Beta-tricalcium phosphate particles as a controlled release carrier of osteogenic proteins for bone tissue engineering

Authors

Junli Hu, Yaping Hou, Hyejin Park, Min Lee

Abstract

Beta-tricalcium phosphate (β-TCP) has been widely used as bone substitutes and delivery carriers of osteogenic proteins. However, low protein carrying capacity and agent burst release profiles of β-TCP limit their usage. This study investigates strategies to enhance protein carrying capacity of β-TCP particles with reduced initial burst by surface etching in citric acid solution or by creating apatite coatings with the simulate body fluid immersion approach. The release kinetics of protein from the modified β-TCP particles was investigated using Nel-like molecule-1 (Nell-1), a novel osteogenic protein, as a model protein. Although chemical etching treatments reduced the initial burst release of protein from the particles, a rapid burst release was observed with high protein dose. In contrast, the burst release of protein was significantly reduced by the apatite coating and a high protein dose was successfully delivered over a prolonged period from the apatite-coated particles. Protein release was further modulated by simultaneously delivering proteins from two different substrates: acid-etched and apatite-coated particles. The bioactivity of the protein was preserved during the loading procedure onto the particles. In addition, protein-loaded particles maintained biological activity in the lyophilized state over 4 weeks. These findings suggest that the protein carrying capacity of β-TCP can be modulated by surface modification, which has a potential for use as a protein carrier with controlled release.

Link to Article

http://dx.doi.org/10.1002/jbm.a.34115

Serum response factor regulates bone formation via IGF-1 and Runx2 signals

Authors

Jianfeng Chen, Kaiyu Yuan, Xia Mao, Joseph M Miano, Hui Wu, Yabing Chen

Abstract

Serum response factor (SRF) plays vital roles in numerous cellular processes; however, the physiological function of SRF in skeletal tissue remains unknown. In several organ systems, SRF regulates the expression of insulin-like growth factor-1 (IGF-1), which is crucial for normal development of mineralized skeleton and bone remodeling throughout life. Here we show that conditional deletion of SRF in osteoblasts by osteocalcin-Cre generated viable mice with normal body size and body weight. In comparison with normal siblings, osteoblast-specific SRF deficient adult mice exhibited a marked decrease in bone mineral density and bone formation rate. Deletion of SRF in primary mouse calvarial osteoblasts reduced cell differentiation and mineralization in vitro. This was accompanied by a decrease in IGF-1 expression and secretion. Addition of IGF-1 in the culture media enhanced osteoblast differentiation in control cells and partially restored the mineralization defect of SRF deficient cells, supporting an important role of SRF in regulating IGF-1 and IGF-1-mediated osteoblast differentiation. IGF-1-induced Akt activation was inhibited in SRF deficient calvarial cells, and enhanced in the SRF overexpressed cells. In addition, SRF deficiency decreased the transcriptional activity of Runx2, the key transcription factor for osteogenesis. Overexpression of SRF induced Runx2 transactivity in control cells, and restored Runx2 transactivity in the SRF deficient cells. Taken together, we conclude that SRF is important for IGF-1-induced osteoblast differentiation and mineralization, via regulating IGF-1 expression and Runx2 transactivity.

Link to Article

http://dx.doi.org/10.1002/jbmr.1607

A Novel Calcium Aluminate-Melatonin Scaffold Enhances Bone Regeneration Within A Calvarial Defect

Authors

William P. Clafshenkel, James L. Rutkowski, Rachelle N. Palchesko, Jared D. Romeo, Ken A. McGowan, Ellen S. Gawalt, Paula A. Witt-Enderby

Abstract

Over 500,000 bone graft or bio-implant procedures are performed annually in the United States. It has been reported that osseous autograft procurement may result in donor site complications and bio-implant allografts have been associated with disease transmission. Ceramic scaffolds are only osteoconductive, limiting their clinical use. The objective of this study was to create a bone filler substitute with regenerating properties similar to natural bone. Therefore, melatonin and platelet-rich plasma (PRP) were utilized for their known osteoinductive properties. It was hypothesized that melatonin and/or PRP would enhance the osteoinductive and osteoconductive properties of calcium aluminate (CA) scaffolds to promote bone regeneration in a model of calvarial defects. The biocompatibility of CA and CA-Mel scaffolds were tested in vitro and in vivo. Data show that CA-Mel scaffolds, in comparison to CA scaffolds, enhanced the adhesion, viability and proliferation of normal human osteoblasts (NHost) cells but not that of NIH 3T3 fibroblasts. Data also showed that human adult mesenchymal stem cells grown on CA or CA-Mel scaffolds showed a time-dependent induction into osteoblasts over 14 days revealed through scanning electron microscopy and by alkaline phosphatase analyses. Implantation of CA-Mel scaffolds into critical size calvarial defects in female, ovariectomized rats showed that the CA-Mel scaffolds were biocompatible, allowed for tissue infiltration and showed evidence of scaffold biodegradation by 3 and 6 months. Bone regeneration, assessed using fluorochrome labeling at 3 and 6 months, was greatest in animals implanted with the CA-Mel scaffold. Overall, results from this study show that CA-Mel scaffolds were osteoconductive and osteoinductive.

Link to Article

http://dx.doi.org/10.1111/j.1600-079X.2012.00989.x

Interferon Gamma Inhibits Adipogenesis in vitro and Prevents Marrow Fat Infiltration in Oophorectomized Mice

Authors

Christopher Vidal, Sandra Bermeo, Wei Li, Dao Chao Huang, Richard Kremer, Gustavo Duque

Abstract

Interferon gamma (IFNγ) has been reported to induce osteoblastogenesis from mesenchymal stem cells (MSC) both in vitro and in vivo. With ageing, adipocytes outnumber osteoblasts within the bone microenvironment leading to a decrease in bone formation. Since both osteoblasts and adipocytes are of mesenchymal origin we hypothesized that IFNγ treatment might negatively affect adipogenesis while stimulates osteoblastogenesis in human MSC. To test this hypothesis, human MSC were induced to differentiate into adipocytes in the presence or absence of osteogenic doses of IFNγ (1, 10, 100 ng/ml). IFNγ-treated MSC showed a decrease in adipocyte differentiation and lipid deposition as compared with vehicle-treated controls. Additionally, adipogenic markers were significantly decreased by IFNγ treatment at the same doses that have been reported to have a strong osteogenic effect in vitro. Furthermore, DNA binding of PPARγ was significantly lower in IFNγ-treated differentiating MSC. Subsequently, ovariectomized C57BL6 mice were treated with osteogenic doses of IFNγ three times a week for 6 weeks. In distal femur, treated mice showed significantly higher hematopoiesis concomitant with lower levels of fat volume/total volume, adipocyte number and expression of adipogenic markers as compared with the vehicle-treated mice. Together, these findings demonstrate that, at osteogenic doses, IFNγ also acts as an inhibitor of adipogenesis in vitro and prevents marrow fat infiltration while favors hematopoiesis in ovariectomized mice.

Link to Article

http://dx.doi.org/10.1002/stem.1063

Histological results after maxillary sinus augmentation with Straumann® BoneCeramic, Bio-Oss®, Puros®, and autologous bone. A randomized controlled clinical trial

Author

Christian Martin Schmitt, Hendrik Doering, Thomas Schmidt, Rainer Lutz, Friedrich Wilhelm Neukam, Karl Andreas Schlegel

Abstract

This investigation focused on a comparison of clinical and histological characteristics after sinus floor augmentation with biphasic calcium phosphate (BCP, Straumann BoneCeramic®), anorganic bovine bone (ABB, Geistlich Bio-Oss®), mineralized cancellous bone allograft (MCBA, Zimmer Puros®), or autologous bone (AB). Thirty consecutive patients with a posterior edentulous maxillary situation and a vertical bone height less than or equal to 4 mm were included in this study. A two-stage procedure was carried out. After augmentation of the maxillary sinus with ABB, BCP, MCBA, or AB followed by a healing period of 5 months, biopsies were taken with simultaneous implant placement. The samples were analyzed using microradiography and histology. Ninety-four implants were placed in the augmented positions and 53 bone biopsies were taken and evaluated. The bone volume fraction of newly formed bone was measured as 30.28 ± 2.16% for BCP, 24.9 ± 5.67% for ABB, 41.74 ± 2.1% for AB, and 35.41 ± 2.78% for MCBA with significant increases in bone volume of AB vs. BCP and ABB, and MCBA vs. ABB samples. Significantly different residual bone substitute material was measured as 15.8 ± 2.1% in the BCP group and 21.36 ± 4.83% in the ABB group. As it provides the highest rate of de novo bone formation, AB can be considered to remain the gold standard in sinus floor augmentation. All tested control materials showed comparable results and are suitable for maxillary sinus augmentation.

Link to Article

http://dx.doi.org/10.1111/j.1600-0501.2012.02431.x

Activation of β-Catenin Signaling in Androgen Receptor–Negative Prostate Cancer Cells

Authors

Xinhai Wan, Jie Liu, Jing-Fang Lu, Vassiliki Tzelepi, Jun Yang, Michael W. Starbuck, Lixia Diao, Jing Wang, Eleni Efstathiou, Elba S. Vazquez, Patricia Troncoso, Sankar N. Maity, and Nora M. Navone

Abstract

To study Wnt/β-catenin in castrate-resistant prostate cancer (CRPC) and understand its function independently of the β-catenin–androgen receptor (AR) interaction. We carried out β-catenin immunocytochemical analysis, evaluated TOP-flash reporter activity (a reporter of β-catenin–mediated transcription), and sequenced the β-catenin gene in MDA prostate cancer 118a, MDA prostate cancer 118b, MDA prostate cancer 2b, and PC-3 prostate cancer cells. We knocked down β-catenin in AR-negative MDA prostate cancer 118b cells and carried out comparative gene-array analysis. We also immunohistochemically analyzed β-catenin and AR in 27 bone metastases of human CRPCs. β-Catenin nuclear accumulation and TOP-flash reporter activity were high in MDA prostate cancer 118b but not in MDA prostate cancer 2b or PC-3 cells. MDA prostate cancer 118a and MDA prostate cancer 118b cells carry a mutated β-catenin at codon 32 (D32G). Ten genes were expressed differently (false discovery rate, 0.05) in MDA prostate cancer 118b cells with downregulated β-catenin. One such gene, hyaluronan synthase 2 (HAS2), synthesizes hyaluronan, a core component of the extracellular matrix. We confirmed HAS2 upregulation in PC-3 cells transfected with D32G-mutant β-catenin. Finally, we found nuclear localization of β-catenin in 10 of 27 human tissue specimens; this localization was inversely associated with AR expression (P = 0.056, Fisher's exact test), suggesting that reduced AR expression enables Wnt/β-catenin signaling. We identified a previously unknown downstream target of β-catenin, HAS2, in prostate cancer, and found that high β-catenin nuclear localization and low or no AR expression may define a subpopulation of men with bone metastatic prostate cancer. These findings may guide physicians in managing these patients.

Link to Article

http://dx.doi.org/10.1158/1078-0432.CCR-11-2521