Histopathological analysis of the synovium in trapeziometacarpal osteoarthritis.

Dorsoradial and anterior oblique ligaments were harvested during surgery in 13 patients with symptomatic trapeziometacarpal osteoarthritis, which had been graded preoperatively by a modified Eaton-Littler radiographic grading. Ligaments, including the periligamentous synovium, were stained with S100 protein, neurotrophic receptor p75, protein gene product 9.5, calcitonin gene related peptide, acetylcholine, substance P, neuropeptide Y, noradrenaline, N-methyl-D-aspartate-receptor and Met/Leu-enkephalin. The synovium was classified as showing no, low-grade or high-grade synovitis. Free nerve endings had higher immunoreactivity for substance P than for N-methyl-D-aspartate-receptor, enkephalin and noradrenaline. The synovial stroma had less immunoreactivity for N-methyl-D-aspartate-receptor than for noradrenaline, substance P and calcitonin gene related peptide. There was no relation between the grade of osteoarthritis and the visual pain analogue scale, synovitis score, immunoreactivity of all antibodies and quantity of free nerve endings or blood vessels. Synovium in trapeziometacarpal joint osteoarthritis produces several neuromediators causing a polymodal neurogenic inflammation and which may serve as biomarkers for osteoarthritis or therapeutic targets.

An expedited approach for sustained delivery of bone morphogenetic protein-7 to bone defects using gene activated fragments of subcutaneous fat.

BACKGROUND: Delivery of bone morphogenetic protein-7 (BMP-7) to bone defects can be improved by applying gene transfer methods. However, traditional ex vivo gene therapy approaches are cumbersome and costly, requiring the extraction and culturing of cells. Therefore, we evaluated a novel, expedited ex vivo BMP-7 gene transfer technology based on the use of fragments of subcutaneous fat tissue. METHODS: We created 5-mm mid-femoral bone defects in the right femora of 23 male, syngeneic Fischer 344 rats. Adipose tissue was harvested from the subcutaneous fat depot of two donor rats. Bone defects were treated with either unmodified fat (control group) or adenovirally BMP-7 transduced fat fragments (treatment group). Healing of bone defects was assessed by radiographs, microcomputed tomography (µCT) and histology at 6 weeks after the implantation of fat tissue fragments. RESULTS: Radiographs, µCT-imaging and histology revealed relevant bone formation in six out of 10 rats treated with BMP-7 activated fat grafts. Two of the defects were bridged. By contrast, femora of the control group receiving unmodified fat did not display signs of osseous healing. BMP-7 gene activated fat treatment led to a significantly higher bone volume (11.18 ± 9.48 mm(3) ) than treatment with unmodified fat grafts (3.19 ± 1.68 mm(3) ) (p = 0.008). CONCLUSIONS: Implantation of BMP-7 gene activated fat tissue fragments can elicit regeneration of large bone defects in rats and could become a clinically expeditious strategy for in vivo bone tissue engineering. However, gene expression must be improved in order to reliably induce osseous bridging of critical-size bone defects. Copyright © 2016 John Wiley & Sons, Ltd.

The effect of BMP-7 gene activated muscle tissue implants on the repair of large segmental bone defects.

BACKGROUND: This study was conducted in order to investigate the effect of Bone Morphogenetic Protein-7 (BMP-7) transduced muscle cells on bone formation and to further develop an innovative abbreviated ex vivo gene therapy for bone repair. As conventional ex vivo gene therapy methods require an elaborative and time-consuming extraction and expansion of cells we evaluated an expedited approach. Fragments of muscle tissue were directly activated by BMP-7 cDNA and implanted into bone defects. METHODS: 25 male, syngeneic Fischer 344 rats were used in the present study. Muscle tissue was harvested from two donor rats and either transduced with an adenovirus carrying the BMP-7 cDNA or remained unmodified. 5mm osseous defects in the right femora of 23 rats were treated with either unmodified muscle tissue (control group) or BMP-7 activated muscle tissue (treatment group). Six weeks after surgery, rat femora were evaluated by radiographs, micro-computed tomography (µCT) and histology. RESULTS: Implantation of BMP-7 activated muscle grafts led to bony bridging in 5 out of 12 defects (41.7%) and to bone formation without bridging in 2 out of 12 defects. In 2 femoral defects of this group radiographs, µCT-imaging and histology did not reveal significant mineralization. Three animals of the BMP-7 treatment group had to be euthanized due to serious wound infection. The bone volume of the treatment group was significantly (p=0.007) higher compared to the control group. CONCLUSION: This study shows that BMP-7 gene activated muscle fragments have the potential to regenerate critical-size segmental bone defects in rats. However, further development of this promising expedited treatment modality is required to improve the healing rate and to investigate if the high infection rate is related to treatment with BMP-7 activated muscle grafts.

Comprehensive histological evaluation of bone implants.

To investigate and assess bone regeneration in sheep in combination with new implant materials classical histological staining methods as well as immunohistochemistry may provide additional information to standard radiographs or computer tomography. Available published data of bone defect regenerations in sheep often present none or sparely labeled histological images. Repeatedly, the exact location of the sample remains unclear, detail enlargements are missing and the labeling of different tissues or cells is absent. The aim of this article is to present an overview of sample preparation, staining methods and their benefits as well as a detailed histological description of bone regeneration in the sheep tibia. General histological staining methods like hematoxylin and eosin, Masson-Goldner trichrome, Movat's pentachrome and alcian blue were used to define new bone formation within a sheep tibia critical size defect containing a polycaprolactone-co-lactide (PCL) scaffold implanted for 3 months (n = 4). Special attention was drawn to describe the bone healing patterns down to cell level. Additionally one histological quantification method and immunohistochemical staining methods are described.

Distribution of sensory nerve endings around the human sinus tarsi: a cadaver study.

The aim of this study was to analyse the pattern of sensory nerve endings and blood vessels around the sinus tarsi. The superficial and deep parts of the fat pads at the inferior extensor retinaculum (IER) as well as the subtalar joint capsule inside the sinus tarsi from 13 cadaver feet were dissected. The distribution of the sensory nerve endings and blood vessels were analysed in the resected specimens as the number per cm(2) after staining with haematoxylin-eosin, S100 protein, low-affinity neurotrophin receptor p75, and protein gene product 9.5 using the classification of Freeman and Wyke. Free nerve endings were the predominant sensory ending (P < 0.001). Ruffini and Golgi-like endings were rarely found and no Pacini corpuscles were seen. Significantly more free nerve endings (P < 0.001) and blood vessels (P = 0.01) were observed in the subtalar joint capsule than in the superficial part of the fat pad at the IER. The deep part of the fat pad at the IER had significantly more blood vessels than the superficial part of the fat pad at the IER (P = 0.012). Significantly more blood vessels than free nerve endings were seen in all three groups (P < 0.001). No significant differences in distribution were seen in terms of right or left side, except for free nerve endings in the superficial part of the fat pad at the IER (P = 0.003). A greater number of free nerve endings correlated with a greater number of blood vessels. The presence of sensory nerve endings between individual fat cells supports the hypothesis that the fat pad has a proprioceptive role monitoring changes and that it is a source of pain in sinus tarsi syndrome due to the abundance of free nerve endings.

Long-bone critical-size defects treated with tissue-engineered polycaprolactone-co-lactide scaffolds: a pilot study on rats.

The aim of this study was to evaluate the osteogenic potential of embroidered, tissue-engineered polycaprolactone-co-lactide (trade name: PCL) scaffolds for the reconstruction of large bone defects. Ten piled-up PCL scaffolds were implanted in femura with a critical size defect of immunodeficient nude rats for 12 weeks [n = 4, group 1: noncoated, group 2: collagen I (coll I), group 3: collagen I/chondroitin sulfate (coll I/CS), and group 4: collagen I/chondroitin sulfate/human mesenchymal stem cells (coll I/CS/hMSC)]. X-ray examination, computer tomography, and histological analyses of the explanted scaffold pads were performed. The quantification of the bone volume ratio showed a significantly higher rate of new bone formation at coll I/CS-coated scaffolds compared with the other groups. Histological investigations revealed that the defect reconstruction started from the peripheral bone ends and incorporated into the scaffold material. Additionally seeded hMSC on coll I/CS-coated scaffolds showed a higher matrix deposition inside the implant but no higher bone formation was observed. These data imply that the coll I/CS-coated PCL scaffolds have the highest potential for treating critical size defects. The scaffolds, being variable in size and structure, can be adapted to any bone defect.

Evaluation of the osteogenic potential and vascularization of 3D poly(3)hydroxybutyrate scaffolds subcutaneously implanted in nude rats.

The aim of this study was to evaluate the osteogenic potential and the vascularization of embroidered, tissue engineered, and cell-seeded 3D poly(3)hydroxybutyrate (PHB) scaffolds in nude rats. Collagen I (coll I)- and collagen I/chondroitin sulfate (coll I/CS)-coated PHB scaffolds were seeded with human mesenchymal stem cells (hMSCs). Proliferation and differentiation were characterized by different biochemical assays in vitro. For animal experiments, the cells were cultivated on coll I- or coll I/CS-coated scaffolds and either expanded or osteogenically differentiated. Scaffolds were piled up to create a 3D scaffold pad and implanted subcutaneously into nude rats. In vitro hMSC showed proliferation and differentiation on PHB scaffolds. Alkaline phosphatase (ALP) and calcium increased in the differentiation medium and in the presence of coll I/CS. In vivo blood vessels were found in the scaffold-stack. Histological/immunohistological analyses of explanted scaffolds showed osteogenic markers such as osteopontin, osteonectin, and coll I around the PHB fibers. Coll I/CS-coated scaffolds with expanded hMSC showed higher values of ALP and calcium than the other combinations. Embroidered PHB scaffolds, coated with extracellular matrix components, provided an adequate environment and, therefore, a template for hMSC which could be differentiated in osteogenic direction.