No abstract

The therapeutic efficacy of drugs often depends on the drug delivery carrier. For efficient delivery of therapeutic proteins, delivery carriers should enable the loading of large doses, sustained release, and retention of the bioactivity of the therapeutic proteins. Here, it is demonstrated that graphene oxide (GO) is an efficient carrier for delivery of therapeutic proteins. Titanium (Ti) substrates are coated with GO through layer-by-layer assembly of positively (GO-NH₃⁺) and negatively (GO-COO⁻) charged GO sheets. Subsequently, a therapeutic protein (bone morphogenetic protein-2, BMP-2) is loaded on the GO-coated Ti substrate with the outermost coating layer of GO-COO⁻ (Ti/GO⁻). The GO coating on Ti substrate enables loading of large doses and the sustained release of BMP-2 with preservation of the structure and bioactivity of the drug. The extent of in vitro osteogenic differentiation of human bone marrow-derived mesenchymal stem cells is higher when they are cultured on Ti/GO- carrying BMP-2 than when they are cultured on Ti with BMP-2. Eight weeks after implantation in mouse models of calvarial defects, the Ti/GO-/BMP-2 implants show more robust new bone formation compared with Ti, Ti/GO-, or Ti/BMP-2 implants. Therefore, GO is an effective carrier for the controlled delivery of therapeutic proteins, such as BMP-2, which promotes osteointegration of orthopedic or dental Ti implants.

Current treatments for wound healing engage in passive healing processes and rarely participate in stimulating skin cell behaviors for active wound healing. Electric potential difference-derived electrical fields (EFs) are known to modulate skin cell behaviors. Here, a piezoelectric dermal patch is developed that can be applied on skin wound site and EF is generated to promote wound healing. The one-directionally aligned zinc oxide nanorodbased piezoelectric patch generates piezoelectric potential upon mechanical deformations induced by animal motion, and induces EF at the wound bed. In vitro and in vivo data demonstrate that the piezoelectric patch promotes the wound healing process through enhanced cellular metabolism, migration, and protein synthesis. This modality may lead to a clinically relevant piezoelectric dermal patch therapy for active wound healing.

Microfracture of cartilage induces migration of bone-marrow-derived mesenchymal stem cells. However, this treatment often results in fibrocartilage regeneration. Growth factors such as bone morphogenetic protein (BMP)-2 induce the differentiation of bone-marrow-derived mesenchymal stem cells into chondrocytes, which can be used for hyaline cartilage regeneration. Here, we tested the hypothesis that long-term delivery of BMP-2 to cartilage defects subjected to microfracture results in regeneration of high-quality hyaline-like cartilage, as opposed to short-term delivery of BMP-2 or no BMP-2 delivery. Heparin-conjugated fibrin (HCF) and normal fibrin were used as carriers for the long- and short-term delivery of BMP-2, respectively. Rabbit articular cartilage defects were treated with microfracture combined with one of the following: no treatment, fibrin, short-term delivery of BMP-2, HCF, or long-term delivery of BMP-2. Eight weeks after treatment, histological analysis revealed that the long-term delivery of BMP-2 group (microfracture + HCF + BMP-2) showed the most staining with alcian blue. A biochemical assay, real-time polymerase chain reaction assay and Western blot analysis all revealed that the long-term delivery of BMP-2 group had the highest glucosaminoglycan content as well as the highest expression level of collagen type II. Taken together, the long-term delivery of BMP-2 to cartilage defects subjected to microfracture resulted in regeneration of hyaline-like cartilage, as opposed to short-term delivery or no BMP-2 delivery. Therefore, this method could be more convenient for hyaline cartilage regeneration than autologous chondrocyte implantation due to its less invasive nature and lack of cell implantation.

Bone morphogenetic protein-2 (BMP-2) induces bone regeneration in a dose-dependent manner, with higher doses of BMP-2 inducing greater bone formation. Previously, we showed that long-term delivery of BMP-2 provides better ectopic bone formation than short-term delivery of an equivalent dose. In the present study, we investigated the efficacy of orthotopic bone formation over a range of BMP-2 doses, using different delivery modes. Heparin-conjugated poly(lactic-co-glycolic acid) nanospheres suspended in fibrin gel were used as a long-term delivery system, and fibrin gel was used as a short-term delivery system. Different doses of BMP-2 were delivered to mouse calvarial defects using either long-term or short-term delivery systems. Eight weeks after treatment, bone regeneration was evaluated by histomorphometry. For both delivery systems, bone regeneration increased as the BMP-2 dose increased up to 1 µg and did not increase beyond this dose. Importantly, at BMP-2 doses higher than 1 µg, long-term delivery resulted in much greater bone formation than short-term delivery. This study shows that long-term delivery of BMP-2 is more effective at enhancing orthotopic bone formation than short-term delivery over a range of doses.

Despite the great potential of cell therapy for ischemic disease, poor cell survival after engraftment in ischemic tissue limits its efficacy. Here we tested a hypothesis that three-dimensionally grafted human umbilical vein endothelial cell (HUVEC) spheroids would exhibit improved angiogenic efficacy following transplantation into mouse ischemic limbs compared with HUVECs prepared by conventional two-dimensional monolayer culture. One day after surgical induction of hindlimb ischemia in athymic mice, HUVECs cultured in monolayer or HUVEC spheroids were transplanted intramuscularly into ischemic limbs. Four weeks after the treatment, in the spheroid HUVEC transplantation group, we observed increased hypoxia-inducible factor-1α expression, decreased apoptosis, and increased HUVEC survival in the ischemic tissue compared with the monolayer HUVEC transplantation group. Transplantation of HUVEC spheroids also resulted in enhanced and prolonged secretion of paracrine factors as well as enhanced expression of factors involved in the recruitment of circulating angiogenic progenitor cells. In summary, transplantation of HUVECs as spheroids enhanced cell survival, increased paracrine factor secretion, and showed a potential as a therapeutic method to treat ischemic tissue damages by promoting angiogenesis.

No abstract

Bone morphogenetic proteins (BMPs) are the most potent osteoinductive growth factors. Clinically utilized BMP-2 uses a type-I collagen scaffold as a carrier. Here we hypothesized that an apatite coating on a type-I collagen scaffold would prolong the BMP-2 release period and enhance bone regeneration in calvarial defects in mice. Apatite coating was achieved by incubating collagen scaffolds in simulated body fluid. BMP-2 release kinetics and bioactivity were evaluated by enzyme-linked immunosorbent assay and alkaline phosphatase activity measurement of cultured osteoblasts. Computed tomography and histomorphometry were performed eight weeks after various doses of BMP-2 were delivered to mouse calvarial defects using either non-modified or apatite-coated collagen scaffolds. Apatite-coated collagen scaffolds released 91.8 ± 11.5% of the loaded BMP-2 over 13 days in vitro, whereas non-modified collagen scaffolds released 98.3 ± 2.2% over the initial one day. The in vivo study showed that BMP-2 delivery with apatite-coated collagen scaffolds resulted in a significantly greater bone formation area and higher bone density than that with non-modified collagen scaffolds. This study suggests that simple apatite coating on collagen scaffolds can enhance the bone regeneration efficacy of BMP-2 released from collagen scaffolds.