Tissue-specific melt electrowritten polymeric scaffolds for coordinated regeneration of soft and hard periodontal tissues

Periodontitis is a chronic inflammatory condition that often causes serious damage to tooth-supporting tissues. The limited successful outcomes of clinically available approaches underscore the need for therapeutics that cannot only provide structural guidance to cells but can also modulate the local immune response. Here, three-dimensional melt electrowritten ( i.e., poly(ε-caprolactone)) scaffolds with tissue-specific attributes were engineered to guide differentiation of human-derived periodontal ligament stem cells (hPDLSCs) and mediate macrophage polarization. The investigated tissue-specific scaffold attributes comprised fiber morphology (aligned vs. random) and highly-ordered architectures with distinct strand spacings (small 250 μm and large 500 μm). Macrophages exhibited an elongated morphology in aligned and highly-ordered scaffolds, while maintaining their round-shape on randomly-oriented fibrous scaffolds. Expressions of periostin and IL-10 were more pronounced on the aligned and highly-ordered scaffolds. While hPDLSCs on the scaffolds with 500 μm strand spacing show higher expression of osteogenic marker (Runx2) over 21 days, cells on randomly-oriented fibrous scaffolds showed upregulation of M1 markers. In an orthotopic mandibular fenestration defect model, findings revealed that the tissue-specific scaffolds ( i.e., aligned fibers for periodontal ligament and highly-ordered 500 μm strand spacing fluorinated calcium phosphate [F/CaP]-coated fibers for bone) could enhance the mimicking of regeneration of natural periodontal tissues

Personalized and Tissue-Specific Melt Electrowritten Scaffolds for Coordinated Regeneration of Soft and Hard Periodontal Tissues

Periodontitis is a chronic inflammatory, bacteria-triggered disorder affecting nearly half of American adults. If left untreated, it leads to severe destruction of the periodontium, i.e., the alveolar bone, periodontal ligament (PDL), and cementum, ultimately leading to tooth loss. Current clinical therapeutic approaches include subgingival scaling, periodontal flap surgery, guided tissue regeneration (GTR), and guided bone regeneration (GBR).1 The formation of new bone, cementum, and periodontal ligament (PDL) is a possible objective of these modalities; however, outcomes are not always predictable, due to varying degrees of destruction and highly complex tissue architecture. This underscores the urgent need for superior strategies to amplify regenerative capacity, regardless of damage severity. Melt electrowriting (MEW) is a novel strategy that allows customization to the desired size, configuration, and architecture of a given scaffold. It allows for the production of consistent fiber morphology and diameter approaching submicron magnitudes, which, in turn, allows for the recapitulation of the tissue-specific cell microenvironment. Thus, a MEW-based approach to form fiber-guiding scaffold would offer a geometric organization for bone-PDL interface. In this dissertation work, we show the potential of highly ordered scaffolds engineered via melt electrowriting (MEW) in guiding soft (PDL) and hard (bone) periodontal tissue regeneration. Our in vitro findings show that the presence of the aligned fibers result in a robust expression of ligamentogenic markers, while the strands’ spacing of MEW constructs play a vital role in the upregulation of osteogenic markers. Moreover, in vitro findings strongly suggest that aligned and porous MEW constructs promote macrophage elongation and further polarization toward M2 macrophage, the prohealing phenotype. Meanwhile, the incorporation of osteoconductive fluorinated calcium phosphate coating, has been shown to positively impact mineralized tissue formation. The fiber morphology of F/CaP-coated scaffolds revealed a unique nanostructured surface depicted as irregular-shaped nanoparticles (~ 50-150 nm) and a homogenous rough layer covering each individual fiber. The presence of a nanostructured F/CaP coating led to a marked upregulation of osteogenic genes and attenuated bacterial growth. The reduced bacterial growth, even with a rougher F/CaP coated surface, suggests an antimicrobial action due to the coating’s composition. Collectively, to regenerate tissue-specific architecture and recapitulate the physiological function of native periodontal tissues, a scaffold with tailored properties and bioactive functional was developed. Altogether, our findings confirmed that the tissue-specific scaffolds with F/CaP-coated, bone compartment, and aligned PDL compartment are biocompatible and lead to periodontal tissue regeneration when implanted in a well-established rat mandibular periodontal fenestration defect modelPHDOral Health SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/169852/1/daghrery_1.pd

Advanced biomaterials for periodontal tissue regeneration

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/174945/1/dvg23501_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/174945/2/dvg23501.pd

Unveiling the potential of melt electrowriting in regenerative dental medicine

For nearly three decades, tissue engineering strategies have been leveraged to devise effective therapeutics for dental, oral, and craniofacial (DOC) regenerative medicine and treat permanent deformities caused by many debilitating health conditions. In this regard, additive manufacturing (AM) allows the fabrication of personalized scaffolds that have the potential to recapitulate native tissue morphology and biomechanics through the utilization of several 3D printing techniques. Among these, melt electrowriting (MEW) is a versatile direct electrowriting process that permits the development of well-organized fibrous constructs with fiber resolutions ranging from micron to nanoscale. Indeed, MEW offers great prospects for the fabrication of scaffolds mimicking tissue specificity, healthy and pathophysiological microenvironments, personalized multi-scale transitions, and functional interfaces for tissue regeneration in medicine and dentistry. Excitingly, recent work has demonstrated the potential of converging MEW with other AM technologies and/or cell-laden scaffold fabrication (bioprinting) as a favorable route to overcome some of the limitations of MEW for DOC tissue regeneration. In particular, such convergency fabrication strategy has opened great promise in terms of supporting multi-tissue compartmentalization and predetermined cell commitment. In this review, we offer a critical appraisal on the latest advances in MEW and its convergence with other biofabrication technologies for DOC tissue regeneration. We first present the engineering principles of MEW and the most relevant design aspects for transition from flat to more anatomically relevant 3D structures while printing highly-ordered constructs. Secondly, we provide a thorough assessment of contemporary achievements using MEW scaffolds to study and guide soft and hard tissue regeneration, and draw a parallel on how to extrapolate proven concepts for applications in DOC tissue regeneration. Finally, we offer a combined engineering/clinical perspective on the fabrication of hierarchically organized MEW scaffold architectures and the future translational potential of site-specific, single-step scaffold fabrication to address tissue and tissue interfaces in dental, oral, and craniofacial regenerative medicine. Statement of significance: Melt electrowriting (MEW) techniques can further replicate the complexity of native tissues and could be the foundation for novel personalized (defect-specific) and tissue-specific clinical approaches in regenerative dental medicine. This work presents a unique perspective on how MEW has been translated towards the application of highly-ordered personalized multi-scale and functional interfaces for tissue regeneration, targeting the transition from flat to anatomically-relevant three-dimensional structures. Furthermore, we address the value of convergence of biofabrication technologies to overcome the traditional manufacturing limitations provided by multi-tissue complexity. Taken together, this work offers abundant engineering and clinical perspectives on the fabrication of hierarchically MEW architectures aiming towards site-specific implants to address complex tissue damage in regenerative dental medicine

Highly tunable bioactive fiber-reinforced hydrogel for guided bone regeneration

One of the most damaging pathologies that affects the health of both soft and hard tissues around the tooth is periodontitis. Clinically, periodontal tissue destruction has been managed by an integrated approach involving elimination of injured tissues followed by regenerative strategies with bone substitutes and/or barrier membranes. Regrettably, a barrier membrane with predictable mechanical integrity and multifunctional therapeutic features has yet to be established. Herein, we report a fiber-reinforced hydrogel with unprecedented tunability in terms of mechanical competence and therapeutic features by integration of highly porous poly(ε-caprolactone) fibrous mesh(es) with well-controlled 3D architecture into bioactive amorphous magnesium phosphate-laden gelatin methacryloyl hydrogels. The presence of amorphous magnesium phosphate and PCL mesh in the hydrogel can control the mechanical properties and improve the osteogenic ability, opening a tremendous opportunity in guided bone regeneration (GBR). Results demonstrate that the presence of PCL meshes fabricated via melt electrowriting can delay hydrogel degradation preventing soft tissue invasion and providing the mechanical barrier to allow time for slower migrating progenitor cells to participate in bone regeneration due to their ability to differentiate into bone-forming cells. Altogether, our approach offers a platform technology for the development of the next-generation of GBR membranes with tunable mechanical and therapeutic properties to amplify bone regeneration in compromised sites. STATEMENT OF SIGNIFICANCE: In this study, we developed a fiber-reinforced hydrogel platform with unprecedented tunability in terms of mechanical competence and therapeutic features for guided bone regeneration. We successfully integrated highly porous poly(ε-caprolactone) [PCL] mesh(es) into amorphous magnesium phosphate-laden hydrogels. The stiffness of the engineered hydrogel was significantly enhanced, and this reinforcing effect could be modulated by altering the number of PCL meshes and tailoring the AMP concentration. Furthermore, the fiber-reinforced hydrogel showed favorable cellular responses, significantly higher rates of mineralization, upregulation of osteogenic-related genes and bone formation. In sum, these fiber-reinforced membranes in combination with therapeutic agent(s) embedded in the hydrogel offer a robust, highly tunable platform to amplify bone regeneration not only in periodontal defects, but also in other craniomaxillofacial sites

Innovations in craniofacial bone and periodontal tissue engineering–from electrospinning to converged biofabrication

From a materials perspective, the pillars for the development of clinically translatable scaffold-based strategies for craniomaxillofacial (CMF) bone and periodontal regeneration have included electrospinning and 3D printing (biofabrication) technologies. Here, we offer a detailed analysis of the latest innovations in 3D (bio)printing strategies for CMF bone and periodontal regeneration and provide future directions envisioning the development of advanced 3D architectures for successful clinical translation. First, the principles of electrospinning applied to the generation of biodegradable scaffolds are discussed. Next, we present on extrusion-based 3D printing technologies with a focus on creating scaffolds with improved regenerative capacity. In addition, we offer a critical appraisal on 3D (bio)printing and multitechnology convergence to enable the reconstruction of CMF bones and periodontal tissues. As a future outlook, we highlight future directions associated with the utilisation of complementary biomaterials and (bio)fabrication technologies for effective translation of personalised and functional scaffolds into the clinics

Self-assembling peptide-laden electrospun scaffolds for guided mineralized tissue regeneration

OBJECTIVES: Electrospun scaffolds are a versatile biomaterial platform to mimic fibrillar structure of native tissues extracellular matrix, and facilitate the incorporation of biomolecules for regenerative therapies. Self-assembling peptide P 11-4 has emerged as a promising strategy to induce mineralization; however, P 11-4 application has been mostly addressed for early caries lesions repair on dental enamel. Here, to investigate P 11-4's efficacy on bone regeneration, polymeric electrospun scaffolds were developed, and then distinct concentrations of P 11-4 were physically adsorbed on the scaffolds. METHODS: P 11-4-laden and pristine (P 11-4-free) electrospun scaffolds were immersed in simulated body fluid and mineral precipitation identified by SEM. Functional groups and crystalline phases were analyzed by FTIR and XRD, respectively. Cytocompatibility, mineralization, and gene expression assays were conducted using stem cells from human exfoliated deciduous teeth. To investigate P 11-4-laden scaffolds potential to induce in vivo mineralization, an established rat calvaria critical-size defect model was used. RESULTS: We successfully synthesized nanofibrous (∼ 500 nm fiber diameter) scaffolds and observed that functionalization with P 11-4 did not affect the fibers' diameter. SEM images indicated mineral precipitation, while FTIR and XRD confirmed apatite-like formation and crystallization for P 11-4-laden scaffolds. In addition, P 11-4-laden scaffolds were cytocompatible, highly stimulated cell-mediated mineral deposition, and upregulated the expression of mineralization-related genes compared to pristine scaffolds. P 11-4-laden scaffolds led to enhanced in vivo bone regeneration after 8 weeks compared to pristine PCL. SIGNIFICANCE: Electrospun scaffolds functionalized with P 11-4 are a promising strategy for inducing mineralized tissues regeneration in the craniomaxillofacial complex

Nanoscale β-TCP-Laden GelMA/PCL Composite Membrane for Guided Bone Regeneration

Major advances in the field of periodontal tissue engineering have favored the fabrication of biodegradable membranes with tunable physical and biological properties for guided bone regeneration (GBR). Herein, we engineered innovative nanoscale beta-tricalcium phosphate (β-TCP)-laden gelatin methacryloyl/polycaprolactone (GelMA/PCL-TCP) photocrosslinkable composite fibrous membranes via electrospinning. Chemo-morphological findings showed that the composite microfibers had a uniform porous network and β-TCP particles successfully integrated within the fibers. Compared with pure PCL and GelMA/PCL, GelMA/PCL-TCP membranes led to increased cell attachment, proliferation, mineralization, and osteogenic gene expression in alveolar bone-derived mesenchymal stem cells (aBMSCs). Moreover, our GelMA/PCL-TCP membrane was able to promote robust bone regeneration in rat calvarial critical-size defects, showing remarkable osteogenesis compared to PCL and GelMA/PCL groups. Altogether, the GelMA/PCL-TCP composite fibrous membrane promoted osteogenic differentiation of aBMSCs in vitro and pronounced bone formation in vivo. Our data confirmed that the electrospun GelMA/PCL-TCP composite has a strong potential as a promising membrane for guided bone regeneration

Highly tunable bioactive fiber-reinforced hydrogel for guided bone regeneration

One of the most damaging pathologies that affects the health of both soft and hard tissues around the tooth is periodontitis. Clinically, periodontal tissue destruction has been managed by an integrated approach involving elimination of injured tissues followed by regenerative strategies with bone substitutes and/or barrier membranes. Regrettably, a barrier membrane with predictable mechanical integrity and multifunctional therapeutic features has yet to be established. Herein, we report a fiber-reinforced hydrogel with unprecedented tunability in terms of mechanical competence and therapeutic features by integration of highly porous poly(ε-caprolactone) fibrous mesh(es) with well-controlled 3D architecture into bioactive amorphous magnesium phosphate-laden gelatin methacryloyl hydrogels. The presence of amorphous magnesium phosphate and PCL mesh in the hydrogel can control the mechanical properties and improve the osteogenic ability, opening a tremendous opportunity in guided bone regeneration (GBR). Results demonstrate that the presence of PCL meshes fabricated via melt electrowriting can delay hydrogel degradation preventing soft tissue invasion and providing the mechanical barrier to allow time for slower migrating progenitor cells to participate in bone regeneration due to their ability to differentiate into bone-forming cells. Altogether, our approach offers a platform technology for the development of the next-generation of GBR membranes with tunable mechanical and therapeutic properties to amplify bone regeneration in compromised sites. STATEMENT OF SIGNIFICANCE: In this study, we developed a fiber-reinforced hydrogel platform with unprecedented tunability in terms of mechanical competence and therapeutic features for guided bone regeneration. We successfully integrated highly porous poly(ε-caprolactone) [PCL] mesh(es) into amorphous magnesium phosphate-laden hydrogels. The stiffness of the engineered hydrogel was significantly enhanced, and this reinforcing effect could be modulated by altering the number of PCL meshes and tailoring the AMP concentration. Furthermore, the fiber-reinforced hydrogel showed favorable cellular responses, significantly higher rates of mineralization, upregulation of osteogenic-related genes and bone formation. In sum, these fiber-reinforced membranes in combination with therapeutic agent(s) embedded in the hydrogel offer a robust, highly tunable platform to amplify bone regeneration not only in periodontal defects, but also in other craniomaxillofacial sites