Biomedical subjects

Moyo C Kruyt

Publications and source records attributed to Moyo C Kruyt.

8 recordsLinked to original sources

PubMed2006

Analysis of ectopic and orthotopic bone formation in cell-based tissue-engineered constructs in goats.

Despite decades of extensive research, the application of cell-based bone tissue engineering in clinically relevant models remains challenging. To improve effectiveness, a better understanding of how the technique should work is crucial. In the current study, we investigated the onset time, rate, location and direction of bone formation in ectopically and orthotopically implanted clinically sized tissue-engineered constructs to gain insight the mechanism behind it. Bone marrow stromal cells (BMSCs) were obtained from 10 goats, culture expanded and cryopreserved. Porous biphasic calcium phosphate (BCP) disks of 17mmx6mm were per-operatively seeded with BMSCs or left empty. Both conditions were implanted intramuscularly and in bilateral critical-sized iliac wing defects. Fluorochromes were administered at 3, 5 and 7 weeks and samples were retrieved after 9 weeks. Histology showed abundant and homogeneous bone formation throughout the intramuscular BMSC samples and little bone in the controls. Histomorphometry and measurements of the fluorochrome labels of the ectopical BMSC samples indicated that osteogenesis started at the periphery and subsequent osteoconduction filled the whole scaffold within 7 weeks. In the orthotopically implanted disks, there was good integration with the surrounding bone, but minimal bone in the center of the implants, in both conditions. Bone was only derived from the interface with the surrounding bone, there was no early bone at the surfaces in contact to soft tissue as was seen in the ectopical samples. Apparently cell survival was minimal and insufficient for relevant additional bone formation. However, the speed of integration with surrounding bone and subsequent bone apposition on the BMSC-seeded orthotopic scaffolds were found to be significantly enhanced, which may be relevant especially in challenging environments.

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PubMed2006

The effect of cell-based bone tissue engineering in a goat transverse process model.

A disadvantage of traditional posterolateral spinal fusion models is that they are highly inefficient for screening multiple conditions. We developed a multiple-condition model that concentrates on the initial process of bone formation from the transverse process and not on a functional fusion. The effect of bone marrow stromal cells (BMSCs) in four different porous ceramic scaffolds was investigated in this setting. Polyacetal cassettes were designed to fit on the goat transverse process and house four different ceramic blocks, i.e: hydroxyapatite (HA) sintered at 1,150 degrees and 1,250 degrees; biphasic calcium phosphate (BCP) and tricalcium phosphate (TCP). Goat BMSCs (n=10) were cultured and per-operatively seeded autologeously on one of two cassettes implanted per animal. The cassettes were bilaterally mounted on the dorsum of decorticated L2-processes for 9 weeks. To asses the dynamics of bone formation, fluorochrome labels were administered and histomorphometry focused on the distribution of bone in the scaffolds. A clear difference in the extent of bone ingrowth was determined for the different scaffold types. An obvious effect of BMSC seeding was observed in three of four scaffold types, especially in scaffold regions adjacent to the overlying muscle. Generally, the BCP and TCP scaffolds showed better osteoconduction and an increased response to BMSCs administration. In conclusion the model provides a reliable and highly efficient method to study bone formation in cell-based tissue engineering. An effect of cell administration was obvious in three of the four scaffold materials.

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PubMed2006

Towards injectable cell-based tissue-engineered bone: the effect of different calcium phosphate microparticles and pre-culturing.

Bone tissue engineering by combining bone marrow stromal cells (BMSCs) with a porous scaffold is a promising technology. Current major challenges are to upscale the technique for clinical application and to improve the handling characteristics. With respect to minimal invasive surgery, moldable and/or injectable formulations are highly preferable. Ceramic microparticles of different HA/TCP formulations (100/0, 70/30, 60/40, 40/60, and 0/100) with varying surface roughness were sieved to select 200 microg aliquots of the 212-300 microm fraction. Goat BMSCs were seeded on different aliquots one week prior to in vivo implantation. These constructs and remaining cells were cultured for one week. By then, the remaining cells were harvested and resuspended in a specific binder: hyaluronic acid, alginate, or blood plasma, combined with aliquots of 60/40 microparticles peroperatively. All constructs were implanted in nude rats (n = 10) and analyzed for their bone yield histomorphometrically after 6 weeks. All precultured constructs showed consistent bone formation of comparable quantity. No significant differences were observed between the different material compositions. Peroperatively prepared constructs hardly showed any bone formation. The present study demonstrated the osteogenic potential of a tissue- engineered bone substitute made of microparticles of various HA/TCP compositions. There was an obvious advantage when the constructs were pre-cultured.

Alginates↗

PubMed2005

A new in vivo screening model for posterior spinal bone formation: comparison of ten calcium phosphate ceramic material treatments.

This study presents a new screening model for evaluating the influence of multiple conditions on the initial process of bone formation in the posterior lumbar spine of a large animal. This model uses cages designed for placement on the decorticated transverse process of the goat lumbar spine. Five conduction channels per cage, each be defined by a different material treatment, are open to both the underlying bone and overlying soft tissue. The model was validated in ten adult Dutch milk goats, with each animal implanted with two cages containing a total of ten calcium phosphate material treatments according to a randomized complete block design. The ten calcium phosphate ceramic materials were created through a combination of material chemistry (BCP, TCP, HA), sintering temperature (low, medium, high), calcination and surface roughness treatments. To monitor the bone formation over time, fluorochrome markers were administered at 3, 5 and 7 weeks and the animals were sacrificed at 9 weeks after implantation. Bone formation in the conduction channels was investigated by histology and histomorphometry of non-decalcified sections using traditional light and epifluorescent microscopy. According to both observed and measured bone formation parameters, materials were ranked in order of increasing magnitude as follows: low sintering temperature BCP (rough and smooth) approximately medium sintering temperature BCP approximately = TCP > calcined low sintering temperature HA > non-calcined low sintering temperature HA > high sintering temperature BCP (rough and smooth) > high sintering temperature HA (calcined and non-calcined). These results agree closely with those obtained in previous studies of osteoconduction and bioactivity of ceramics thereby validating the screening model presented in this study.

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PubMed2004

Optimization of bone-tissue engineering in goats.

Successful bone-tissue engineering (TE) has been reported for various strategies to combine cells with a porous scaffold. In particular, the period after seeding until implantation of the constructs may vary between hours and several weeks. Differences between these strategies can be reduced to (a) the presence of extracellular matrix, (b) the differentiation status of the cells, and (c) the presence of residual potentially immunogenic serum proteins. These parameters are investigated in two types of calcium phosphate scaffolds in a goat model of ectopic bone formation. Culture-expanded bone-marrow stromal cells from eight goats were seeded onto two types of hydroxyapatite granules: HA60/400 (60% porosity, 400-microm average pore size) and HA70/800. Scaffolds seeded with cells and control scaffolds were cultured for 6 days in medium containing autologous or semisynthetic serum, in the presence or absence of dexamethasone. Other scaffolds were seeded with cells just before implantation in medium with or without serum. All conditions were implanted autologously in the paraspinal muscles. After 12 weeks, bone had formed in 87% of all TE constructs, as demonstrated by histology. Histomorphometry indicated significantly more bone in the HA70/800 scaffolds. Furthermore, a significant advantage in bone formation was found when the constructs had been cultured for 6 days. In conclusion, both scaffold characteristics (porosity) and TE strategy (culturing of the constructs) were demonstrated to be important for bone TE.

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PubMed2004

Osteogenicity of autologous bone transplants in the goat.

BACKGROUND: Little is known about the specific mechanisms that make autologous graft bone (AG) superior to the current alternatives. A potential mechanism is the active bone formation by the osteoprogenitor cells within the AG. However, whether these cells survive the transplantation is questionable, especially in nonvascularized, clinically sized grafts. In the present study, we investigated the role of viability in AG implanted ectopically and orthotopically in the goat. METHODS: Eight goats were operated on twice. At the first operation, pieces of vital or devitalized autologous cortical bone were implanted in the paraspinal muscles. Eight weeks later, corticocancellous plugs were taken from the femoral condyles, morselized, and reimplanted as either vital or devitalized orthotopic grafts. The goats received fluorochrome labels at 5, 7, and 9 weeks after the first operation. At 12 weeks, the goats were killed, and the samples were examined histologically. RESULTS: Ectopically, new bone had formed in both the vital and devitalized grafts. In the vital grafts, all three fluorochrome labels were present, indicating an early osteogenic mechanism. Within the devitalized grafts, only the 9-week label was observed. Histomorphometry indicated significantly more new bone in the vital grafts (10.3% vs. 1.7% in the devitalized grafts, P <0.01). Orthotopically, both vital and devitalized grafts showed new bone. Again, graft viability was advantageous in terms of new bone formation (14.5% vs. 9.3%, P <0.02). CONCLUSION: The cells inside the autologous bone transplants most likely survived transplantation and were capable of initiating and sustaining new bone formation.

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PubMed2004

Optimization of bone tissue engineering in goats: a peroperative seeding method using cryopreserved cells and localized bone formation in calcium phosphate scaffolds.

BACKGROUND: Bone tissue engineering by combining cultured bone marrow stromal cells with a porous scaffold is a promising alternative for the autologous bone graft. Drawbacks of the technique include the delay necessary for cell culture and the complicated logistics. We investigated methods to bypass these drawbacks. Furthermore, we investigated the localization of bone formation inside the scaffold. METHODS: Bone marrow stromal cells from seven goats were culture expanded and cryopreserved. One week before surgery, some of the cells were thawed, cultured, and seeded on porous calcium phosphate scaffolds. The constructs were cultured for another week until implantation. The remaining cryopreserved cells were thawed just before implantation and peroperatively resuspended in plasma before combining with the scaffold. Scaffolds impregnated with fresh bone marrow, devitalized cultured constructs, and empty scaffolds served as controls. All samples were implanted in the back muscles of the goats for 9 weeks. RESULTS: Histologic examination showed minimal (<1%) bone in the empty and devitalized scaffolds, 4.2 +/- 5.1 bone area percent in the bone marrow samples, and significantly more bone in both the cultured and peroperatively seeded constructs (11.7 +/- 2.5 and 14.0 +/- 2.0%). The peripheral 350 microm of the implants contained significantly less bone. CONCLUSION: Peroperative preparation of osteogenic constructs with cryopreserved cells is feasible. These constructs yield substantially more bone than the scaffolds alone or scaffolds impregnated with fresh bone marrow. Bone deposition is much less on the scaffold periphery.

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PubMed2004

Bone tissue engineering in a critical size defect compared to ectopic implantations in the goat.

Since the application of the autologous bone graft, the need for an alternative has been recognized. Tissue engineering (TE) of bone by combining bone marrow stromal cells (BMSCs) with a porous scaffold, is considered a promising technique. In this study we investigated the potential of tissue engineered bone to heal a critical sized defect in the goat. Orthotopic bone formation was compared to ectopic bone formation in comparable constructs. TE constructs were prepared from goat BMSCs and porous biphasic calcium phosphate ceramic scaffolds. These constructs and scaffolds without cells were implanted paired in critical sized iliac wing defects. Comparable samples were implanted intramuscularly. After 9 (n=7) and 12 (n=8) weeks implantation, the samples were analyzed histomorphometrically. After 9-weeks implantation in the iliac wing defect, significantly more bone apposition was found in the TE condition. After 12 weeks, the defects were almost completely filled with bone, but no significant advantage of TE was determined anymore. This contrasted with the intramuscular samples where TE implants showed significantly more bone at both time points. In conclusion, bone TE is feasible in critical sized defects. However, when appropriate osteoconductive/inductive materials are applied the effect of cell seeding may be temporary.

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