Extrusion-based 3D Bioprinting of Bioactive and Piezoelectric Scaffolds for Treating Critical Soft Tissue Wounds.

OBJECTIVE: This study focuses on developing bioactive piezoelectric scaffolds that could deliver bioelectrical cues to potentially treat injuries to soft tissues such as skeletal muscles and promote muscle regeneration. APPROACH: To address the underexplored aspect of bioelectrical cues in skeletal muscle tissue engineering (SMTE), we developed piezoelectric bioinks based on natural bioactive materials such as alginate, gelatin, and chitosan. Extrusion-based 3D bioprinting was utilized to develop scaffolds that mimic muscle stiffness and generate electrical stimulation when subjected to forces. The biocompatibility of these scaffolds was tested with C2C12 muscle cell line. RESULTS: The bioinks demonstrated suitable rheological properties for 3D bioprinting, resulting in high-resolution composite alginate-gelatin-chitosan scaffolds with good structural fidelity. The scaffolds exhibited a 42-60 kPa stiffness, similar to muscles. When a controlled force of 5 N was applied to the scaffolds at a constant frequency of 4 Hz, they generated electrical fields and impulses (charge), indicating their suitability as a standalone scaffold to generate electrical stimulation and instill bioelectrical cues in the wound region. The cell viability and proliferation test results confirm the scaffold's biocompatibility with C2C12s and the benefit of piezoelectricity in promoting muscle cell growth kinetics. Our study indicates that our piezoelectric bioinks and scaffolds offer promise as autonomous electrical stimulation-generating regenerative therapy for SMTE. INNOVATION: A novel approach for treating skeletal muscle wounds was introduced by developing a bioactive electroactive scaffold capable of autonomously generating electrical stimulation without stimulators and electrodes. This scaffold offers a unique approach to enhancing skeletal muscle regeneration through bioelectric cues, addressing a major gap in the SMTE, i.e., fibrotic tissue formation due to delayed muscle regeneration. CONCLUSION: A piezoelectric scaffold was developed, providing a promising solution for promoting skeletal muscle regeneration. This development can potentially address skeletal muscle injuries and offer a unique approach to facilitating skeletal muscle wound healing.

High-Density Porous Polyethylene Implant Cranioplasty: A Systematic Review of Outcomes.

Porous polyethylene has been widely used in craniofacial reconstruction due to its biomechanical properties and ease of handling. The objective of this study was to perform a systematic review of the literature to summarize outcomes utilizing high-density porous polyethylene (HDPP) implants in cranioplasty. A literature search of PubMed, Cochrane Library, and Scopus databases was conducted to identify original studies with HDPP cranioplasty from inception to March 2023. Non-English articles, commentaries, absent indications or outcomes, and nonclinical studies were excluded. Data on patient demographics, indications, defect size and location, outcomes, and patient satisfaction were extracted. Summary statistics were calculated using weighted averages based on the available reported data. A total of 1089 patients involving 1104 cranioplasty procedures with HDPP were identified. Patients' mean age was 44.0 years (range 2 to 83 y). The mean follow-up duration was 32.0 months (range 2 wk to 8 y). Two studies comprising 17 patients (1.6%) included only pediatric patients. Alloplastic cranioplasty was required after treatment of cerebrovascular diseases (50.9%), tumor excision (32.0%), trauma (11.4%), trigeminal neuralgia/epilepsy (3.4%), and others such as abscesses/cysts (1.4%). The size of the defect ranged from 3 to 340 cm 2 . An overall postoperative complication rate of 2.3% was identified, especially in patients who had previously undergone surgery at the same site. When data were available, contour improvement and high patient satisfaction were reported in 98.8% and 98.3% of the patients. HDPP implants exhibit favorable outcomes for reconstruction of skull defects. Higher complication rates may be anticipated in secondary cranioplasty cases.

Extrusion 3D (Bio)Printing of Alginate-Gelatin-Based Composite Scaffolds for Skeletal Muscle Tissue Engineering.

Volumetric muscle loss (VML), which involves the loss of a substantial portion of muscle tissue, is one of the most serious acute skeletal muscle injuries in the military and civilian communities. The injured area in VML may be so severely affected that the body loses its innate capacity to regenerate new functional muscles. State-of-the-art biofabrication methods such as bioprinting provide the ability to develop cell-laden scaffolds that could significantly expedite tissue regeneration. Bioprinted cell-laden scaffolds can mimic the extracellular matrix and provide a bioactive environment wherein cells can spread, proliferate, and differentiate, leading to new skeletal muscle tissue regeneration at the defect site. In this study, we engineered alginate−gelatin composite inks that could be used as bioinks. Then, we used the inks in an extrusion printing method to develop design-specific scaffolds for potential VML treatment. Alginate concentration was varied between 4−12% w/v, while the gelatin concentration was maintained at 6% w/v. Rheological analysis indicated that the alginate−gelatin inks containing 12% w/v alginate and 6% w/v gelatin were most suitable for developing high-resolution scaffolds with good structural fidelity. The printing pressure and speed appeared to influence the printing accuracy of the resulting scaffolds significantly. All the hydrogel inks exhibited shear thinning properties and acceptable viscosities, though 8−12% w/v alginate inks displayed properties ideal for printing and cell proliferation. Alginate content, crosslinking concentration, and duration played significant roles (p < 0.05) in influencing the scaffolds' stiffness. Alginate scaffolds (12% w/v) crosslinked with 300, 400, or 500 mM calcium chloride (CaCl2) for 15 min yielded stiffness values in the range of 45−50 kPa, i.e., similar to skeletal muscle. The ionic strength of the crosslinking concentration and the alginate content significantly (p < 0.05) affected the swelling and degradation behavior of the scaffolds. Higher crosslinking concentration and alginate loading enhanced the swelling capacity and decreased the degradation kinetics of the printed scaffolds. Optimal CaCl2 crosslinking concentration (500 mM) and alginate content (12% w/v) led to high swelling (70%) and low degradation rates (28%) of the scaffolds. Overall, the results indicate that 12% w/v alginate and 6% w/v gelatin hydrogel inks are suitable as bioinks, and the printed scaffolds hold good potential for treating skeletal muscle defects such as VML.

3D-Printed Piezoelectric Porous Bioactive Scaffolds and Clinical Ultrasonic Stimulation Can Help in Enhanced Bone Regeneration.

This paper presents a comprehensive effort to develop and analyze first-of-its-kind design-specific and bioactive piezoelectric scaffolds for treating orthopedic defects. The study has three major highlights. First, this is one of the first studies that utilize extrusion-based 3D printing to develop design-specific macroporous piezoelectric scaffolds for treating bone defects. The scaffolds with controlled pore size and architecture were synthesized based on unique composite formulations containing polycaprolactone (PCL) and micron-sized barium titanate (BaTiO3) particles. Second, the bioactive PCL-BaTiO3 piezoelectric composite formulations were explicitly developed in the form of uniform diameter filaments, which served as feedstock material for the fused filament fabrication (FFF)-based 3D printing. A combined method comprising solvent casting and extrusion (melt-blending) was designed and deemed suitable to develop the high-quality PCL-BaTiO3 bioactive composite filaments for 3D printing. Third, clinical ultrasonic stimulation (US) was used to stimulate the piezoelectric effect, i.e., create stress on the PCL-BaTiO3 scaffolds to generate electrical fields. Subsequently, we analyzed the impact of scaffold-generated piezoelectric stimulation on MC3T3 pre-osteoblast behavior. Our results confirmed that FFF could form high-resolution, macroporous piezoelectric scaffolds, and the poled PCL-BaTiO3 composites resulted in the d33 coefficient in the range of 1.2-2.6 pC/N, which is proven suitable for osteogenesis. In vitro results revealed that the scaffolds with a mean pore size of 320 µm resulted in the highest pre-osteoblast growth kinetics. While 1 Hz US resulted in enhanced pre-osteoblast adhesion, proliferation, and spreading, 3 Hz US benefited osteoblast differentiation by upregulating important osteogenic markers. This study proves that 3D-printed bioactive piezoelectric scaffolds coupled with US are promising to expedite bone regeneration in orthopedic defects.

Patient-specific 3D printed Poly-ether-ether-ketone (PEEK) dental implant system.

Fused Filament Fabrication (FFF)-based 3D printing is an efficient technique for developing medical implants, but it is not very useful in developing small yet mechanically robust design-specific fixtures such as dental implants (<15 mm). Specifically, it is challenging to 3D print robust Polyetheretherketone (PEEK) small implants due to PEEK's high melting temperature and melt viscosity. However, in this study, we efficiently utilize high-temperature FFF to develop the first-of-its-kind patient-specific robust PEEK dental implants with high print resolution. Specifically, we explore the effects of critical FFF processing conditions on the mechanical properties of the implants and subsequently determine an optimized set of processing conditions that are essential in developing durable dental implant systems. Our results indicate that the 3D printed dental implants exhibit good fatigue properties and suffice the clinical and industrial requirements for dental implants. Furthermore, we prove that the 3D printed implants exhibit adequate mechanical durability even after simulated (accelerated) aging of 30 years.

Influence of Fused Deposition Modelling Nozzle Temperature on the Rheology and Mechanical Properties of 3D Printed ß-Tricalcium Phosphate (TCP)/Polylactic Acid (PLA) Composite.

The primary goal of this study is to develop and analyze 3D printed structures based on a well-known composite known as ß-Tricalcium Phosphate (TCP)- polylactic acid (PLA). There are some interesting aspects of this study. First, we developed 3D printable TCP-PLA composite filaments in-house, with high reproducibility, by a one-step process method using a single screw extruder. Second, we explored the physicochemical properties of the developed TCP-PLA composite filaments. Third, we investigated the effect of an FDM-based nozzle temperature of 190 °C, 200 °C, 210 °C, and 220 °C on the composite's crystallinity and rheological and mechanical properties. Results confirmed the successful development of constant-diameter TCP-PLA composite filaments with a homogeneous distribution of TCP particles in the PLA matrix. We observed that a higher nozzle temperature in the FDM process increased the crystallinity of the printed PLA and TCP-PLA structures. As a result, it also helped to enhance the mechanical properties of the printed structures. The rheological studies were performed in the same temperature range used in the actual FDM process, and results showed an improvement in rheological properties at higher nozzle temperatures. The bare polymer and the composite polymer-ceramic melts exhibited lower viscosity and less rigidity at higher nozzle temperatures, which resulted in enhancing the polymer melt flowability and interlayer bonding between the printed layers. Overall, our results confirmed that 3D printable TCP-PLA filaments could be made in-house, and optimization of the nozzle temperature is essential to developing 3D printed composite parts with favorable mechanical properties.

Fused Filament Fabrication (Three-Dimensional Printing) of Amorphous Magnesium Phosphate/Polylactic Acid Macroporous Biocomposite Scaffolds.

The ultimate goal of this paper is to develop novel ceramic-polymer-based biocomposite orthopedic scaffolds with the help of additive manufacturing. Specifically, we incorporate a bioceramic known as amorphous magnesium phosphate (AMP) into polylactic acid (PLA) with the help of the melt-blending technique. Magnesium phosphate (MgP) was chosen as the bioactive component as previous studies have confirmed its favorable biomaterial properties, especially in orthopedics. Special care was taken to develop constant diameter AMP-PLA composite filaments, which would serve as feedstock for a fused filament fabrication (FFF)-based three-dimensional (3D) printer. Before the filaments were used for FFF, a thorough set of characterization protocols comprising of phase analysis, microstructure evaluations, thermal analysis, rheological analysis, and in vitro degradation determinations was performed on the biocomposites. Scanning electron microscopy (SEM) results confirmed a homogenous dispersion of AMP particles in the PLA matrix. Rheological studies demonstrated good printability behavior of the AMP-PLA filaments. In vitro degradation studies indicated a faster degradation rate in the case of AMP-PLA filaments as compared to the single phase PLA filaments. Subsequently, the filaments were fed into an FFF setup, and tensile bars and design-specific macroporous AMP-PLA scaffolds were printed. The biocomposite exhibited favorable mechanical properties. Furthermore, in vitro cytocompatibility results revealed higher pre-osteoblast cell attachment and proliferation on AMP-PLA scaffolds as compared to single-phase PLA scaffolds. Altogether, this study provides a proof of concept that design-specific bioactive AMP-PLA biocomposite scaffolds fabricated by FFF can be potential candidates as medical implants in orthopedics.

Antibacterial calcium phosphate composite cements reinforced with silver-doped magnesium phosphate (newberyite) micro-platelets.

This article demonstrates our efforts in developing and evaluating all-ceramic, biodegradable composites of calcium phosphate cements (CPCs) reinforced with silver (Ag)-doped magnesium phosphate (MgP) crystals. Two primary goals of this study were to 1) enhance CPC's poor mechanical properties with micro-platelet reinforcement, and 2) impart antibacterial functionalities in composites with the aim to inhibit surgical site infections (SSI). The work embodies three novel features. First, as opposed to well-known reinforcements with whisker or fiber-like morphology, we explored micro-platelets for the first time as the strengthening phase in the CPC matrix. Second, in contrast to conventional polymeric or calcium phosphate (CaP) fibrous reinforcements, newberyite (NB, MgHPO4.3H2O) micro-platelets belonging to the less explored yet promising MgP family, were evaluated as reinforcements for the first time. Third, NB micro-platelets were doped with Ag+ ions (AgNB, Ag content: 2 wt%) for enhancing antibacterial functionalities. Results indicated that 1 wt% of AgNB micro-platelet incorporation in the CPC matrix enhanced the compressive and flexural strengths by 200% and 140% respectively as compared to the un-reinforced ones. Besides, antibacterial assays revealed effective bactericidal functionalities (>99% bacteria kill) of the AgNB reinforced CPCs against Escherichia coli. Finally, cytocompatibility studies confirmed favorable pre-osteoblast cell proliferation and differentiation in vitro. Hence, this effort was successful in developing a self-setting and injectable AgNB reinforced CPC composition with favorable mechanical and antibacterial properties.

Magnetic Calcium Phosphate Cement for Hyperthermia Treatment of Bone Tumors.

This article reports, for the first time, the 'proof-of-concept' results on magnetic monetite (CaHPO4)-based calcium phosphate cements (CPCs) compositions developed for the hyperthermia treatment of bone tumors. Hyperthermia involves the heating of a tumor within a temperature range of 40-45 °C, inducing apoptosis in the tumor cells. This process holds promising potential in the field of cancer treatment and has been proven to be more effective than conventional therapeutics. Hence, we aimed to develop cement compositions that are capable of the hyperthermia treatment of bone tumors. To achieve that central goal, we incorporated iron oxide (Fe3O4), a ferromagnetic material, into monetite and hypothesized that, upon the application of a magnetic field, magnetite will generate heat and ablate the tumor cells near the implantation site. The results confirmed that an optimized content of magnetite incorporation in monetite can generate heat in the range of 40-45 °C upon the application of a magnetic field. Furthermore, the compositions were bioactive and cytocompatible with an osteoblastic cell line.

Bioactive amorphous magnesium phosphate-polyetheretherketone composite filaments for 3D printing.

OBJECTIVE: The aim of this study was to develop bioactive and osseointegrable polyetheretherketone (PEEK)-based composite filaments melt-blended with novel amorphous magnesium phosphate (AMP) particles for 3D printing of dental and orthopedic implants. MATERIALS AND METHODS: A series of materials and biological analyses of AMP-PEEK were performed. Thermal stability, thermogravimetric and differential scanning calorimetry curves of as-synthesized AMP were measured. Complex viscosity, elastic modulus and viscous modulus were determined using a rotational rheometer. In vitro bioactivity was analyzed using SBF immersion method. SEM, EDS and XRD were used to study the apatite-forming ability of the AMP-PEEK filaments. Mouse pre-osteoblasts (MC3T3-E1) were cultured and analyzed for cell viability, proliferation and gene expression. For in vivo analyses, bare PEEK was used as the control and 15AMP-PEEK was chosen based on its in vitro cell-related results. After 4 or 12 weeks, animals were euthanized, and the femurs were collected for micro-computed tomography (µ-CT) and histology. RESULTS: The collected findings confirmed the homogeneous dispersion of AMP particles within the PEEK matrix with no phase degradation. Rheological studies demonstrated that AMP-PEEK composites are good candidates for 3D printing by exhibiting high zero-shear and low infinite-shear viscosities. In vitro results revealed enhanced bioactivity and superior pre-osteoblast cell function in the case of AMP-PEEK composites as compared to bare PEEK. In vivo analyses further corroborated the enhanced osseointegration capacity for AMP-PEEK implants. SIGNIFICANCE: Collectively, the present investigation demonstrated that AMP-PEEK composite filaments can serve as feedstock for 3D printing of orthopedic and dental implants due to enhanced bioactivity and osseointegration capacity.

Microwave processing of calcium phosphate and magnesium phosphate based orthopedic bioceramics: A state-of-the-art review.

The main theme of this paper is to review microwave-assisted synthesis and processing of calcium and magnesium phosphate bioceramics. Microwave processing of advanced materials has been an active field of research for the last three decades and has been already reviewed in the literature. Microwave processing of bioceramics is being pursued for almost the same period of time. Unfortunately, to the best of our knowledge, we are not aware of any comprehensive review in the literature. Our group has been a significant contributor to the field, and we feel that it is an appropriate time for reviewing the state-of-the-art of the field. The paper is divided into several sections. After rationalizing the motivation behind writing this paper in the introduction, the second section builds on some fundamental aspects of microwave-matter interactions. The third section, representing the synthesis aspects, is subdivided into five sub-sections focusing on various calcium and magnesium phosphates in both crystalline and amorphous forms. The fourth section focuses on magnesium phosphate-based bioceramics. The fifth and the sixth section describe results on the utility of microwave assistance in developing multi-functional coatings on medical implants and orthopedic cements respectively. The subsequent section reviews results on microwave sintering of calcium and magnesium phosphates. The paper concludes with remarks on unresolved issues and future directions of research. It is expected that this comprehensive review on the interdisciplinary topic will further propel the exploration of other novel applications of microwave technology in processing biomaterials by a diverse group of scientists and engineers. STATEMENT OF SIGNIFICANCE: 1. This review highlights the broad-spectrum capabilities of microwave applications in processing orthopedic bioceramics. 2. The article covers "processing" in the broadest sense of the word, comprising of material synthesis, sintering, coating formation, and setting of orthopedic cements. It also expands beyond conventional calcium phosphates to include the emergent family of magnesium phosphates. 3. In vitro/in vivo responses of microwave-processed bioceramics are discussed thus providing an integral understanding of biological aspects of these materials. 4. The comprehensive review on this interdisciplinary topic will help researchers in various disciplines to appreciate the significance and usefulness of microwaves in biomaterials processing. Further, we also believe that it will propel the exploration of other novel applications of microwave technology in the biomaterials sector.

Silver (Ag) doped magnesium phosphate microplatelets as next-generation antibacterial orthopedic biomaterials.

This article reports for the first time our successful result in the synthesis of antibacterial single-phase newberyite (NB, MgHPO4 .3H2 O), an important magnesium phosphate (MgP) bioceramic. The prime novelty lies in the fact that we explore novel MgPs as next-generation orthopedic biomaterials as opposed to conventional calcium phosphates (CaP). While NB has already shown great promise, unlike its competitor struvite (ST, MgNH4 PO4 .6H2 O), NB is not intrinsically antibacterial. Given the havoc created by surgical site infections (SSI) in orthopedics, it would be worthwhile to explore if antibacterial NB can be synthesized cost-effectively. To accomplish that central goal, we used silver ion (Ag+ ) containing precursor solutions and exposed those to microwave irradiation. This action resulted in the rapid synthesis of NB microplatelets. Besides, three other specific objectives are addressed. First, Ag-doping was optimized to preserve the single-phase nature for sustained dopant release. Second, Ag+ release kinetics against common infection causing bacterial strains was analyzed. Finally, we inspected for any harmful effect of Ag-doped NB on MC3T3 preosteoblasts. Interestingly, the single-phase nature of NB microplatelets can be retained until 2 wt % Ag-doping and they exhibit good antibacterial and cytocompatible properties. Even though 3 wt % Ag-doped compositions (composites) were 100% antibacterial; they were cytotoxic.

Smart Injectable Self-Setting Monetite Based Bioceramics for Orthopedic Applications.

The present study is the first of its kind dealing with the development of a specific bioceramic which qualifies as a potential material in hard-tissue replacements. Specifically, we report the synthesis and evaluation of smart injectable calcium phosphate bone cement (CPC) which we believe will be suitable for various kinds of orthopedic and spinal-fusion applications. The smart nature of this next generation orthopedic implant is attained by incorporating piezoelectric barium titanate (BT) particles into monetite-based (dicalcium phosphate anhydrous, DCPA) CPC composition. The main goal is to take advantage of the piezoelectric properties of BT, as electromechanical effect plays a vital role in fracture healing at the defect site and bone integration with the implant. Furthermore, radiopacity of BT would help in easy detection of the CPC presence at the fracture site during surgery. Results reveal that BT addition favors important properties of bone cement such as good compressive strength, injectability, bioactivity, biocompatibility, and even washout resistance. Most importantly, the self-setting nature of the bone cements are not compromised with BT incorporation. The in vitro results confirm that the developed bone-cement abides by the standard orthopedic requirements making it apt for real-time prosthetic materials.

Microwave assisted coating of bioactive amorphous magnesium phosphate (AMP) on polyetheretherketone (PEEK).

Polyetheretherketone (PEEK) with great thermal and chemical stability, desirable mechanical properties and promising biocompatibility is being widely used as orthopedic and dental implant materials. However, the bioinert surface of PEEK can hinder direct osseointegration between the host tissue and PEEK based implants. The important signatures of this paper are as follows. First, we report for the formation of osseointegrable amorphous magnesium phosphate (AMP) coating on PEEK surface using microwave energy. Second, coatings consist of nano-sized AMP particles with a stacked thickness of 800nm. Third, coatings enhance bioactivity in-vitro and induce significantly high amount of bone-like apatite coating, when soaked in simulated body fluid (SBF). Fourth, the as-deposited AMP coatings present no cytotoxicity effects and are beneficial for cell adhesion at early stage. Finally, the high levels of expression of osteocalcin (OCN) in cells cultured on AMP coated PEEK samples indicate that AMP coatings can promote new bone formation and hence osseointegration.

Single-Phase, Antibacterial Trimagnesium Phosphate Hydrate Coatings on Polyetheretherketone (PEEK) Implants by Rapid Microwave Irradiation Technique.

This Article reports the fabrication and evaluation of single-phase, silver-doped trimagnesium phosphate hydrate (Ag-TMPH) nanosheet coatings on polyetheretherketone (PEEK), a well-known material used to fabricate orthopedic and spinal implants. While PEEK has better biomechanical compatibility with bone compared to metallic implants, it is also quite inert. Therefore, it is a common practice to coat PEEK implants with conventional calcium phosphates (CaPs) to enhance cell attachment, proliferation and differentiation. As opposed to well-studied CaP compounds, relatively less-explored magnesium phosphates (MgPs) are also becoming interesting orthopedic biomaterials and is the prime focus in this research. The novel aspects of this paper are as follows. First, we report developing TMPH coatings within minutes with the help of microwave irradiation technology. Microwave irradiation plays an important role in the coating formation with accelerated kinetics. Scanning electron microscopy (SEM) confirmed the fabrication of approximately 650 nm thick TMPH coatings. The coatings resulted in submicron level surface roughness and in vitro cell studies confirmed enhanced MC3T3 cell adhesion within 4 h on such surfaces. The coatings also resulted in significant apatite formation after immersing in simulated body fluid for 7 days. Second, multifunctionality was achieved by doping TMPH coatings with Ag, thus rendering the coatings antibacterial. The antibacterial properties were evaluated against two most common infection-causing bacterial strains-Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. The results indicated good bacterial resistance and bactericidal properties of the Ag-TMPH coatings. Third, in spite of Ag doping, the single-phase nature of the coatings were retained (without forming composite systems) with the help of the low-processing temperature of the microwave irradiation. The inductive coupled plasma technique confirmed that the doped single-phase TMPH coatings supported a uniform and controlled release of Ag+ ions over a period of 3 weeks. MTT assay evaluations and SEM micrographs confirmed no signs of cytotoxicity and healthy proliferation of cells in all cases. Quantitative real time PCR (qRT-PCR) indicated a significant rise in collagen (Col1) and osteocalcin (OCN) gene expression levels in the case of TMPH coated PEEK. Thus, microwave irradiation was successfully employed in forming multifunctional, that is, bioactive, cytocompatible, and antibacterial MgP coatings on PEEK.