Publications by authors named "Joost D de Bruijn"

62 Publications

Macrophage polarization plays roles in bone formation instructed by calcium phosphate ceramics.

J Mater Chem B 2020 03;8(9):1863-1877

Kuros Biosciences BV, Prof. Bronkhorstlaan 10, 3723 MB Bilthoven, The Netherlands and MERLN Institute, Maastricht University, The Netherlands.

To investigate the roles of macrophages in material-instructed bone formation, two calcium phosphate (TCP) ceramics with the same chemistry but various scales of surface topography were employed in this study. After being implanted subcutaneously in FVB mice for 8 weeks, TCPs (TCP ceramics with submicron surface topography) gave rise to bone formation, while TCPb (TCP ceramics with micron surface topography) did not, showing the crucial role of surface topography scale in material-instructed bone formation. Depletion of macrophages with liposomal clodronate (LipClod) blocked such bone formation instructed by TCPs, confirming the role of macrophages in material-instructed bone formation. Macrophage cells (i.e. RAW 264.7 cells) cultured on TCPs in vitro polarized to tissue repair macrophages as evidenced by gene expression and cytokine production, while polarizing to pro-inflammatory macrophages on TCPb. Submicron surface topography of TCP ceramics directed macrophage polarization via PI3K/AKT pathways with the synergistic regulation of integrin β1. Finally, the tissue repair macrophage polarization on TCPs resulted in osteogenic differentiation of mesenchymal stem cells in vitro. At early implantation in FVB mice, TCPs recruited more macrophages which polarized towards tissue repair macrophages with time. The present data demonstrate the important roles of macrophage polarization in bone formation instructed by calcium phosphate ceramics.
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http://dx.doi.org/10.1039/c9tb02932jDOI Listing
March 2020

MagnetOs, Vitoss, and Novabone in a Multi-endpoint Study of Posterolateral Fusion: A True Fusion or Not?

Clin Spine Surg 2020 07;33(6):E276-E287

Surgical and Orthopaedic Research Laboratories, Prince of Wales Clinical School, University of New South Wales, Sydney, NSW, Australia.

Study Design: This study was a multi-endpoint analysis of bone graft substitutes implanted as a standalone graft in a clinically relevant Ovine model of instrumented posterolateral spinal fusion (PLF).

Objective: The objective of this study was to obtain high-quality evidence on the efficacy of commercial bone graft substitutes compared with autograft in instrumented PLF using a state-of-the-art model with a complete range of assessment techniques.

Summary Of Background Data: Preclinical and clinical data on the quality of spinal fusions obtained with bone graft substitutes are often limited. Calcium phosphates with submicron topography have shown promising results in PLF, as these are able to induce bone formation in tissues distant from the host bone, which facilitates bony union.

Methods: Nine female, skeletally mature sheep (4-5 y) underwent posterior pedicle screw/rods instrumented PLF at L2-L3 and L4-L5 using the following bone graft materials as a standalone graft per spinal segment: (1) biphasic calcium phosphate with submicron topography (BCP<µm), (2) 45S5 Bioglass (BG), and (3) collagen-β-tricalcium phosphate with a 45S5 Bioglass adjunct (TCP/BG). Autograft bone (AB) was used as a positive control treatment. Twelve weeks after implantation, the spinal segments were evaluated by fusion assessment (manual palpation, x-ray, micro-computed tomography, and histology), fusion mass volume quantification (micro-computed tomography), range of motion (ROM) testing, histologic evaluation, and histomorphometry.

Results: Fusion assessment revealed equivalence between AB and BCP<µm by all fusion assessment methods, whereas BG and TCP/BG led to significantly inferior results. Fusion mass volume was highest for BCP<µm, followed by AB, BG, and TCP/BG. ROM testing determined equivalence for spinal levels treated with AB and BCP<µm, while BG and TCP/BG exhibited higher ROM. Histologic evaluation revealed substantial bone formation in the intertransverse regions for AB and BCP<µm, whereas BG and TCP/BG grafts contained fibrous tissue and minimal bone formation. Histologic observations were supported by the histomorphometry data.

Conclusions: This study reveals clear differences in efficacy between commercially available bone graft substitutes, emphasizing the importance of clinically relevant animal models with multiendpoint analyses for the evaluation of bone graft materials. The results corroborate the efficacy of calcium phosphate with submicron topography, as this was the only material that showed equivalent performance to autograft in achieving spinal fusion.
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http://dx.doi.org/10.1097/BSD.0000000000000920DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7337107PMC
July 2020

Bone Morphogenetic Protein 4 Gene Therapy in Mice Inhibits Myeloma Tumor Growth, But Has a Negative Impact on Bone.

JBMR Plus 2020 Jan 22;4(1):e10247. Epub 2019 Nov 22.

Department of Clinical and Molecular Medicine, Faculty of Medicine Norwegian University of Science and Technology (NTNU) Trondheim Norway.

Multiple myeloma is characterized by accumulation of malignant plasma cells in the bone marrow. Most patients suffer from an osteolytic bone disease, caused by increased bone degradation and reduced bone formation. Bone morphogenetic protein 4 (BMP4) is important for both pre- and postnatal bone formation and induces growth arrest and apoptosis of myeloma cells. BMP4-treatment of myeloma patients could have the potential to reduce tumor growth and restore bone formation. We therefore explored BMP4 gene therapy in a human-mouse model of multiple myeloma where humanized bone scaffolds were implanted subcutaneously in RAG2 γCmice. Mice were treated with adeno-associated virus serotype 8 BMP4 vectors (AAV8-BMP4) to express BMP4 in the liver. When mature BMP4 was detectable in the circulation, myeloma cells were injected into the scaffolds and tumor growth was examined by weekly imaging. Strikingly, the tumor burden was reduced in AAV8-BMP4 mice compared with the AAV8-CTRL mice, suggesting that increased circulating BMP4 reduced tumor growth. BMP4-treatment also prevented bone loss in the scaffolds, most likely due to reduced tumor load. To delineate the effects of BMP4 overexpression on bone per se, without direct influence from cancer cells, we examined the unaffected, non-myeloma femurs by μCT. Surprisingly, the AAV8-BMP4 mice had significantly reduced trabecular bone volume, trabecular numbers, as well as significantly increased trabecular separation compared with the AAV8-CTRL mice. There was no difference in cortical bone parameters between the two groups. Taken together, BMP4 gene therapy inhibited myeloma tumor growth, but also reduced the amount of trabecular bone in mice. Our data suggest that care should be taken when considering using BMP4 as a therapeutic agent. © 2019 The Authors. published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.
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http://dx.doi.org/10.1002/jbm4.10247DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6957984PMC
January 2020

Biphasic calcium phosphate with submicron surface topography in an model of instrumented posterolateral spinal fusion.

JOR Spine 2018 Dec 28;1(4):e1039. Epub 2018 Nov 28.

Kuros Biosciences BV Bilthoven The Netherlands.

As spinal fusions require large volumes of bone graft, different bone graft substitutes are being investigated as alternatives. A subclass of calcium phosphate materials with submicron surface topography has been shown to be a highly effective bone graft substitute. In this work, a commercially available biphasic calcium phosphate (BCP) with submicron surface topography (MagnetOs; Kuros Biosciences BV) was evaluated in an model of instrumented posterolateral fusion. The material was implanted stand-alone, either as granules (BCP) or as granules embedded within a fast-resorbing polymeric carrier (BCP) and compared to autograft bone (AG). Twenty-five adult, female Merino sheep underwent posterolateral fusion at L2-3 and L4-5 levels with instrumentation. After 6, 12, and 26 weeks, outcomes were evaluated by manual palpation, range of motion (ROM) testing, micro-computed tomography, histology and histomorphometry. Fusion assessment by manual palpation 12 weeks after implantation revealed 100% fusion rates in all treatment groups. The three treatment groups showed a significant decrease in lateral bending at the fusion levels at 12 weeks ( < 0.05) and 26 weeks ( < 0.001) compared to the 6 week time-point. Flexion-extension and axial rotation were also reduced over time, but statistical significance was only reached in flexion-extension for AG and BCP between the 6 and 26 week time-points ( < 0.05). No significant differences in ROM were observed between the treatment groups at any of the time-points investigated. Histological assessment at 12 weeks showed fusion rates of 75%, 92%, and 83% for AG, BCP and BCP, respectively. The fusion rates were further increased 26 weeks postimplantation. Similar trends of bone growth were observed by histomorphometry. The fusion mass consisted of at least 55% bone for all treatment groups 26 weeks after implantation. These results suggest that this BCP with submicron surface topography, in granules or putty form, is a promising alternative to autograft for spinal fusion.
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http://dx.doi.org/10.1002/jsp2.1039DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6686792PMC
December 2018

Efficacy of a synthetic calcium phosphate with submicron surface topography as autograft extender in lapine posterolateral spinal fusion.

J Biomed Mater Res B Appl Biomater 2019 08 7;107(6):2080-2090. Epub 2019 Jan 7.

Kuros Biosciences BV, Bilthoven, the Netherlands.

Posterolateral spinal fusion (PLF) is a common procedure in orthopedic surgery that is performed to fuse adjacent vertebrae to reduce symptoms related to spinal conditions. In the current study, a novel synthetic calcium phosphate with submicron surface topography was evaluated as an autograft extender in a validated rabbit model of PLF. Fifty-nine skeletally mature New Zealand white rabbits were divided into three groups and underwent single-level intertransverse process PLF at L4-5 using (1) autologous bone graft (ABG) alone or in a 1:1 combination with (2) calcium phosphate granules (ABG/BCP ), or (3) granules embedded in a fast-resorbing polymeric carrier (ABG/BCP ). After 6, 9, and 12 weeks, animals were sacrificed and spinal fusion was assessed by manual palpation, Radiographs, micro-CT, mechanical testing (12 weeks only), histology, and histomorphometry. Based on all endpoints, all groups showed a gradual progression in bone formation and maturation during time, leading to solid fusion masses between the transverse processes after 12 weeks. Fusion assessments by manual palpation, radiography and histology were consistent and demonstrated equivalent fusion rates between groups, with high bilateral fusion rates after 12 weeks. Mechanical tests after 12 weeks indicated substantially lower range of motion for all groups, compared to non-operated controls. By histology and histomorphometry, the gradual formation and maturation of bone in the fusion mass was confirmed for each graft type. With these results, we describe the equivalent performance between autograft and a novel calcium phosphate material as an autograft extender in a rabbit model of PLF using an extensive range of evaluation techniques. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 2080-2090, 2019.
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http://dx.doi.org/10.1002/jbm.b.34301DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6690075PMC
August 2019

Modulating Bone Regeneration in Rabbit Condyle Defects with Three Surface-Structured Tricalcium Phosphate Ceramics.

ACS Biomater Sci Eng 2018 Sep 26;4(9):3347-3355. Epub 2018 Jul 26.

Kuros Biosciences BV, 3723 MB Bilthoven, The Netherlands.

Tricalcium phosphate (TCP) ceramics are used as bone void fillers because of their bioactivity and resorbability, while their performance in bone regeneration and material resorption vary with their physical properties (e.g., the dimension of the crystal grain). Herein, three TCP ceramic bone substitutes (TCP-S, TCP-M, and TCP-L) with gradient crystal grain size (0.77 ± 0.21 μm for TCP-S, 1.21 ± 0.35 μm for TCP-M and 4.87 ± 1.90 μm for TCP-L), were evaluated in a well-established rabbit lateral condylar defect model (validated with sham) with respect to bone formation and material resorption up to 26 weeks. Surface structure-dependent bone regeneration was clearly shown after 4 weeks implantation with TCP-S having most mineralized bone (20.2 ± 3.4%), followed by TCP-M (14.0 ± 3.5%), sham (8.1 ± 4.2%), and TCP-L (6.6 ± 2.6%). Afterward, the amount of mineralized bone was similar in all the three groups, but bone marrow and material resorption varied. After 26 weeks, TCP-S induced most bone tissue formation (mineralized bone + bone marrow) (61.6 ± 7.8%) and underwent most material resorption (80.1 ± 9.0%), followed by TCP-M (42.9 ± 5.2% and 61.4 ± 8.0% respectively), TCP-L (28.3 ± 5.5% and 45.6 ± 9.7% respectively), and sham (25.7 ± 4.2%). Given the fact that the three ceramics are chemically identical, the results indicate that the surface structure (especially, the crystal grain size) of TCP ceramics can greatly tune their bone regeneration potential and the material resorption in rabbit condyle defect model, with the submicron surface structured TCP ceramic performing the best.
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http://dx.doi.org/10.1021/acsbiomaterials.8b00630DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6134343PMC
September 2018

Variation of the bone forming ability with the physicochemical properties of calcium phosphate bone substitutes.

Biomater Sci 2017 Dec;6(1):136-145

Biomaterials Science and Technology, MIRA Institute, University of Twente, 7500 AE, Enschede, the Netherlands.

Because of their bioactive properties and chemical similarity to the inorganic component of bone, calcium phosphate (CaP) materials are widely used for bone regeneration. Six commercially available CaP bone substitutes (Bio-Oss, Actifuse, Bi-Ostetic, MBCP, Vitoss and chronOs) as well as two tricalcium phosphate (TCP) ceramics with either a micron-scale (TCP-B) or submicron-scale (TCP-S) surface structure are characterized and their bone forming potential is evaluated in a canine ectopic implantation model. After 12 weeks of implantation in the paraspinal muscle of four beagles, sporadic bone (0.1 ± 0.1%) is observed in two Actifuse implants (2/4), limited bone (2.1 ± 1.4%) in four MBCP implants (4/4) and abundant bone (21.6 ± 4.5%) is formed in all TCP-S implants (4/4). Bone is not observed in any of the Bio-Oss, Bi-Ostetic, Vitoss, chronOs and TCP-B implants (0/4). When correlating the bone forming potential with the physicochemical properties of each material, we observe that the physical characteristics (e.g. grain size and micropore size at the submicron scale) might be the dominant trigger of material directed bone formation via specific mechanotransduction, instead of protein adsorption, surface mineralization and calcium ion release.
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http://dx.doi.org/10.1039/c7bm00717eDOI Listing
December 2017

Genetically engineered mesenchymal stromal cells produce IL-3 and TPO to further improve human scaffold-based xenograft models.

Exp Hematol 2017 07 26;51:36-46. Epub 2017 Apr 26.

Department of Experimental Hematology, Cancer Research Centre Groningen, University Medical Centre Groningen, University of Groningen, Groningen, The Netherlands. Electronic address:

Recently, NOD-SCID IL2Rγ (NSG) mice were implanted with human mesenchymal stromal cells (MSCs) in the presence of ceramic scaffolds or Matrigel to mimic the human bone marrow (BM) microenvironment. This approach allowed the engraftment of leukemic samples that failed to engraft in NSG mice without humanized niches and resulted in a better preservation of leukemic stem cell self-renewal properties. To further improve our humanized niche scaffold model, we genetically engineered human MSCs to secrete human interleukin-3 (IL-3) and thrombopoietin (TPO). In vitro, these IL-3- and TPO-producing MSCs were superior in expanding human cord blood (CB) CD34 hematopoietic stem/progenitor cells. MLL-AF9-transduced CB CD34 cells could be transformed efficiently along myeloid or lymphoid lineages on IL-3- and TPO-producing MSCs. In vivo, these genetically engineered MSCs maintained their ability to differentiate into bone, adipocytes, and other stromal components. Upon transplantation of MLL-AF9-transduced CB CD34 cells, acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) developed in engineered scaffolds, in which a significantly higher percentage of myeloid clones was observed in the mouse compartments compared with previous models. Engraftment of primary AML, B-cell ALL, and biphenotypic acute leukemia (BAL) patient samples was also evaluated, and all patient samples could engraft efficiently; the myeloid compartment of the BAL samples was better preserved in the human cytokine scaffold model. In conclusion, we show that we can genetically engineer the ectopic human BM microenvironment in a humanized scaffold xenograft model. This approach will be useful for functional study of the importance of niche factors in normal and malignant human hematopoiesis.
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http://dx.doi.org/10.1016/j.exphem.2017.04.008DOI Listing
July 2017

Topography of calcium phosphate ceramics regulates primary cilia length and TGF receptor recruitment associated with osteogenesis.

Acta Biomater 2017 07 27;57:487-497. Epub 2017 Apr 27.

Complex Tissue Regeneration Department, MERLN Institute for Technology Inspired Regenerative Medicine, Maastricht University, Universiteitsingel 40, 6229 ER, The Netherlands; Xpand Biotechnology BV, Bilthoven, The Netherlands; National Engineering Research Center for Biomaterials, Sichuan University, 610064 Chengdu, China. Electronic address:

The surface topography of synthetic biomaterials is known to play a role in material-driven osteogenesis. Recent studies show that TGFβ signalling also initiates osteogenic differentiation. TGFβ signalling requires the recruitment of TGFβ receptors (TGFβR) to the primary cilia. In this study, we hypothesize that the surface topography of calcium phosphate ceramics regulates stem cell morphology, primary cilia structure and TGFβR recruitment to the cilium associated with osteogenic differentiation. We developed a 2D system using two types of tricalcium phosphate (TCP) ceramic discs with identical chemistry. One sample had a surface topography at micron-scale (TCP-B, with a bigger surface structure dimension) whilst the other had a surface topography at submicron scale (TCP-S, with a smaller surface structure dimension). In the absence of osteogenic differentiation factors, human bone marrow stromal cells (hBMSCs) were more spread on TCP-S than on TCP-B with alterations in actin organization and increased primary cilia prevalence and length. The cilia elongation on TCP-S was similar to that observed on glass in the presence of osteogenic media and was followed by recruitment of transforming growth factor-β RII (p-TGFβ RII) to the cilia axoneme. This was associated with enhanced osteogenic differentiation of hBMSCs on TCP-S, as shown by alkaline phosphatase activity and gene expression for key osteogenic markers in the absence of additional osteogenic growth factors. Similarly, in vivo after a 12-week intramuscular implantation in dogs, TCP-S induced bone formation while TCP-B did not. It is most likely that the surface topography of calcium phosphate ceramics regulates primary cilia length and ciliary recruitment of p-TGFβ RII associated with osteogenesis and bone formation. This bioengineering control of osteogenesis via primary cilia modulation may represent a new type of biomaterial-based ciliotherapy for orthopedic, dental and maxillofacial surgery applications.

Statement Of Significance: The surface topography of synthetic biomaterials plays important roles in material-driven osteogenesis. The data presented herein have shown that the surface topography of calcium phosphate ceramics regulates mesenchymal stromal cells (e.g., human bone marrow mesenchymal stromal cells, hBMSCs) with respect to morphology, primary cilia structure and TGFβR recruitment to the cilium associated with osteogenic differentiation in vitro. Together with bone formation in vivo, our results suggested a new type of biomaterial-based ciliotherapy for orthopedic, dental and maxillofacial surgery by the bioengineering control of osteogenesis via primary cilia modulation.
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http://dx.doi.org/10.1016/j.actbio.2017.04.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5489417PMC
July 2017

Cells responding to surface structure of calcium phosphate ceramics for bone regeneration.

J Tissue Eng Regen Med 2017 11 8;11(11):3273-3283. Epub 2017 Feb 8.

Department of Tissue Regeneration, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, 7500AE Enschede, the Netherlands.

Surface structure largely affects the inductive bone-forming potential of calcium phosphate (CaP) ceramics in ectopic sites and bone regeneration in critical-sized bone defects. Surface-dependent osteogenic differentiation of bone marrow stromal cells (BMSCs) partially explained the improved bone-forming ability of submicron surface structured CaP ceramics. In this study, we investigated the possible influence of surface structure on different bone-related cells, which may potentially participate in the process of improved bone formation in CaP ceramics. Besides BMSCs, the response of human brain vascular pericytes (HBVP), C2C12 (osteogenic inducible cells), MC3T3-E1 (osteogenic precursors), SV-HFO (pre-osteoblasts), MG63 (osteoblasts) and SAOS-2 (mature osteoblasts) to the surface structure was evaluated in terms of cell proliferation, osteogenic differentiation and gene expression. The cells were cultured on tricalcium phosphate (TCP) ceramics with either micron-scaled surface structure (TCP-B) or submicron-scaled surface structure (TCP-S) for up to 14 days, followed by DNA, alkaline phosphatase (ALP) and quantitative polymerase chain reaction gene assays. HBVP were not sensitive to surface structure with respect to cell proliferation and osteogenic differentiation, but had downregulated angiogenesis-related gene expression (i.e. vascular endothelial growth factor) on TCP-S. Without additional osteogenic inducing factors, submicron-scaled surface structure enhanced ALP activity and osteocalcin gene expression of human (h)BMSCs and C2C12 cells, favoured the proliferation of MC3T3-E1, MG63 and SAOS-2, and increased ALP activity of MC3T3-E1 and SV-HFO. The results herein indicate that cells with osteogenic potency (either osteogenic inducible cells or osteogenic cells) could be sensitive to surface structure and responded to osteoinductive submicron-structured CaP ceramics in cell proliferation, ALP production or osteogenic gene expression, which favour bone regeneration. Copyright © 2017 John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/term.2236DOI Listing
November 2017

Comparison of Two Moldable Calcium Phosphate-Based Bone Graft Materials in a Noninstrumented Canine Interspinous Implantation Model.

Tissue Eng Part A 2017 12 28;23(23-24):1310-1320. Epub 2017 Feb 28.

1 Biomaterial Science and Technology, MIRA Institute, University of Twente , Enschede, The Netherlands .

There is a continuing search for novel synthetic materials as an alternative to autologous bone grafting. Different technologies are explored to promote bone formation, which include the addition of BioGlass™ particles in calcium phosphate (CaP)-based materials and the use of surface modification in the form of submicron surface topographies. In this work, we aimed at comparing the bone formation in a noninstrumented canine interspinous model of moldable formulations of a submicron-surface structured tricalcium phosphate/alkylene oxide copolymer (CaP/AOC) or a tricalcium phosphate/BioGlass/collagen (CaP/BG/C) bone graft material. Intramuscular implantation was carried out as well to evaluate soft tissue responses. Eight mature male mongrel dogs underwent single-level, noninstrumented interspinous implantation, where the bone graft materials were implanted at either side of the spinous processes (L3-L4), with separation by the interspinous ligament ensuring comparison of both materials in each animal (n = 8 per material). The materials were also implanted in paraspinal muscle pouches. Animals were euthanized 12 weeks after surgery and the lumbar spines excised and intramuscular implants retrieved. Undecalcified sections were prepared for histological evaluation and histomorphometry was performed to quantify bone formation and material resorption. After 12 weeks, all submicron structured CaP/AOC implants showed abundant bone formation in the (L3-L4) interspinous space (20.8% ± 6.8%), whereas bone was not found in the CaP/BG/C implants (0% ± 0%). Intramuscularly, the CaP/AOC material triggered significant bone formation (12.0% ± 7.8%), whereas CaP/BG/C did not form any bone. In both the spinal and muscular sites, resorption of the CaP/AOC material was evident by a decrease in Feret diameter of the CaP granules as well as in their histological surface compared with the starting material, whereas CaP/BG/C material had a milder resorption. This study shows that a submicron-surface structured CaP/AOC bone graft material has superior bone-forming properties in both an interspinous implantation model and intramuscularly, as compared with a CaP/BG/C bone graft material.
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http://dx.doi.org/10.1089/ten.TEA.2016.0347DOI Listing
December 2017

Establishing human leukemia xenograft mouse models by implanting human bone marrow-like scaffold-based niches.

Blood 2016 12 12;128(25):2949-2959. Epub 2016 Oct 12.

Department of Experimental Hematology, Cancer Research Center Groningen, University Medical Center Groningen, University of Groningen, The Netherlands.

To begin to understand the mechanisms that regulate self-renewal, differentiation, and transformation of human hematopoietic stem cells or to evaluate the efficacy of novel treatment modalities, stem cells need to be studied in their own species-specific microenvironment. By implanting ceramic scaffolds coated with human mesenchymal stromal cells into immune-deficient mice, we were able to mimic the human bone marrow niche. Thus, we have established a human leukemia xenograft mouse model in which a large cohort of patient samples successfully engrafted, which covered all of the important genetic and risk subgroups. We found that by providing a humanized environment, stem cell self-renewal properties were better maintained as determined by serial transplantation assays and genome-wide transcriptome studies, and less clonal drift was observed as determined by exome sequencing. The human leukemia xenograft mouse models that we have established here will serve as an excellent resource for future studies aimed at exploring novel therapeutic approaches.
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http://dx.doi.org/10.1182/blood-2016-05-719021DOI Listing
December 2016

Submicron-surface structured tricalcium phosphate ceramic enhances the bone regeneration in canine spine environment.

J Orthop Res 2016 11 7;34(11):1865-1873. Epub 2016 Mar 7.

Xpand Biotechnology BV, Bilthoven, The Netherlands.

Calcium phosphate ceramics with submicron-scaled surface structure can trigger bone formation in non-osseous sites and are expected to enhance bone formation in spine environment. In this study, two tricalcium phosphate ceramics having either a submicron-scaled surface structure (TCP-S) or a micron-scaled one (TCP-B) were prepared and characterized regarding their physicochemical properties. Granules (size 1-2 mm) of both materials were implanted on either left or right side of spinous process, between the two lumbar vertebrae (L3-L4), and in paraspinal muscle of eight beagles. After 12 weeks of implantation, ectopic bone was observed in muscle in TCP-S explants (7.7 ± 3.7%), confirming their ability to inductively form bone in non-osseous sites. In contrast, TCP-B implants did not lead to bone formation in muscle. Abundant bone (34.1 ± 6.6%) was formed within TCP-S implants beside the two spinous processes, while limited bone (5.1 ± 4.5%) was seen in TCP-B. Furthermore, the material resorption of TCP-S was more pronounced than that of TCP-B in both the muscle and spine environments. The results herein indicate that the submicron-scaled surface structured tricalcium phosphate ceramic could enhance bone regeneration as compared to the micron-scaled one in spine environment. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1865-1873, 2016.
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http://dx.doi.org/10.1002/jor.23201DOI Listing
November 2016

Preparation and mechanical properties of photo-crosslinked poly(trimethylene carbonate) and nano-hydroxyapatite composites.

Clin Hemorheol Microcirc 2015 ;60(1):3-11

MIRA Institute for Biomedical Technology and Technical Medicine, and Department of Biomaterials Science and Technology, University of Twente, Enschede, The Netherlands.

Composite materials of photo-crosslinked poly(trimethylene carbonate) and nanoscale hydroxyapatite were prepared and their mechanical characteristics for application as orbital floor implants were assessed. The composites were prepared by solvent casting poly(trimethylene carbonate) macromers with varying amounts of nano-hydroxyapatite and subsequent photo-crosslinking. The incorporation of the nano-hydroxyapatite into the composites was examined by thermogravimetric analysis, scanning electron microscopy and gel content measurements. The mechanical properties were investigated by tensile testing and trouser tearing experiments. Our results show that nano-hydroxyapatite particles can readily be incorporated into photo-crosslinked poly(trimethylene carbonate) networks. Compared to the networks without nano-hydroxyapatite, incorporation of 36.3 wt.% of the apatite resulted in an increase of the E modulus, yield strength and tensile strength from 2.2 MPa to 51 MPa, 0.5 to 1.4 N/mm2 and from 1.3 to 3.9 N/mm2, respectively. We found that composites containing 12.4 wt.% nano-hydroxyapatite had the highest values of strain at break, toughness and average tear propagation strength (376% , 777 N/mm2 and 3.1 N/mm2, respectively).
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http://dx.doi.org/10.3233/CH-151936DOI Listing
June 2016

Effect of particle size on osteoinductive potential of microstructured biphasic calcium phosphate ceramic.

J Biomed Mater Res A 2015 Jun 16;103(6):1919-29. Epub 2014 Sep 16.

State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China.

Material factors such as chemistry, surface microstructure and geometry have shown their influence on osteoinduction of calcium phosphate ceramics. Hereby we report that osteoinduction of a micro-structured biphasic calcium phosphate ceramic (BCP) has a relation with the particle sizes. BCP particles with the size of 212-300 µm, 106-212 µm, 45-106 µm, and smaller than 45 µm were prepared and implanted in paraspinal muscle of dogs for 12 weeks. Histological evaluation of the explants showed abundant bone in all samples with particle size of 212-300 µm, 106-212 µm, and 45-106 µm, while no bone was seen in any sample having particle size smaller than 45 µm. Bone was formed as early as 3 weeks after implantation in implants having BCP particles bigger than 45 µm and the volume of the formed bone was similar among the implants with particles larger than 45 µm after 12 weeks implantation. The results herein show that a size limitation of microstructured calcium phosphate ceramic particles for osteoinduction. It is most likely that the particle size affect inductive bone formation via macroporous structures for body fluid infiltration, cell/tissue ingrowth and angiogenesis.
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http://dx.doi.org/10.1002/jbm.a.35325DOI Listing
June 2015

Microporous calcium phosphate ceramics driving osteogenesis through surface architecture.

J Biomed Mater Res A 2015 Mar 18;103(3):1188-99. Epub 2014 Jul 18.

Department of Tissue Regeneration, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, P.O. Box 217, 7500AE, Enschede, The Netherlands; Key Laboratory of Advanced Technologies of Materials, Ministry of Education, School of Materials Science and Engineering, Southwest Jiaotong University, Chengdu, People's Republic of China.

The presence of micropores in calcium phosphate (CaP) ceramics has shown its important role in initiating inductive bone formation in ectopic sites. To investigate how microporous CaP ceramics trigger osteoinduction, we optimized two biphasic CaP ceramics (i.e., BCP-R and BCP-S) to have the same chemical composition, equivalent surface area per volume, comparable protein adsorption, similar ion (i.e., calcium and phosphate) exchange and the same surface mineralization potential, but different surface architecture. In particular, BCP-R had a surface roughness (Ra) of 325.4 ± 58.9 nm while for BCP-S it was 231.6 ± 35.7 nm. Ceramic blocks with crossing or noncrossing channels of 250, 500, 1000, and 2000 µm were implanted in paraspinal muscle of dogs for 12 weeks. The percentage of bone volume in the channels was not affected by the type of pores (i.e., crossing vs. closed) or their size, but it was greatly influenced by the ceramic type (i.e., BCP-R vs. BCP-S). Significantly, more bone was formed in the channels of BCP-R than in those of BCP-S. Since the two CaP ceramics differed only in their surface architecture, the results hereby demonstrate that microporous CaP ceramics may induce ectopic osteogenesis through surface architecture.
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http://dx.doi.org/10.1002/jbm.a.35272DOI Listing
March 2015

Osteoclast resorption of beta-tricalcium phosphate controlled by surface architecture.

Biomaterials 2014 Aug 11;35(26):7441-51. Epub 2014 Jun 11.

MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, 7522 NB Enschede, Netherlands; Xpand Biotechnology BV, 3723 MB Bilthoven, Netherlands; School of Engineering and Materials Science (SEMS), Queen Mary University of London, E1 4NS London, United Kingdom.

A resorbable bone graft substitute should mimic native bone in its capacity to support bone formation and be remodeled by osteoclasts (OCl) or other multinucleated cells such as foreign body giant cells (FBGC). We hypothesize that by changing the scale of surface architecture of beta-tricalcium phosphate (TCP), cellular resorption can be influenced. CD14(+) monocyte precursors were isolated from human peripheral blood (n = 4 independent donors) and differentiated into OCl or FBGC on the surface of TCP discs comprising either submicron- or micron-scale surface topographical features (TCPs and TCPb, respectively). On submicrostructured TCPs, OCl survived, fused, differentiated, and extensively resorbed the substrate; however, on microstructured TCPb, OCl survival, TRAP activation, and fusion were attenuated. Importantly, no resorption was observed on microstructured TCPb. By confocal microscopy, OCl formed on TCPs contained numerous actin rings allowing for resorption, but not on TCPb. In comparison, FBGC could not resorb either TCP material, suggesting that osteoclast-specific machinery is necessary to resorb TCP. By tuning surface architecture, it appears possible to control osteoclast resorption of calcium phosphate. This approach presents a useful strategy in the design of resorbable bone graft substitutes.
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http://dx.doi.org/10.1016/j.biomaterials.2014.05.048DOI Listing
August 2014

Liposomal clodronate inhibition of osteoclastogenesis and osteoinduction by submicrostructured beta-tricalcium phosphate.

Biomaterials 2014 Jun 31;35(19):5088-97. Epub 2014 Mar 31.

Xpand Biotechnology B.V., The Netherlands.

Bone graft substitutes such as calcium phosphates are subject to the innate inflammatory reaction, which may bear important consequences for bone regeneration. We speculate that the surface architecture of osteoinductive β-tricalcium phosphate (TCP) stimulates the differentiation of invading monocyte/macrophages into osteoclasts, and that these cells may be essential to ectopic bone formation. To test this, porous TCP cubes with either submicron-scale surface architecture known to induce ectopic bone formation (TCPs, positive control) or micron-scale, non-osteoinductive surface architecture (TCPb, negative control) were subcutaneously implanted on the backs of FVB strain mice for 12 weeks. Additional TCPs samples received local, weekly injections of liposome-encapsulated clodronate (TCPs + LipClod) to deplete invading monocyte/macrophages. TCPs induced osteoclast formation, evident by positive tartrate resistant acid phosphatase (TRAP) cytochemical staining and negative macrophage membrane marker F4/80 immunostaining. No TRAP positive cells were found in TCPb or TCPs + LipClod, only F4/80 positive macrophages and foreign body giant cells. TCPs stimulated subcutaneous bone formation in all implants, while no bone could be found in TCPb or TCPs + LipClod. In agreement, expression of bone and osteoclast gene markers was upregulated in TCPs versus both TCPb and TCPs + LipClod, which were equivalent. In summary, submicron-scale surface structure of TCP induced osteoclastogenesis and ectopic bone formation in a process that is blocked by monocyte/macrophage depletion.
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http://dx.doi.org/10.1016/j.biomaterials.2014.03.013DOI Listing
June 2014

The size of surface microstructures as an osteogenic factor in calcium phosphate ceramics.

Acta Biomater 2014 Jul 27;10(7):3254-63. Epub 2014 Mar 27.

Department of Tissue Regeneration, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, P.O. Box 217, 7500AE Enschede, The Netherlands; Xpand Biotechnology BV, Bilthoven, The Netherlands. Electronic address:

The microporosity of calcium phosphate (CaP) ceramics has been shown to have an essential role in osteoinduction by CaP ceramics after ectopic implantation. Here we show that it is not the microporosity but the size of surface microstructural features that is the most likely osteogenic factor. Two tricalcium phosphate (TCP) ceramics, namely TCP-S and TCP-B, were fabricated with equivalent chemistry and similar microporosity but different sizes of surface microstructural features. TCP-S has a grain size of 0.99 ± 0.20 μm and a micropore size of 0.65 ± 0.25 μm, while TCP-B displays a grain size of 3.08 ± 0.52 μm and a micropore size of 1.58 ± 0.65 μm. In vitro, both cell proliferation and osteogenic differentiation were significantly enhanced when human bone marrow stromal cells were cultured on TCP-S without any osteogenic growth factors, compared to TCP-B ceramic granules. The possible involvement of direct contact between cells and the TCP ceramic surface in osteogenic differentiation is also shown with a trans-well culture model. When the ceramic granules were implanted in paraspinal muscle of dogs for 12 weeks, abundant bone was formed in TCP-S (21 ± 10% bone in the available space), whereas no bone was formed in any of the TCP-B implants. The current in vitro and in vivo data reveal that the readily controllable cue, i.e. the size of the surface microstructure, could be sufficient to induce osteogenic differentiation of mesenchymal stem cells, ultimately leading to ectopic bone formation in calcium phosphate ceramics.
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http://dx.doi.org/10.1016/j.actbio.2014.03.021DOI Listing
July 2014

In vitro and in vivo bioactivity assessment of a polylactic acid/hydroxyapatite composite for bone regeneration.

Biomatter 2014 17;4:e27664. Epub 2014 Jan 17.

Department of Tissue Regeneration; University of Twente; Enschede, The Netherlands.

Synthetic bone graft substitutes based on composites consisting of a polymer and a calcium-phosphate (CaP) ceramic are developed with the aim to satisfy both mechanical and bioactivity requirements for successful bone regeneration. In the present study, we have employed extrusion to produce a composite consisting of 50 wt.% poly(D,L-lactic acid) (PLA) and 50 wt.% nano-sized hydroxyapatite (HA) powder, achieving homogeneous distribution of the ceramic within the polymeric phase. In vitro, in both a simulated physiological saline (SPS) and a simulated body fluid (SBF), a greater weight loss was observed for PLA/HA than for PLA particles upon 12-week immersion. Furthermore, in SPS, a continuous release of calcium and phosphate from the composite was measured, whereas in SBF, decrease of the amount of the two ions in the solution was observed both for PLA and PLA/HA accompanied with the formation of a CaP layer on the surface. In vitro characterization of the composite bioactivity was performed by culturing human mesenchymal stromal cells (hMSCs) and assessing proliferation and osteogenic differentiation, with PLA as a control. Both PLA/HA composite and PLA control were shown to support hMSCs proliferation over a period of two weeks. In addition, the composite significantly enhanced alkaline phosphatase (ALP) activity of hMSCs in osteogenic medium as compared with the polymer control. A novel implant design was employed to develop implants from dense, extruded materials, suitable for testing osteoinductivity in vivo. In a preliminary study in dogs, PLA/HA composite implants induced heterotopic bone formation upon 12-week intramuscular implantation in all animals, in contrast to PLA control, which was not osteoinductive. Unlike in vitro, a more pronounced degradation of PLA was observed in vivo as compared with PLA/HA composite.
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http://dx.doi.org/10.4161/biom.27664DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3961484PMC
January 2015

Zinc in calcium phosphate mediates bone induction: in vitro and in vivo model.

Acta Biomater 2014 Jan 17;10(1):477-85. Epub 2013 Oct 17.

Xpand Biotechnology BV, Prof. Bronkhorstlaan 10, Bld 48, 3723 MB Bilthoven, The Netherlands.

Zinc-containing tricalcium phosphate (Zn-TCP) was synthesized to investigate the role of zinc in osteoblastogenesis, osteoclastogenesis and in vivo bone induction in an ectopic implantation model. Zinc ions were readily released in the culture medium. Zn-TCP with the highest zinc content enhanced the alkaline phosphatase activity of human bone marrow stromal cells and tartrate-resistant acid phosphatase activity, as well as multinuclear giant cell formation of RAW264.7 monocyte/macrophages. RAW264.7 cultured with different dosages of zinc supplements in medium with or without zinc-free TCP showed that zinc could influence both the activity and the formation of multinuclear giant cells. After a 12-week implantation in the paraspinal muscle of canines, de novo bone formation and bone incidence increased with increasing zinc content in Zn-TCP - up to 52% bone in the free space. However, TCP without zinc induced no bone formation. Although the observed bone induction cannot be attributed to zinc release alone, these results indicate that zinc incorporated in TCP can modulate bone metabolism and render TCP osteoinductive, indicating to a novel way to enhance the functionality of this synthetic bone graft material.
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http://dx.doi.org/10.1016/j.actbio.2013.10.011DOI Listing
January 2014

Influence of polymer molecular weight in osteoinductive composites for bone tissue regeneration.

Acta Biomater 2013 Dec 31;9(12):9401-13. Epub 2013 Jul 31.

Xpand Biotechnology BV, Bilthoven, The Netherlands. Electronic address:

In bone tissue regeneration, certain polymer and calcium-phosphate-based composites have been reported to enhance some biological surface phenomena, facilitating osteoinduction. Although the crucial role of inorganic fillers in heterotopic bone formation by such materials has been shown, no reports have been published on the potential effects the polymer phase may have. The present work starts from the assumption that the polymer molecular weight regulates the fluid uptake, which determines the hydrolysis rate and the occurrence of biological surface processes. Here, two composites were prepared by extruding two different molecular weight L/D,L-lactide copolymers with calcium phosphate apatite. The lower molecular weight copolymer allowed larger fluid uptake in the composite thereof, which was correlated with a higher capacity to adsorb proteins in vitro. Further, the large fluid absorption led to a quicker composite degradation that generated rougher surfaces and enhanced ion release. Following intramuscular implantation in sheep, only the composite with the lower molecular weight polymer could induce heterotopic bone formation. Besides influencing the biological potential of composites, the molecular weight also regulated their viscoelastic behaviour under cyclic stresses. The results lead to the conclusion that designing biomaterials with appropriate physico-chemical characteristics is crucial for bone tissue regeneration in mechanical load-bearing sites.
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http://dx.doi.org/10.1016/j.actbio.2013.07.026DOI Listing
December 2013

Comparing autograft, allograft, and tricalcium phosphate ceramic in a goat instrumented posterolateral fusion model.

Tissue Eng Part C Methods 2013 Nov 30;19(11):821-8. Epub 2013 May 30.

1 Department of Orthopaedics, University Medical Center Utrecht , Utrecht, The Netherlands .

The most common application of bone grafts is spinal fusion surgery, in which the use of iliac crest autograft is the gold standard. Harvesting of autograft, however, requires an extra surgical procedure, which is associated with additional morbidity. Allograft is the well-known alternative, but it is generally considered less effective in posterior fusions. Therefore, the need for an effective alternative remains. Recently, it was shown that ceramics can be endowed with biologically instructive properties by changing the basic parameters of the material. In this study, we compared a novel tricalcium phosphate ceramic (TCP) to iliac crest autograft and allograft, in instrumented posterolateral fusions in a goat model. A total of nine goats were included, who underwent a two-level lumbar fusion. Each side of the spine was randomized into one type of graft: iliac crest autograft; fresh-frozen allograft; TCP alone; or TCP combined with local autograft (50:50). The fusion rates after 16 weeks were comparable between the groups (autograft 3/8, allograft 4/8, TCP 4/8, and TCP/local autograft 5/8). Calculation of the fusion volume on computed tomography images, showed significantly greater volume in the control groups (autograft 7.8 mL and allograft 8.9 mL) compared with the groups with TCP (TCP 6.1 mL and TCP/local autograft 6.0 mL). No adverse tissue response was seen on histological analysis and TCP was almost completely resorbed. The results demonstrate that TCP is capable of achieving fusion at a similar rate to iliac crest autograft in posterolateral fusions, while almost completely resorbing within 16 weeks. Despite the lower fusion volume, the TCP is a promising alternative circumventing the disadvantages of autograft and allograft.
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http://dx.doi.org/10.1089/ten.TEC.2012.0576DOI Listing
November 2013

Controlling dynamic mechanical properties and degradation of composites for bone regeneration by means of filler content.

J Mech Behav Biomed Mater 2013 Apr 29;20:162-72. Epub 2013 Jan 29.

Xpand Biotechnology BV, Bilthoven, 3723 MB, The Netherlands.

Bone tissue is a dynamic composite system that adapts itself, in response to the surrounding daily (cyclic) mechanical stimuli, through an equilibrium between growth and resorption processes. When there is need of synthetic bone grafts, the biggest issue is to support bone regeneration without causing mechanically-induced bone resorption. Apart from biological properties, such degradable materials should initially support and later leave room to bone formation. Further, dynamic mechanical properties comparable to those of bone are required. In this study we prepared composites comprising calcium phosphate and L-lactide/D-lactide copolymer in various content ratios using the extrusion method. We evaluated the effect of the inorganic filler amount on the polymer phase (i.e. on the post-extrusion intrinsic viscosity). We then studied their in vitro degradation and dynamic mechanical properties (in dry and humid conditions). By increasing the filler content, we observed significant decrease of the intrinsic viscosity of the polymer phase during the extrusion process. Composites containing higher amounts of apatite had faster degradation, and were also mechanically stiffer. But, due to the lower intrinsic viscosity of their polymer phase, they had larger damping properties. Besides this, higher amounts of apatite also rendered the composites more hydrophilic letting them absorb more water and causing them the largest decrease in stiffness. These results show the importance of filler content in controlling the properties of such composites. Further, in this study we observed that the viscoelastic properties of the composite containing 50wt% apatite were comparable to those of dry human cortical bone.
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http://dx.doi.org/10.1016/j.jmbbm.2013.01.012DOI Listing
April 2013

Continuous and uninterrupted oxygen tension influences the colony formation and oxidative metabolism of human mesenchymal stem cells.

Tissue Eng Part C Methods 2013 Jan 12;19(1):68-79. Epub 2012 Oct 12.

School of Engineering and Materials Science, Queen Mary University of London, London, United Kingdom.

Mesenchymal stem cells (MSCs) are an attractive cell source for tissue engineering applications due to their multipotentiality and increased expansion potential compared to mature cells. However, the full potential of MSCs for cellular therapies is not realised, due, in part, to premature proliferative senescence and impaired differentiation capacity following expansion under 20% oxygen. Bone marrow MSCs reside under reduced oxygen levels (4%-7% oxygen), thus this study investigates the effects of uninterrupted physiological oxygen tensions (2%, 5%) on MSC expansion and subsequent differentiation. Expansion potential was evaluated from colony formation efficiency, population-doubling rates, and cellular senescence. Colony formation was significantly reduced under 5% oxygen compared to 2% and 20% oxygen. Population-doubling time was initially shorter with 20% oxygen, but subsequently no significant differences in doubling time were detected between the oxygen conditions. MSCs expanded with 20% oxygen contained a greater proportion of senescent cells than those under physiological oxygen levels, indicated by a three to fourfold increase in β-galactosidase staining. This may be related to the approximately twofold enhanced mitochondrial oxygen consumption under this culture condition. Chondrogenic differentiation was achieved following expansion at each oxygen condition. However, osteogenesis was only achieved for cells expanded and differentiated at 20% oxygen, indicated by alkaline phosphatase activity and alizarin red staining. These studies demonstrate that uninterrupted hypoxia may enhance long-term MSC expansion, but results in a population with impaired osteogenic differentiation potential. Thus, novel differentiation conditions are required to enable differentiation to nonchondrogenic lineages using hypoxia-cultured MSCs.
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http://dx.doi.org/10.1089/ten.TEC.2011.0734DOI Listing
January 2013

Reconstructing the human hematopoietic niche in immunodeficient mice: opportunities for studying primary multiple myeloma.

Blood 2012 Jul 31;120(3):e9-e16. Epub 2012 May 31.

Departments of Cell Biology.

Interactions within the hematopoietic niche in the BM microenvironment are essential for maintenance of the stem cell pool. In addition, this niche is thought to serve as a sanctuary site for malignant progenitors during chemotherapy. Therapy resistance induced by interactions with the BM microenvironment is a major drawback in the treatment of hematologic malignancies and bone-metastasizing solid tumors. To date, studying these interactions was hampered by the lack of adequate in vivo models that simulate the human situation. In the present study, we describe a unique human-mouse hybrid model that allows engraftment and outgrowth of normal and malignant hematopoietic progenitors by implementing a technology for generating a human bone environment. Using luciferase gene marking of patient-derived multiple myeloma cells and bioluminescent imaging, we were able to follow pMM cells outgrowth and to visualize the effect of treatment. Therapeutic interventions in this model resulted in equivalent drug responses as observed in the corresponding patients. This novel human-mouse hybrid model creates unprecedented opportunities to investigate species-specific microenvironmental influences on normal and malignant hematopoietic development, and to develop and personalize cancer treatment strategies.
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http://dx.doi.org/10.1182/blood-2012-03-414920DOI Listing
July 2012

In vivo performance of microstructured calcium phosphate formulated in novel water-free carriers.

Acta Biomater 2012 Jul 7;8(7):2759-69. Epub 2012 Apr 7.

Xpand Biotechnology BV, MB Bilthoven, The Netherlands.

Osteoinductive calcium phosphate (CaP) ceramics can be combined with polymeric carriers to make shapeable bone substitutes as an alternative to autologous bone; however, carriers containing water may degrade the ceramic surface microstructure, which is crucial to bone formation. In this study five novel tricalcium phosphate (TCP) formulations were designed from water-free polymeric binders and osteoinductive TCP granules of different particle sizes (500-1000 μm for moldable putty forms, and 150-500 μm for flowable paste forms). The performance of these novel TCP formulations was studied and compared with control TCP granules alone (both 150-500 and 500-1000 μm). In vitro the five TCP formulations were characterized by their carrier dissolution times and TCP mineralization kinetic profiles in simulated body fluid. In vivo the formulations were implanted in the dorsal muscle and a unicortical femoral defect (Ø=5 mm) of dogs for 12 weeks. The TCP formulation based on a xanthan gum-glycerol carrier exhibited fast carrier dissolution (1 h) and TCP mineralization (7 days) in vitro, but induced inflammation and showed little ectopic bone formation. This carrier chemistry was thus found to disrupt the early cellular response related to osteoinduction by microstructured TCP. TCP formulations based on carboxymethyl cellulose-glycerol and Polyoxyl 15-hydroxystearate-Pluronic(®) F127 allowed the in vitro surface mineralization of TCP by day 7 and produced the highest level of orthotopic bone bridging and ectopic bone formation, which was equivalent to the control. These results demonstrate that water-free carriers can preserve the chemistry, microstructure, and performance of osteoinductive CaP ceramics.
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http://dx.doi.org/10.1016/j.actbio.2012.04.007DOI Listing
July 2012