Het meten van wandschuifspanningen:tussentijds rapport

Een literatuuronderzoek naar methodes om wandschuifspanningen te meten en experimentele toetsing van de electrochemische massa transport technie

The cytocompatibility and early osteogenic characteristics of an injectable calcium phosphate cement.

In this study, the cytocompatibility and early osteogenic characteristics of rat bone marrow cells (RBMCs) on injectable calcium phosphate (CaP) cement (Calcibon) were investigated. In addition to unmodified CaP cement discs, 2 other treatments were given to the discs: preincubation in MilliQ and sintering at different temperatures. After primary culture, RBMCs were dropwise seeded on the discs and cultured for 12 days. The samples were evaluated in terms of cell viability, morphology (live and dead assays and scanning electron microscopy (SEM)), cell proliferation (deoxyribonucleic acid (DNA) analyses), early cell differentiation (alkaline phosphatase (ALP) activity), and physicochemical analyses (xray diffraction (XRD)). The live and dead, DNA, and SEM results showed that Calcibon discs without any additional treatment were not supporting osteoblast-like cells in vitro. There were fewer cells, and cell layers were detached from the disc surface. Therefore, different preincubation periods and sintering temperatures were evaluated to improve the cytocompatibility of the CaP cement. Preincubating discs in MilliQ for periods of 1, 4, 8, and 12 weeks resulted in the hydrolysis of a-tri calcium phosphate (TCP) into an apatite-like structure with some b-TCP, as shown with XRD, but the material was not cytocompatible. Sintering the discs between 8008C and 11008C resulted in conversion of a-TCP to b-TCP with some hydroxyapatite and an increase in crystallinity. Eventually, the discs sintered at 11008C achieved better cell attachment, more-abundant cell proliferation, and earlier differentiation than other sintered (6008C, 8008C, and 10008C), preincubated, and unmodified specimens. On basis of our results, we conclude that in vivo results with CaP-based cements do not guarantee in vitro applicability. Furthermore, unmodified Calcibon is not cytocompatible in vitro, although preincubation of the material results in a more-favorable cell response, sintering of the material at 11008C results in the best osteogenic properties. In contrast to in vivo studies, the Calcibon CaP cement is not suitable as a scaffold for cellbased tissue-engineering strategies

Bone regenerative properties of injectable PGLA-CaP composite with TGF-beta1 in a rat augmentation model.

The aim of this study was to examine the bone augmentation properties of an injectable composite consisting of PLGA microspheres/CaP cement (20/80), and the additional effect of loading PLGA microspheres with TGF-β1 (200 ng). For this purpose, PLGA/CaP composites (control) and PLGA/CaP composites loaded with TGF-β1 (test group) were injected on top of the skulls of 24 Wistar rats. Each rat received 2 materials from the same experimental group, and in total 48 implants were placed (n = 8). After 2, 4, and 8 weeks the results were evaluated histologically and histomorphometrically. The contact length between the implants and newly formed bone increased in time, and was significantly higher for the TGF-β1-loaded composites after 2 weeks. Also, bone formation was significantly higher for the TGF-β1-loaded composites (18.5% ± 3) compared to controls (7.21% ± 5) after 8 weeks of implantation. Immunohistochemical staining demonstrated massive inflammatory infiltrates in both groups, particularly at 2 weeks, which decreased substantially at 4 and 8 weeks. In conclusion, injectable PLGA/CaP composites stimulated bone augmentation in a rat model. The addition of TGF-β1 to the composite significantly increased bone contact at 2 weeks and enhanced new bone formation at 8 weeks

Early-stage macroporosity enhancement in calcium phosphate cements by inclusion of poly(N-vinylpyrrolidone) particles as a porogen

The incorporation of poly(DL-lactic-co-glycolic acid) (PLGA) particles into calcium phosphate cements (CPCs) is an effective strategy to enhance CPC macroporosity and degradation. However, bone regeneration is hindered until hydrolytic PLGA degradation starts a few weeks after implantation. Additionally, CPC and CPC/PLGA injectability and cohesion are suboptimal. In the current study, poly(N-vinylpyrrolidone) (PVP), a water-soluble polymer, was incorporated as a porogen in CPC and CPC/PLGA composites to enhance handling properties and early-stage macroporosity formation. Further, the effect of PVP molecular weight (Mw) and particle size was studied. The results showed that PVP incorporation increased both injectability and cohesion of the CPC pastes, especially with addition of high Mw PVP. Moreover, the in vitro degradation studies revealed that incorporation of PVP induced an initial mass loss during the first week of incubation. In combination with PLGA, small PVP particles induced a higher mass loss at an early stage than large PVP particles, but this effect was no longer apparent after 4 weeks of incubation. In contrast, the incorporation of low Mw PVP had a stronger effect on in vitro degradation in the long term compared to high Mw. Finally, the presence of PLGA porogens appeared to be necessary for adequate CPC degradation.Peer ReviewedPostprint (published version

Persistent transcriptional responses show the involvement of feed-forward control in a repeated dose toxicity study

Chemical carcinogenesis, albeit complex, often relies on modulation of transcription through activation or repression of key transcription factors. While analyzing extensive networks may hinder the biological interpretation, one may focus on dynamic network motifs, among which persistent feed-forward loops (FFLs) are known to chronically influence transcriptional programming. Here, to investigate the relevance a FFL-oriented approach in depth, we have focused on aflatoxin B1-induced transcriptomic alterations during distinct states of exposure (daily administration during 5 days followed by a non-exposed period) of human hepatocytes, by exploring known interactions in human transcription. Several TF-coding genes were persistently deregulated after washout of AFB1. Oncogene MYC was identified as the prominent regulator and driver of many FFLs, among which a FFL comprising MYC/HIF1A was the most recurrent. The MYC/HIF1A FFL was also identified and validated in an independent set as the master regulator of metabolic alterations linked to initiation and progression of carcinogenesis, i.e. the Warburg effect, possibly as result of persistent intracellular alterations arising from AFB1 exposure (nuclear and mitochondrial DNA damage, oxidative stress, transcriptional activation by secondary messengers). In summary, our analysis shows the involvement of FFLs as modulators of gene expression suggestive of a carcinogenic potential even after termination of exposure

Osteogenic differentiation driven by osteoclasts and macrophages

Introduction Osteoclasts are bone-resorbing cells closely related to bone turnover, whereas different macrophage subtypes contribute to bone fracture healing. As osteoclasts and macrophages share the same hematopoietic origin, the difference between both cell types on osteoblast coupling, crosstalk extent and consequent bone formation remains poorly understood. This study compares the potential of primary cells that are routinely considered as osteoclast and macrophage cultures on their ability to support osteogenic differentiation of human mesenchymal stromal cells (hMSCs). Methods Human Peripheral Blood Mononuclear Cells (hPBMCs) were used to obtain macrophage or osteoclast cultures using appropriate stimulatory factors. With different seeding densities of hPBMCs, conditioned media from macrophage or osteoclast cultures were harvested for comparative evaluation of effects thereof on the osteogenic differentiation of hMSCs. Specific cytological staining was used to qualitatively evaluate macrophage and osteoclast cultures. Additionally, quantitative data on hMSC proliferation, osteogenic differentiation and mineralization were obtained via biochemical assays. Results Conditioned medium from osteoclast cultures obtained via low hPBMCs seeding densities, but not from high hPBMCs seeding densities or macrophages, stimulated hMSC osteogenic differentiation and mineralization. Upon cellular crosstalk, both pre-differentiated osteoclasts and non-polarized macrophages equally supported early hMSC osteogenic differentiation and mineralization, as confirmed by increased alkaline phosphatase levels within 7 days and increased calcium content within 14 days in comparison with undifferentiated controls. Initial hPBMCs seeding density strongly influences osteoclastogenesis and the paracrine effect of the resultant osteoclast population on the osteogenic differentiation of hMSCs. In addition, only in indirect coculture, macrophages provide similar stimulatory effects as pre-differentiated osteoclasts on the osteogenic differentiation of MSCs and mineralization. Conclusion Our results demonstrate stimulatory effects of osteoclast conditioned medium on hMSC osteogenic differentiation, depending on initial hPBMC seeding density. In addition, we show that osteoclast and macrophage cultures contain pools of polarized macrophages, which may be involved in the osteogenic effects. Our data provide insight into bone tissue engineering approaches by using multicellular interactions related to bone remodeling and healing for the in vitro modulation of osteogenic differentiation

INFLUENCE OF SURFACE MICROSTRUCTURE AND CHEMISTRY ON OSTEOINDUCTION AND OSTEOCLASTOGENESIS BY BIPHASIC CALCIUM PHOSPHATE DISCS

It has been reported that surface microstructural dimensions can influence the osteoinductivity of calcium phosphates (CaPs), and osteoclasts may play a role in this process. We hypothesised that surface structural dimensions of </= 1 mum trigger osteoinduction and osteoclast formation irrespective of macrostructure (e.g., concavities, interconnected macropores, interparticle space) or surface chemistry. To test this, planar discs made of biphasic calcium phosphate (BCP: 80 % hydroxyapatite, 20 % tricalcium phosphate) were prepared with different surface structural dimensions - either ~ 1 mum (BCP1150) or ~ 2-4 mum (BCP1300) - and no macropores or concavities. A third material was made by sputter coating BCP1150 with titanium (BCP1150Ti), thereby changing its surface chemistry but preserving its surface structure and chemical reactivity. After intramuscular implantation in 5 dogs for 12 weeks, BCP1150 formed ectopic bone in 4 out of 5 samples, BCP1150Ti formed ectopic bone in 3 out of 5 samples, and BCP1300 formed no ectopic bone in any of the 5 samples. In vivo, large multinucleated osteoclast-like cells densely colonised BCP1150, smaller osteoclast-like cells formed on BCP1150Ti, and osteoclast-like cells scarcely formed on BCP1300. In vitro, RAW264.7 cells cultured on the surface of BCP1150 and BCP1150Ti in the presence of osteoclast differentiation factor RANKL (receptor activator for NF-kappaB ligand) proliferated then differentiated into multinucleated osteoclast-like cells with positive tartrate resistant acid phosphatase (TRAP) activity. However, cell proliferation, fusion, and TRAP activity were all significantly inhibited on BCP1300. These results indicate that of the material parameters tested - namely, surface microstructure, macrostructure, and surface chemistry - microstructural dimensions are critical in promoting osteoclastogenesis and triggering ectopic bone formation