Regenerative engineering of long bones using the small molecule forskolin.

Bone grafting procedures have become increasingly common in the United States, with approximately 500,000 cases occurring each year at a societal cost exceeding $2.4 billion. Recombinant human bone morphogenetic proteins (rhBMPs) are therapeutic agents that have been widely used by orthopedic surgeons to stimulate bone tissue formation alone and when paired with biomaterials. However, significant limitations such as immunogenicity, high production cost, and ectopic bone growth from these therapies remain. Therefore, efforts have been made to discover and repurpose osteoinductive small-molecule therapeutics to promote bone regeneration. Previously, we have demonstrated that a single-dose treatment with the small-molecule forskolin for just 24 h induces osteogenic differentiation of rabbit bone marrow-derived stem cells in vitro, while mitigating adverse side effects attributed with prolonged small-molecule treatment schemes. In this study, we engineered a composite fibrin-PLGA [poly(lactide-co-glycolide)]-sintered microsphere scaffold for the localized, short-term delivery of the osteoinductive small molecule, forskolin. In vitro characterization studies showed that forskolin released out of the fibrin gel within the first 24 h and retained its bioactivity toward osteogenic differentiation of bone marrow-derived stem cells. The forskolin-loaded fibrin-PLGA scaffold was also able to guide bone formation in a 3-mo rabbit radial critical-sized defect model comparable to recombinant human bone morphogenetic protein-2 (rhBMP-2) treatment, as demonstrated through histological and mechanical evaluation, with minimal systemic off-target side effects. Together, these results demonstrate the successful application of an innovative small-molecule treatment approach within long bone critical-sized defects.

Biodegradable Nanofiber Bone-Tissue Scaffold as Remotely-Controlled and Self-Powering Electrical Stimulator.

Electrical stimulation (ES) has been shown to induce and enhance bone regeneration. By combining this treatment with tissue-engineering approaches (which rely on biomaterial scaffolds to construct artificial tissues), a replacement bone-graft with strong regenerative properties can be achieved while avoiding the use of potentially toxic levels of growth factors. Unfortunately, there is currently a lack of safe and effective methods to induce electrical cues directly on cells/tissues grown on the biomaterial scaffolds. Here, we present a novel bone regeneration method which hybridizes ES and tissue-engineering approaches by employing a biodegradable piezoelectric PLLA (Poly(L-lactic acid)) nanofiber scaffold which, together with externally-controlled ultrasound (US), can generate surface-charges to drive bone regeneration. We demonstrate that the approach of using the piezoelectric scaffold and US can enhance osteogenic differentiation of different stem cells in vitro, and induce bone growth in a critical-sized calvarial defect in vivo. The biodegradable piezoelectric scaffold with applied US could significantly impact the field of tissue engineering by offering a novel biodegradable, battery-free and remotely-controlled electrical stimulator.

Skeletal Muscle Regenerative Engineering.

Skeletal muscles have the intrinsic ability to regenerate after minor injury, but under certain circumstances such as severe trauma from accidents, chronic diseases or battlefield injuries the regeneration process is limited. Skeletal muscle regenerative engineering has emerged as a promising approach to address this clinical issue. The regenerative engineering approach involves the convergence of advanced materials science, stem cell science, physical forces, insights from developmental biology, and clinical translation. This article reviews recent studies showing the potential of the convergences of technologies involving biomaterials, stem cells and bioactive factors in concert with clinical translation, in promoting skeletal muscle regeneration. Several types of biomaterials such as electrospun nanofibers, hydrogels, patterned scaffolds, decellularized tissues, and conductive matrices are being investigated. Detailed discussions are given on how these biomaterials can interact with cells and modulate their behavior through physical, chemical and mechanical cues. In addition, the application of physical forces such as mechanical and electrical stimulation are reviewed as strategies that can further enhance muscle contractility and functionality. The review also discusses established animal models to evaluate regeneration in two clinically relevant muscle injuries; volumetric muscle loss (VML) and muscle atrophy upon rotator cuff injury. Regenerative engineering approaches using advanced biomaterials, cells, and physical forces, developmental cues along with insights from immunology, genetics and other aspects of clinical translation hold significant potential to develop promising strategies to support skeletal muscle regeneration.

Repositioning Tacrolimus: Evaluation of the Effect of Short-Term Tacrolimus Treatment on Osteoprogenitor Cells and Primary Cells for Bone Regenerative Engineering.

Small-molecule-based bone regenerative engineering is an encouraging strategy for repair and regeneration of skeletal tissue. Using osteogenic small molecules for engineering bone tissue has several potential benefits over polypeptide-based approaches. Interestingly, hundreds of such small molecules possess the capability to promote osteogenesis, and several of these are already approved by the FDA for use in other applications, indicating their safety for human use. However, the need for their use at a high frequency and/or duration, due to their short half-life and nonspecificity, is still problematic. We, and others, have identified several non-FDA-approved small-molecule-based compounds that induce long-lasting osteogenic effects following short-term (<24 h) treatment. In this study, however, we have performed a proactive screen to investigate and compare the osteogenic effects of several preselected FDA-approved small-molecule drugs in vitro using osteoprogenitor MC3T3-E1 cells. Our results demonstrate that treatment with the small-molecule drug tacrolimus (FK-506) for 24 h significantly enhanced long-lasting osteogenic responses in both osteoprogenitor cells and primary cell cultures. In addition, we tested whether a short-term treatment with FK-506 is able to induce osteogenic differentiation of cells seeded on a polymeric scaffold in vitro. Using an osteogenic small molecule that has long-lasting effects despite a short duration of exposure to cells may alleviate the undesirable effects often seen with many osteogenic small molecules.

The roles of ions on bone regeneration.

Bone scientists are actively investigating a range of methods to promote skeletal tissue regeneration. A review of recent literature has revealed that several ions are uniquely capable of inducing stem cell differentiation down desired lineages. There exists enormous promise for these ions to be used in bone regenerative medicine. Given that these ions can be released from biodegradable polymeric materials, their long-term delivery can be achieved through a variety of controlled-release strategies compared with the relatively few options available for expensive and fragile polypeptide-based growth factors. In this review, we highlight the developments in using ions in conjunction with biomaterials for bone regeneration.

Engineered Bone Tissue with Naturally-Derived Small Molecules.

Natural products remain the best resources of drugs and drug leads. Recently, there is growing recognition that identifying new small molecules to promote bone regeneration is a laudable translational goal. In fact, new approaches for bone repair and regeneration that involve inexpensive naturally-derived compounds would have an important impact on the treatment of bone disorders and injuries. Over the past several decades, a number of naturally-derived small molecules with the potential of regenerating bone tissue (i.e. osteoinductive) have been reported in the literature. Here, we review the current literature, paying attention to the prospects for natural small molecule-based bone regenerative engineering. We also review various delivery strategies of the selected naturally- derived small molecules for bone regenerative engineering applications.

Harnessing cAMP signaling in musculoskeletal regenerative engineering.

This paper reviews the most recent findings in the search for small molecule cyclic AMP analogues regarding their potential use in musculoskeletal regenerative engineering.