Anh-Vu Do

3D Printing of Scaffolds for Tissue Regeneration Applications

The current need for organ and tissue replacement, repair, and regeneration for patients is continually growing such that supply is not meeting demand primarily due to a paucity of donors as well as biocompatibility issues leading to immune rejection of the transplant. In order to overcome these drawbacks, scientists have investigated the use of scaffolds as an alternative to transplantation. These scaffolds are designed to mimic the extracellular matrix (ECM) by providing structural support as well as promoting attachment, proliferation, and differentiation with the ultimate goal of yielding functional tissues or organs. Initial attempts at developing scaffolds were problematic and subsequently inspired an interest in 3D printing as a mode for generating scaffolds. Utilizing three-dimensional printing (3DP) technologies, ECM-like scaffolds can be produced with a high degree of complexity, where fine details can be included at a micrometer level. In this Review, the criteria for printing viable and functional scaffolds, scaffolding materials, and 3DP technologies used to print scaffolds for tissue engineering are discussed. Creating biofunctional scaffolds could potentially help to meet the demand by patients for tissues and organs without having to wait or rely on donors for transplantation.

Chemically modified RNA activated matrices enhance bone regeneration

There exists a dire need for improved therapeutics to achieve predictable bone regeneration. Gene therapy using non-viral vectors that are safe and efficient at transfecting target cells is a promising approach to overcoming the drawbacks of protein delivery of growth factors. Here, we investigated the transfection efficiency, cytotoxicity, osteogenic potential and in vivo bone regenerative capacity of chemically modified ribonucleic acid (cmRNA) (encoding BMP-2) complexed with polyethylenimine (PEI) and made comparisons with PEI complexed with conventional plasmid DNA (encoding BMP-2). The polyplexes were fabricated at an amine (N) to phosphate (P) ratio of 10 and characterized for transfection efficiency using human bone marrow stromal cells (BMSCs). The osteogenic potential of BMSCs treated with these polyplexes was validated by determining the expression of bone-specific genes, osteocalcin and alkaline phosphatase as well as through the detection of bone matrix deposition. Using a calvarial bone defect model in rats, it was shown that PEI-cmRNA (encoding BMP-2)-activated matrices promoted significantly enhanced bone regeneration compared to PEI-plasmid DNA (BMP-2)-activated matrices. Our proof of concept study suggests that scaffolds loaded with non-viral vectors harboring cmRNA encoding osteogenic proteins may be a powerful tool for stimulating bone regeneration with significant potential for clinical translation.

Controlled and Sequential Delivery of Fluorophores from 3D Printed Alginate-PLGA Tubes

Controlled drug delivery systems, that include sequential and/or sustained drug delivery, have been utilized to enhance the therapeutic effects of many current drugs by effectively delivering drugs in a time-dependent and repeatable manner. In this study, with the aid of 3D printing technology, a novel drug delivery device was fabricated and tested to evaluate sequential delivery functionality. With an alginate shell and a poly(lactic-co-glycolic acid) (PLGA) core, the fabricated tubes displayed sequential release of distinct fluorescent dyes and showed no cytotoxicity when incubated with the human embryonic kidney (HEK293) cell line or bone marrow stromal stem cells (BMSC). The controlled differential release of drugs or proteins through such a delivery system has the potential to be used in a wide variety of biomedical applications from treating cancer to regenerative medicine.

Long-term release of a thiobenzamide from a backbone functionalized poly(lactic acid)

Hydrogen sulfide is emerging as a critically important molecule in medicine, yet there are few methods for the long-term delivery of molecules that degrade to release H2S. In this paper the first long-term release of a thiobenzamide that degrades to release H2S is described. A series of polymers were synthesized by the copolymerization of L-lactide and a lactide functionalized with 4-hydroxythiobenzamide. A new method to attach functional groups to a derivative of L-lactide is described based on the addition of a thiol to an α,β-unsaturated lactide using catalytic I2. This reaction proceeded under mild conditions and did not ring-open the lactone. The copolymers had molecular weights from 8 to 88 kg mol-1 with PDIs below 1.50. Two sets of microparticles were fabricated from a copolymer; the average diameters of the microparticles were 0.53 and 12 μm. The degradation of the smaller microparticles was investigated in buffered water to demonstrate the slow release of thiobenzamide over 4 weeks. Based on the ability to synthesize polymers with different loadings of thiobenzamide and that thiobenzamide is a known precursor to H2S, these particles provide a polymer-based method to deliver H2S over days to weeks.

Regeneration of bone using nanoplex delivery of FGF-2 and BMP-2 genes in diaphyseal long bone radial defects in a diabetic rabbit model.

Abstract Bone fracture healing impairment related to systemic diseases such as diabetes can be addressed by growth factor augmentation. We previously reported that growth factors such as fibroblast growth factor‐2 (FGF‐2) and bone morphogenetic protein‐2 (BMP‐2) work synergistically to encourage osteogenesis in vitro. In this report, we investigated if BMP‐2 and FGF‐2 together can synergistically promote bone repair in a leporine model of diabetes mellitus, a condition that is known to be detrimental to union. We utilized two kinds of plasmid DNA encoding either BMP‐2 or FGF‐2 formulated into polyethylenimine (PEI) complexes. The fabricated nanoplexes were assessed for their size, charge, in vitro cytotoxicity, and capacity to transfect human bone marrow stromal cells (BMSCs). Using diaphyseal long bone radial defects in a diabetic rabbit model it was demonstrated that co‐delivery of PEI‐(pBMP‐2 + pFGF‐2) embedded in collagen scaffolds resulted in a significant improvement in bone regeneration compared to PEI‐pBMP‐2 embedded in collagen scaffolds alone. This study demonstrated that scaffolds loaded with PEI‐(pBMP‐2 + pFGF‐2) could be an effective way of promoting bone regeneration in patients with diabetes. Graphical abstract Figure. No Caption available.

3D printing technologies for 3D scaffold engineering

Abstract 3D printing technologies are a promising and viable means of fabricating replicable and functional scaffolds capable of promoting tissue regeneration. Printing technologies such as direct 3D printing, 3D-Bioplotter printing, fused deposition modeling, selective laser sintering, stereolithography, indirect 3D printing, and electrospinning are commonly used 3D printing techniques for creating scaffolds. These techniques have shown success in pre-clinical settings with respect to regenerating various tissue types, and have designed implantable structures investigated in clinical trials for bone defect restoration, pulmonary disease, dysphagia, and lung cancer. Continued advancement and hybrid approaches using different material combinations, printing methodologies, and controlled drug delivery will further advance the progress of 3D printing technologies toward developing scaffolds capable of being utilized in the clinic. An overview of these 3D printing technologies and their recent developments in the field of tissue engineering are discussed, and demonstrate the promise of the recent advancements made in 3D printing.

Controlled drug delivery from 3D printed two-photon polymerized poly(ethylene glycol) dimethacrylate devices

ABSTRACT Controlled drug delivery systems have been utilized to enhance the therapeutic effects of many drugs by delivering drugs in a time‐dependent and sustained manner. Here, with the aid of 3D printing technology, drug delivery devices were fabricated and tested using a model drug (fluorophore: rhodamine B). Poly(ethylene glycol) dimethacrylate (PEGDMA) devices were fabricated using a two‐photon polymerization (TPP) system and rhodamine B was homogenously entrapped inside the polymer matrix during photopolymerization. These devices were printed with varying porosity and morphology using varying printing parameters such as slicing and hatching distance. The effects of these variables on drug release kinetics were determined by evaluating device fluorescence over the course of one week. These PEGDMA‐based structures were then investigated for toxicity and biocompatibility in vitro, where MTS assays were performed using a range of cell types including induced pluripotent stem cells (iPSCs). Overall, tuning the hatching distance, slicing distance, and pore size of the fabricated devices modulated the rhodamine B release profile, in each case presumably due to resulting changes in the motility of the small molecule and its access to structure edges. In general, increased spacing provided higher drug release while smaller spacing resulted in some occlusion, preventing media infiltration and thus resulting in reduced fluorophore release. The devices had no cytotoxic effects on human embryonic kidney cells (HEK293), bone marrow stromal stem cells (BMSCs) or iPSCs. Thus, we have demonstrated the utility of two‐photon polymerization to create biocompatible, complex miniature devices with fine details and tunable release of a model drug.