aligned nanofibers in the inner layer and random nanofibers in the outer layer. Rolipram, a small molecule that can enhance cAMP activity in neurons and suppress inflammatory responses, was immobilized onto nanofibers. To test the therapeutic effects of nanofibrous scaffolds, the nanofibrous scaffolds loaded with rolipram were used to bridge the hemisection lesion in 8-week old athymic rats. The scaffolds with rolipram increased axon growth through the scaffolds and in the lesion, promoted angiogenesis through the scaffold, and decreased the population of astrocytes and chondroitin sulfate proteoglycans in the lesion. Locomotor scale rating analysis showed that the scaffolds with rolipram significantly improved hindlimb function after 3 weeks. This study demonstrated that nanofibrous scaffolds offered a valuable platform for drug delivery for spinal cord regeneration.
doi:10.1002/adfm.200901889
PMCID: PMC3558949  PMID: 23378825
Nanofibers; spinal cord; drug delivery; regeneration

Results 1-25 (146059)