Metabolic bone diseases, such as osteoporosis and osteopetrosis, result from an imbalanced bone remodeling process. In vitro bone models are often used to investigate either bone formation or resorption independently, while in vivo, these processes are coupled. Combining these processes in a co-culture is challenging as it requires finding the right medium components to stimulate each cell type involved without interfering with the other cell type's differentiation. Furthermore, differentiation stimulating factors often comprise growth factors in supraphysiological concentrations, which can overshadow the cell-mediated crosstalk and coupling. To address these challenges, we aimed to recreate the physiological bone remodeling process, which follows a specific sequence of events starting with cell activation and bone resorption by osteoclasts, reversal, followed by bone formation by osteoblasts. We used a mineralized silk fibroin scaffold as a bone-mimetic template, inspired by bone's extracellular matrix composition and organization. Our model supported osteoclastic resorption and osteoblastic mineralization in the specific sequence that represents physiological bone remodeling. We also demonstrated how culture variables, such as different cell ratios, base media, and the use of osteogenic/osteoclast supplements, and the application of mechanical load, can be adjusted to represent either a high bone turnover system or a self-regulating system. The latter system did not require the addition of osteoclastic and osteogenic differentiation factors for remodeling, therefore avoiding growth factor use. Our in vitro model for bone remodeling has the potential to reduce animal experiments and advance in vitro drug development for bone remodeling pathologies like osteoporosis. By recreating the physiological bone remodeling cycle, we can investigate cell-cell and cell-matrix interactions, which are essential for understanding bone physiology and pathology. Furthermore, by tuning the culture variables, we can investigate bone remodeling under various conditions, potentially providing insights into the mechanisms underlying different bone disorders.
It is well known that environmental cues such as mechanical loading and/or cell culture medium composition affect tissue-engineered constructs resembling natural bone. These studies are mostly based on an initial setting of the influential parameter that will not be further changed throughout the study. Through the growth of the cells and the deposition of the extracellular matrix (ECM) the initial environmental conditions of the cells will change, and with that also the loads on the cells will change. This study investigates how changes of mechanical load or media composition during culture influences the differentiation and ECM production of mesenchymal stromal cells seeded on porous 3D silk fibroin scaffolds. ECM formation, ECM mineralization and cell differentiation in 3D tissue-engineered bone were analyzed using microscopic tools. Our results suggest that mechanical stimuli are necessary to differentiate human mesenchymal stromal cells of both bone marrow and adipose tissue origin into ECM producing osteoblasts which ultimately become ECM-embedded osteocytes. However, the influence of this stimulus seems to fade quickly after the onset of the culture. Constructs which were initially cultured under mechanical loading continued to deposit minerals at a similar growth rate once the mechanical stimulation was stopped. On the other hand, cell culture medium supplementation with FBS was identified as an extremely potent biochemical cue that influences the mechanosensitivity of the cells with regards to cell differentiation, ECM secretion and mineral deposition. Only through a thorough understanding on these influences over time will we be able to predictably control tissue development in vitro.
Novel biomaterials are being developed and studied, intended to be applied as bone graft substitute materials. Typically, these materials are being tested in in vitro setups, where among others their cytotoxicity and alkaline phosphatase activity (as a marker for osteoblastic differentiation) are being evaluated. However, it has been reported that in vitro tests correlate poorly with in vivo results and therefore many promising biomaterials may not reach the clinic as a bone graft substitute product. One of the reasons for the poor correlation, may be the minimal complexity of the in vitro tests, as compared to the in vivo environment. Ex vivo models, mimicking the natural tissue environment whilst maintaining control of culture parameters, may be a promising alternative to assess biomaterials for bone formation. Assess the possibility of an ex vivo culture platform to test biomaterials on their potential to stimulate new bone formation. Osteochondral plugs (cylinders n=10, Ø 10 mm, height 15 mm) were drilled from fresh porcine knees, from the slaughterhouse. A bone defect (Ø 6 mm) was created and which was filled with a biomaterial graft (S53P4 bioactive glass (n=3); collagen sponges loaded with BMP-2 (n=3, as positive control)) or kept empty (n=4). The explants were cultured in custom-made two-chamber bioreactors for six weeks (LifeTec Group BV). Cartilage and bone were physically separated, similar to the in vivo situation, by a sealing ring. The two tissues were cultured in separate compartments, allowing for specific culture medium for each tissue. Medium was changed every 2–3 days and weekly micro computed tomography (µCT) images were obtained to longitudinally monitor the formation of new bone. An MTT assay was performed on half of the samples after six weeks of culture. The other samples were fixed for histology, to determine which cells were present after six weeks. The MTT metabolic assay showed that a number of cells in the bone were viable after six weeks. The further away from the border, the fewer living cells were observed. The cells in the cartilage also survived. No significant bone formation was observed with µCT in either of groups, even though abundant bone formation was expected in the BMP-2 group. Explanations of the negative results of the positive group might be that too few viable cells remain after six weeks, or that the cells that are still present are not able to form bone. No significant bone formation was observed in the bone defects in osteochondral explants that were cultured with, or without, biomaterials for six weeks. However, the platform showed that it is capable to successfully culture osteochondral explants for six weeks. Histology needs to be performed to evaluate which cells were present at the end of the culture and this will be compared to the cells present directly after drilling the explants.
Bone tissue engineering has the intent to grow bone copies in the laboratory that could be used either for bone regeneration or as model systems to study bone physiology and pathology. Bone marrow- or adipose derived derived mesenchymal stromal cells are commonly used as they have been shown to be capable to differentiate into osteoblasts and depositing a calcium phosphate rich extracellular matrix. However, real bone is more than that: there are commonly three cell types described that are essential contributors to the tissue's native function: osteoblasts, osteocytes and osteoclasts. While all three cell types are being investigated separately, co-cultures of them including their precursors and inactive forms still provide a huge challenge these days, both in terms of culturing and (quantitative) evaluation. In addition, the matrix deposited by the osteoblasts
Based on these results, 25 knees (76 %) were rated excellent, 5 knees (15 %) were rated good, 2 knees (6 %) were rated fair and one patient (3 %) had a poor result. Complications were one temporary peroneal palsy, one luxation of the spacer due to insufficient extensor mechanism and one fracture of the tibia due to substantial primary metaphyseal bone loss.
Minimal Invasive Surgery (MIS) in total knee arthroplasty (TKA) has gained much attention in the scientific community and the public in the last few years. There still exists confusion in the related terminology and different surgical techniques are recommended. Cost effectiveness and risk/benefit analysis are not available at the moment. There still remains controversy whether these new techniques represent only a modern trend or the future of TKA. MIS Unicondylar replacement has shown significant faster rehabilitation but the same reproducible radiographic and clinical results compared with the conventional open technique. In Oct 2003 we have started using MIS TKA in our hospital. After a significant learning curve the decision was made to do only MIS TKA from Nov. 2004 up to now. More than 300 cases were performed. Only few definite data are available at this stage. In 20% of the patients we performed the so called quad sparing (QS) technique. This offers a less invasive but very demanding and time consuming approach, where most of the surgery has to be performed from the side using complete new side cutting instruments. In the majority of our patients (80%) we performed a modified mini midvastus (MMI) approach, using standard 4 in 1 front cutting instruments. Electromagnetic navigation (EM) might be a helpful tool for MIS surgery in TKA. We have limited experience with this new EM navigation system in combination with the new MIS TKA surgical techniques. In a pilot study with two groups of patients the direct comparison between QS and MMI was evaluated. Clinical evaluation was performed by two scores (KSS and WOMAC) and five additional functional tests including straight leg raising, active motion, raising a chair, stair climbing and functional gait analysis. Testing was performed pre-op and at 1, 6 and 12 weeks post-op. Patients and investigators were blinded to the surgical technique (either QS or MMI). The average OR time was 92 min (70 to 130) for MMI and 110 (85 to 165) for QS respectively. There were no complications in the MMI and 1 (wound healing) in the QS group. There were no differences in the different scores and in the functional tests between the groups at any time. There is still controversy in the benefit-risk analysis for the different minimal invasive techniques. In our hospital the MIS future for TKA has already started. Patients’ satisfaction and significant earlier rehabilitation are the key advantages of these new surgical techniques. The much easier MMI technique is now the standard. Only in selected cases the more demanding QS technique is performed. According to the learning curve these new MIS techniques are for specialized surgeons only and require additional training programmes. Despite these facts, we do believe that MIS is the future of TKA surgery.
These clinical effects were sustained over the entire follow-up. At the end of study, 53% of iloprost patients showed healing of at least one BME affected bone as compared to only 19% of Tramadol patients. Regression of subchondral lesions occurred in 4 iloprost patients. No serious adverse events occurred; however, three Tramadol patients discontinued the treatment prematurely due to adverse events.