Microbial nanocellulose biotextiles for a circular materials economy.

The synthesis and bottom-up assembly of nanocellulose by microbes offers unique advantages to tune and meet key design criteria-rapid renewability, low toxicity, scalability, performance, and degradability-for multi-functional, circular economy textiles. However, development of green processing methods that meet these criteria remains a major research challenge. Here, we harness microbial biofabrication of nanocellulose and draw inspiration from ancient textile techniques to engineer sustainable biotextiles with a circular life cycle. The unique molecular self-organization of microbial nanocellulose (MC) combined with bio-phosphorylation with a lecithin treatment yields a compostable material with superior mechanical and flame-retardant properties. Specifically, treatment of MC with a lecithin-phosphocholine emulsion makes sites available to modulate cellulose cross-linking through hydroxyl, phosphate and methylene groups, increasing the interaction between cellulose chains. The resultant bioleather exhibits enhanced tensile strength and high ductility. Bio-phosphorylation with lecithin also redirects the combustion pathway from levoglucosan production towards the formation of foaming char as an insulating oxygen barrier, for outstanding flame retardance. Controlled color modulation is demonstrated with natural dyes. Life cycle impact assessment reveals that MC bioleather has up to an order of magnitude lower carbon footprint than conventional textiles, and a thousandfold reduction in the carcinogenic impact of leather production. Eliminating the use of hazardous substances, these high performance materials disrupt linear production models and strategically eliminate its toxicity and negative climate impacts, with widespread application in fashion, interiors and construction. Importantly, the biotextile approach developed in this study demonstrates the potential of biofabrication coupled with green chemistry for a circular materials economy.

Nanofiber-based transforming growth factor-ß3 release induces fibrochondrogenic differentiation of stem cells.

Fibrocartilage is typically found in regions subject to complex, multi-axial loads and plays a critical role in musculoskeletal function. Mesenchymal stem cell (MSC)-mediated fibrocartilage regeneration may be guided by administration of appropriate chemical and/or physical cues, such as by culturing cells on polymer nanofibers in the presence of the chondrogenic growth factor TGF-ß3. However, targeted delivery and maintenance of effective local factor concentrations remain challenges for implementation of growth factor-based regeneration strategies in clinical settings. Thus, the objective of this study was to develop and optimize the bioactivity of a biomimetic nanofiber scaffold system that enables localized delivery of TGF-ß3. To this end, we fabricated TGF-ß3-releasing nanofiber meshes that provide sustained growth factor delivery and demonstrated their potential for guiding synovium-derived stem cell (SDSC)-mediated fibrocartilage regeneration. TGF-ß3 delivery enhanced cell proliferation and synthesis of relevant fibrocartilaginous matrix in a dose-dependent manner. By designing a scaffold that eliminates the need for exogenous or systemic growth factor administration and demonstrating that fibrochondrogenesis requires a lower growth factor dose compared to previously reported, this study represents a critical step towards developing a clinical solution for regeneration of fibrocartilaginous tissues. STATEMENT OF SIGNIFICANCE: Fibrocartilage is a tissue that plays a critical role throughout the musculoskeletal system. However, due to its limited self-healing capacity, there is a significant unmet clinical need for more effective approaches for fibrocartilage regeneration. We have developed a nanofiber-based scaffold that provides both the biomimetic physical cues, as well as localized delivery of the chemical factors needed to guide stem cell-mediated fibrocartilage formation. Specifically, methods for fabricating TGF-ß3-releasing nanofibers were optimized, and scaffold-mediated TGF-ß3 delivery enhanced cell proliferation and synthesis of fibrocartilaginous matrix, demonstrating for the first time, the potential for nanofiber-based TGF-ß3 delivery to guide stem cell-mediated fibrocartilage regeneration. This nanoscale delivery platform represents an exciting new strategy for fibrocartilage regeneration.

Estrogen Promotes Mandibular Condylar Fibrocartilage Chondrogenesis and Inhibits Degeneration via Estrogen Receptor Alpha in Female Mice.

Temporomandibular joint degenerative disease (TMJ-DD) is a chronic form of TMJ disorder that specifically afflicts people over the age of 40 and targets women at a higher rate than men. Prevalence of TMJ-DD in this population suggests that estrogen loss plays a role in the disease pathogenesis. Thus, the goal of the present study was to determine the role of estrogen on chondrogenesis and homeostasis via estrogen receptor alpha (ERα) during growth and maturity of the joint. Young and mature WT and ERαKO female mice were subjected to ovariectomy procedures and then given placebo or estradiol treatment. The effect of estrogen via ERα on fibrocartilage morphology, matrix production, and protease activity was assessed. In the young mice, estrogen via ERα promoted mandibular condylar fibrocartilage chondrogenesis partly by inhibiting the canonical Wnt signaling pathway through upregulation of sclerostin (Sost). In the mature mice, protease activity was partly inhibited with estrogen treatment via the upregulation and activity of protease inhibitor 15 (Pi15) and alpha-2-macroglobulin (A2m). The results from this work provide a mechanistic understanding of estradiol on TMJ growth and homeostasis and can be utilized for development of therapeutic targets to promote regeneration and inhibit degeneration of the mandibular condylar fibrocartilage.

Enhanced activation and expansion of T cells using mechanically soft elastomer fibers.

Practical deployment of cellular therapies requires effective platforms for producing clinically relevant numbers of high-quality cells. This report introduces a materials-based approach to improving activation and expansion of T cells, which are rapidly emerging as an agent for treating cancer and a range of other diseases. Electrospinning is used to create a mesh of poly(ε-caprolactone) fibers, which is used to present activating ligands to CD3 and CD28, which activate T cells for expansion. Incorporation of poly(dimethyl siloxane) elastomer into the fibers reduces substrate rigidity and enhances expansion of mixed populations of human CD4+ and CD8+ T cells. Intriguingly, this platform also rescues expansion of T cells isolated from CLL patients, which often show limited responsiveness and other features resembling exhaustion. By simplifying the process of cell expansion, compared to current bead-based platforms, and improving T cell expansion, the system introduced here may accelerate development of cellular immunotherapy.

Designing the stem cell microenvironment for guided connective tissue regeneration.

Adult mesenchymal stem cells (MSCs) are an attractive cell source for regenerative medicine because of their ability to self-renew and their capacity for multilineage differentiation and tissue regeneration. For connective tissues, such as ligaments or tendons, MSCs are vital to the modulation of the inflammatory response following acute injury while also interacting with resident fibroblasts to promote cell proliferation and matrix synthesis. To date, MSC injection for connective tissue repair has yielded mixed results in vivo, likely due to a lack of appropriate environmental cues to effectively control MSC response and promote tissue healing instead of scar formation. In healthy tissues, stem cells reside within a complex microenvironment comprising cellular, structural, and signaling cues that collectively maintain stemness and modulate tissue homeostasis. Changes to the microenvironment following injury regulate stem cell differentiation, trophic signaling, and tissue healing. Here, we focus on models of the stem cell microenvironment that are used to elucidate the mechanisms of stem cell regulation and inspire functional approaches to tissue regeneration. Recent studies in this frontier area are highlighted, focusing on how microenvironmental cues modulate MSC response following connective tissue injury and, more importantly, how this unique cell environment can be programmed for stem cell-guided tissue regeneration.

Advances in biologic augmentation for rotator cuff repair.

Rotator cuff tear is a very common shoulder injury that often necessitates surgical intervention for repair. Despite advances in surgical techniques for rotator cuff repair, there is a high incidence of failure after surgery because of poor healing capacity attributed to many factors. The complexity of tendon-to-bone integration inherently presents a challenge for repair because of a large biomechanical mismatch between the tendon and bone and insufficient regeneration of native tissue, leading to the formation of fibrovascular scar tissue. Therefore, various biological augmentation approaches have been investigated to improve rotator cuff repair healing. This review highlights recent advances in three fundamental approaches for biological augmentation for functional and integrative tendon-bone repair. First, the exploration, application, and delivery of growth factors to improve regeneration of native tissue are discussed. Second, applications of stem cell and other cell-based therapies to replenish damaged tissue for better healing are covered. Finally, this review will highlight the development and applications of compatible biomaterials to both better recapitulate the tendon-bone interface and improve delivery of biological factors for enhanced integrative repair.

Cellular interactions regulate stem cell differentiation in tri-culture.

Currently, the mechanism governing the regeneration of the soft tissue-to-bone interface, such as the transition between the anterior cruciate ligament (ACL) and bone, is not known. Focusing on the ACL-to-bone insertion, this study tests the novel hypothesis that interactions between cells from the ligament (fibroblasts) and bone (osteoblasts) initiate interface regeneration. Specifically, these heterotypic cell interactions direct the fibrochondrogenic differentiation of interface-relevant cell populations, defined here as ligament fibroblasts and bone marrow stromal cells (BMSC). The objective of this study is to examine the effects of heterotypic cellular interactions on BMSC or fibroblast growth and biosynthesis, as well as expression of fibrocartilage-relevant markers in tri-culture. The effects of cell-cell physical contact and paracrine interactions between fibroblasts and osteoblasts were also determined. It was found that, in tri-culture with fibroblasts and osteoblasts, BMSC exhibited greater fibrochondrogenic potential than ligament fibroblasts. The growth of BMSC decreased while proteoglycan production and TGF-ß3 expression increased. Moreover, tri-culture regulated BMSC response via paracrine factors, and interestingly, fibroblast-osteoblast contact further promoted proteoglycan and TGF-ß1 synthesis as well as induced SOX9 expression in BMSC. Collectively, the findings of this study suggest that fibroblast-osteoblast interactions play an important role in regulating the stem cell niche for fibrocartilage regeneration, and the mechanisms of these interactions are directed by paracrine factors and augmented with direct cell-cell contact.

Multilayer scaffolds in orthopaedic tissue engineering.

PURPOSE: The purpose of this study was to summarize the recent developments in the field of tissue engineering as they relate to multilayer scaffold designs in musculoskeletal regeneration. METHODS: Clinical and basic research studies that highlight the current knowledge and potential future applications of the multilayer scaffolds in orthopaedic tissue engineering were evaluated and the best evidence collected. Studies were divided into three main categories based on tissue types and interfaces for which multilayer scaffolds were used to regenerate: bone, osteochondral junction and tendon-to-bone interfaces. RESULTS: In vitro and in vivo studies indicate that the use of stratified scaffolds composed of multiple layers with distinct compositions for regeneration of distinct tissue types within the same scaffold and anatomic location is feasible. This emerging tissue engineering approach has potential applications in regeneration of bone defects, osteochondral lesions and tendon-to-bone interfaces with successful basic research findings that encourage clinical applications. CONCLUSIONS: Present data supporting the advantages of the use of multilayer scaffolds as an emerging strategy in musculoskeletal tissue engineering are promising, however, still limited. Positive impacts of the use of next generation scaffolds in orthopaedic tissue engineering can be expected in terms of decreasing the invasiveness of current grafting techniques used for reconstruction of bone and osteochondral defects, and tendon-to-bone interfaces in near future.

Combined effects of chemical priming and mechanical stimulation on mesenchymal stem cell differentiation on nanofiber scaffolds.

Functional tissue engineering of connective tissues such as the anterior cruciate ligament (ACL) remains a significant clinical challenge, largely due to the need for mechanically competent scaffold systems for grafting, as well as a reliable cell source for tissue formation. We have designed an aligned, polylactide-co-glycolide (PLGA) nanofiber-based scaffold with physiologically relevant mechanical properties for ligament regeneration. The objective of this study is to identify optimal tissue engineering strategies for fibroblastic induction of human mesenchymal stem cells (hMSC), testing the hypothesis that basic fibroblast growth factor (bFGF) priming coupled with tensile loading will enhance hMSC-mediated ligament regeneration. It was observed that compared to the unloaded, as well as growth factor-primed but unloaded controls, bFGF stimulation followed by physiologically relevant tensile loading enhanced hMSC proliferation, collagen production and subsequent differentiation into ligament fibroblast-like cells, upregulating the expression of types I and III collagen, as well as tenasin-C and tenomodulin. The results of this study suggest that bFGF priming increases cell proliferation, while mechanical stimulation of the hMSCs on the aligned nanofiber scaffold promotes fibroblastic induction of these cells. In addition to demonstrating the potential of nanofiber scaffolds for hMSC-mediated functional ligament tissue engineering, this study yields new insights into the interactive effects of chemical and mechanical stimuli on stem cell differentiation.

The guidance of stem cell differentiation by substrate alignment and mechanical stimulation.

Mesenchymal stem cells (MSC) represent a promising and clinically relevant cell source for tissue engineering applications. As such, guiding MSCs toward specific lineages and maintaining these phenotypes have been particularly challenging as the contributions of mechanical, chemical and structural cues to the complex differentiation process are largely unknown. To fully harness the potential of MSCs for regenerative medicine, a systematic investigation into the individual and combined effects of these stimuli is needed. In addition, unlike chemical stimulation, for which temporal and concentration gradients are difficult to control, mechanical stimulation and scaffold-based cues may be relatively more biomimetic and can be applied with greater control to ensure fidelity in MSC differentiation. The objective of this study is to investigate the role of nanofiber matrix alignment and mechanical stimulation on MSC differentiation, focusing on elucidating the relative contribution of each parameter in guided regeneration of functional connective tissues. It is observed that nanofiber alignment directs MSC response to physiological loading and that fibroblastic differentiation requires a combination of physiologically-relevant cell-material interactions in conjunction with mechanical stimulation. Importantly, the results of this study reveal that systemic and readily controllable cues, such as scaffold alignment and optimized mechanical stimulation, are sufficient to drive MSC differentiation, without the need for additional chemical stimuli. Moreover, these findings yield a set of fundamental design rules that can be readily applied to connective tissue regeneration strategies.

Biomimetic scaffold design for functional and integrative tendon repair.

Rotator cuff tears represent the most common shoulder injuries in the United States. The debilitating effect of this degenerative condition coupled with the high incidence of failure associated with existing graft choices underscores the clinical need for alternative grafting solutions. The 2 critical design criteria for the ideal tendon graft would require the graft to not only exhibit physiologically relevant mechanical properties but also be able to facilitate functional graft integration by promoting the regeneration of the native tendon-to-bone interface. Centered on these design goals, this review will highlight current approaches to functional and integrative tendon repair. In particular, the application of biomimetic design principles through the use of nanofiber- and nanocomposite-based scaffolds for tendon tissue engineering will be discussed. This review will begin with nanofiber-based approaches to functional tendon repair, followed by a section highlighting the exciting research on tendon-to-bone interface regeneration, with an emphasis on implementation of strategic biomimicry in nanofiber scaffold design and the concomitant formation of graded multi-tissue systems for integrative soft-tissue repair. This review will conclude with a summary and discussion of future directions.

Orthopedic interface tissue engineering for the biological fixation of soft tissue grafts.

Interface tissue engineering is a promising new strategy aimed at the regeneration of tissue interfaces and ultimately enabling the biological fixation of soft tissue grafts used in orthopedic repair and sports medicine. Many ligaments and tendons with direct insertions into subchondral bone exhibit a complex enthesis consisting of several distinct yet continuous regions of soft tissue, noncalcified fibrocartilage, calcified fibrocartilage, and bone. Regeneration of this multi-tissue interface will be critical for functional graft integration and improving long-term clinical outcome. This review highlights current knowledge of the structure-function relationship at the interface, the mechanism of interface regeneration, and the strategic biomimicry implemented in stratified scaffold design for interface tissue engineering and multi-tissue regeneration. Potential challenges and future directions in this emerging field are also discussed. It is anticipated that interface tissue engineering will lead to the design of a new generation of integrative fixation devices for soft tissue repair, and it will be instrumental for the development of integrated musculoskeletal tissue systems with biomimetic complexity and functionality.

Development of controlled matrix heterogeneity on a triphasic scaffold for orthopedic interface tissue engineering.

Biological fixation of orthopedic soft tissue grafts to bone poses a significant clinical challenge. The clinical success of soft tissue-based grafts for anterior cruciate ligament (ACL) reconstruction is limited by the lack of functional graft integration with subchondral bone. Soft tissues such as the ACL connect to subchondral bone via a complex interface whereby three distinct tissue regions (ligament, fibrocartilage, and bone) work in concert to facilitate load transfer from soft to hard tissue while minimizing stress concentration at the interface. Although a fibrovascular tissue forms at the graft-to-bone interface following surgery, this tissue is nonphysiologic and represents a weak link between the graft and bone. We propose that the re-establishment of the native multi-tissue interface is essential for biological graft fixation. In vivo observations and our in vitro monolayer co-culture results suggest that osteoblast-fibroblast interaction is important for interface regeneration. This study focuses on the design of a triphasic scaffold system mimicking the multi-tissue organization of the native ACL-to-bone interface and the evaluation of osteoblast-fibroblast interactions during three-dimensional co-culture on the triphasic scaffold. We found that the triphasic scaffold supported cell proliferation, migration and phenotypic matrix production while maintaining distinct cellular regions and phase-specific extracellular matrix deposition over time. This triphasic scaffold is designed to guide the eventual reestablishment of an anatomically oriented and mechanically functional fibrocartilage interfacial region directly on biological and synthetic soft tissue grafts. The results of this study demonstrate the feasibility of multi-tissue regeneration on a single scaffold, and the potential of interface tissue engineering to enable the biological fixation of soft tissue grafts to bone.

Polymer-ceramic composite scaffold induces osteogenic differentiation of human mesenchymal stem cells.

One of the design goals of the ideal tissue-engineered bone graft is ostoinductivity, the ability to induce the osteogenic differentiation of mesenchymal stem cells and progenitor cells. In this study, we evaluated the osteoinductive potential of a polymer-ceramic composite in vitro. This composite has been shown to be biodegradable, osteoconductive, and osteointegrative in previous studies. It is hypothesized that this composite will enhance osteoblastic differentiation in human mesenchymal stem cells (hMSCs), and that this inductive potential is substrate-dependent. Human MSCs were cultured on PLGA-BG composite scaffolds and their growth and differentiation were assessed over a four-week period. Composite scaffolds of PLGA and hydroxyapatite (HA), and hMSC cultures treated with osteogenic medium served as controls. It was found that hMSCs grown on PLGA-BG composite scaffolds expressed osteogenic markers without osteogenic media stimulation. In addition, alkaline phosphatase (ALP) activity peaked significantly earlier on the PLGA-BG composite compared to that on the PLGA scaffolds. The findings of this study collectively demonstrate the osteoinductivity of the PLGA-BG composite and its potential as a bone tissue engineering scaffold.

Controlled release of PRP-derived growth factors promotes osteogenic differentiation of human mesenchymal stem cells.

Platelet-rich plasma (PRP) has been gaining increasing popularity in orthopedics and oral and maxillofacial surgery because of its potential efficacy in enhancing bone regeneration. To maximally augment bone healing using PRP and to control the bioavailability of the relevant growth factors, we have designed an alginate hydrogel-based PRP-delivery system. The bioactivity of the growth factors released from PRP carriers was evaluated by determining the ability of these factors to induce osteogenic differentiation of human mesenchymal stem cells (hMSCs). Specifically, monolayers of hMSCs were incubated with the PRP-containing hydrogel carriers over a two-week culture period. Osteoblast-like cells treated with the hydrogel carriers served as controls. The growth and osteogenic differentiation (alkaline phosphatase activity and mineralization) of hMSCs was determined. The results showed that PRP-derived growth factors released from hydrogel carriers stimulated the osteogenic differentiation of hMSCs and most significantly, the cellular response was carrier type-dependent. Future studies will focus on in vitro and in vivo testing of the efficacy of hydrogel-based PRP release systems.

Differential growth factor retention by platelet-rich plasma composites.

PURPOSE: This study evaluates the temporal sequence and growth factor release from platelet-rich plasma (PRP) combined with different bone substitutes (BS), to identify an optimal substrate for extended growth factor retention. MATERIALS AND METHODS: PRP was clotted with bovine thrombin or thrombin receptor activator peptide-6 (TRAP). In addition, PRP was clotted using Allogro (Ceramed, Lakewood, CO), BioGlass (Mo-Sci, Rolla, MN), or BioOss (Osteohealth, Shirley, NY). The effects of media exchange and BS on platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF beta) release were quantified via enzyme-linked immunosorbent assay. RESULTS: At day 1, the thrombin group released 36% more PDGF than the TRAP group and 80% more than the BS groups. At 7 days, PDGF release was the greatest for the TRAP group. PDGF release was minimal for all groups at day 14, with BS groups retaining 60% more PDGF than thrombin clots. Similarly, the thrombin group released the greatest amount of TGF beta (81.4% of the total), whereas TRAP and BS groups released significantly less TGF beta at day 1. Compared with thrombin, TRAP retained 39.2% more TGF beta, whereas BS groups retained even greater levels (Allogro, 54.3%; BioOss, 45.8%; BioGlass, 67.0%). No significant difference in TGF beta release was observed among the substitutes after day 1. The BS groups continued to retain TGF beta after 14 days, whereas all TGF beta in the thrombin clots was depleted. CONCLUSIONS: PRP preparation with thrombin results in a large, immediate release of growth factors that could be lost into the interstitium in vivo. TRAP-BS may prove more efficacious than thrombin in sustaining growth factor levels critical for the cascade of events leading to bone formation.

Activation of platelet-rich plasma using thrombin receptor agonist peptide.

PURPOSE: This study proposes an alternative preparation method of platelet-rich plasma (PRP). Specifically, we compare the use of thrombin receptor agonist peptide-6 (TRAP) and bovine thrombin as a clotting agent in the preparation of PRP. MATERIALS AND METHODS: PRP was prepared by centrifugation and clotted with thrombin or TRAP. In vitro clotting times were monitored as a function of TRAP concentration, and clot retraction was determined by measuring clot diameter over time. Following the optimization of TRAP concentration, experiments were repeated with the addition of several commercially available bone substitutes. The release of PRP-relevant growth factors as a function of PRP preparation was also determined. RESULTS: The most rapid polymerization of PRP takes place with the addition of thrombin, followed by TRAP/Allogro (Ceramed, Lakewood, CO), TRAP/BioGlass (Mo-Sci, Rolla, MN), TRAP/BioOss (Osteohealth, Shirley, NY), and TRAP alone. Thrombin caused considerable clot retraction (43%), whereas TRAP alone resulted in only 15% retraction. TRAP/Allogro, TRAP/BioOss, and TRAP/BioGlass all exhibited minimal retraction (8%). CONCLUSIONS: The use of TRAP to activate clot formation in the preparation of PRP may be a safe alternative to bovine thrombin. It results in an excellent working time and significantly less clot retraction than the currently available methods of PRP production.

Three-dimensional, bioactive, biodegradable, polymer-bioactive glass composite scaffolds with improved mechanical properties support collagen synthesis and mineralization of human osteoblast-like cells in vitro.

In the past decade, tissue engineering-based bone grafting has emerged as a viable alternative to biological and synthetic grafts. The biomaterial component is a critical determinant of the ultimate success of the tissue-engineered graft. Because no single existing material possesses all the necessary properties required in an ideal bone graft, our approach has been to develop a three dimensional (3-D), porous composite of polylactide-co-glycolide (PLAGA) and 45S5 bioactive glass (BG) that is biodegradable, bioactive, and suitable as a scaffold for bone tissue engineering (PLAGA-BG composite). The objectives of this study were to examine the mechanical properties of a PLAGA-BG matrix, to evaluate the response of human osteoblast-like cells to the PLAGA-BG composite, and to evaluate the ability of the composite to form a surface calcium phosphate layer in vitro. Structural and mechanical properties of PLAGA-BG were measured, and the formation of a surface calcium phosphate layer was evaluated by surface analysis methods. The growth and differentiation of human osteoblast-like cells on PLAGA-BG were also examined. A hypothesis was that the combination of PLAGA with BG would result in a biocompatible and bioactive composite, capable of supporting osteoblast adhesion, growth and differentiation, with mechanical properties superior to PLAGA alone. The addition of bioactive glass granules to the PLAGA matrix resulted in a structure with higher compressive modulus than PLAGA alone. Moreover, the PLAGA-BA composite was found to be a bioactive material, as it formed surface calcium phosphate deposits in a simulated body fluid (SBF), and in the presence of cells and serum proteins. The composite supported osteoblast-like morphology, stained positively for alkaline phosphatase, and supported higher levels of Type I collagen synthesis than tissue culture polystyrene controls. We have successfully developed a degradable, porous, polymer bioactive glass composite possessing improved mechanical properties and osteointegrative potential compared to degradable polymers of poly(lactic acid-glycolic acid) alone. Future work will focus on the optimization of the composite scaffold for bone tissue-engineering applications and the evaluation of the 3-D composite in an in vivo model.

Silk matrix for tissue engineered anterior cruciate ligaments.

A silk-fiber matrix was studied as a suitable material for tissue engineering anterior cruciate ligaments (ACL). The matrix was successfully designed to match the complex and demanding mechanical requirements of a native human ACL, including adequate fatigue performance. This protein matrix supported the attachment, expansion and differentiation of adult human progenitor bone marrow stromal cells based on scanning electron microscopy, DNA quantitation and the expression of collagen types I and III and tenascin-C markers. The results support the conclusion that properly prepared silkworm fiber matrices, aside from providing unique benefits in terms of mechanical properties as well as biocompatibility and slow degradability, can provide suitable biomaterial matrices for the support of adult stem cell differentiation toward ligament lineages. These results point toward this matrix as a new option for ACL repair to overcome current limitations with synthetic and other degradable materials.

Integrin expression by human osteoblasts cultured on degradable polymeric materials applicable for tissue engineered bone.

The use of biodegradable polymers in the field of orthopaedic surgery has gained increased popularity, as surgical pins and screws, and as potential biological scaffolds for repairing cartilage and bone defects. One such group of polymers that has gained considerable attention are the polyesters, poly(lactide-co-glycolide) (PLAGA) and polylactic acid (PLA), because of their minimal tissue inflammatory response, favorable biocompatibility and degradation characteristics. The objective of this study was to evaluate human osteoblastic cell adherence and growth on PLAGA and PLA scaffolds by examining integrin receptor (alpha2, alpha3, alpha4, alpha5, alpha6 and beta1) expression. Primary human osteoblastic cells isolated from trabecular bone adhered efficiently to both PLAGA and PLA, with the rate of adherence on PLAGA comparable to that of control tissue culture polystyrene (TCPS), and significantly higher than on PLA polymers at 3, 6 and 12 h. Human osteoblastic phenotypic expression, alkaline phosphatase (ALP) activity was positive on both degradable matrices, whereas osteocalcin levels were significantly higher on cells grown on PLAGA than on PLA composites. Interestingly, the integrin subunits, alpha2, alpha3, alpha4, alpha5, alpha6 and beta1 were all expressed at higher levels by osteoblasts cultured on PLAGA than those on PLA as analyzed by westerns blots and by flow cytometry. Among the integrins, alpha2, beta5 and beta1 showed the greatest difference in levels between the two surfaces. Thus, both PLA and PLAGA support osteoblastic adhesion and its accompanying engagement of integrin receptor and expression of osteocalcin and ALP. However PLAGA consistently appeared to be a better substrate for osteoblastic cells based on these parameters. This study is one of the first to investigate the ability of primary human osteoblastic cells isolated from trabecular bone to adhere to the biodegradable polymers PLAGA and PLA, and to examine the expression of their key adhesion receptors (integrins) on these substrates.