Robert Gassner

The role of oxygen termination of nanocrystalline diamond on immobilisation of BMP-2 and subsequent bone formation

Medical implants are increasingly often inserted into bone of frail patients, who are advanced in years. Due to age, severe trauma or pathology-related bone changes, osseous healing at the implant site is frequently limited. We were able to demonstrate that coating of endosseous implants with nanocrystalline diamond (NCD) allows stable functionalization by means of physisorption with BMP-2. Strong physisorption was shown to be directly related to the unique properties of NCD, and BMP-2 in its active form interacted strongly when NCD was oxygen-terminated. The binding of the protein was monitored under physiological conditions by single molecule force spectroscopy, and the respective adsorption energies were further substantiated by force-field-calculations. Implant surfaces refined in such a manner yielded enhanced osseointegration in vivo, when inserted into sheep calvaria. Our results further suggest that this technical advancement can be readily applied in clinical therapies with regard to bone healing, since primary human mesenchymal stromal cells strongly activated the expression of osteogenic markers when being cultivated on NCD physisorbed with physiological amounts of BMP-2.

Human Bone Marrow Hosts Polyfunctional Memory CD4+ and CD8+ T Cells with Close Contact to IL-15–Producing Cells

Recently, a key role in memory T cell homing and survival has been attributed to the bone marrow (BM) in mice. In the human BM, the repertoire, function, and survival niches of CD4+ and CD8+ T cells have not yet been elucidated. In this study, we demonstrate that CD4+ and CD8+ effector memory T cells accumulate in the human BM and are in a heightened activation state as revealed by CD69 expression. BM-resident memory T cells produce more IFN-γ and are frequently polyfunctional. Immunofluorescence analysis revealed that CD4+ and CD8+ T cells are in the immediate vicinity of IL-15–producing BM cells, suggesting a close interaction between these two cell types and a regulatory role of IL-15 on T cells. Accordingly, IL-15 induced an identical pattern of CD69 expression in peripheral blood CD4+ and CD8+ T cell subsets. Moreover, the IL-15–inducible molecules Bcl-xL, MIP-1α, MIP-1β, and CCR5 were upregulated in the human BM. In summary, our results indicate that the human BM microenvironment, in particular IL-15–producing cells, is important for the maintenance of a polyfunctional memory CD4+ and CD8+ T cell pool.

The impact of aging on memory T cell phenotype and function in the human bone marrow.

Recently, the BM has been shown to play a key role in regulating the survival and function of memory T cells. However, the impact of aging on these processes has not yet been studied. We demonstrate that the number of CD4+ and CD8+ T cells in the BM is maintained during aging. However, the composition of the T cell pool in the aged BM is altered with a decline of naïve and an increase in TEM cells. In contrast to the PB, a highly activated CD8+CD28– T cell population, which lacks the late differentiation marker CD57, accumulates in the BM of elderly persons. IL‐6 and IL‐15, which are both increased in the aged BM, efficiently induce the activation, proliferation, and differentiation of CD8+ T cells in vitro, highlighting a role of these cytokines in the age‐dependent accumulation of highly activated CD8+CD28– T cells in the BM. Yet, these age‐related changes do not impair the maintenance of a high number of polyfunctional memory CD4+ and CD8+ T cells in the BM of elderly persons. In summary, aging leads to the accumulation of a highly activated CD8+CD28– T cell population in the BM, which is driven by the age‐related increase of IL‐6 and IL‐15. Despite these changes, the aged BM is a rich source of polyfunctional memory T cells and may thus represent an important line of defense to fight recurrent infections in old age.

Traumatic intracranial haemorrhage in conscious patients with facial fractures – A review of 1959 cases

OBJECTIVEnFacial fracture patients who are conscious with a Glasgow Coma Scale (GCS) score of 15 in the absence of clinical neurological abnormalities are commonly not expected to have suffered severe intracranial pathology. However, high velocity impact may result in intracranial haemorrhage in different compartments.nnnMETHODSnOver a 7-year period, 1959 facial fracture patients with GCS scores of 15 and the absence of neurological abnormalities were analysed. In 54 patients (2.8%) computed tomography scans revealed the presence of accompanying intracranial haemorrhage (study group). These patients were compared with the 1905 patients without intracranial haemorrhage (control group).nnnRESULTSnUnivariate analysis identified accompanying vomiting/nausea and seizures, cervical spine injuries, cranial vault and basal skull fractures to be significantly associated with intracranial bleeding. In multivariate analysis the risk was increased nearly 25-fold if an episode of vomiting/nausea had occurred. Seizures increased the risk of bleeding more than 15-fold. The mean functional outcome of the study group according to the Glasgow Outcome Scale was 4.7+/-0.7.nnnCONCLUSIONnIntracranial haemorrhage cannot be excluded in patients with facial fractures despite a GCS score of 15 and normal findings following neurological examination. Predictors, such as vomiting/nausea or seizures, skull fractures and closed head injuries, enhance the likelihood of an intracranial haemorrhage and have to be considered.

Complications related to midfacial fractures: operative versus non-surgical treatment

The treatment of midfacial fractures depends on the dislocation of the fracture and patient-related limitations. Surgical treatment risks iatrogenic complications. In 740 patients with midfacial fractures, the age, sex, fracture type, concomitant injuries, cause of accident and the decision to use operative or non-surgical treatment were recorded. Follow-up was performed 6 and 12 months after the injury. In 41% the fractures were isolated; they were multiple in 59%. Initially, hypaesthesia of the infraorbital nerve was present in 10% of the single and 16% of the multiple fracture patients. Surgical treatment was performed in 57% of the single and in 75% of the multiple fracture patients. Women underwent surgical treatment considerably less frequently than men. After 6 and 12 months, significantly more complications were present in the surgically treated cohort. Nerve disturbances and meteorosensitivity were most prominent. These results, together with previous findings, indicate that there is a need for prospective clinical investigations that fulfil the criteria of evidence-based medicine to generate guidelines for decision making in trauma surgery. In the meantime, the decision to use surgical treatment for midfacial fractures has to be made carefully.

Bone marrow T cells from the femur are similar to iliac crest derived cells in old age and represent a useful tool for studying the aged immune system

BackgroundCD4+ and CD8+ T cells reside in the human bone marrow (BM) and show a heightened activation state. However, only small sample sizes are available from sources such as the iliac crest. Larger samples can be obtained from the femur in the course of hip replacement surgery. It was therefore the goal of the present study to compare the phenotype and function of BM T cells from different sources from elderly persons and to investigate how femur derived bone marrow T cells can serve as a tool to gain a better understanding of the role of adaptive immune cells in the BM in old age.ResultsBone marrow mononuclear cells (BMMC) were isolated from either the iliac crest or the femur shaft. As expected the yield of mononuclear cells was higher from femur than from iliac crest samples. There were no phenotypic differences between BMMC from the two sources. Compared to PBMC, both BM sample types contained fewer naïve and more antigen experienced CD4+ as well as CD8+ T cells, which, in contrast to peripheral cells, expressed CD69. Cytokine production was also similar in T cells from both BM types. Larger sample sizes allowed the generation of T cell lines from femur derived bone marrow using non-specific as well as specific stimulation. The phenotype of T cell lines generated by stimulation with OKT-3 and IL-2 for two weeks was very similar to the one of ex vivo BM derived T cells. Such lines can be used for studies on the interaction of different types of BM cells as shown by co-culture experiments with BM derived stromal cells. Using CMVNLV specific T cell lines we additionally demonstrated that BM samples from the femur are suitable for the generation of antigen specific T cell lines, which can be used in studies on the clonal composition of antigen specific BM T cells.ConclusionIn conclusion, our results demonstrate that BMMC from the femur shaft are a useful tool for studies on the role of T cells in the BM in old age.

A domestic porcine model for studying the effects of radiation on head and neck cancers.

BACKGROUNDnRadiation therapy (RT) of the head and neck region is often accompanied by serious side effects. Research in this area is needed to improve treatment outcomes and ameliorate therapy tolerance. Laboratory rodents are barely matching todays clinical standards in RT research. Yet domestic swine (Sus scrofa domestica) have previously proved suitable for various advanced tests in clinical research and training. We therefore investigated whether S. scrofa domestica is also appropriate for irradiation of the mandible.nnnSTUDY DESIGNnA common scheme for irradiation treatment of S. scrofa domestica mandibles in a split-mouth design was acquired by applying computed tomography (CT) scanning under sedation. Basing on close anatomic resemblance, a standard treatment plan comprising 2 opposed irradiation fields could be accomplished.nnnRESULTSnRT was carried out in a clinical environment with 2xa0×xa09xa0Gy. The resulting operating procedure facilitated complication-free sedation, transport, positioning, CT scanning, and effective irradiation.nnnCONCLUSIONnBased on common standards applied for RT in humans, domestic pigs can be employed to progress RT clinical research. Due to their human-like anatomy, physiology, size, and weight, the swine model is expedient for advancing experimental RT of the head and neck area.