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The faithful engineering of complex human tissues such as the bone/soft tissue/mineralized tissue interface in periodontal tissues requires innovative molecular cues in conjunction with tailored scaffolds. To address the loss of periodontal bone and connective tissues following periodontal disease, we have generated a polydopamine and collagen coated electrospun PLGA-PCL (PP) scaffold enriched with the small molecule mediator PFI-2 (PP-PFI-pDA-COL-PFI). In vitro 3D studies using PDL progenitors revealed that the PP-PFI-pDA-COL-PFI scaffold substantially enhanced Alizarin Red staining, increased Ca/P ratios 4-fold, and stimulated cell proliferation more than 12-fold compared to PP-controls, suggestive of its potential for mineralized tissue engineering. When applied in our experimental periodontitis model, the PP-PFI-pDA-COL-PFI scaffold resulted in a substantial 34% reduction in alveolar bone defect height, a 25% root-length gain in periodontal attachment, and the formation of highly ordered regenerated acellular cementum twice as thick as in controls. Explaining the mechanism of PFI-2 mineralized tissue regeneration in periodontal tissues, PFI-2 inhibited SETD7-mediated β-Catenin protein methylation and increased β-Catenin nuclear localization. Together, dual-level PFI-2 incorporation into a degradable, dopamine/collagen coated PLGA/PCL scaffold backbone resulted in the regeneration of the tripartite periodontal complex with unprecedented fidelity, including periodontal attachment and new formation of mineralized tissues in inflamed periodontal environments.

Chemotherapy promotes phosphatidylserine (PS) externalization in tumors undergoing apoptosis, forms an immunosuppressive tumor microenvironment (TME), and inhibits dendritic cell (DC) maturation and antigen presentation by binding PS receptors expressed in DCs, thereby limiting naive T cell education and activation. In this study, we demonstrate a selective nanocarrier system composed of annexin A5-labeled poly (lactide-co-glycolide) nanoparticles (PLGA_NPs) encapsulating tumor specific antigen or neoantigen, to target apoptotic tumor cells expressing PS as an innate immune checkpoint inhibitor (ICI) that induces active cancer immunotherapy. Moreover, PLGA_NPs enhanced tumor-specific antigen-based cytotoxic T cell immunity via the original function of DCs by converting the tumor antigen-rich environment. Therefore, chemotherapy combined with an immunomodulatory nanocarrier system demonstrated an enhanced anticancer immune response by increasing survival rates, immune-activating cells, and pro-inflammatory cytokines in the spleen and TME. In contrast, the tumor mass, immune-suppressive cells, and anti-inflammatory cytokines were decreased. Furthermore, the combination of a nanocarrier system with other ICIs against large tumors showed therapeutic efficacy by immunosuppression in the TME and further amplified the anticancer immunity of interferon gamma

Bispecific nanoparticles (NPs) are conjugated with two antibodies that enhance T cell cytotoxicity by sequentially targeting CD3 and tumor-specific proteins. This interaction redirects T cells to specific tumor antigens and activates them to lyse tumor cells by blocking two different signaling pathways simultaneously. This study developed NP-based bispecific T-cell engagers (nanoBiTEs), which are R848-loaded bispecific poly(lactic-co-glycolic acid) NPs decorated with anti-CD3 antibody targeting T cells and anti-PD-L1 antibody targeting PD-L1 ligands (bis-R848-PLGA-NPs). Bis-R848-PLGA-NPs enhance the immunogenic response in destroying cancer cells by restoring the T cell effector functions. These interactions allow T cells to come in close proximity to the tumor cells. Finally, the release of R848 from PLGA-NPs activates dendritic cells, enhancing T cell activation. In vitro results show maximum internalization of bis-R848-PLGA-NPs in SK-OV3 and B16F10 cell lines, attributed to high PD-L1 expression in both cells. Furthermore, bis-R848-PLGA-NPs-treated CD8

A robust regimen for inducing allogeneic transplantation tolerance involves pre-emptive recipient treatment with donor splenocytes (SP) rendered apoptotic by 1-ethyl-3-(3′-dimethylaminopropyl)-carbodiimide(ECDI) treatment. However, such a regimen is limited by availability of donor cells, cost of cell procurement, and regulatory hurdles associated with cell-based therapies. Nanoparticles (NP) delivering donor antigens are a promising alternative for promoting transplantation tolerance. Here, we used a B6.C-H-2bm12(bm12) to C57BL/6(B6) skin transplant model involving a defined major histocompatibility antigen mismatch to investigate design parameters of poly(lactide-co-glycolide) (PLG) NPs delivering peptides containing the donor antigen for optimizing skin allograft survival. We showed that an epitope-containing short peptide (P1) was more effective than a longer peptide (P2) at providing graft protection. Importantly, the NP and P1 complex (NP-ECDI-P1) resulted in a significant expansion of graft-infiltrating Tregs. Interestingly, in comparison to donor ECDI-SP that provided indefinite graft protection, NP-ECDI-P1 targeted different splenic phagocytes and skin allografts in these recipients harbored significantly more graft-infiltrating CD8+IFN-γ+ cells. Collectively, the current study provides initial engineering parameters for a cell-free and biocompatible NP-peptide platform for transplant immunoregulation. Moreover, it also provides guidance to future NP engineering endeavors to recapitulate the effects of donor ECDI-SP as a goal for maximizing tolerance efficacy of NP formulations.

Bone defect repair is a challenging clinical problem in musculoskeletal system, especially in orthopaedic disorders such as steroid associated osteonecrosis (SAON). Magnesium (Mg) as a biodegradable metal with properly mechanical properties has been investigating for a long history. In this study, Mg powder, poly (lactide-co-glycolide) (PLGA), β-tricalcium phosphate (β-TCP) were the elements to formulate a novel porous PLGA/TCP/Mg (PTM) scaffolds using low temperature rapid prototyping (LT-RP) technology. The physical characterization of PTM scaffold and Mg ions release were analyzed in vitro. The osteogenic and angiogenic properties of PTM scaffolds, as well as the biosafety after implantation were assessed in an established SAON rabbit model. Our results showed that the PTM scaffold possessed well-designed bio-mimic structure and improved mechanical properties. Findings of dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and micro-computed tomography (micro CT)-based angiography indicated that PTM scaffold could increase blood perfusion and promote new vessel ingrowth at 4 weeks after surgery, meanwhile, a plenty of newly formed vessels with well-architective structure were observed at 8 weeks. Correspondingly, at 12 weeks after surgery, micro-CT, histological and mechanical properties analysis showed that PTM could significant enhance new bone formation and strengthen newly formed bone mechanical properties. The mean bone volume in PTM group was 56.3% greater than that in PT group. Biosafety assessments from 0 to 12 weeks after implantation did not induce increase in serum Mg ions concentration, and immune response, liver and kidney function parameters were all at normal level. These findings suggested that the PTM scaffold had both osteogenic and angiogenic abilities which were synergistic effect in enhancing new bone formation and strengthen newly formed bone quality in SAON. In summary, PTM scaffolds are promising composite biomaterials for repairing challenging bone defect that would have great potential for its clinical translation.

Immunotherapy is one of the most promising approaches to inhibit tumor growth and metastasis by activating host immune functions. However, the arising problems such as low immune response caused by complex tumor microenvironment and extremely systemic immune storm still limit the clinical applications of immunotherapy. Here, we construct Poly I: C-encapsulated poly (lactic-co-glycolic acid) nanoparticles (PLP NPs) with a slow release profile. A biomimetic system (MPLP), which loads PLP NPs on the surface of bone marrow-derived macrophage (BMDM) via the maleimide-thiol conjugation, is synthesized to effectively deliver PLP, control drug release and activate the tumor-specific immune response in situ. The results show that PLP NPs loading does not affect the activity and function of BMDM. Then, BMDM acts as a living cell drug vehicle and promotes the accumulation of PLP NPs in tumors, where Poly I: C is released from PLP NPs and reprograms BMDM into tumoricidal M1 macrophage. Furthermore, MPLP triggers potent antitumor immune responses in vivo and effectively inhibits local and metastatic tumors without causing adverse pathological immune reactions. This study offers an inspiration to facilitate clinical translation through the delivery of drugs by living immune cells for future anticancer therapy.

A near-infrared (NIR) light-triggered drug delivery platform is produced by incorporating SrCl2 and BP nanosheets (BPs) into poly(lactic-co-glycolic acid) (PLGA) for bone regeneration. The fabricated BP-SrCl2/PLGA microspheres show efficient NIR absorption and photothermal effects due to the BPs. The NIR-triggered release behavior of Sr2+ by flawing the PLGA shells is investigated and the microspheres exhibit excellent cell viability and biodegradability. Implantation of the BP-SrCl2/PLGA microspheres into a rat femoral defect demonstrates good tissue compatibility and excellent bone regeneration capacity under NIR light irradiation. Our study indicates that local release of Sr2+ at optimal time periods controlled by NIR irradiation improves bone regeneration significantly and this NIR-triggered drug delivery system composed of BPs is suitable for therapies requiring precise control at specific time.

A range of magnesium ions (Mg2+) used has demonstrated osteogenic tendency in vitro. Hence, we propose to actualize this concept by designing a new system to precisely control the Mg2+ delivery at a particular concentration in vivo in order to effectively stimulate in-situ bone regeneration. To achieve this objective, a monodisperse core-shell microsphere delivery system comprising of poly (lactic-co-glycolic acid) (PLGA) biopolymer, alginate hydrogel, and magnesium oxide nano-particles has been designed by using customized microfluidic capillary device. The PLGA-MgO sponge-like spherical core works as a reservoir of Mg2+ while the alginate shell serves as physical barrier to control the outflow of Mg2+ at ∼50 ppm accurately for 2 weeks via its adjustable surface micro-porous network. With the aid of controlled release of Mg2+, the new core-shell microsphere system can effectively enhance osteoblastic activity in vitro and stimulate in-situ bone regeneration in vivo in terms of total bone volume, bone mineral density (BMD), and trabecular thickness after operation. Interestingly, the Young's moduli of formed bone on the core-shell microsphere group have been restored to ∼96% of that of the surrounding matured bone. These findings indicate that the concept of precisely controlled release of Mg2+ may potentially apply for in-situ bone regeneration clinically.

This study aims to develop a novel surface modification technology to prolong the survival time of pancreatic islets in a xenogenic transplantation model, using 3,4-dihydroxyphenethylamine (DOPA) conjugated poly(lactide-co-glycolide)-poly(ethylene glycol) (PLGA-PEG) nanoparticles (DOPA-NPs) carrying immunosuppressant FK506 (FK506/DOPA-NPs). The functionalized DOPA-NPs formed a versatile coating layer for antigen camouflage without interfering the viability and functionality of islets. The coating layer effectively preserved the morphology and viability of islets in a co-culture condition with xenogenic lymphocytes for 7 days. Interestingly, the mean survival time of islets coated with FK506/DOPA-NPs was significantly higher as compared with that of islets coated with DOPA-NPs (without FK506) and control. This study demonstrated that the combination of surface camouflage and localized low dose of immunosuppressant could be an effective approach in prolonging the survival of transplanted islets. This newly developed platform might be useful for immobilizing various types of small molecules on therapeutic cells and biomaterial surface to improve the therapeutic efficacy in cell therapy and regenerative medicine.

Steroid-associated osteonecrosis (SAON) often requires surgical core decompression (CD) in the early stage for removal of necrotic bone to facilitate repair where bone grafts are needed for filling bone defect and avoiding subsequent joint collapse. In this study, we developed a bioactive composite scaffold incorporated with icariin, a unique phytomolecule that can provide structural and mechanical support and facilitate bone regeneration to fill into bone defects after surgical CD in established SAON rabbit model. An innovative low-temperature 3D printing technology was used to fabricate the poly (lactic-co-glycolic acid)/β-calcium phosphate/icariin (PLGA/TCP/Icariin, PTI) scaffold. The cytocompatibility of the PTI scaffold was tested in vitro, and the osteogenesis properties of PTI scaffolds were assessed in vivo in the SAON rabbit models. Our results showed that the fabricated PTI scaffold had a well-designed biomimic structure that was precisely printed to provide increased mechanical support and stable icariin release from the scaffold for bone regeneration. Furthermore, our in vivo study indicated that the PTI scaffold could enhanced the mechanical properties of new bone tissues and improved angiogenesis within the implanted region in SAON rabbit model than those of PLGA/TCP (PT) scaffold. The underlying osteoblastic mechanism was investigated using MC3T3-E1 cells in vitro and revealed that icariin could facilitate MC3T3-E1 cells ingrowth into the PTI scaffold and regulate osteoblastic differentiation. The PTI scaffold exhibited superior biodegradability, biocompatibility, and osteogenic capability compared with those of PT scaffold. In summary, the PTI composite scaffold which incorporated bioactive phyto-compounds is a promising potential strategy for bone tissue engineering and regeneration in patients with challenging SAON.

Nanocarriers (NCs) help improve the performance of therapeutics, but their removal by phagocytes in the liver, spleen, tissues, etc. diminishes this potential. Although NC functionalization with polyethylene glycol (PEG) lowers interaction with phagocytes, it also reduces interactions with tissue cells. Coating NCs with CD47, a protein expressed by body cells to avoid phagocytic removal, offers an alternative. Previous studies showed that coating CD47 on non-targeted NCs reduces phagocytosis, but whether this alters binding and endocytosis of actively-targeted NCs remains unknown. To evaluate this, we used polymer NCs targeted to ICAM-1, a receptor overexpressed in many diseases. Co-coating of CD47 on anti-ICAM NCs reduced macrophage phagocytosis by ∼50% for up to 24 h, while increasing endothelial-cell targeting by ∼87% over control anti-ICAM/IgG NCs. Anti-ICAM/CD47 NCs were endocytosed via the CAM-mediated pathway with efficiency similar (0.99-fold) to anti-ICAM/IgG NCs. Comparable outcomes were observed for NCs targeted to PECAM-1 or transferrin receptor, suggesting broad applicability. When injected in mice, anti-ICAM/CD47 NCs reduced liver and spleen uptake by ∼30–50% and increased lung targeting by ∼2-fold (∼10-fold over IgG NCs). Therefore, co-coating NCs with CD47 and targeting moieties reduces macrophage phagocytosis and improves targeted uptake. This strategy may significantly improve the efficacy of targeted drug NCs.

Drug delivery to the brain is limited by poor penetration of pharmaceutical agents across the blood-brain barrier (BBB), within the brain parenchyma, and into specific cells of interest. Nanotechnology can overcome these barriers, but its ability to do so is dependent on nanoparticle physicochemical properties including surface chemistry. Surface chemistry can be determined by a number of factors, including by the presence of stabilizing surfactant molecules introduced during the formulation process. Nanoparticles coated with poloxamer 188 (F68), poloxamer 407 (F127), and polysorbate 80 (P80) have demonstrated uptake in BBB endothelial cells and enhanced accumulation within the brain. However, the impact of surfactants on nanoparticle fate, and specifically on brain extracellular diffusion or intracellular targeting, must be better understood to design nanotherapeutics to efficiently overcome drug delivery barriers in the brain. Here, we evaluated the effect of the biocompatible and commonly used surfactants cholic acid (CHA), F68, F127, P80, and poly (vinyl alcohol) (PVA) on poly (lactic- co -glycolic acid)-poly (ethylene glycol) (PLGA-PEG) nanoparticle transport to and within the brain. The inclusion of these surfactant molecules decreases diffusive ability through brain tissue, reflecting the surfactant's role in encouraging cellular interaction at short length and time scales. After in vivo administration, PLGA-PEG/P80 nanoparticles demonstrated enhanced penetration across the BBB and subsequent internalization within neurons and microglia . Surfactants incorporated into the formulation of PLGA-PEG nanoparticles therefore represent an important design parameter for controlling nanoparticle fate within the brain.

It is an ambitious target to improve overall Hepatocellular Carcinoma therapeutic effects. Recently, MW ablation has emerged as a powerful thermal ablation technique, affording favorable survival with excellent local tumor control. To achieve better therapeutic effects of MW ablation, MW sensitizers are prepared for enhanced MW ablation to preferentially heat tumor territory. However, it is still not practicable for treatment of the orthotopic transplantation tumor. Herein, biocompatible and degradable methoxy poly(ethylene glycol)-poly(lactic-co-glycolic acid) (mPEG-PLGA) microcapsules with hierarchical structure have been designed for microwave-induced tumor therapy. Chemical drug doxorubicin hydrochloride (DOX·HCl), microwave (MW) sensitizers and CT imaging contrast MoS2 nanosheets and MR imaging contrast Fe3O4 nanoparticles are co-incorporated into the microcapsules. In vitro/vivo MR/CT dual-modal imaging results prove the potential application for guiding synergetic therapy and predicting post-therapy tumor progression in the orthotopic transplantation tumor model. After blocking the tumor-feeding arteries, these microcapsules not only exclude the cooling effect by cutting off the blood flow but also enhance MW heating conversion at tumor site. The focused MW heating makes microcapsules mollescent or ruptured and releases DOX·HCl from the microcapsules, achieving the controlled release of drugs for chemical therapy. Compared with MW ablation, 29.4% increase of necrosis diameter of normal liver in rabbit is obtained under MW ablation combined with transcatheter arterial blocking, and the average size of necrosis and inhibition rate of VX-2 liver orthotopic transplantation tumor in rabbit has increased by 129.33% and 73.46%. Moreover, it is proved that the superselectively arterial administration of the as-prepared microcapsules has no recognizable toxicity on the animals. Therefore, this research provides a novel strategy for the construction of MW-induced microcapsules for orthotopic transplantation tumor ablation with the properties of MW sensitizing, superselective arterial blocking, control release and enhanced accumulation of DOX·HCl, and MR/CT dual-modal imaging, which exhibits great potential applications in the field of HCC therapy.

Random skin flaps are commonly used in plastic and reconstructive surgery for patients suffering from severe or large scale wounds or in facial reconstruction. However, skin flaps are sometimes susceptible to partial or complete necrosis at the distal parts of the flaps due to insufficient blood perfusion in the defected area. In order to improve neovascularization in skin flaps, we developed an exogenous growth factor (GF) delivery platform comprised of coacervate-coated poly(lactic-co-glycolic acid) (PLGA) nanofibers. We used a coacervate that is a self-assembled complex of poly(ethylene argininyl aspartate diglyceride) (PEAD) polycation, heparin, and cargo GFs (i.e., vascular endothelial growth factor (VEGF) and/or transforming growth factor beta 3 (TGF-β3)). The coacervate was coated onto a nanofibrous PLGA membrane for co-administration of dual GFs. In vitro proliferation of human dermal fibroblasts and endothelial tube formation using human umbilical vein endothelial cells indicated an enhanced bioactivity of released GFs when both VEGF and TGF-β3 were incorporated into coacervate-coated PLGA nanofibers (Coa-Dual NFs). Moreover, an in vivo study using a mouse skin flap model demonstrated that implantation of Coa-Dual NF reduced necrosis and enhanced blood perfusion in skin flap areas after 10 days, as compared to any single GF-loaded coacervate/PLGA fiber (Coa-Single NF) along with direct administration of the other GF onto the defect site. Moreover, Coa-Dual NFs exhibited a well-composed skin appendage and a significantly higher number of blood vessels. Based upon these results, we conclude that Coa-Dual NFs may stimulate cellular activity by enhancing the bioactivity of the released GF, leading to a synergetic effect of dual GFs for reducing necrosis in the random skin flaps. Therefore, Coa-Dual NFs could be a valuable drug delivery platform for a variety of potential clinical applications for skin tissue regeneration applications.

Biomimetic materials that target the immune system and generate an anti-tumor responses hold promise in augmenting cancer immunotherapy. These synthetic materials can be engineered and optimized for their biodegradability, physical parameters such as shape and size, and controlled release of immune-modulators. As these new platforms enter the playing field, it is imperative to understand their interaction with existing immunotherapies since single-targeted approaches have limited efficacy. Here, we investigate the synergy between a PLGA-based artificial antigen presenting cell (aAPC) and a checkpoint blockade molecule, anti-PD1 monoclonal antibody (mAb). The combination of antigen-specific aAPC-based activation and anti-PD-1 mAb checkpoint blockade induced the greatest IFN-γ secretion by CD8+ T cells in vitro. Combination treatment also acted synergistically in an in vivo murine melanoma model to result in delayed tumor growth and extended survival, while either treatment alone had no effect. This was shown mechanistically to be due to decreased PD-1 expression and increased antigen-specific proliferation of CD8+ T cells within the tumor microenvironment and spleen. Thus, biomaterial-based therapy can synergize with other immunotherapies and motivates the translation of biomimetic combinatorial treatments.

Obesity and overweight, the most serious health problems, are associated with chronic metabolic complications such as type 2 diabetes, insulin resistance, and nonalcoholic fatty liver disease (NAFLD). However, current pharmacological therapies for obesity are challenged by potential side effects, low effectiveness, and low aqueous solubility, which limit their clinical application. Here, we develop nifedipine-loaded nanoparticles (NFD-NPs) that alleviate obesity-related metabolic dysfunction to be used as instruments for translational medicine. Nanoparticles (NPs) composed of poly (lactic-co-glycolic acid) (PLGA) not only enhance water solubility of hydrophobic nifedipine (NFD), a calcium channel blocker, without modifying the chemical structure of NFD for intravenous administration, but also allow prolonged release of NFD in vivo. NFD-NPs do not show cytotoxicity and reduce palmitate-induced protein inclusions and endoplasmic reticulum stress in human hepatoma HepG2 cells. Importantly, tail-vein injection of NFD-NPs into diet-induced obese mice results in sustained retention of NFD-NPs in the liver and suppression of metabolic derangements associated with NAFLD by enhancing autophagic clearance through Ca2+/calmodulin-dependent kinase II (CaMKII) phosphorylation, consequently decreasing diet-induced insulin resistance and improving glucose tolerance. Our findings offer new clinical tools for NP-mediated pharmaceutical strategies to treat NAFLD and its related metabolic dysfunction.

Nanoparticle (NP)-based approaches to cancer drug delivery are challenged by the heterogeneity of the enhanced permeability and retention (EPR) effect in tumors and the premature attrition of payload from drug carriers during circulation. Here we show that such challenges can be overcome by a magnetophoretic approach to accelerate NP delivery to tumors. Payload-bearing poly(lactic-co-glycolic acid) NPs were converted into polymer-iron-oxide nanocomposites (PINCs) by attaching colloidal Fe3O4 onto the surface, via a simple surface modification method using dopamine polymerization. PINCs formed stable dispersions in serum-supplemented medium and responded quickly to magnetic field gradients above 1 kG/cm. Under the field gradients, PINCs were rapidly transported across physical barriers and into cells and captured under flow conditions similar to those encountered in postcapillary venules, increasing the local concentration by nearly three orders of magnitude. In vivo magnetophoretic delivery enabled PINCs to accumulate in poorly vascularized subcutaneous SKOV3 xenografts that did not support the EPR effect. In vivo magnetic resonance imaging, ex vivo fluorescence imaging, and tissue histology all confirmed that the uptake of PINCs was higher in tumors exposed to magnetic field gradients, relative to negative controls.

Despite gaining increasing attention, chelation of multiple active targeting ligands greatly increase the formation probability of protein corona, disabling active targeting. To overcome it, a synergistic retention strategy of RGD-mediated active targeting and radiofrequency (RF) electromagnetic field-enhanced permeability has been proposed here. It is validated that such a special synergistic retention strategy can promote more poly lactic-co-glycolic acid (PLGA)-based capsules encapsulating camptothecin (CPT) and solid DL-menthol (DLM) to enter and retain in tumor in vitro and in vivo upon exposure to RF irradiation, receiving an above 8 fold enhancement in HeLa retention. Moreover, the PLGA-based capsules can respond RF field to trigger the entrapped DLM to generate solid-liquid-gas (SLG) tri-phase transformation for enhancing RF ablation and CPT release. Therefore, depending on the enhanced RF ablation and released CPT and the validated synergistic retention effect, the inhibitory outcome for tumor growth has gained an over 10-fold improvement, realizing RF ablation & chemotherapy synergistic treatment against HeLa solid tumor, which indicates a significant promise in clinical RF ablation.

Despite considerable advances in prostate cancer research, there is a major need for a systemic delivery platform that efficiently targets anti-cancer drugs to sites of disseminated prostate cancer while minimizing host toxicity. In this proof-of-principle study, human mesenchymal stem cells (MSCs) were loaded with poly(lactic-co-glycolic acid) (PLGA) microparticles (MPs) that encapsulate the macromolecule G114, a thapsigargin-based prostate specific antigen (PSA)-activated prodrug . G114-particles (~950nm in size) were internalized by MSCs, followed by the release of G114 as an intact prodrug from loaded cells. Moreover, G114 released from G114 MP-loaded MSCs selectively induced death of the PSA-secreting PCa cell line, LNCaP. Finally, G114 MP-loaded MSCs inhibited tumor growth when used in proof-of-concept co-inoculation studies with CWR22 PCa xenografts, suggesting that cell-based delivery of G114 did not compromise the potency of this pro-drug in-vitro or in-vivo. This study demonstrates a potentially promising approach to assemble a cell-based drug delivery platform, which inhibits cancer growth in-vivo without the need of genetic engineering. We envision that upon achieving efficient homing of systemically infused MSCs to cancer sites, this MSC-based platform may be developed into an effective, systemic ‘Trojan Horse’ therapy for targeted delivery of therapeutic agents to sites of metastatic PCa.

A major hindrance in islet transplantation as a feasible therapeutic approach for patients with type 1 diabetes is the insufficient oxygenation of the grafts, which results in cell death in portions of the implant. Here we address this limitation through the application of oxygen-generating microparticles (MP) and a fibrin-conjugated heparin/VEGF collagen scaffold to support cell survival by using a β cell line and pancreatic rat islets. MP are composed of a polyvinylpyrrolidone/hydrogen peroxide (PVP/H2O2) core and poly(D,L-lactide-co-glycolide) (PLGA) shell, along with immobilized catalase on the shell. The presence of MP is sufficient to reduce hypoxia-induced cell dysfunction and death for both cell types, resulting in localization of hypoxia-inducible factor (HIF-1α) into the cytoplasm and enhanced metabolic function. After co-transplantation of MP and a reduced islet mass (250 islet equivalents) within an angiogenic scaffold in the omental pouch of streptozotocin-induced diabetic nude mice, we have observed significantly promoted graft function as evidenced by improved blood glucose levels, body weight, glucose tolerance, serum C-peptide, and graft revascularization. These results suggest that the developed platform has great potential to enhance the efficacy for implants in cases where the cell dosage is critical for efficacy, such as islet transplantation and ischemic tissues.

Curcumin (Cur) has been demonstrated to have wide pharmacological window including anti-oxidant and anti-inflammatory properties. However, phototoxicity under sunlight exposure and poor biological availability limits its applicability. We have synthesized biodegradable and non-toxic polymer-poly (lactic-co-glycolic) acid (PLGA) encapsulated formulation of curcumin (PLGA-Cur-NPs) of 150 nm size range. Photochemically free curcumin generates ROS, lipid peroxidation and induces significant UVA and UVB mediated impaired mitochondrial functions leading to apoptosis/necrosis and cell injury in two different origin cell lines viz., mouse fibroblasts-NIH-3T3 and human keratinocytes-HaCaT as compared to PLGA-Cur-NPs. Molecular docking studies suggested that intact curcumin from nanoparticles, bind with BAX in BIM SAHB site and attenuate it to undergo apoptosis while upregulating anti-apoptotic genes like BCL2. Real time studies and western blot analysis with specific phosphorylation inhibitor of ERK1 and AKT1/2/3 confirm the involvement of ERK/AKT signaling molecules to trigger the survival cascade in case of PLGA-Cur-NPs. Our finding demonstrates that low level sustained release of curcumin from PLGA-Cur-NPs could be a promising way to protect the adverse biological interactions of photo-degradation products of curcumin upon the exposure of UVA and UVB. Hence, the applicability of PLGA-Cur-NPs could be suggested as prolonged radical scavenging ingredient in curcumin containing products.

Several factors are involved in angiogenesis. To form new blood vessels, we fabricated vehicles carrying an angiogenesis-related peptide (apelin) and gene (vascular endothelial growth factor (VEGF)165) that were internalized by human mesenchymal stem cells (hMSCs). These non-toxic poly-(DL)-lactic-co-glycolic acid (PLGA) nanoparticles (NPs) easily entered hMSCs without cytotoxicity. The negatively charged outer surface of PLGA NPs can be easily complexed with highly positively charged polyethylenimine (PEI) to deliver genes into cells. PLGA NPs complexed with PEI could be coated with negatively charged VEGF plasmid DNA and loaded with apelin. The physical characteristics of these PLGA NPs were determined by size distribution, gel retardation, and morphological analyses. Transfection of VEGF-coated apelin-loaded PLGA NPs resulted in the differentiation of hMSCs into endothelial cells and vascular formation in Matrigel in vitro. Following injection of hMSCs transfected with these PLGA NPs into an ischemic hind limb mouse model, these cells differentiated into endothelial cells and accelerated neovascularization.

Insufficient neovascularization is associated with high levels of resorption and necrosis in autologous and engineered fat grafts. We tested the hypothesis that incorporating angiogenic growth factor into a scaffold–stem cell construct and implanting this construct around a vascular pedicle improves neovascularization and adipogenesis for engineering soft tissue flaps. Poly(lactic-co-glycolic-acid/polyethylene glycol (PLGA/PEG) microspheres containing vascular endothelial growth factor (VEGF) were impregnated into collagen-chitosan scaffolds seeded with human adipose-derived stem cells (hASCs). This setup was analyzed in vitro and then implanted into isolated chambers around a discrete vascular pedicle in nude rats. Engineered tissue samples within the chambers were harvested and analyzed for differences in vascularization and adipose tissue growth. In vitro testing showed that the collagen-chitosan scaffold provided a supportive environment for hASC integration and proliferation. PLGA/PEG microspheres with slow-release VEGF had no negative effect on cell survival in collagen-chitosan scaffolds. In vivo, the system resulted in a statistically significant increase in neovascularization that in turn led to a significant increase in adipose tissue persistence after 8 weeks versus control constructs. These data indicate that our model—hASCs integrated with a collagen-chitosan scaffold incorporated with VEGF-containing PLGA/PEG microspheres supported by a predominant vascular vessel inside a chamber—provides a promising, clinically translatable platform for engineering vascularized soft tissue flap. The engineered adipose tissue with a vascular pedicle could conceivably be transferred as a vascularized soft tissue pedicle flap or free flap to a recipient site for the repair of soft-tissue defects.

Dual responsive nanoparticles are developed for co-delivery of multiple anticancer drugs to target the drug resistance mechanisms of cancer stem-like cells (CSCs). The nanoparticles consist of four polymers approved by the Food and Drug Administration (FDA) for medical use: Poly(D,L-lactide-co-glycolide) (PLGA), Pluronic F127 (PF127), chitosan, and hyaluronic acid (HA). By combining PLGA and PF127 together, more stable and uniform-sized nanoparticles can be obtained than using PLGA or PF127 alone. The HA is used for not only actively targeting CSCs to reduce their drug resistance due to dormancy (i.e., slow metabolism), but also replacing the commonly used poly(vinyl alcohol) as a stabilizing agent to synthesize the nanoparticles using the double-emulsion approach and to allow for acidic pH-triggered drug release and thermal responsiveness. Besides minimizing drug efflux from CSCs, the nanoparticles encapsulated with doxorubicin hydrochloride (DOX, hydrophilic) and irinotecan (CPT, hydrophobic) to inhibit the activity of topoisomerases II and I, respectively, can fight against the CSC drug resistance associated with their enhanced DNA repair and anti-apoptosis. Ultimately, the two drugs-laden nanoparticles can be used to efficiently destroy the CSCs both in vitro and in vivo with up to ~500 times of enhancement compared to the simple mixture of the two drugs.

Tissue engineering is a promising strategy for the repair of large-scale bone defects, in which scaffolds and growth factors are two critical issues influencing the efficacy of bone regeneration. Unfortunately, the broad application of growth factors is limited by their poor stability in the scaffolds. In the present study, the strictly controlled expression of human bone morphogenetic protein-4 (hBMP-4) in the presence of doxycycline is achieved by adding an hBMP-4 gene fragment into a non-viral artificial restructuring plasmid vector (pSTAR) to form the pSTAR-hBMP-4 plasmid (phBMP-4). Furthermore, the controlled release of phBMP-4 is obtained with an electroactive tissue engineering scaffold, generated by combining a triblock copolymer of poly( l -lactic acid)-block-aniline pentamer-block-poly( l -lactic acid) (PLA-AP) with poly(lactic-co-glycolic acid)/hydroxyapatite (PLGA/HA). This PLGA/HA/PLA-AP/phBMP-4 composite scaffold, with controlled gene release and Dox-regulated gene expression upon electrical stimulation, operating synergistically, exhibits an improved cell proliferation ability, enhanced osteogenesis differentiation in vitro, and effective bone healing in vivo in a rabbit radial defect model. Taking these results together, the proposed smart PLGA/HA/PLA-AP/phBMP-4 scaffold lays a solid theoretical and experimental basis for future applications of such multi-functional materials in bone tissue engineering to help patients in need.

Nano-sized particles (NPs) of various materials have been extensively used as therapeutic and diagnostic agents, drug delivery systems, and biomedical devices. However, the biological impacts of NP exposure during early embryogenesis on following development and next generations have not been investigated. Here, we demonstrated that polylactic-co-glycolic acid (PLGA)-NPs were not toxic and did not perturb development of preimplantation mouse embryos in vitro. Moreover, subsequent fetal development in vivo after embryo transfer proceeded normally and healthy pups were born without any genetic aberrations, suggesting biosafety of PLGA-NPs during developmental processes. TRITC-labeled PLGA-NPs, named TRITC nano-tracer (TnT) were used to visualize the successful delivery of the NPs into sperms, oocytes and early embryos. Various molecular markers for early embryogenesis demonstrated that TnT treatment at various developmental stages did not compromise embryo development to the blastocyst. mRNA-Seq analyses reinforced that TnT treatment did not significantly affect mRNA landscapes of blastocysts which undergo embryo implantation critical for following developmental processes. Moreover, when 2-cell embryos exposed to TnT were transferred into pseudopregnant recipients, healthy offspring were born without any distinct morphologic and chromosomal abnormalities. TnT treatment did not affect the sex ratio of the exposed embryos after birth. When mated with male mice, female mice that were exposed to TnT during early embryogenesis produced a comparable number of pups as control females. Furthermore, the phenotypes of the offspring of mice experienced TnT at their early life clearly demonstrated that TnT did not elicit any negative transgenerational effects on mammalian development.

Bioengineering strategies to enhance the natural targeting function of nanocarriers would expand their therapeutic applications. Here, we designed bioengineered stem cell membrane-functionalized nanocarriers (BSMNCs) harboring C-X-C chemokine receptor type 4 (CXCR4) to achieve robust targeting and also to increase their retention time in ischemic tissue. Stem cell membrane coated nanocarrier (SMNCs) or poly (lactic-co -glycolic acid) (PLGA) nanocarriers (PNCs) and BSMNCs were prepared by functionalizing PNCs with human adipose-derived stem cells (hASCs) membranes and hASCs engineered to overexpress CXCR4-receptor, respectively. The functionalization of PNCs with stem cell membranes derived from hASCs significantly enhance the nanocarrier penetration across endothelial cell barrier compare to PNCs. In addition, stem cell membrane functionalization on PNCs also significantly decreased the nanoparticles uptake in J774 (murine) and THP (human) macrophages respectively from 84% to 76%–29% and 24%. Interestingly, BSMNCs showed much higher level of accumulation in ischemic tissue than SMNCs. Systemic retro-orbital injection of BSMNCs loaded with VEGF into mice with hindlimb ischemia resulted substantially enhancement of blood reperfusion , muscle repair, and limb salvage compared to animals treated with SMNCs loaded with similar concentration of VEGF. The reported strategy could be used to create biocompatible and custom-tailored biomimetic nanoparticles with various hybrid functionalities, which may overcome the limitations of current nanoparticle-based therapeutic and imaging platforms.

There are disadvantages in Elastic Stable Intramedullary Nailing (ESIN) of forearm-shaft fractures, such as the need of implant removal. Biodegradable Intramedullary Nailing (BIN) is a new technique developed for these fractures. We hypothesized that there is no difference in rotational ROM between the patients treated by BIN vs. ESIN. A randomized, controlled clinical trial included patients, aged 5–15 years, requiring surgery for forearm-shaft fractures. Biodegradable polylactide-co-glycolide (PLGA) nails (Activa IM-Nail™, Bioretec Ltd., Finland) were used in 19 and titanium nails (TEN ®, SynthesDePuy Ltd., USA) in 16 patients. Rotational ROM of forearm after two years was the primary outcome. Elbow and wrist ROM, pain and radiographic bone healing were secondary outcomes. Forearm rotation was mean 162° and 151° in BIN and ESIN groups, respectively (P = 0.201). No difference between the groups was found in any other ROMs. Three cases in the ESIN vs. none in the BIN group reported pain (P = 0.113). There was no clinically significant residual angulation in radiographs . Two adolescents in the BIN group vs. none in the ESIN (P = 0.245) were excluded because of implant failure ; another two with complete bone union suffered from re-injury. Therefore, satisfactory implant stability among older children needs to be studied.

Nogo-B receptor (NgBR) plays fundamental roles in regulating angiogenesis, vascular development, and the epithelial-mesenchymal transition (EMT) of cancer cells. However, the therapeutic effect of NgBR blockade on tumor vasculature and malignancy is unknown, investigations on which requires an adequate delivery system for small interfering RNA against NgBR (NgBR siRNA). Here a surface charge switchable polymeric nanoparticle that was sensitive to the slightly acidic tumor microenvironment was developed for steady delivery of NgBR siRNA to tumor tissues. The nanoformulation was constructed by conjugating 2, 3-dimethylmaleic anhydride (DMMA) molecules to the surface amines of micelles formed by cationic co-polymer poly(lactic-co-glycolic acid)2-poly(ethylenimine) and subsequent absorption of NgBR siRNAs. The nanoparticles remained negatively charged in physiological condition and smartly converted to positive surface charge due to tumor-acidity-activated shedding of DMMA. The charge conversion facilitated cellular uptake of siRNAs and in turn efficiently depleted the expression of NgBR in tumor-bearing tissues. Silencing of NgBR suppressed endothelial cell migration and tubule formation, and reverted the EMT process of breast cancer cells. Delivery of the nanoformulation to mice bearing orthotopic breast carcinoma showed no effect on tumor growth, but led to remarkable decrease of distant metastasis by normalizing tumor vessels and suppressing the EMT of breast cancer cells. This study demonstrated that NgBR is a promising therapeutic target in abnormal tumor vasculature and aggressive cancer cells, and the tumor-responsive nanoparticle with the feature of charge transformation offers great potential for tumor-specific delivery of gene therapeutics.

Steroid-associated osteonecrosis (SAON) may lead to joint collapse and subsequent joint replacement. Poly lactic-co-glycolic acid/tricalcium phosphate (P/T) scaffold providing sustained release of icaritin (a metabolite of Epimedium-derived flavonoids) was investigated as a bone defect filler after surgical core-decompression (CD) to prevent femoral head collapse in a bipedal SAON animal model using emu (a large flightless bird). The underlying mechanism on SAON was evaluated using a well-established quadrupedal rabbit model. Fifteen emus were established with SAON, and CD was performed along the femoral neck for the efficacy study. In this CD bone defect, a P/T scaffold with icaritin (P/T/I group) or without icaritin (P/T group) was implanted while no scaffold implantation was used as a control. For the mechanistic study in rabbits, the effects of icaritin and composite scaffolds on bone mesenchymal stem cells (BMSCs) recruitment, osteogenesis, and anti-adipogenesis were evaluated. Our efficacy study showed that P/T/I group had the significantly lowest incidence of femoral head collapse, better preserved cartilage and mechanical properties supported by more new bone formation within the bone tunnel. For the mechanistic study, our in vitro tests suggested that icaritin enhanced the expression of osteogenesis related genes COL1α, osteocalcin, RUNX2, and BMP-2 while inhibited adipogenesis related genes C/EBP-ß, PPAR-γ, and aP2 of rabbit BMSCs. Both P/T and P/T/I scaffolds were demonstrated to recruit BMSCs both in vitro and in vivo but a higher expression of migration related gene VCAM1 was only found in P/T/I group in vitro. In conclusion, both efficacy and mechanistic studies show the potential of a bioactive composite porous P/T scaffold incorporating icaritin to enhance bone defect repair after surgical CD and prevent femoral head collapse in a bipedal SAON emu model.

A drug-eluting stent (DES) is one of the commonly used treatment techniques in percutaneous coronary intervention (PCI). Sirolimus (SRL) has been widely used for DES as a drug for suppressing neointimal hyperplasia causing restenosis. Phytoncides (PTC) are compounds released from trees and plants, and their solutions contain monoterpenoids such as α-pinene, careen, and myrceen. Some studies have reported that these components exhibit antioxidant, antimicrobial, and anti-inflammatory activities. We hypothesized that PTC may become an alternative drug to SRL for DES, exhibiting alleviated side effects as compared to SRL. A PTC-incorporated stent was compared with an SRL-incorporated stent in terms of physicochemical, pharmacokinetic, and biological properties. In in vitro studies, the effects of each drug on cells were investigated. The results showed that both drugs exhibited similar cytotoxicity, anti-inflammation, and antiproliferation effects. However, these effects resulted from different mechanisms associated with cells, as seen in the immunofluorescence result. An in vivo assay showed that the lumen area was significantly larger and the neointimal area was significantly smaller in SRL- and PTC-loaded stents compared to a drug-unloaded stent. These results suggest that phytoncide can be a feasible alternative drug to SRL for advanced DES although more studies are needed.

siRNA-loaded nanoparticles (NPs) administered systemically can overcome the poor stability and rapid elimination of free double-stranded RNA in circulation, resulting in increased tumor accumulation and efficacy. siRNA against osteopontin (siOPN), a protein involved in breast cancer development, was encapsulated in poly(D,L-lactic-co-glycolic acid) NPs by a double emulsion solvent diffusion (DESD) technique. We also compared the effect of polyethylenimine (PEI) molecular weight (800 Da and 25 kDa), used as the counter-ion for siRNA complexation, on the physicochemical properties of the NPs, cytotoxicity, and cellular uptake. NPs prepared by the DESD technique were obtained at the desired size (∼170 nm) using both types of PEIs, and were characterized with a neutral surface charge, high encapsulation yield (up to ∼60%), siOPN concentration of 5.6–8.4 μg/mg, stability in physiologic conditions in vitro and in vivo, and long-term shelf-life stability (> 3 years). The NPs prepared using both PEIs exhibited no cytotoxicity in primary smooth muscle culture, and no detrimental effect on mice liver enzymes following their IV administration. Following cellular uptake and biodistribution studies, the therapeutic potential of the NPs was demonstrated by a significant decrease of tumor progression and size in an ectopic xenograft model of mammary carcinoma in mice.

Antigen-specific treatments are highly desirable for autoimmune diseases in contrast to treatments which induce systemic immunosuppression. A novel antigen-specific therapy has been developed which, when administered semi-therapeutically, is highly efficacious in the treatment of the mouse model for multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). The treatment uses dual-sized, polymeric microparticles (dMPs) loaded with specific antigen and tolerizing factors for intra- and extra-cellular delivery, designed to recruit and modulate dendritic cells toward a tolerogenic phenotype without systemic release. This approach demonstrated robust efficacy and provided complete protection against disease. Therapeutic efficacy required encapsulation of the factors in controlled-release microparticles and was antigen-specific. Disease blocking was associated with a reduction of infiltrating CD4+ T cells, inflammatory cytokine-producing pathogenic CD4+ T cells, and activated macrophages and microglia in the central nervous system. Furthermore, CD4+ T cells isolated from dMP-treated mice were anergic in response to disease-specific, antigen-loaded splenocytes. Additionally, the frequency of CD86hiMHCIIhi dendritic cells in draining lymph nodes of EAE mice treated with Ag-specific dMPs was reduced. Our findings highlight the efficacy of microparticle-based drug delivery platform to mediate antigen-specific tolerance, and suggest that such a multi-factor combinatorial approach can act to block autoimmunity.

Drug resistance and toxicity are major obstacles in cancer chemotherapy. Combination therapies can overcome resistance, and synergies can minimize dosing. Polymer nanocarriers are interesting vehicles for cancer therapeutics for their delivery and tumor targeting abilities. We synthesized a multi-layered polymer nanoparticle (MLNP), comprising of poly(lactic-co-glycolic acid) with surface polyethyleneimine and functional peptides, for targeted drug and gene delivery. We confirmed the particle's ability to inhibit tumor growth through synergistic action of the drug and gene product. MLNPs achieved transfection levels similar to lipofectamine, while maintaining minimal cytotoxicity. The particles delivered camptothecin (CPT), and plasmid encoding TNF related apoptosis inducing ligand (pTRAIL) (CT MLNPs), and synergistically inhibited growth of multiple cancer cells in vitro. The synergy of co-delivering CPT and pTRAIL via CT MLNPs was confirmed using the Chou-Talalay method: the combination index (CI) values at 50% inhibition ranged between 0.31 and 0.53 for all cell lines. Further, co-delivery with MLNPs resulted in a 3.1-15 fold reduction in CPT and 4.7-8.0 fold reduction in pTRAIL dosing. CT MLNPs obtained significant HCT116 growth inhibition in vivo compared to monotherapy. These results support our hypothesis that MLNPs can deliver both small molecules and genetic agents towards synergistically inhibiting tumor growth.

Stem cell-based therapy has been proposed as an enabling alternative not only for the treatment of diseases but also for the regeneration of tissues beyond complex surgical treatments or tissue transplantation. In this study, we approached a conceptual platform that can integrate stem cells into a multiscale patterned substrate for bone regeneration. Inspired by human bone tissue, we developed hierarchically micro- and nanopatterned transplantable patches as synthetic extracellular matrices by employing capillary force lithography in combination with a surface micro-wrinkling method using a poly(lactic-co-glycolic acid) (PLGA) polymer. The multiscale patterned PLGA patches were highly flexible and showed higher tissue adhesion to the underlying tissue than did the single nanopatterned patches. In response to the anisotropically multiscale patterned topography, the adhesion and differentiation of human mesenchymal stem cells (hMSCs) were sensitively controlled. Furthermore, the stem cell patch composed of hMSCs and transplantable PLGA substrate promoted bone regeneration in vivo when both the micro- and nanotopography of the substrate surfaces were synergistically combined. Thus, our study concludes that multiscale patterned transplantable stem cell patches may have a great potential for bone regeneration as well as for various regenerative medicine approaches.

Combination cancer therapy has emerged as crucial approach for achieving superior anti-cancer efficacy. In this study, we developed a strategy by localized co-delivery of PLK1shRNA/polylysine-modified polyethylenimine (PEI-Lys) complexes and doxorubicin (DOX) using biodegradable, thermosensitive PLGA-PEG-PLGA hydrogels for treatment of osteosarcoma. When incubated with osteosarcoma Saos-2 and MG-63 cells, the hydrogel containing PLK1shRNA/PEI-Lys and DOX displayed significant synergistic effects in promoting the apoptosis of osteosarcoma cells in vitro. After subcutaneous injection of the hydrogel containing PLK1shRNA/PEI-Lys and DOX beside the tumors of nude mice bearing osteosarcoma Saos-2 xenografts, the hydrogels exhibited superior antitumor efficacy in vivo compared to the hydrogels loaded with PLK1shRNA/PEI-Lys or DOX alone. It is noteworthy that the combination treatment in vivo led to almost complete suppression of tumor growth up to 16 days, significantly enhanced PLK1 silencing, higher apoptosis of tumor masses, as well as increased cell cycle regulation. Additionally, ex vivo histological analysis of major organs of the mice indicated that the localized treatments showed no obvious damage to the organs, suggesting lower systemic toxicity of the treatments. Therefore, the strategy of localized, sustained co-delivery of PLK1shRNA and DOX by using the biodegradable, injectable hydrogel may have potential for efficient clinical treatment of osteosarcoma.

Components of the blood have been proposed as potential therapeutic targets for improving cellular regeneration after injury and neurodegenerative disease. In this work, thrombin is shown to increase endogenous neural progenitor proliferation in the intact murine spinal cord. A local injection of heparin before a spinal cord injury reduces cell proliferation and astrogliogenesis associated with scarring. We sought to create depot-formulations of PLGA microsphere and Pluronic F-127 for sustained local delivery of two thrombin inhibitors, heparin and hirudin. Each hydrogel depot-formulation showed delayed drug release compared to microspheres or hydrogel alone. Animals with a lateral demyelination lesion showed a reduction in CD68+ macrophages when treated with hirudin-loaded PLGA/F-127 gels compared to control and heparin-treated animals. Moreover, hirudin-loaded materials showed an accelerated recovery in coordinated stepping and increased oligodendrocyte densities. Together, these data demonstrate that controlled delivery of hirudin accelerates functional recovery from a demyelination lesion in the spinal cord.

Mesenchymal stem cells (MSCs) are multipotent cells capable of proliferating and differentiating into several lineages. In regenerative medicine, their potential as a resource for tissue-replacement therapy is receiving much attention. However, transplanting MSCs to repair larger bone defects in animal models has so far proved disappointing. Here we report on the healing of both critical-sized (5 mm) and massive (15 mm) full-thickness femur defects in rats by implanting a uniquely fabricated PLGA scaffold seeded with MSCs pre-differentiated in vitro into cartilage-forming chondrocytes (MSC-DCs). This strategy closely mimics endochondral ossification, the process by which long bones develop in nature. It is thought that because the transplanted MSC-DCs induced natural bone formation, the defect size was not critical to the outcome. Crucially, after 8 weeks the mean biomechanical strength of femora with the massive 15 mm implant reached 75% that of a normal rat femur, while in the case of 5 mm implants there was no significant difference. Successful healing was also highly reproducible, with bone union occurring in all treated animals examined radiologically 8 or 16 weeks after surgery.

The purification of human adipose-derived stem cells (hADSCs) from human adipose tissue cells (stromal vascular fraction) was investigated using membrane filtration through poly(lactide-co-glycolic acid)/silk screen hybrid membranes. Membrane filtration methods are attractive in regenerative medicine because they reduce the time required to purify hADSCs (i.e., less than 30 min) compared with conventional culture methods, which require 5-12 days. hADSCs expressing the mesenchymal stem cell markers CD44, CD73, and CD90 were concentrated in the permeation solution from the hybrid membranes. Expression of the surface markers CD44, CD73, and CD99 on the cells in the permeation solution from the hybrid membranes, which were obtained using 18 mL of feed solution containing 50 × 10⁴ cells, was statistically significantly higher than that of the primary adipose tissue cells, indicating that the hADSCs can be purified in the permeation solution by the membrane filtration method. Cells expressing the stem cell-associated marker CD34 could be successfully isolated in the permeation solution, whereas CD34⁺ cells could not be purified by the conventional culture method. The hADSCs in the permeation solution demonstrated a superior capacity for osteogenic differentiation based on their alkali phosphatase activity, their osterix gene expression, and the results of mineralization analysis by Alizarin Red S and von Kossa staining compared with the cells from the suspension of human adipose tissue. These results suggest that the hADSCs capable of osteogenic differentiation preferentially permeate through the hybrid membranes.

A major cross-cutting problem for glioma therapy is the poor extravasation and penetration of the payload drug in target glioma parenchyma. Here, to overcome these obstacles, a tumor vessel recognizing and tumor penetrating system is developed by functionalizating the poly (ethyleneglycol)-poly (L-lactic-co-glycolic acid) nanoparticles with an iNGR moiety (iNGR-NP). The nanoparticulate formulation is expected to achieve specific deep penetration in the tumor tissue by initially binding to aminopeptidase N, with iNGR proteolytically cleaved to CRNGR, and then bind with neuropilin-1 to mediate deep penetration in the tumor parenchyma. iNGR-NP exhibits significantly enhanced cellular uptake in human umbilical vein endothelial cells, improves the anti-proliferation and anti-tube formation abilities of paclitaxel in vitro. Following intravenous administration, iNGR-NP present favorable pharmacokinetic and tumor homing profiles. Glioma distribution and penetration assays confirm that iNGR-NP achieve the highest accumulation and deepest penetration at the glioma sites. The anti-glioma efficacy of paclitaxel-loaded iNGR-NP is verified by its improved anti-angiogenesis activity and the significantly prolonged survival time in mice bearing intracranial glioma. These evidences highlight the potential of iNGR-decorated nanoparticles in overcoming the leading edge problem in anti-glioma drug delivery.

Dentin regeneration remains a great challenge in clinic. Dental pulp stem cells (DPSCs) actively contribute to dentinogenesis, which is orchestrated by a spectrum of signaling factors. However, the exact mechanism underlying the reparative dentin regeneration process is largely unknown and the application of DPSCs in the repair of dentin defect is thus limited. Here, using a rat reparative dentin regeneration model, we observed that DPSCs underwent a proliferation phase followed by a differentiation phase after dental injury. A transient elevation of nephroblastoma overexpressed (NOV, or CCN3) expression correlated with this progressive dental tissue restoration process. Further studies revealed that over-expression of CCN3 promoted human DPSCs proliferation via activation of Notch. Moreover, using cocultured cells (DPSCs/CCN3 and DPSCs) in vitro and the cocultured cells-poly (lactic-co-glycolic acid) (PLGA) scaffold complex in vivo, we demonstrated that CCN3 was capable of promoting mineralization in a non-cell autonomous manner through promoting secretion of BMP2. CCN3 can promote dentinogenesis by coordinating proliferation and odontoblastic differentiation of DPSCs via modulating Notch and BMP2 signaling pathways and CCN3 is a promising therapeutic target in dentin tissue engineering.

Nanoparticles of biodegradable polymers (NPs) have been widely used for drug delivery. However, there has been little research on their fate after internalized into the cells. We show in this research by using docetaxel as a model anticancer drug, which is formulated in the cholic acid conjugated nanoparticles of poly(lactic-co-glycolic acid (PLGA NPs) that the NPs induce autophagy of the cancer cells and thus may hinder the advantages of the nanomedicine. Moreover, we show both in vitro and in vivo that co-administration of autophagy inhibitors such as 3-methyladenine (3-MA) and Chloroquine (CQ) could greatly enhance the therapeutic effects of the nanoparticle formulation. The IC50 values of the drug formulated in the PLGA NPs after 24 h treatment with no autophagy inhibitor or in combination with 10 mm 3-MA or 30 μm CQ are 38.27 ± 1.23, 6.7 ± 1.05, 4.78 ± 1.75 μg/mL, which implie 5.7 or 8,0 fold efficient by the autophagy inhibition respectively. Moreover, both the volume and the weight of the shrunk tumor of the mice after 20 day treatment with the PLGA NPs formulation combined with 3-MA or CQ are found to be only about a half in comparison with the treatment with the PLGA NPs formulation alone. In this research, we reported such a new mechanism of cancer cells to have PLGA NPs captured and degraded by auto-lysosomes. The findings provide advanced knowledge for development of nanomedicine for clinical application.

We report a strategy to make use of poly(lactic-co-glycolic acid) nanoparticle (PLGA NPs) for co-delivery of docetaxel (DTX) as a model anticancer drug together with vitamin E TPGS. The latter plays a dual role as a pore-forming agent in the nanoparticles that may result in smaller particle size, higher drug encapsulation efficiency and faster drug release, and also as a bioactive agent that could inhibit P-glycoprotein to overcome multi-drug resistance of the cancer cells, The DTX-loaded PLGA NPs of 0, 10, 20 and 40% TPGS were prepared by the nanoprecipitation method and then characterized for their size and size distribution, surface morphology, physical status and encapsulation efficiency of the drug in the NPs. All four NPs were found of size ranged 100-120 nm and EE ranged 85-95% at drug loading level around 10%. The in vitro evaluation showed that the 48 h IC50 values of the free DTX and the DTX-loaded PLGA NPs of 0, 10, 20% TPGS were 2.619 and 0.474, 0.040, 0.009 μg/mL respectively, which means that the PLGA NPs formulation could be 5.57 fold effective than the free DTX and that the DTX-loaded PLGA NPs of 10 or 20% TPGS further be 11.85 and 52.7 fold effective than the DTX-loaded PLGA NPs of no TPGS (therefore, 66.0 and 284 fold effective than the free DTX). Xenograft tumor model and immunohistological staining analysis further confirmed the advantages of the strategy of co-delivery of anticancer drugs with TPGS by PLGA NPs.

M cells, the key players of the mucosal immunity induction, are one of the intestinal barriers for the efficient delivery of vaccines to mucosal immune tissues. To overcome the barrier, we have developed an efficient oral vaccine carrier that constitutes poly (lactic-co-glycolic acid) (PLGA) microparticle coated with M cell targeting peptide. In this study, a membrane protein B of Brachyspira hyodysenteriae (BmpB) as a model vaccine against swine dysentery was loaded into porous PLGA microparticles (MPs). The PLGA MPs were further coated with the water-soluble chitosan (WSC) conjugated with M cell homing peptide (CKS9) to prepare BmpB-CKS9-WSC-PLGA MPs. Oral immunization of BmpB vaccine with CKS9-WSC-PLGA MPs in mice showed elevated secretory IgA responses in the mucosal tissues and systemic IgG antibody responses, providing a complete immune response. Specifically, the immunization with these MPs demonstrated to induce both Th1- and Th2-type responses based on elevated IgG1 and IgG2a titers. The elevated immune responses were attributed to the enhanced M cell targeting and transcytosis ability of CKS9-WSC-PLGA MPs to Peyer's patch regions. The high binding affinity of CKS9-WSC-PLGA MPs with the M cells to enter into the Peyer's patch regions of mouse small intestine was investigated by closed ileal loop assay and it was further confirmed by confocal laser scanning microscopy. These results suggest that the M cell targeting approach used in this study is a promising tool for targeted oral vaccine delivery.