One-Pot Synthesis of Bioinspired Peptide-Decorated Apatite Nanoparticles for Nanomedicine.

Hybrid organic-inorganic bio-inspired apatite nanoparticles (NPs) are attractive for biomedical applications and especially in nanomedicine. Unfortunately, their applications in nanomedicine are limited by their broad particle size distributions and uncontrolled drug loading due to their multistep synthesis process.  Besides, very few attempts at exposing bioactive peptides on apatite NPs are made. In this work, an original one-pot synthesis of well-defined bioactive hybrid NPs composed of a mineral core of bioinspired apatite surrounded by an organic corona of bioactive peptides is reported. Dual stabilizing-bioactive agents, phosphonated polyethylene glycol-peptide conjugates, are prepared and directly used during apatite precipitation i) to form the organic corona during apatite precipitation, driving the size and shape of resulting hybrid NPs with colloidal stabilization and ii) to expose peptide moieties (RGD or YIGSR sequences) at the NPs periphery in view of conferring additional surface properties to enhance their interaction with cells. Here, the success of this approach is demonstrated, the functionalized NPs are fully characterized by Fourier-transform infrared, Raman, X-ray diffraction, solid and liquid state NMR, transmission electron microscopy, and dynamic light scattering, and their interaction with fibroblast cells is followed, unveiling a synergistic proliferative effect.

Controlled Self-Assembly of DNA-Mimicking Nanotubes to Form a Layer-by-Layer Scaffold for Homeostatic Tissue Constructs.

Various biomaterial scaffolds have been developed for improving stem cell anchorage and function in tissue constructs for in vitro and in vivo uses. Growth factors are typically applied to scaffolds to mediate cell differentiation. Conventionally, growth factors are not strictly localized in the scaffolds; thus, they may leak into the surrounding environment, causing undesired side effects on tissues or cells. Hence, there is a need for improved tissue construct strategies based on highly localized drug delivery and a homeostatic microenvironment. This study developed an injectable nanomatrix (NM) scaffold with a layer-by-layer structure inside each nanosized fiber of the scaffold based on controlled self-assembly at the molecular level. The NM was hierarchically assembled from Janus base nanotubes (JBNTs), matrilin-3, and transforming growth factor ß-1 (TGF-ß1) via bioaffinity. JBNTs, which form the NM backbone, are novel DNA-inspired nanomaterials that mimic the natural helical nanostructures of collagens. The chondrogenic factor, TGF-ß1, was enveloped in the inner layer inside the NM fibers to prevent its release. Matrilin-3 was incorporated into the outer layer to create a cartilage-mimicking microenvironment and to maintain tissue homeostasis. Interestingly, human mesenchymal stem cells (hMSCs) had a strong preference to anchor along the NM fibers and formed a localized homeostatic microenvironment. Therefore, this NM has successfully generated highly organized structures via molecular self-assembly and achieved localized drug delivery and stem cell anchorage for homeostatic tissue constructs.

Endogenous FGF-2 levels impact FGF-2/BMP-2 growth factor delivery dosing in aged murine calvarial bone defects.

Bone repair in elderly mice has been shown to be improved or negatively impacted by supplementing the highly osteogenic bone morphogenetic protein-2 (BMP-2) with fibroblast growth factor-2 (FGF-2). To better predict the outcome of FGF-2 supplementation, we investigated whether endogenous levels of FGF-2 play a role in optimal dosing of FGF-2 for augmenting BMP-2 activity in elderly mice. In vivo calvarial bone defect studies in Fgf2 knockout mice with wildtype controls were conducted with the growth factors delivered in a highly localized manner from a biomimetic calcium phosphate/polyelectrolyte multilayer coating applied to a bone graft substitute. Endogenous FGF-2 levels were measured in old mice versus young and found to decrease with age. Optimal dosing for improving bone defect repair correlated with levels of endogenous FGF-2, with a larger dose of FGF-2 required to have a positive effect on bone healing in the Fgf2 knockout mice. The same dose in wildtype old mice, with higher levels of FGF-2, promoted chondrogenesis and increased osteoclast activity. The results suggest a personalized medicine approach, based on a knowledge of endogenous levels of FGF-2, should guide FGF-2 supplementation in order to avoid provoking excessive bone resorption and cartilage formation, both of which inhibited calvarial bone repair.

Self-assembled biomimetic Nano-Matrix for stem cell anchorage.

Mesenchymal stem cells (MSCs) have been widely applied in biomedicine due to their ability to differentiate into many different cell types and their ability to synthesize a broad spectrum of growth factors and cytokines that directly and indirectly influence other cells in their vicinity. To guide MSC infiltration to a bone fracture site, we developed a novel self-assembled Nano-Matrix which can be used as an injectable scaffold to repair bone fractures. The Nano-Matrix is formed by Janus base nanotubes (JBNTs) and fibronectin (FN). JBNTs are nucleobase-derived nanotubes mimicking collagen fibers, and FN is one of the cell adhesive glycoproteins which is responsible for cell-extracellular matrix interactions and guides stem cell migration and differentiation to desired cells types. Here, we demonstrated the successful fabrication and characterization of the JBNT/FN Nano-Matrix as well as its excellent bioactivity that encouraged human MSC migration and adhesion. This work lays a solid foundation for using the Nano-Matrix as an injectable approach to improve MSC retention and function during bone fracture healing.

Controlled M1-to-M2 transition of aged macrophages by calcium phosphate coatings.

Older adults suffer from weakened and delayed bone healing due to age-related alterations in bone cells and in the immune system. Given the interaction between the immune system and skeletal cells, therapies that address deficiencies in both the skeletal and the immune system are required to effectively treat bone injuries of older patients. The sequence of macrophage activation observed in healthy tissue repair involves a transition from a pro-inflammatory state followed by a pro-reparative state. In older patients, inflammation is slower to resolve and impedes healing. The goal of this study was to design a novel drug delivery system for temporal guidance of the polarization of macrophages using bone grafting materials. A biomimetic calcium phosphate coating (bCaP) physically and temporally separated the pro-inflammatory stimulus interferon-gamma (IFNγ) from the pro-reparative stimulus simvastatin (SIMV). Effective doses were identified using a human monocyte line (THP-1) and testing culminated with bone marrow macrophages obtained from old mice. Sequential M1-to-M2 activation was achieved with both cell types. These results suggest that this novel immunomodulatory drug delivery system holds potential for controlling macrophage activation in bones of older patients.

Cell Type Influences Local Delivery of Biomolecules from a Bioinspired Apatite Drug Delivery System.

Recently, the benefit of step-wise sequential delivery of fibroblast growth factor-2 (FGF-2) and bone morphogenetic protein-2 from a bioinspired apatite drug delivery system on mouse calvarial bone repair was demonstrated. The thicknesses of the nanostructured poly-l-Lysine/poly-l-Glutamic acid polyelectrolyte multilayer (PEM) and the bone-like apatite barrier layer that make up the delivery system, were varied. The effects of the structural variations of the coating on the kinetics of cell access to a cytotoxic factor delivered by the layered structure were evaluated. FGF-2 was adsorbed into the outer PEM, and cytotoxic antimycin-A (AntiA) was adsorbed to the substrate below the barrier layer to detect the timing of the cell access. While MC3T3-E1 osteoprogenitor cells accessed AntiA after three days, the RAW 264.7 macrophage access occurred within 4 h, unless the PEM layer was removed, in which case the results were reversed. Pits were created in the coating by the RAW 264.7 macrophages and initiated delivery, while the osteoprogenitor cell access to drugs occurred through a solution-mediated coating dissolution, at junctions between the islands of crystals. Macrophage-mediated degradation is therefore a mechanism that controls drug release from coatings containing bioinspired apatite.

* Calvarial Bone Regeneration Is Enhanced by Sequential Delivery of FGF-2 and BMP-2 from Layer-by-Layer Coatings with a Biomimetic Calcium Phosphate Barrier Layer.

A drug delivery coating for synthetic bone grafts has been developed to provide sequential delivery of multiple osteoinductive factors to better mimic aspects of the natural regenerative process. The coating is composed of a biomimetic calcium phosphate (bCaP) layer that is applied to a synthetic bone graft and then covered with a poly-l-Lysine/poly-l-Glutamic acid polyelectrolyte multilayer (PEM) film. Bone morphogenetic protein-2 (BMP-2) was applied before the coating process directly on the synthetic bone graft and then, bCaP-PEM was deposited followed by adsorption of fibroblast growth factor-2 (FGF-2) into the PEM layer. Cells access the FGF-2 immediately, while the bCaP-PEM temporally delays the cell access to BMP-2. In vitro studies with cells derived from mouse calvarial bones demonstrated that Sca-1 and CD-166 positive osteoblast progenitor cells proliferated in response to media dosing with FGF-2. Coated scaffolds with BMP-2 and FGF-2 were implanted in mouse calvarial bone defects and harvested at 1 and 3 weeks. After 1 week in vivo, proliferation of cells, including Sca-1+ progenitors, was observed with low dose FGF-2 and BMP-2 compared to BMP-2 alone, indicating that in vivo delivery of FGF-2 activated a similar population of cells as shown by in vitro testing. At 3 weeks, FGF-2 and BMP-2 delivery increased bone formation more than BMP-2 alone, particularly in the center of the defect, confirming that the proliferation of the Sca-1 positive osteoprogenitors by FGF-2 was associated with increased bone healing. Areas of bone mineralization were positive for double fluorochrome labeling of calcium and alkaline phosphatase staining of osteoblasts, along with increased TRAP+ osteoclasts, demonstrating active bone formation distinct from the bone-like collagen/hydroxyapatite scaffold. In conclusion, the addition of a bCaP layer to PEM delayed access to BMP-2 and allowed the FGF-2 stimulated progenitors to populate the scaffold before differentiating in response to BMP-2, leading to improved bone defect healing.

Implant-guided supracrestal alveolar bone growth using scaffolds, BMP-2, and novel scaffold-retaining device.

OBJECTIVE: To evaluate the efficacy of various scaffold systems and a Ti scaffold-retaining device with and without non-glycosylated rhBMP-2 (BMP-2) for increasing the vertical alveolar bone growth in the intra-oral mini-pig model. METHODS: Forty-eight Straumann Bone Level implants with hydrophilic (SLActive) surfaces were partially embedded in mandibles of 12 adult mini-pigs with the shoulder of the implant located 3 mm above the bone crest. Twenty-four implants were placed in conjunction with BMP-2 (50 µg) incorporated within resorbable scaffolds. Twenty-four additional control implants were placed with scaffolds only. Scaffolds were placed around the implant and stabilized with a newly developed Ti "umbrella" scaffold retainer. Scaffolds included (i) HA-coated collagen (Healos); (ii) biphasic HA/ß-TCP crystals (Straumann Bone Ceramic, SBC); and (iii) SBC crystals infused with polyethylene glycol (PEG) hydrogel. Eight test and control pairs for each scaffold group were implanted. At 9 weeks, soft tissue healing was assessed and the extent of new vertical bone was evaluated with microCT and histomorphometry. RESULTS: microCT analysis revealed a mean of 167 ± 47 mm3 new supracrestal mineralized tissue volume formation around the test sites where BMP-2 was released from the scaffold whereas the control group (no BMP-2) showed a significantly lower mineralized tissue volume of 106 ± 55 mm3 . The SBC+BMP-2 group had the highest mineralized tissue volume of 189 ± 36 mm3 . Histomorphometry showed bone-to-implant contact of 54.5% for the test groups and 33.3% for the control groups and new vertical bone growth of 2.2 ± 1.0 and 1.0 ± 0.9 mm, respectively. The SBC+BMP-2 group again demonstrated the best outcome (2.7 ± 0.4 mm). The qualitative scoring of soft tissue dehiscence showed that the presence of BMP-2 yielded far superior outcomes, 0.63 vs. 1.75 for all control implant sites (with scores ranging from 0, reflecting no soft dehiscence, to 4, showing a completely exposed umbrella). CONCLUSION: The release of BMP-2 from a SBC scaffold adjacent to a hydrophilic, rough Ti implant and scaffold retention umbrella consistently regenerated the greatest volume and height of new vertical bone along the length of the implant.

The US/China workshop: Regulation, standards, and innovation IV, organized by the Chinese Society for Biomaterials (CSBM) and the US Society for Biomaterials (SFB) in Minneapolis 2017.

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Layer-by-layer nanoparticle platform for cancer active targeting.

Nanoparticles as drug delivery carriers have been investigated over the last few decades, particularly for cancer treatment. The rationale in developing such nanoparticles is to maximize drug efficacy while minimizing toxic side effects. This can be most effectively achieved through target specific drug delivery. A novel biocompatible nanoparticle platform prepared using the core-shell self-assembly technique is reported. The core consists of calcium phosphate which is biocompatible and pH-sensitive, and the shell is composed of biocompatible polymers (hyaluronic acid, CD44 targeting moiety; and chitosan, physical cross-linker). Cisplatin was selected as a model drug and incorporated between the core and the shell. The nanoparticle composition was optimized for high serum stability and low protein binding. These nanoparticles demonstrated target specific delivery in human lung cancer cells (which overexpress CD44 receptors). The targeting ability of the nanoparticles was confirmed with an 8-fold increase of drug efficacy (IC50) compared to cisplatin. Furthermore, the pH-sensitive core of the nanoparticle platform led to controlled drug release through destabilization in acidic conditions. This platform technology provides a simple approach for the design of targeted biocompatible nanoparticles for cancer therapy.

Enhanced differentiation of dental pulp cells cultured on microtubular polymer scaffolds in vitro.

Dental caries (tooth decay) is the most common chronic disease. Dental tissue engineering is a promising alternative approach to alleviate the shortcomings of the currently available restorative materials. Mimicking the natural extracellular matrix (ECM) could enhance the performance of tissue engineering scaffolds. In this study, we developed microtubular (~20 µm diameter) polymethyl methacrylate (PMMA) scaffolds resembling the tubular (~2.5 µm diameter) structure of dentin, the collagen-based mineralized tissue that forms the major portion of teeth, to study the effect of scaffold architecture on differentiation of mouse dental pulp cells in vitro. Flat (control), plasma-treated solid and microtubular PMMA scaffolds with densities of 240±15, 459±51 and 480±116 tubules/mm2 were first characterized using scanning electron microscopy and contact angle measurements. Dental pulp cells were cultured on the surface of the scaffolds for up to 21 days and examined using various assays. Cell proliferation and mineralization were examined using Alamar Blue and Xylenol Orange (XO) staining assays, respectively. The differentiation of pulp cells into odontoblasts was examined by immunostaining for Nestin and by quantitative PCR analysis for dentin matrix protein 1 (Dmp1), dentin sialophosphoprotein (Dspp) and osteocalcin (Ocn). Our results showed that the highest tubular density scaffolds significantly (p<0.05) enhanced differentiation of pulp cells into odontoblasts as compared to control flat scaffolds, as evidenced by increased expression of Nestin (5.4x). However, mineralization was suppressed on all surfaces, possibly due to low cell density. These results suggest that the microtubular architecture may be a desirable feature of scaffolds developed for clinical applications. LAY SUMMARY: Regenerative engineering of diseased or traumatized tooth structure could avoid the deficiencies of traditional dental restorative (filling) materials. Cells in the dental pulp have the potential to differentiate to dentin-producing odontoblast cells. Furthermore, cell-supporting scaffolds that mimic a natural extracellular matrix (ECM) are known to influence behavior of progenitor cells. Accordingly, we hypothesized that a dentin-like microtubular scaffold would enhance differentiation of dental pulp cells. The hypothesis was proven true and differentiation to odontoblasts increased with increasing density of the microtubules. However, mineralization was suppressed, possibly due to a low density of cells. The results demonstrate the potential benefits of a microtubular scaffold design to promote odontoblast cells for regeneration of dentin.

Human biofield therapy does not affect tumor size but modulates immune responses in a mouse model for breast cancer.

OBJECTIVE: To assess the effect of human biofield therapy, an integrative medicine modality, on the development of tumors and metastasis, and immune function in a mouse breast cancer model. METHODS: Mice were injected with 66cl4 mammary carcinoma cells. In study one, mice received biofield therapy after cell injection. In study two, mice were treated by the biofield practitioner only prior to cell injection. Both studies had two control groups of mock biofield treatments and phosphate-buffered saline injection. Mice were weighed and tumor volume was determined. Blood samples were collected and 32 serum cytokine/chemokine markers were measured. Spleens/popliteal lymph nodes were isolated and dissociated for fluorescent-activated cell sorting (FACS) analysis of immune cells or metastasis assays in cell culture. RESULTS: No significant differences were found in weight, tumor size or metastasis. Significant effects were found in the immune responses in study one but no additional effects were found in study two. In study one, human biofield treatment significantly reduced percentage of CD4(+)CD44loCD25(+) and percentage of CD8(+) cells, elevated by cancer in the lymph nodes, to control levels determined by FACS analysis. In the spleen, only CD11b(+) macrophages were increased with cancer, and human biofield therapy significantly reduced them. Of 11 cytokines elevated by cancer, only interferon-γ, interleukin-1, monokine induced by interfer-γ, interleukin-2 and macrophage inflammatory protein-2 were significantly reduced to control levels with human biofield therapy. CONCLUSION: Human biofield therapy had no significant effect on tumor size or metastasis but produced significant effects on immune responses apparent in the down-regulation of specific lymphocytes and serum cytokines in a mouse breast cancer model.

Comparison of bone morphogenetic protein-2 delivery systems to induce supracrestal bone guided by titanium implants in the rabbit mandible.

OBJECTIVE: To compare the ability of three non-glycosylated/rhBMP-2 (BMP-2) delivery systems to induce supracrestal bone. MATERIAL AND METHODS: Thirty-six custom SLActive dental implants were partially embedded in transverse orientation into the posterior mandibles of 18 adult rabbits with the head of the implant located 3 mm supracrestal. Delivery of BMP-2 (30 µg) from the following materials was studied: (1) Ti implant + BMP-2 with hydroxyapatite (HA)-coated collagen (Col/HA) scaffold, (2) Ti implant with Col/HA infused with PEG hydrogel + BMP-2, or (3) Ti implant with HA/ß-TCP/PEG hydrogel scaffold + BMP-2. Scaffolds were secured with a metal "umbrella." Non-BMP-2 contralateral controls were included. MicroCT imaging and histological analysis was performed after 10 weeks to assess new supracrestal bone formation. In vitro BMP-2 release studies were conducted. RESULTS: All treatment groups displayed new supracrestal bone formation. Ti + BMP-2 with Col/HA (3.0 ± 0.2 mm) and Ti with Col/HA/PEG hydrogel + BMP-2 (2.7 ± 0.4 mm) had significantly greater (P < 0.05) outcomes than without BMP-2. Maximum bone volume occurred in the Ti implant with HA/ß-TCP/PEG hydrogel scaffold + BMP-2 group. CONCLUSIONS: The use of an implant system composed of a partially inserted Ti implant, adjacent scaffold and scaffold stabilizer resulted in the formation of new supracrestal bone across all test groups with and without BMP-2. Delivery of BMP-2 directly from the Ti implant increased bone height, BIC and bone volume as compared to no BMP-2 when a Col/HA was used, but did not improve performance of the HA/ß-TCP/PEG scaffold.

Age-Related Changes in FGF-2, Fibroblast Growth Factor Receptors and ß-Catenin Expression in Human Mesenchyme-Derived Progenitor Cells.

FGF-2 stimulates preosteoblast replication, and knockout of the FGF-2 gene in mice resulted in osteopenia with age, associated with decreased Wnt-ß-Catenin signaling. In addition, targeted expression of FGF-2 in osteoblast progenitors increased bone mass in mice via Wnt-ß-Catenin signaling. We posited that diminution of the intrinsic proliferative capacity of human mesenchyme-derived progenitor cells (HMDPCs) with age is due in part to reduction in FGF-2. To test this hypothesis HMDPCs from young (27-38), middle aged (47-56), and old (65-76) female human subjects were isolated from bone discarded after orthopedic procedures. HMDPCs cultures were mostly homogeneous with greater than 90% mesenchymal progenitor cells, determined by fluorescence-activated cell sorting. There was a progressive decrease in FGF-2 and FGFR1 mRNA and protein in HMDPCs with age. Since FGF-2 activates ß-catenin, which can enhance bone formation, we also assessed its age-related expression in HMDPCs. An age-related decrease in total-ß-Catenin mRNA and protein expression was observed. However there were increased levels of p-ß-Catenin and decreased levels of activated-ß-Catenin in old HMDSCs. FGF-2 treatment increased FGFR1 and ß-Catenin protein, reduced the level of p-ß-Catenin and increased activated-ß-Catenin in aged HMDPCs. In conclusion, reduction in FGF-2 expression could contribute to age-related impaired function of HMDPCs via modulation of Wnt-ß-catenin signaling.

Design and characterization of calcium phosphate ceramic scaffolds for bone tissue engineering.

OBJECTIVES: Our goal is to review design strategies for the fabrication of calcium phosphate ceramic scaffolds (CPS), in light of their transient role in bone tissue engineering and associated requirements for effective bone regeneration. METHODS: We examine the various design options available to meet mechanical and biological requirements of CPS and later focus on the importance of proper characterization of CPS in terms of architecture, mechanical properties and time-sensitive properties such as biodegradability. Finally, relationships between in vitro versus in vivo testing are addressed, with an attempt to highlight reliable performance predictors. RESULTS: A combinatory design strategy should be used with CPS, taking into consideration 3D architecture, adequate surface chemistry and topography, all of which are needed to promote bone formation. CPS represent the media of choice for delivery of osteogenic factors and anti-infectives. Non-osteoblast mediated mineral deposition can confound in vitro osteogenesis testing of CPS and therefore the expression of a variety of proteins or genes including collagen type I, bone sialoprotein and osteocalcin should be confirmed in addition to increased mineral content. CONCLUSIONS: CPS are a superior scaffold material for bone regeneration because they actively promote osteogenesis. Biodegradability of CPS via calcium and phosphate release represents a unique asset. Structural control of CPS at the macro, micro and nanoscale and their combination with cells and polymeric materials is likely to lead to significant developments in bone tissue engineering.

Carboxymethyl Hyaluronan-Stabilized Nanoparticles for Anticancer Drug Delivery.

Carboxymethyl hyaluronic acid (CMHA) is a semisynthetic derivative of HA that is recognized by HA binding proteins but contains an additional carboxylic acid on some of the 6-hydroxyl groups of the N-acetyl glucosamine sugar units. These studies tested the ability of CMHA to stabilize the formation of calcium phosphate nanoparticles and evaluated their potential to target therapy resistant, CD44(+)/CD24(-/low) human breast cancer cells (BT-474EMT). CMHA stabilized particles (nCaP(CMHA)) were loaded with the chemotherapy drug cis-diamminedichloroplatinum(II) (CDDP) to form nCaP(CMHA)CDDP. nCaP(CMHA)CDDP was determined to be poorly crystalline hydroxyapatite, 200 nm in diameter with a -43 mV zeta potential. nCaP(CMHA)CDDP exhibited a two-day burst release of CDDP that tapered resulting in 86% release by 7 days. Surface plasmon resonance showed that nCaP(CMHA)CDDP binds to CD44, but less effectively than CMHA or hyaluronan. nCaP(CMHA-AF488) was taken up by CD44(+)/CD24(-) BT-474EMT breast cancer cells within 18 hours. nCaP(CMHA)CDDP was as cytotoxic as free CDDP against the BT-474EMT cells. Subcutaneous BT-474EMT tumors were more reproducibly inhibited by a near tumor dose of 2.8 mg/kg CDDP than a 7 mg/kg dose nCaP(CMHA)CDDP. This was likely due to a lack of distribution of nCaP(CMHA)CDDP throughout the dense tumor tissue that limited drug diffusion.

Therapeutic Touch Has Significant Effects on Mouse Breast Cancer Metastasis and Immune Responses but Not Primary Tumor Size.

Evidence-based integrative medicine therapies have been introduced to promote wellness and offset side-effects from cancer treatment. Energy medicine is an integrative medicine technique using the human biofield to promote well-being. The biofield therapy chosen for study was Therapeutic Touch (TT). Breast cancer tumors were initiated in mice by injection of metastatic 66cl4 mammary carcinoma cells. The control group received only vehicle. TT or mock treatments were performed twice a week for 10 minutes. Two experienced TT practitioners alternated treatments. At 26 days, metastasis to popliteal lymph nodes was determined by clonogenic assay. Changes in immune function were measured by analysis of serum cytokines and by fluorescent activated cells sorting (FACS) of immune cells from the spleen and lymph nodes. No significant differences were found in body weight gain or tumor size. Metastasis was significantly reduced in the TT-treated mice compared to mock-treated mice. Cancer significantly elevated eleven cytokines. TT significantly reduced IL-1-a, MIG, IL-1b, and MIP-2 to control/vehicle levels. FACS demonstrated that TT significantly reduced specific splenic lymphocyte subsets and macrophages were significantly elevated with cancer. Human biofield therapy had no significant effect on primary tumor but produced significant effects on metastasis and immune responses in a mouse breast cancer model.

Effects of low dose FGF-2 and BMP-2 on healing of calvarial defects in old mice.

There is an age-associated reduction in the bone healing activity of bone morphogenetic protein-2 (BMP-2) that is currently addressed by administering higher doses of BMP-2 in elderly patients. The unwanted medical complications from high dose BMP-2 motivated this investigation to determine whether the addition of a low dose of fibroblast growth factor 2 (FGF-2) could enhance the ability of a lower dose of BMP-2 to heal calvarial bone defects in old mice (18-20 months old). FGF-2 (5 ng) and BMP-2 (2 µg) were administered by a controlled release two-phase biomaterial scaffold placed into the bone defect. FGF-2 released more rapidly and completely in vitro than BMP-2 (40% vs 2%). In vivo, both BMP-2 and FGF-2+BMP-2 groups formed more new bone in calvarial defects than scaffold alone (p < 0.001) or FGF-2 only groups (p < 0.01). The overall total volume of new bone was not statistically increased by the addition of FGF-2 to BMP-2 as measured by microCT, but the pattern of bone deposition was different. In old mice, but not young, there was enhanced bony fill in the central bone defect area when the BMP-2 was supplemented with FGF-2. Histological analysis of the center of the defect revealed an increased bone volume (%BV/TV (p = 0.004)) from the addition of FGF-2. These studies suggest that combining a low dose of FGF-2 with a low dose of BMP-2 has the potential to increase bone healing in old mice relative to BMP-2 alone.

Lithium-end-capped polylactide thin films influence osteoblast progenitor cell differentiation and mineralization.

End-capping by covalently binding functional groups to the ends of polymer chains offers potential advantages for tissue engineering scaffolds, but the ability of such polymers to influence cell behavior has not been studied. As a demonstration, polylactide (PLA) was end-capped with lithium carboxylate ionic groups (hPLA13kLi) and evaluated. Thin films of the hPLA13kLi and PLA homopolymer were prepared with and without surface texturing. Murine osteoblast progenitor cells from collagen 1α1 transgenic reporter mice were used to assess cell attachment, proliferation, differentiation, and mineralization. Measurement of green fluorescent protein expressed by these cells and xylenol orange staining for mineral allowed quantitative analysis. The hPLA13kLi was biologically active, increasing initial cell attachment and enhancing differentiation, while reducing proliferation and strongly suppressing mineralization, relative to PLA. These effects of bound lithium ions (Li(+) ) had not been previously reported, and were generally consistent with the literature on soluble additions of lithium. The surface texturing generated here did not influence cell behavior. These results demonstrate that end-capping could be a useful approach in scaffold design, where a wide range of biologically active groups could be employed, while likely retaining the desirable characteristics associated with the unaltered homopolymer backbone.