Tissue Engineering Strategies in Periodontal Regeneration: an Update

Introduction

Periodontium consists of fibrous periodontal ligament which attaches cementum of the tooth to the alveolar bone. The bulk of Periodontal ligament contains neurovascular elements. Cyclic masticatory forces between two mineralized bodies are distributed by the relative motion between the tooth and bone with the help of this soft tissue. These short term physiological forces allow continuous adaptation of the bone-PDL-cementum complex.

In particular, angulated PDLs with spatiotemporal organizations between the teeth and the alveolar bone significantly contribute to masticatory/occlusal stress absorptions and distributions, as well as the optimization of mineralized tissue remodeling for tooth-periodontium complexes . Therefore, perpendicular/oblique PDL orientations to the tooth-root surfaces add to the functionalization and revitalization of tooth-supportive biofunctional structures. Spatiotemporal compartmentalization is a critical requirement for micron-scaled multiple tissue regeneration and functional restoration. However instabilities in multiple tissue interfaces or the loss of their skeletal-supportive functions can be induced by diseases or traumatic injuries of the musculoskeletal systems.

Periodontitis, a highly prevalent inflammatory infectious disease, commonly induces tissue destruction of the periodontal complex in humans. This disease is initiated by bacterial products such as lipopolysaccharide (LPS), which can stimulate cytokines to signal precursor cells to differentiate and activate osteoclastic cells and/or the periodontal inflammatory process by bacterial biofilm. The therapeutic knowledge is currently limited to sub-micron-scaled interfaces and systemic compartmentalization to mimic periodontal structures and functions for the re-establishment of tooth-supportive functions.

This brief review provides importance of 3D printing techniques & approaches in regenerating ligamant-bone complexes by regulating spatiotemporal cell organizations. Some techniques currently being used to produce scaffolds are direct 3D printing, fused deposition modelling, stereolithography, selective laser sintering,etc. Advantages of using 3D printing include the ability to fabricate versatile scaffolds with complex shapes capable of homogenous cell distribution, and the ability to imitate the extracellular matrix (ECM). However, the availability of biomaterials with the stability and desired properties for 3D printing of scaffolds is restricted depending on the printing technology used. Another disadvantage is the production time that it takes to fabricate scaffolds, which greatly increases as the scaffold design becomes more and more precise and intricate.

Methods and Strategies

Heat-Mediated 3D Fabrication

Fabrication by heat energy combines pre-fabricated polymer layers into simple three dimensional structures by raising the polymer above its glass transition temperature and fusing the softened layers together with applied pressure.11 It includes several techniques like selective laser sintering, fused deposition modelling, 3-D plotting, etc.

Selective Laser Sintering/melting

University of Texas in 1989 developed Selective laser sintering (SLS) technique. CO2 laser beam is used in this technique which selectively fuses powdered material by scanning cross-sections generated from a 3D digital description of the part on the surface of a powder bed. After each cross-section is scanned, the powder bed is lowered by one layer thickness, a new layer of material is applied on top, and the process is repeated until the part is completed.

The integration of computational design and SLS techniques enables the ability to fabricate scaffolds that have anatomically shaped external architectures and porous interior structure. FDA clearance was recently awarded for the use of SLS to process medical grade polyether ether ketone (PEEK) to make custom craniofacial implants. More recently, SLM was used to create the first patient-specific, ready for implantation titanium mandible that accepts dental implants to support a mandibular denture.13

Fused Deposition Modeling(FDM)

This technique uses a moving nozzle to extrude a fiber of polymeric material from which the physical model is built layer by layer. Polylactic Acid (PLA) is currently applied in FDM mainly due to its biocompatibility and good thermal & physical properties. When primary human fibroblasts were cultured in these scaffolds, they proliferated and produced extracellular matrix14, Hutmacher et al. evaluated compressive strength of each printed group and it was compatible with that of human cancellous bone.

While FDM exhibits high pattern resolution in the xy-plane, it is limited in the z-direction by the diameter of the extruded polymer filament that defines layer thickness and corresponding pore height. Further, high processing temperatures limit the biomaterials that are compatible with the method. However, FDM capabilities are expanding with new developments such as multi-phase jet solidification (MJS), a technique that allows simultaneous extrusion of multiple melted materials.16

Light Mediated Fabrication

An UV laser is used to solidify the exposed polymer regions while leaving the remaining areas in liquid form. The movable table then drops by a sufficient amount to cover the solid polymer with another layer of liquid resin. The process is repeated to create the desired shape. As with SLS, stereolithography is limited in resolution by laser beam diameter to approximately 250 μm, although small-spot laser systems have demonstrated the production of smaller (70 μm) features.17 Two different methods of irradiation may be applied to stereolithography, laser-based stereolithography and digital light projection stereolithography.

The laser-based method is a direct write approach in which a computer-manipulated laser beam fabricates structures in a vector-by-vector, bottom-up manner. In digital light projection, the UV light source is projected on a transparent surface at the bottom of a vat, which holds the photosensitive resin; an entire layer of material is simultaneously polymerized upon light exposure.

In initial attempts involving this approach, a physical mask was applied to define the specific pattern to be illuminated during light projection stereolithography.18 Stereolithography enables significant freedom of design and is capable of fabricating minimum features sizes on the micrometer scale; although some stereolithography systems are capable of preparing structures with ≤5 µm features, most commercial systems prepare structures with ≥50 µm features.19

Pressure Assisted Microsyringe

The pressure-assisted microsyringe (PAM) technique was developed at the Interdepartmental Research Centre “E.Piaggio” at the University of Pisa. It is based on the use of microsyringe that allows the deposition of a wide range of polymers, as well as hydrogels. A stage controlled microsyringe delivery system deposits a stream of polymer dissolved in solvent through a 10–20 μm glass capillary needle.23

Adhesive –Mediated Fabrication

3-D printing

The first report using 3-D printing technology for periodontal tissue regeneration demonstrated that this technology represents a promising approach to the design and manufacture of complicated, unpredictable geometries.9

The main advantage of this technique is its ability to produce an implant directly from 3D data in one step without using an additional mould. The matrices generated by 3D printing are seeded with patient-derived cells and eventually implanted into the body. Besides ceramics, scaffolds from polymers can be fabricated with 3D printing process.20

In addition to preclinical studies, a human case study using a patient-specific scaffold manufactured by 3D printing attempted to treat a large periodontal defect and to regenerate periodontal complexes (bone-PDL-cementum),21 Chen Ho Park et al demonstrated that the different angulated microgroove patterns on 3D printed scaffolds can control the orientation of ligamentous cell bundles with high manufacturing reproducibility. This simple strategy provides the topographical platform to precisely form functional architectures for 3D organizations of fibrous connective tissues.3

One interesting derivation of 3D bioprinting is the integration of microelectronic and mechatronic components. For example, a bio-bot is a walking robot powered by the contraction of a strip of mammalian skeletal muscle cells24

Materials

3DP materials include calcium polyphosphate and PVA , HA and TCP , TCP , TCP with SrO and MgO doping , HA and apatite–wollastonite glass ceramic with water-based binder , calcium phosphate with collagen in binder , PLGA , and Farringtonite powder (Mg3(PO4)2) . Materials used in indirect 3DP gelatin preforms replaced with PCL and chitosan .22

Specific Forms Of Materials And Suitable 3D-Printing Processes.24

Form Examples 3D-printing processes

Solidifiable fluid Photopolymer resins, temperature sensitive polymers, ion cross-linkable hydrogels, ceramic paste, etc. Stereolithography (SLA)

Polyjet

Digital light processing (DLP)

Micro-extrusion

Non-brittle filament Thermoplastics, e.g., ABS, PLA, and PCL Fused deposition modeling (FDM)

Laminated thin sheet Paper, Plastic sheet,

Metal foil Paper lamination technology (PLT)

Laminated object manufacturing (LOM) Ultrasonic consolidation (UC)

Fine powder Plastic fine powder, ceramic powder, Selective laser sintering/melting (SLS/SLM) Electron beam melting (EBM) Laser engineered net shaping (LENS) [40] Direct metal deposition (DMD) Colorjet printing (CJP)

Notes: ABS—acrylonitrile-butadiene-styrene; PLA—polylactic acid; PCL—polycaprolactone.

Future Perspectives

It is still a long way from transformation of theoretical knowledge to implement it into clinical practice. A prolonged delay in this standardization would make regulatory work even more complicated, especially with the currently trending and transforming 3D bioprinting technologies, because the definition of “medical device” may soon be redefined.24 In a nutshell, proper computer aided system with newer & emerging techniques can change the future of dental system.