An in-depth diagnostic exploration of an inflammation and necrosis syndrome in a population of newborn piglets.

Inflammation and loss of tail integrity can be reasons for serious impairment of animal welfare and one of the major challenges facing modern pig farming. Evidence from practice increasingly suggests that tail lesions might be caused not only by tail biting but also by inflammation and necrosis, which can occur without any action from other pigs. Such changes are not limited to the tail but can also be observed in the ears, heels and soles, claw coronary bands, teats, navel, vulva and face. To describe inflammatory and necrotic manifestations in newborn piglets, all 146 piglets from 11 sows were clinically examined not later than 2 h after birth. In addition, the tail base of 30 randomly selected piglets out of the 146 was histo-pathologically examined as one of the most conspicuously affected body parts. Over 80% of the newborns showed affections in the tail base, claw wall and heels. In 65-87% of the animals, the coronary bands, teats, the face and the ears were affected. None of the 146 piglets was completely free from pathological manifestations. On average, the piglets were affected in six out of nine body parts simultaneously. Histological examinations showed that clear alterations in the skin were already manifested around the time of birth in all examined piglets. Alterations were characterised by the occurrence of numerous lymphocytes and granulocytes throughout the entire subepithelial connective tissue, predominantly in perivascular and perifollicular localisation but also within directly subepithelial glandular ducts and diffusely within the subepithelial connective tissue. In the majority of individuals, the epithelial structure was intact. This concurrence of symptoms in the newborns indicates a primarily endogenous aetiology of an inflammation and necrosis syndrome. Further studies in diverse herd contexts are necessary to establish the conditions for the emergence of such a syndrome and develop welfare indicators.

Manipulation of osteoclastogenesis: Bioactive multiphasic silica/collagen composites and their effects of surface and degradation products.

The intent of the present study was to demonstrate that multiphasic silica/collagen xerogels are able to manipulate cellular processes. These xerogels were prepared by a sol-gel approach allowing the incorporation of mineral phases. The resulting nanocomposites are designed as biomaterial for bone regeneration. Human osteoclasts derived from peripheral blood mononuclear cells were cultured both indirectly and directly, either in presence of different xerogel types or on their surface, to investigate the factor with the main influence on osteoclastogenesis. To this end, the incorporation of a third phase to silica/collagen xerogels was used to affect osteoclastogenesis. In cell culture, ambient ion conditions controlled by both the degradation products of the xerogel and the bioactivity-dependent ion release and reprecipitation were shown to have the main effect on osteoclast specific enzyme tartrate-resistant acid phosphatase (TRAP) 5b. Late stage of osteoclastogenesis characterized by resorption was strongly dependent on the xerogels composition. Surface chemistry of the xerogels was displayed to play an important role in osteoclast resorption. Biphasic silica/collagen xerogels and triphasic xerogels with calcium carbonate offered widespread resorbed areas, whereas hydroxyapatite containing xerogels showed distinctly reduced resorption. The incorporation of strontium carbonate and phosphate, respectively, as third phase changed TRAP 5b activity dose-dependently and inhibited resorption within 21 days. Quantitative evaluation on osteoclast differentiation was carried out using biochemical methods (TRAP 5b, cathepsin K) and was supported by confocal laser scanning microscopy and scanning electron microscopy (SEM). Qualitative estimation of resorption was carried out by SEM.

Visfatin alters the cytokine and matrix-degrading enzyme profile during osteogenic and adipogenic MSC differentiation.

OBJECTIVES: Age-related bone loss is associated with bone marrow adiposity. Adipokines (e.g., visfatin, resistin, leptin) are adipocyte-derived factors with immunomodulatory properties and might influence differentiation of bone marrow-derived mesenchymal stem cells (MSC) in osteoarthritis (OA) and osteoporosis (OP). Thus, the presence of adipokines and MMPs in bone marrow and their effects on MSC differentiation were analyzed. METHODS: MSC and ribonucleic acid (RNA) were isolated from femoral heads after hip replacement surgery of OA or osteoporotic femoral neck fracture (FF) patients. Bone structural parameters were evaluated by microcomputed tomography (µCT). MSC were differentiated towards adipocytes or osteoblasts with/without adipokines. Gene expression (adipokines, bone marker genes, MMPs, TIMPs) and cytokine production was evaluated by realtime-polymerase chain reaction (realtime-PCR) and enzyme-linked immunosorbent assay (ELISA). Matrix mineralization was quantified using Alizarin red S staining. RESULTS: µCT showed an osteoporotic phenotype of FF compared to OA bone (reduced trabecular thickness and increased ratio of bone surface vs volume of solid bone). Visfatin and leptin were increased in FF vs OA. Visfatin induced the secretion of IL-6, IL-8, and MCP-1 during osteogenic and adipogenic differentiation. In contrast to resistin and leptin, visfatin increased MMP2 and MMP13 during adipogenesis. In osteogenically differentiated cells, MMPs and TIMPs were reduced by visfatin. Visfatin significantly increased matrix mineralization during osteogenesis, whereas collagen type I expression was reduced. CONCLUSION: Visfatin-mediated increase of matrix mineralization and reduced collagen type I expression could contribute to bone fragility. Visfatin is involved in impaired bone remodeling at the adipose tissue/bone interface through induction of proinflammatory factors and dysregulated MMP/TIMP balance during MSC differentiation.

New Insights into the Neural Differentiation Potential of Canine Adipose Tissue-Derived Mesenchymal Stem Cells.

Adipose tissue-derived stem cells (ASCs) can be obtained from different adipose tissue sources within the body. It is an abundant cell pool, easily accessible, suitable for cultivation and expansion in vitro and preparation for therapeutic approaches. Amongst these therapeutic approaches are tissue engineering and nervous system disorders such as spinal cord injuries. For such treatment, ASCs have to be reliably differentiated in to the neuronal direction. Therefore, we investigated the neural differentiation potential of ASCs using protocols with neurogenic inductors such as valproic acid and forskolin, while dog brain tissue served as control. Morphological changes could already be noticed 1 h after neuronal induction. Gene expression analysis revealed that the neuronal markers nestin and ßIII-tubulin as well as MAP2 were expressed after induction of neuronal differentiation. Additionally, the expression of the neurotrophic factors NGF, BDNF and GDNF was determined. Some of the neuronal markers and neurotrophic factors were already expressed in undifferentiated cells. Our findings point out that ASCs can reliably be differentiated into the neuronal lineage; therefore, these cells are a suitable cell source for cell transplantation in disorders of the central nervous system. Follow-up studies would show the clinical benefit of these cells after transplantation.

Strontium substitution in apatitic CaP cements effectively attenuates osteoclastic resorption but does not inhibit osteoclastogenesis.

UNLABELLED: Strontium ions were discovered to exert a dual effect on bone turnover, namely an inhibition of cell-driven bone resorption and a simultaneous stimulation of new bone tissue formation. A variety of strontium containing calcium phosphate bone cements (SrCPC) have been developed to benefit from both effects to locally support the healing of osteoporotic bone defects. While the stimulating effect of strontium modification on bone forming cells has been demonstrated in a number of studies, this study focuses on the inhibition and/or reduction of osteoclastogenesis and osteoclastic resorption by a strontium substituted calcium phosphate bone cement (SrCPC). Human peripheral blood mononuclear cells (PBMC) were differentiated into osteoclasts in the presence of different Sr(2+)-concentrations as well as on the surface of SrCPC disks. Osteoclastogenesis of PBMC was shown to be merely unaffected by medium Sr(2+)-concentrations comparable to those released from SrCPC in vitro (0.05-0.15mM). However, an altering effect of 0.1mM strontium on the cytoskeleton of osteoclast-like cells was shown. In direct contact to SrCPC disks, these cells exhibited typical morphological features and osteoclast markers on both RNA and protein level were formed. However, calcium phosphate resorption was significantly decreased on strontium-containing cements in comparison to a strontium-free control. This was accompanied by an intracellular accumulation of strontium that increased with substrate strontium content as demonstrated by Time of Flight Secondary Ion Mass Spectrometry (ToF-SIMS). This study illustrates that SrCPC do not inhibit osteoclastogenesis but significantly attenuate osteoclastic substrate resorption in vitro. STATEMENT OF SIGNIFICANCE: Strontium ions have been shown to promote bone formation and inhibit bone resorption. Therefore strontium is successfully used in the treatment of osteoporosis and also inspired the development of strontium-containing strontium/calcium phosphate bone cements (SrCPC). Studies have shown the positive effects of SrCPC on bone formation, however, the inhibiting effect of strontium on bone resorption in the context of such cements has not been shown so far. We found that the formation of bone-resorbing osteoclasts is not inhibited, but that their resorption activity is decreased in contact to SrCPC. The former is important since those cells play an important role in the bone cell signaling. The latter is a key requirement in osteoporosis therapy, which addresses excess bone resorption.

Intraperitoneal and subcutaneous injections of the TLR9 agonist ODN 1668 in rats: brain inflammatory responses are related to peripheral IL-6 rather than interferons.

Subcutaneous or intraperitoneal administration of Toll-like receptor (TLR)-9 agonist, ODN 1668 caused moderate fever and anorexia. In comparison to stimulation of other intracellular TLRs, activation of TLR9 did not result in pronounced peripheral induction of interferons, but rather induced interleukin-6. Expression of cytokines (TNFα, IL-1ß) and inducible forms of enzymes for prostaglandin E2 synthesis occurred in the brain, in conjunction with a moderate activation of the transcription factors STAT3 and NF-IL6 in brain endothelial cells. The lack of a septic-like state in ODN 1668-treated rats reinforces the therapeutic value of this drug.

Amniotic fluid derived stem cells give rise to neuron-like cells without a further differentiation potential into retina-like cells.

Amniotic fluid contains heterogeneous cell types and has become an interesting source for obtaining fetal stem cells. These stem cells have a high proliferative capacity and a good differentiation potential and may thus be suitable for regenerative medicine. As there is increasing evidence, that these stem cells are also able to be directed into the neural lineage, in our study we investigated the neuronal and glial differentiation potential of these cells, so that they may also be applied to cure degenerative diseases of the retina. Mesenchymal stem cells were isolated from routine prenatal amniocentesis at 15 to 18 weeks of pregnancy of human amniotic fluid and expanded in the cell culture. Cells were cultivated according to standard procedures for mesenchymal stem cells and were differentiated along the neural lineage using various protocols. Furthermore, it was also tried to direct them into cell types of the retina as well as into endothelial cells. Cells of more than 72 amniotic fluid samples were collected and characterized. While after induction neural-like phenotypes could actually be detected, which was confirmed using neural marker proteins such as GFAP and ßIII tubulina further differentiation into retinal like cells could not reliably be shown. These data suggest that amniotic fluid derived cells are an interesting cell source, which may also give rise to neural-like cells. However, a more specific differentiation into neuronal and glial cells could not unequivocally be shown, so that further investigations have to becarried out.

Effect of extracorporeal shock wave on proliferation and differentiation of equine adipose tissue-derived mesenchymal stem cells in vitro.

Mesenchymal stem cells are regarded as common cellular precursors of the musculoskeletal tissue and are responsible for tissue regeneration in the course of musculoskeletal disorders. In equine veterinary medicine extracorporeal shock wave therapy (ESWT) is used to optimize healing processes of bone, tendon and cartilage. Nevertheless, little is known about the effects of the shock waves on cells and tissues. Thus, the aim of this study was to investigate the influence of focused ESWT on the viability, proliferation, and differentiation capacity of adipose tissue-derived mesenchymal stem cells (ASCs) and to explore its effects on gap junctional communication and the activation of signalling cascades associated with cell proliferation and differentiation. ASCs were treated with different pulses of focused ESWT. Treated cells showed increased proliferation and expression of Cx43, as detected by means of qRT-PCR, histological staining, immunocytochemistry and western blot. At the same time, cells responded to ESWT by significant activation (phosphorylation) of Erk1/2, detected in western blots. No significant effects on the differentiation potential of the ASCs were evident. Taken together, the present results show significant effects of shock waves on stem cells in vitro.

Tenogenic differentiation of equine adipose-tissue-derived stem cells under the influence of tensile strain, growth differentiation factors and various oxygen tensions.

Mesenchymal stem cells have become extremely interesting for regenerative medicine and tissue engineering in the horse. Stem cell therapy has been proven to be a powerful and successful instrument, in particular for the healing of tendon lesions. We pre-differentiated equine adipose-tissue-derived stem cells (ASCs) in a collagen I gel scaffold by applying tensile strain, growth differentiation factors (GDFs) and various oxygen tensions in order to determine the optimal conditions for in vitro differentiation toward the tenogenic lineage. We compared the influence of 3% versus 21% oxygen tension, the use of GDF 5, GDF 6 and GDF 7 and the application of uniaxial tensile strain versus no mechanical stimulation on differentiation results as evaluated by cell morphology and by the expression of the tendon-relevant genes collagen I, collagen III, cartilage oligomeric matrix protein and scleraxis. The best results were obtained with an oxygen tension of 21%, tensile stimulation and supplementation with GDF 5 or GDF 7. This approach raises the hope that the in vivo application of pre-differentiated stem cells will improve healing and recovery time in comparison with treatment involving undifferentiated stem cells.

Calcium phosphate phases integrated in silica/collagen nanocomposite xerogels enhance the bioactivity and ultimately manipulate the osteoblast/osteoclast ratio in a human co-culture model.

A human co-culture model of osteoblasts and osteoclasts, derived from bone marrow stromal cells and monocytes respectively, was used to characterize the influence of biomaterial modification on the bioactivity and ultimately the ratio of bone-forming to bone-resorbing cells cultivated directly on the surface. Nanocomposites of silica and collagen have been shown to function as skeletal structures in nature and were reproduced in vitro by using a sol-gel approach. The resulting xerogels exhibit a number of features that make it a valuable system for the development of innovative materials for bone substitution applications. In the present study, the incorporation of different calcium phosphate phases in silica/collagen-based gels was demonstrated to enhance the bioactivity of these samples. This ability of the biomaterial to precipitate calcium phosphate on the surface when incubated in simulated body fluids or cell culture medium is generally considered to an advantageous property for bone substitution materials. By co-cultivating human osteoblasts and osteoclasts up to 42 days on the xerogels, we demonstrate that the long-term ratio of these cell types depends on the level of bioactivity of the substrate samples. Biphasic silica/collagen xerogels exhibited comparably low bioactivity but encouraged proliferation of osteoblasts in comparison to osteoclast formation. A balanced ratio of both cell types was detected for moderately bioactive triphasic xerogels with 5% calcium phosphate. However, enhancing the bioactivity of the xerogel samples by increasing the calcium phosphate phase percentage to 20% resulted in a diminished number of osteoblasts in favor of osteoclast formation. Quantitative evaluation was carried out by biochemical methods (calcium, DNA, ALP, TRAP 5b) as well as RT-PCR (ALP, BSP II, OC, RANKL, TRAP, CALCR, VTNR, CTSK), and was supported by confocal laser scanning microscopy (cell nuclei, actin, CD68, TRAP) as well as scanning electron microscopy.

Amniotic-Fluid Stem Cells: Growth Dynamics and Differentiation Potential after a CD-117-Based Selection Procedure.

Amniotic fluid (AF) has become an interesting source of fetal stem cells. However, AF contains heterogeneous and multiple, partially differentiated cell types. After isolation from the amniotic fluid, cells were characterized regarding their morphology and growth dynamics. They were sorted by magnetic associated cell sorting using the surface marker CD 117. In order to show stem cell characteristics such as pluripotency and to evaluate a possible therapeutic application of these cells, AF fluid-derived stem cells were differentiated along the adipogenic, osteogenic, and chondrogenic as well as the neuronal lineage under hypoxic conditions. Our findings reveal that magnetic associated cell sorting (MACS) does not markedly influence growth characteristics as demonstrated by the generation doubling time. There was, however, an effect regarding an altered adipogenic, osteogenic, and chondrogenic differentiation capacity in the selected cell fraction. In contrast, in the unselected cell population neuronal differentiation is enhanced.

Hydrolyzed fish collagen induced chondrogenic differentiation of equine adipose tissue-derived stromal cells.

Adipose-derived stromal cells (ADSCs) are multipotent cells which, in the presence of appropriate stimuli, can differentiate into various lineages such as the osteogenic, adipogenic and chondrogenic. In this study, we investigated the effect of transforming growth factor beta 1 (TGF-ß1) in comparison to hydrolyzed fish collagen in terms of the chondrogenic differentiation potential of ADSCs. ADSCs were isolated from subcutaneous fat of horses by liposuction. Chondrogenesis was investigated using a pellet culture system. The differentiation medium was either supplemented with TGF-ß1 (5 ng/ml) or fish collagen (0.5 mg/ml) for a 3 week period. After the 3 weeks in vitro differentiation, RT-PCR and histological staining for proteoglycan synthesis and type II collagen were performed to evaluate the degree of chondrogenic differentiation and the formation of cartilaginous extracellular matrix (ECM). The differentiation of ADSCs induced by TGF-ß1 showed a high expression of glycosaminoglycan (GAG). Histological analysis of cultures stimulated by hydrolyzed fish collagen demonstrated an even higher GAG expression than cultures stimulated under standard conditions by TGF-ß1. The expression of cartilage-specific type II collagen and Sox9 was about the same in both stimulated cultures. In this study, chondrogenesis was as effectively induced by hydrolyzed fish collagen as it was successfully induced by TGF-ß1. These findings demonstrated that hydrolyzed fish collagen alone has the potential to induce and maintain ADSCs-derived chondrogenesis. These results support the application of ADSCs in equine veterinary tissue engineering, especially for cartilage repair.

NADPH-diaphorase activity and NO synthase expression in the olfactory epithelium of the bovine.

NADPH-diaphorase (NADPH-d) staining of the bovine olfactory epithelium was compared with the immunohistochemical localization of nitric oxide synthase (NOS), soluble guanylyl cyclase, and cGMP (cyclic guanosine 3',5'-monophosphate). Out of the three isoforms, only the inducible NOS (NOS-II) was found at the epithelial surface correlating with the strong labelling for NADPH-d. In contrast, light diaphorase staining associated with deeper epithelial regions did not coincide with any NOS immunoreactivity. As there is overlapping expression of NOS-II, soluble guanylyl cyclase and cGMP at the luminal surface morphologically occupied by dendritic knobs of olfactory receptor neurons and microvillar endings of supporting cells, the nitric oxide (NO)/cGMP pathway is likely to be involved in modulating the odour signals during olfactory transduction.

Neuronal characteristics of amniotic fluid derived cells after adenoviral transformation.

Efficient transformation of primary human amniocytes by E1 gene functions of human adenovirus serotype 5 (Ad5) yield in stable cell lines, which exhibit morphological features of epithelial like cells. A thorough investigation using immunocytochemistry confirmed the expression of epithelial cell markers. The analysis also revealed the expression of neuronal and glial marker proteins, such as nestin, vimentin, A2B5 and GFAP. Using RT-PCR, transcripts of the neurotrophic factors nerve growth factor (NGF), brain derived neurotrophic factor (BDNF), glial cell line derived neurotrophic factor (GDNF), and neurotrophin 3 (NT-3) could be detected. Neurotrophic factors could also be detected in the cell culture supernatants of transformed amniocytes. In line with previous experimental data on a human Ad5 E1-transformed embryonal kidney cell line (HEK-293), the results suggest a co-expression of epithelial and neuronal marker proteins in E1-transformed human amniotic fluid derived cells and thus a preferential transformation into neuronal-like cells.

Diagnosis of osteoporosis with vitamin k as a new biochemical marker.

Osteoporosis is a metabolic bone disease characterized by reduced bone quality and quantity. As a consequence, patients are at risk for fractures, subsequent immobility, and higher mortality especially among elder patients. Because of the high incidence of complications and the associated financial burden for the health system, new parameters for diagnostic and therapeutic purposes are urgently needed. In this regard, research focused on vitamin K as a biochemical bone marker has provided promising results. Vitamin K represents an important enzyme-cofactor for the posttranslational modification and activation of several proteins involved in bone metabolism. Vitamin K has been proven to be a valuable diagnostic as well as therapeutic parameter especially in osteoporosis. Patients with osteoporosis have been shown to have decreased levels of vitamin K. Further, regular intake of vitamin K may increase bone mineral density (BMD), thereby lowering the fracture risk. Yet vitamin K alone may not sufficiently indicate the mineral status of the bone. However, the usefulness of a combination of several biochemical bone markers as improved surrogate markers of bone metabolism has been shown recently. Therefore, this review will focus on the significance and importance of vitamin K for bone metabolism. Beyond this, aspects on the current and prospective use of vitamin K as well as other newly developed biochemical bone markers will be discussed.

[Elution kinetics and antimicrobial effects of gentamicin- and clindamycin-loaded bone cements in vitro]. / In-vitro-Untersuchungen zu Elutionskinetiken und antimikrobiellen Wirkungen von Gentamicin- und Clindamycin-haltigen Knochenzementen.

AIMS: In arthroplasty and particulary in revision surgery antibiotic-loaded bone cements are applied. The purpose of the current study was to analyse the kinetics of antibiotic release and antimicrobial effects of the gentamicin- and clindamycin-loaded bone cement Versabond. MATERIALS AND METHODS: Test cylinders of bone cements, containing gentamicin and clindamycin in different concentrations, were investigated in vitro with regards to the kinetics of antibiotic release by applying static and continuous elution methods. Antimicrobial effects of antibiotic-loaded bone cement were tested in an infection model with different strains of Staphylococcus aureus and Staphylococcus epidermidis and live/dead dye plus fluorescence microscopy. For the surface analysis of antibiotic-loaded test cylinders contaminated with Staphylococcus epidermidis, scanning electron microscopy was used. RESULTS: The measurement of static and continuous elution revealed initially higher rates of gentamicin release in comparison with clindamycin. The gentamicin amount decreased within 24 h, whereas clindamycin-loaded test cylinders showed a prolonged release. After 24 h exclusive avital bacteria on antibiotic-loaded cement cylinders were seen, after 72 h on gentamicin-loaded cement bacterial growth was again detected. Application of antibiotics in bone cement revealed an inhomogeneous surface of the bone cement cylinder. CONCLUSIONS: On application of clindamycin in bone cement the features of increased antibiotic uptake and antimicrobial effect compared with gentamicin-loaded bone cement are observed.

Enhanced alpha 1(I) mRNA expression in frozen shoulder and dupuytren tissue.

The purpose of this study has been to investigate collagen I and III synthesis during the fibrosing stage of frozen shoulder and Dupuytren samples in comparison to normal capsule tissue. - By using the quantitative PCR significantly increased levels of alpha 1(I) mRNA transcription in samples of frozen shoulder (p = 0.016) and Duypuytren (p = 0.041) could be demonstrated, whereas alpha 2(I) and alpha 1(III) chains have shown the same mRNA levels as in normal capsule tissue. - Despite an enhancement of alpha 1(I) mRNA transcription in frozen shoulder and Dupuytren samples the intracellular precursor procollagen I and extracellular mature collagen I was detected immunohistochemically in reduced levels. - The structural alteration of collagen I assembly might be caused by disturbed post-translation from the polypeptide chains into the triple helices procollagen I though alpha 1(I) mRNA transcription was significantly increased and alpha 2(I) mRNA transcription was in normal range. Fibroblasts might release high quantities of free alpha 1(I) polypeptide chains or (alpha 1(I)) 3 homotrimer into the extracellular space during the fibrosing stage of frozen shoulder and Dupuytren disease. - In all samples neither differences of alpha 1(III) mRNA transcription nor differences of immunohistochemical staining intensity of collagen III could be seen. This might result from apoptosis of myofibroblasts in the final phase of the fibrosing processes. - The stimulating effect of insulin-like growth factor type I (IGF-I) to induce fibrosis in connective tissue such as scarlet is known. In all patients suffering from frozen shoulder and Dupuytren disease the serum IGF-I level was in a normal range and the IGF-I receptor - (IGFR-I) mRNA transcription in the samples was also in the same level compared with normal capsule tissue.

[Reaming debris: a source of vital cells! First results of human specimens]. / Bohrmehl: Eine Quelle vitaler Zellen! Erste Ergebnisse von humanen Proben.

Reaming debris is generated in the course of intramedullary reaming of long bones. Up to now there has been little information about the composition of reaming debris. Especially, it remains to be elucidated if reaming debris contains vital cells. The goal of the present vitro investigation has been the harvest of cells from human reaming debris and the subsequent characterization of the cells. 21 specimens of human reaming debris have been investigated. Each specimen has been divided into two parts. One part has been examined by means of transmission electron microscopy while the other part of each specimen has been transferred into culture dishes. The developing cell cultures were characterized by using FACS analysis and were incubated within osteogenic, adipogenic and chondrogenic differentiation media. The results of electron microscopy have revealed the presence of both, vital cells and massively altered cells. Cell growth occurred after initial plating of all specimens. The cells which were grown within the culture dishes could be characterized as mesenchymal stem cells on the basis of their morphology, differentiation capacity and antigen profile. Based upon these results reaming debris has to be regarded as a source of vital mesenchymal stem cells. It remains to be elucidated, if reaming debris can be used as an alternative to bone tissue grafting.

Glial fibrillary acidic protein and myelin basic protein as markers for the immunochemical detection of bovine central nervous tissue in heat-treated meat products.

Because bovine central nervous tissue (CNT) is the main risk material in transmission of the infective agent of bovine spongiform encephalopathy, a suitable test is needed to enforce the ban on CNT in human foodstuffs in the United States and the European Union and to ensure that meat products are free of CNT To detect bovine CNT in heat-treated meat products, we used immunohistochemistry and Western blots with antibodies against glial fibrillary acidic protein (GFAP) and myelin basic protein (MBP). Both antigens were resistant to processing methods used for meat products. The anti-GFAP antibody showed a high degree of tissue specificity, whereas the anti-MBP antibody had high species specificity, clearly differentiating between porcine and bovine CNT Therefore, immunochemistry performed with both proteins provides an effective means for detecting bovine CNT in meat products.

Effects of platelet growth factors on human mesenchymal stem cells and human endothelial cells in vitro.

The aim of the present in vitro study has been to investigate the effects of a enriched platelet derived growth factors on proliferation and migration of human endothelial and mesenchymal stem cells and on osteogenic differentiation of stem cells. Platelet rich plasma has been produced, yielding a four time higher number of thrombocytes than normal plasma. Degranulation of platelets has been performed by means of calcium and thrombin. Plasma has served as a control, whereas plasma in combination with calcium and thrombin was used to distinguish the difference between calcium and/or thrombin mediated effects and growth factor induced effects on the cells. The observed enhanced proliferation and migration of endothelial cells towards the platelet derived growth factors was driven by the plasma component of these preparations. However PDGF solely stimulated the migration and proliferation of mesenchymal stem cells. The increased osteogenic differentiation of growth factor treated mesenchymal stem cells was mostly driven by the high level of calcium used for the platelets degranulation. In summery, the different components of platelet derived growth factors work together to influence human endothelial and mesenchymal stem cells. This is of special clinically interest regarding the stimulation of bone healing in orthopaedic and traumatic surgery.