Neuromuscular electrical stimulation to combat cognitive aging in people with spinal cord injury: protocol for a single case experimental design study.

INTRODUCTION: Individuals with spinal cord injury (SCI) can experience accelerated cognitive aging. Myokines (factors released from muscle cells during contractions), such as brain-derived neurotrophic factor (BDNF), are thought to have beneficial effects on cognition. Neuromuscular electrical stimulation (NMES) was shown to elicit a large release of myokines. However, the effects of NMES on cognitive function have not been studied. OBJECTIVE: To present the study protocol for a clinical trial evaluating the effects of NMES aimed at improving cognition and BDNF. METHODS: A replicated randomized three-phases single-case experimental design (SCED) with sequential multiple baseline time series and a single-armed prospective trial will be conducted with 15 adults with chronic SCI (> 12 months after injury) above L1 neurological level undergoing 30-min quadriceps NMES, 3 days per week for 12 weeks. MAIN STUDY ENDPOINTS: Primary endpoint is cognitive performance (assessed by a smartphone test) conducted three times per week during the baseline phase with random duration of 3 to 8 weeks, the intervention phase of 12 weeks, and the follow-up phase of 3 weeks after a no measurement rest period of 12 weeks. Secondary endpoints are changes in BDNF levels and cognitive performance measured before the baseline period, before and after intervention and after a 12 weeks follow-up. CONCLUSION: This will be the first study investigating the effects of 12 weeks NMES on both cognition and BDNF levels in individuals with SCI. The SCED results provide information on individual treatment effect courses which may direct future research. TRIAL REGISTRATION: ClinicalTrials.gov (NCT05822297, 12/01/2023).

Damage Control Orthopaedics Induced Less Trauma-Induced Coagulopathy than Early Total Care in a Porcine Polytrauma Model.

INTRODUCTION: Coagulopathic disorders (CDs) complicate treatment in polytraumatised patients. Against this background, operative strategies for fracture management are controversial in this cohort. This study therefore investigated the effects of two established operative concepts, early total care (ETC) and damage control orthopaedics (DCO), on CD in a large-animal polytrauma (PT) model. METHODS: Twenty-two animals (Sus scrofa domesticus) sustained PT involving blunt-chest trauma, liver laceration, bilateral femur fracture, and pressure-controlled haemorrhagic shock. After resuscitation, animals were allocated to ETC (n = 8), DCO (n = 8), or served as a non-traumatised control group (CG, n = 6). Animals were ventilated and monitored under ICU standards for 72 h. Blood samples were collected at baseline and post-trauma after 1.5, 2.5, 24, 48, and 72 h. Plasminogen activator inhibitor-1 (PAI-1) and thrombin-antithrombin (TAT) complex concentrations were determined by ELISA. RESULTS: Compared to the CG, ETC and DCO subjects had significantly increased plasma concentrations of PAI-1 after 2.5 h (CG vs. ETC: p = 0.0050, CG vs. DCO: p = 0.0016). Furthermore, the ETC group showed significantly increased plasma PAI-1 concentrations after 24 h compared to the CG and DCO groups (CG vs. ETC: p = 0.0002, DCO vs. ETC: p = 0.0004). During the later clinical course, concentrations of TAT were significantly increased in the ETC group compared to the CG and DCO group after 72 h (CG vs. ETC: p = 0.0290, DCO vs. ETC: p = 0.0322). CONCLUSION: PT is strongly associated with CD in the early post-traumatic course. In comparison to DCO, ETC appeared to be negatively associated with CD. Future studies must investigate this impact, especially in those patients admitted with trauma-induced coagulopathy, to improve outcomes.

Polytherapy versus monotherapy in the treatment of tibial non-unions: a retrospective study.

BACKGROUND: Treating tibial non-unions efficiently presents a challenge for orthopaedic trauma surgeons. The established gold standard involves implanting autologous bone graft with adequate fixation, but the addition of biologicals according to the so-called diamond concept has become increasingly popular in the treatment of non-unions. Previous studies have indicated that polytherapy, which involves implanting mesenchymal stem cells, bioactive factors and osteoconductive scaffolds, can improve bone healing. This study aims to evaluate the efficacy of polytherapy compared with monotherapy in treating tibial non-unions of varying severity. MATERIALS AND METHODS: Data from consecutive tibial non-unions treated between November 2014 and July 2023 were retrospectively analysed. The Non Union Scoring System (NUSS) score before non-union surgery, and the Radiographic Union Score for Tibial fractures (RUST), scored at 1, 3, 6, 9, 12 and 18 months post-surgery, were recorded. Initially, a comparison was made between the polytherapy and monotherapy groups. Subsequently, patients receiving additional surgical non-union treatment were documented, and the frequency of these treatments was tallied for a subsequent per-treatment analysis. RESULTS: A total of 34 patients were included and divided into a polytherapy group (n = 15) and a monotherapy group (n = 19). The polytherapy group demonstrated a higher NUSS score (44 (39, 52) versus 32 (29, 43), P = 0.019, z = -2.347) and a tendency towards a higher success rate (93% versus 68%, P = 0.104) compared with the monotherapy group. For the per-treatment analysis, 44 treatments were divided into the polytherapy per-treatment group (n = 20) and the monotherapy per-treatment group (n = 24). The polytherapy per-treatment group exhibited a higher NUSS score (48 (43, 60) versus 38 (30, 50), P = 0.030, z = -2.173) and a higher success rate (95% versus 58%, P = 0.006) than the monotherapy per-treatment group. Within the monotherapy per-treatment group, the NUSS score displayed excellent predictive performance (AUC = 0.9143). Setting the threshold value at 48, the sensitivity and specificity were 100.0% and 70.0%, respectively. CONCLUSIONS: Polytherapy is more effective than monotherapy for severe tibial non-unions, offering a higher success ratio. The NUSS score supports decision-making in treating tibial non-unions. LEVEL OF EVIDENCE: Level III.

MiRNAs as Potential Regulators of Enthesis Healing: Findings in a Rodent Injury Model.

MicroRNAs (miRNAs) are short non-coding RNA sequences with the ability to inhibit the expression of a target mRNA at the post-transcriptional level, acting as modulators of both the degenerative and regenerative processes. Therefore, these molecules constitute a potential source of novel therapeutic tools. In this study, we investigated the miRNA expression profile that presented in enthesis tissue upon injury. For this, a rodent enthesis injury model was developed by creating a defect at a rat's patellar enthesis. Following injury, explants were collected on days 1 (n = 10) and 10 (n = 10). Contra lateral samples (n = 10) were harvested to be used for normalization. The expression of miRNAs was investigated using a "Fibrosis" pathway-focused miScript qPCR array. Later, target prediction for the aberrantly expressed miRNAs was performed by means of the Ingenuity Pathway Analysis, and the expression of mRNA targets relevant for enthesis healing was confirmed using qPCRs. Additionally, the protein expression levels of collagens I, II, III, and X were investigated using Western blotting. The mRNA expression pattern of EGR1, COL2A1, RUNX2, SMAD1, and SMAD3 in the injured samples indicated their possible regulation by their respective targeting miRNA, which included miR-16, -17, -100, -124, -133a, -155 and -182. Furthermore, the protein levels of collagens I and II were reduced directly after the injury (i.e., day 1) and increased 10 days post-injury, while collagens III and X showed the opposite pattern of expression.

Dysregulation of Wnt signaling in bone of type 2 diabetes mellitus and diabetic Charcot arthropathy.

BACKGROUND: Type 2 diabetes mellitus (T2DM) patients show a markedly higher fracture risk and impaired fracture healing when compared to non-diabetic patients. However in contrast to type 1 diabetes mellitus, bone mineral density in T2DM is known to be normal or even regionally elevated, also known as diabetic bone disease. Charcot arthropathy is a severe and challenging complication leading to bone destruction and mutilating bone deformities. Wnt signaling is involved in increasing bone mineral density, bone homeostasis and apoptotic processes. It has been shown that type 2 diabetes mellitus is strongly associated with gene variants of the Wnt signaling pathway, specifically polymorphisms of TCF7L2 (transcription factor 7 like 2), which is an effector transcription factor of this pathway. METHODS: Bone samples of 19 T2DM patients and 7 T2DM patients with additional Charcot arthropathy were compared to 19 non-diabetic controls. qPCR analysis for selected members of the Wnt-signaling pathway (WNT3A, WNT5A, catenin beta, TCF7L2) and bone gamma-carboxyglutamate (BGLAP, Osteocalcin) was performed and analyzed using the 2-ΔΔCt- Method. Statistical analysis comprised one-way analysis of variance (ANOVA). RESULTS: In T2DM patients who had developed Charcot arthropathy WNT3A and WNT5A gene expression was down-regulated by 89 and 58% compared to healthy controls (p < 0.0001). TCF7L2 gene expression showed a significant reduction by 63% (p < 0.0001) and 18% (p = 0.0136) in diabetic Charcot arthropathy. In all diabetic patients BGLAP (Osteocalcin) was significantly decreased by at least 59% (p = 0.0019). CONCLUSIONS: For the first time with this study downregulation of members of the Wnt-signaling pathway has been shown in the bone of diabetic patients with and without Charcot arthropathy. This may serve as future therapeutic target for this severe disease.

A Microwell-Based Intestinal Organoid-Macrophage Co-Culture System to Study Intestinal Inflammation.

The mammalian intestinal epithelium contains more immune cells than any other tissue, and this is largely because of its constant exposure to pathogens. Macrophages are crucial for maintaining intestinal homeostasis, but they also play a central role in chronic pathologies of the digestive system. We developed a versatile microwell-based intestinal organoid-macrophage co-culture system that enables us to recapitulate features of intestinal inflammation. This microwell-based platform facilitates the controlled positioning of cells in different configurations, continuous in situ monitoring of cell interactions, and high-throughput downstream applications. Using this novel system, we compared the inflammatory response when intestinal organoids were co-cultured with macrophages versus when intestinal organoids were treated with the pro-inflammatory cytokine TNF-α. Furthermore, we demonstrated that the tissue-specific response differs according to the physical distance between the organoids and the macrophages and that the intestinal organoids show an immunomodulatory competence. Our novel microwell-based intestinal organoid model incorporating acellular and cellular components of the immune system can pave the way to unravel unknown mechanisms related to intestinal homeostasis and disorders.

Transcript-Activated Coatings on Titanium Mediate Cellular Osteogenesis for Enhanced Osteointegration.

Osteointegration is one of the most important factors for implant success. Several biomolecules have been used as part of drug delivery systems to improve implant integration into the surrounding bone tissue. Chemically modified mRNA (cmRNA) is a new form of therapeutic that has been used to induce bone healing. Combined with biomaterials, cmRNA can be used to develop transcript-activated matrices for local protein production with osteoinductive potential. In this study, we aimed to utilize this technology to create bone morphogenetic protein 2 (BMP2) transcript-activated coatings for titanium (Ti) implants. Therefore, different coating methodologies as well as cmRNA incorporation strategies were evaluated. Three different biocompatible biomaterials were used for the coating of Ti, namely, poly-d,l-lactic acid (PDLLA), fibrin, and fibrinogen. cmRNA-coated Ti disks were assayed for transfection efficiency, cmRNA release, cell viability and proliferation, and osteogenic activity in vitro. We found that cmRNA release was significantly delayed in Ti surfaces previously coated with biomaterials. Consequently, the transfection efficiency was greatly improved. PDLLA coating improved the transfection efficiency in a concentration-dependent manner. Lower PDLLA concentration used for the coating of Ti resulted in higher transfection efficiency. Fibrin and fibrinogen coatings showed even higher transfection efficiencies compared to all PDLLA concentrations. In those disks, not only the expression was up to 24-fold higher but also the peak of maximal expression was delayed from 24 h to 5 days, and the duration of expression was also extended until 7 days post-transfection. For fibrin, higher transfection efficiencies were obtained in the coatings with the lowest thrombin amounts. Accordingly, fibrinogen coatings gave the best results in terms of cmRNA transfection. All biomaterial-coated Ti surfaces showed improved cell viability and proliferation, though this was more noticeable in the fibrinogen-coated disks. The latter was also the only coating to support significant amounts of BMP2 produced by C2C12 cells in vitro. Osteogenesis was confirmed using BMP2 cmRNA fibrinogen-coated Ti disks, and it was dependent of the cmRNA amount present. Alkaline phosphatase (ALP) activity of C2C12 increased when using fibrinogen coatings containing 250 ng of cmRNA or more. Similarly, mineralization was also observed that increased with increasing cmRNA concentration. Overall, our results support fibrinogen as an optimal material to deliver cmRNA from titanium-coated surfaces.

Wound fluid under occlusive dressings from diabetic patients show an increased angiogenic response and fibroblast migration.

INTRODUCTION: Metabolic diseases like diabetes mellitus often show prolonged healing and chronic wounds. Occlusive wound dressings are known to support wound closure by creating a moist environment which supports collagen synthesis, epithelialization and angiogenesis. We aimed to assess the effect of occlusion on diabetic wound fluid on the cellular level regarding fibroblast activity and angiogenetic response. MATERIAL AND METHODS: 22 split skin donor sites from 22 patients (11 patients with diabetes mellitus) were treated with occlusive dressings intraoperatively. On day 3, fluid and blood serum samples were harvested while changing the dressings. The influence of wound fluid on fibroblasts was assessed by measuring metabolic activity (Alamar Blue assay, Casey Counter), cell stress/death (LDH assay) and migration (in vitro wound healing assay) of fibroblasts. Angiogenesis of endothelial cells (HUVEC) was analyzed with the tube formation assay. Furthermore, a Magnetic Luminex Assay for multi-cytokines detection was performed focusing on inflammatory and pro-angiogenetic cytokines. RESULTS: The influence of wound fluid under occlusive dressings from diabetic patients showed a significantly increased angiogenic response and fibroblast migration compared to the non-diabetic patient group. Additionally, cell stress was increased in the diabetic group. Cytokine analysis showed an increase in VEGF-A in the diabetic group. CONCLUSION: Occlusive dressings may stimulate regenerative effects in diabetic wounds. Our in-vitro study shows the influence of wound fluid under occlusive dressings from diabetic patients on angiogenesis, migration and proliferation of fibroblasts, which are essential modulators of wound healing and scar modulation.

Measures of infection prevention and incidence of SARS-CoV-2 infections in cancer patients undergoing radiotherapy in Germany, Austria and Switzerland.

PURPOSE: COVID-19 infection has manifested as a major threat to both patients and healthcare providers around the world. Radiation oncology institutions (ROI) deliver a major component of cancer treatment, with protocols that might span over several weeks, with the result of increasing susceptibility to COVID-19 infection and presenting with a more severe clinical course when compared with the general population. The aim of this manuscript is to investigate the impact of ROI protocols and performance on daily practice in the high-risk cancer patients during this pandemic. METHODS: We addressed the incidence of positive COVID-19 cases in both patients and health care workers (HCW), in addition to the protective measures adopted in ROIs in Germany, Austria and Switzerland using a specific questionnaire. RESULTS: The results of the questionnaire showed that a noteworthy number of ROIs were able to complete treatment in SARS-CoV­2 positive cancer patients, with only a short interruption. The ROIs reported a significant decrease in patient volume that was not impacted by the circumambient disease incidence, the type of ROI or the occurrence of positive cases. Of the ROIs 16.5% also reported infected HCWs. About half of the ROIs (50.5%) adopted a screening program for patients whereas only 23.3% also screened their HCWs. The range of protective measures included the creation of working groups, instituting home office work and protection with face masks. Regarding the therapeutic options offered, curative procedures were performed with either unchanged or moderately decreased schedules, whereas palliative or benign radiotherapy procedures were more often shortened. Most ROIs postponed or cancelled radiation treatment for benign indications (88.1%). The occurrence of SARS-CoV­2 infections did not affect the treatment options for curative procedures. Non-university-based ROIs seemed to be more willing to change their treatment options for curative and palliative cases than university-based ROIs. CONCLUSION: Most ROIs reported a deep impact of SARS-CoV­2 infections on their work routine. Modification and prioritization of treatment regimens and the application of protective measures preserved a well-functioning radiation oncology service and patient care.

Cytokine and immune cell profiling in the cerebrospinal fluid of patients with neuro-inflammatory diseases.

BACKGROUND: Cytokines play multiple roles during neuro-inflammatory processes and several cytokines have been studied in the context of specific diseases. This study provides a comprehensive picture of cerebrospinal fluid (CSF) changes during neuro-inflammation by analyzing multiple cytokines in combination with immune cell subsets and standard CSF parameters. METHODS: Using multiplex assays, we simultaneously measured 36 cytokines (CCL1-3, CCL7, CCL8, CCL11, CCL13, CCL19, CCL20, CCL22-27, CXCL1, CXCL2, CXCL5, CXCL6, CXCL8, CXCL9, CXCL11-13, CXCL16, CX3CL1, IL2, IL4, IL6, IL10, IL16, GM-CSF, IFNγ, MIF, TNFα, and MIB1ß) in the CSF and serum of 75 subjects. Diagnoses included clinically isolated syndrome and relapsing-remitting multiple sclerosis (MS, n = 18), secondary progressive MS (n = 8), neuro-syphilis (n = 6), Lyme neuro-borreliosis (n = 13), bacterial and viral meningitis (n = 20), and patients with non-inflammatory neurological diseases (NIND, n = 10). Cytokine concentrations were correlated with CSF standard parameters and CSF immune cell subsets (CD4 and CD8 T cells, B cells, plasmablasts, monocytes, and NK cells) quantified by flow cytometry. RESULTS: We observed increased levels of multiple cytokines (26/36) in patients with neuro-inflammatory diseases when compared to NIND that consistently correlated with CSF cell count and QAlbumin. Most CSF cytokine concentrations correlated with each other, but correlations between CSF and serum values were scarce (3/36). Within the CSF compartment, CXCL13 showed a strong association with B cells when analyzing all patients, as well as patients with an intact blood-brain barrier (BBB). NK cells positively correlated with CSF concentrations of multiple cytokines (22/36) when analyzing all patients. These correlations were maintained when looking at patients with a disrupted BBB but not detectable in patients with an intact BBB. CONCLUSIONS: Under conditions of neuro-inflammation, multiple CSF cytokines are regulated in parallel and most likely produced locally. A combined increase of CSF CXCL13 levels and B cells occurs under conditions of an intact BBB. Under conditions of a disrupted BBB, CSF NK cells show significantly increased values and seem to have a major contribution to overall inflammatory processes, reflected by a strong correlation with multiple cytokines. Future studies are necessary to address the exact kinetics of these cytokines during neuro-inflammation and their relation to specific diseases phenotypes.

A Growth Factor-Free Co-Culture System of Osteoblasts and Peripheral Blood Mononuclear Cells for the Evaluation of the Osteogenesis Potential of Melt-Electrowritten Polycaprolactone Scaffolds.

Scaffolds made of biodegradable biomaterials are widely used to guide bone regeneration. Commonly, in vitro assessment of scaffolds' osteogenesis potential has been performed predominantly in monoculture settings. Hence, this study evaluated the potential of an unstimulated, growth factor-free co-culture system comprised of osteoblasts (OB) and peripheral blood mononuclear cells (PBMC) over monoculture of OB as an in vitro platform for screening of bone regeneration potential of scaffolds. Particularly, this study focuses on the osteogenic differentiation and mineralized matrix formation aspects of cells. The study was performed using scaffolds fabricated by means of a melt electrowriting (MEW) technique made of medical-grade polycaprolactone (PCL), with or without a surface coating of calcium phosphate (CaP). Qualitative results, i.e., cell morphology by fluorescence imaging and matrix mineralization by von Kossa staining, indicated the differences in cell behaviours in response to scaffolds' biomaterial. However, no obvious differences were noted between OB and OB+PBMC groups. Hence, quantitative investigation, i.e., alkaline phosphatase (ALP), tartrate-resistant acid phosphatase (TRAP) activities, and gene expression were quantitatively evaluated by reverse transcription-polymerase chain reaction (RT-qPCR), were evaluated only of PCL/CaP scaffolds cultured with OB+PBMC, while PCL/CaP scaffolds cultured with OB or PBMC acted as a control. Although this study showed no differences in terms of osteogenic differentiation and ECM mineralization, preliminary qualitative results indicate an obvious difference in the cell/non-mineralized ECM density between scaffolds cultured with OB or OB+PBMC that could be worth further investigation. Collectively, the unstimulated, growth factor-free co-culture (OB+PBMC) system presented in this study could be beneficial for the pre-screening of scaffolds' in vitro bone regeneration potential prior to validation in vivo.

Impact of Periprosthetic Fibroblast-Like Cells on Osteoclastogenesis in Co-Culture with Peripheral Blood Mononuclear Cells Varies Depending on Culture System.

Co-culture studies investigating the role of periprosthetic fibroblasts (PPFs) in inflammatory osteoclastogenesis reveal contrary results, partly showing an osteoprotective function of fibroblasts and high OPG expression in monolayer. These data disagree with molecular analyses of original periosteolytic tissues. In order to find a more reliable model, PPFs were co-cultivated with peripheral blood mononuclear cells (PBMCs) in a transwell system and compared to conventional monolayer cultures. The gene expression of key regulators of osteoclastogenesis (macrophage colony-stimulating factor (MCSF), receptor activator of NF-κB ligand (RANK-L), osteoprotegerin (OPG), and tumor necrosis factor alpha (TNFα)) as well as the ability of bone resorption were analyzed. In monolayer co-cultures, PPFs executed an osteoprotective function with high OPG-expression, low RANK-L/OPG ratios, and a resulting inhibition of osteolysis even in the presence of MCSF and RANK-L. For transwell co-cultures, profound changes in gene expression, with a more than hundredfold decrease of OPG and a significant upregulation of TNFα were observed. In conclusion, we were able to show that a change of culture conditions towards a transwell system resulted in a considerably more osteoclastogenic gene expression profile, being closer to findings in original periosteolytic tissues. This study therefore presents an interesting approach for a more reliable in vitro model to examine the role of fibroblasts in periprosthetic osteoclastogenesis in the future.

Development, Characterization and In Vitro Biological Properties of Scaffolds Fabricated From Calcium Phosphate Nanoparticles.

Ceramic materials mimic the mineral composition of native bone and feature osteoconductive properties; they are therefore used to regenerate bone tissue. Much research focuses on increasing the porosity and pore interconnectivity of ceramic scaffolds to increase osteoconductivity, cell migration and cell-cell interaction. We aimed to fabricate biocompatible 3D-scaffolds featuring macro- and microporous calcium phosphates with high pore interconnection. Nanoparticles of hydroxyapatite (HA) and calcium deficient hydroxyapatite (CDHA) were synthesized by wet chemical precipitation. Scaffolds were produced from them by the replication polymeric foam technique. Solid content and sintering temperature were varied. Nanoparticles and scaffolds were characterized regarding morphology, chemical and mineral composition, porosity and mechanical properties. Biocompatibility, cell attachment and distribution were evaluated in vitro with human adipose mesenchymal stem cells. Scaffolds with total porosity of 71%-87%, pores in the range of 280-550 µm and connectivity density up to 43 mm-3 were obtained. Smaller pore sizes were obtained at higher sintering temperature. High solid content resulted in a decrease of total porosity but increased interconnectivity. Scaffolds 50HA/50ß-TCP featured superior interconnectivity and mechanical properties. They were bioactive and biocompatible. High HA solid content (40 wt.%) in the HA pure scaffolds was negative for cell viability and proliferation, while in the 50HA/50ß-TCP composite scaffolds it resulted more biocompatible.

Immobilization of Denosumab on Titanium Affects Osteoclastogenesis of Human Peripheral Blood Monocytes.

Immobilization of proteins has been examined to improve implant surfaces. In this study, titanium surfaces were modified with nanofunctionalized denosumab (cDMAB), a human monoclonal anti-RANKL IgG. Noncoding DNA oligonucleotides (ODN) served as linker molecules between titanium and DMAB. Binding and release experiments demonstrated a high binding capacity of cDMAB and continuous release. Human peripheral mononuclear blood cells (PBMCs) were cultured in the presence of RANKL/MCSF for 28 days and differentiated into osteoclasts. Adding soluble DMAB to the medium inhibited osteoclast differentiation. On nanofunctionalized titanium specimens, the osteoclast-specific TRAP5b protein was monitored and showed a significantly decreased amount on cDMAB-titanium in PBMCs + RANKL/MCSF. PBMCs on cDMAB-titanium also changed SEM cell morphology. In conclusion, the results indicate that cDMAB reduces osteoclast formation and has the potential to reduce osteoclastogenesis on titanium surfaces.

The posttraumatic activation of CD4+ T regulatory cells is modulated by TNFR2- and TLR4-dependent pathways, but not by IL-10.

Platelets modulate the immune system following injury by interacting with CD4+ T regulatory cells (CD4+ Tregs). The underlying mechanisms remain unsolved. We hypothesize paracrine interactions via Tumor necrosis factor-alpha (TNFα)-, Toll like receptor-4 (TLR4)-, and Interleukin-10 (IL-10). In the murine burn injury model, CD4+ Treg activation pathways were selectively addressed using TNFR2-, TLR4- and IL-10-deficient mice. The CD4+ Treg signalling molecule PKC-θ was analyzed using phospho-flow cytometry to detect rapid cell activation. Thromboelastometry (ROTEM®) was used to assess platelet activation. Injury induced significant early activation of CD4+ Tregs, disruption of TNFR2 and TLR4 activation pathways resulted in lower activity. The disruption of IL-10 crosstalk had no significant impact. Selective disruption of paracrine interactions is associated with changes in posttraumatic hemostasis parameters. TNFR2- and TLR4-dependent pathways modulate the activation of CD4+ Tregs following trauma. In contrast, we did not observe a role of IL-10 in the posttraumatic activation of CD4+ Tregs. ONE SENTENCE SUMMARY: TLR4- and TNFR2-dependent mechanisms, but not IL-10-dependent pathways, modulate the anti-inflammatory response of CD4+ Tregs following trauma.

Dose-dependent effects of peroxisome proliferator-activated receptors ß/δ agonist on systemic inflammation after haemorrhagic shock.

INTRODUCTION: PPARß/δ agonists are known to modulate the systemic inflammatory response after sepsis. In this study, inflammation modulation effects of PPARß/δ are investigated using the selective PPARß/δ agonist (GW0742) in a model of haemorrhagic shock (HS)-induced sterile systemic inflammation. METHODS: Blood pressure-controlled (35±5mmHg) HS was performed in C57/BL6 mice for 90min. Low-dose GW0742 (0.03mg/kg/BW) and high-dose GW0742 (0.3mg/kg/BW) were then administered at the beginning of resuscitation. Mice were sacrificed 6h after induction of HS. Plasma levels of IL-6, IL-1ß, IL-10, TNFα, KC, MCP-1, and GM-CSF were determined by ELISA. Myeloperoxidase (MPO) activity in pulmonary and liver tissues was analysed with standardised MPO kits. RESULTS: In mice treated with high-dose GW0742, plasma levels of IL-6, IL-1ß, and MCP-1 were significantly increased compared to the control group mice. When compared to mice treated with low-dose GW0742 plasma levels of IL-6, IL-1ß, GM-CSF, KC, and MCP-1 were significantly elevated in high-dose-treated mice. Low-dose GW0742 treatment was associated with a non-significant downtrend of inflammatory factors in mice with HS. No significant changes of MPO activity in lung and liver were observed between the control group and the GW0742 treatment groups. CONCLUSION: This study identified dose-dependent effects of GW0742 on systemic inflammation after HS. While high-dose GW0742 substantially enhanced the systemic inflammatory response, low-dose GW0742 led to a downtrend of pro-inflammation cytokine expression. The exact mechanisms are yet unknown and need to be assessed in further studies.

Whole-body computed tomography in severely injured patients.

PURPOSE OF REVIEW: To provide an update on the relevant and recent studies on whole-body computed tomography (WBCT) imaging of severely injured patients. RECENT FINDINGS: The advantages of WBCT in time saving, diagnostic accuracy and even in survival have been proven in numerous studies. WBCT can also be beneficial in haemodynamically unstable major trauma patients. The CT scanner should be located close to the emergency department or even in the trauma room. The issue of radiation is still quite important, however, iterative as well as split-bolus protocols can nowadays reduce radiation significantly. The question: which trauma patient should receive WBCT and which not is not yet solved sufficiently. Postmortem WBCT has a promising potential to promptly define the definitive cause of death of trauma victims comparably to traditional autopsy. SUMMARY: On account of the recent advances, whole-body CT has become a crucial part of the initial in-hospital assessment of severely injured patients. It is recommended as the standard radiological tool for the emergency diagnostic work-up in major trauma patients.

Healthcare IT Utilization and Penetration among Physicians: Novel IT Solutions in Healthcare - Use and Acceptance in Hospitals.

BACKGROUND: Healthcare IT (HIT) increasingly gains public attention and clinical daily relevance. A growing number of patients and physicians increasingly relies on IT services to monitor and support well-being and recovery both in their private and professional environment. This is assumed to develop rapidly in the upcoming years. OBJECTIVE: This study examines the current status of HIT, its use and penetration among physicians in hospitals and researches utilization as well as future expectations regarding HIT. METHODS: Physicians in Germany, Austria and Switzerland were addressed via e-mail to answer a standardized Internet-based questionnaire consisting of 17 multiple-choice and 3 open text questions. Parameters were evaluated in 5 categories: general use, frequency, acceptance, IT needs and future expectations. RESULTS: An overall 234 physicians (response rate 83.6%) with a median age of 45 (range 25-60) responded and filled out the entire online questionnaire. A significant correlation between parameters gender, age and level of training (resident, specialist, consultant etc.) was proven. The professional, medical employment of technology shows a strong correlation with age as well as level of training. Whereas increasing age among physicians is associated with a decreasing level of application of HIT, a higher training level is accompanied by an increasing level of professional application of IT services and tools within the healthcare context. Routine employment of HIT is regarded as a necessary and positive standard. Most users assume the importance of HIT to strongly grow in the future in comparison to current use. A clear lack of trust towards data security and storage is recognized on both patient and physician sides. Needs are currently satisfied by employing privately acquired IT in the professional setup rather than the hospitals'. Future expectations from HIT show a clear demand for interoperability and exchangeability of data. CONCLUSIONS: The results display a clear gap between demand and expectations of IT for medical purposes. The rate of use of HIT applications generally correlates with age, gender as well as role within the hospital and type of employment within the healthcare sector. The current offering does not satisfy the needs of healthcare professionals.

Co-Culture with Human Osteoblasts and Exposure to Extremely Low Frequency Pulsed Electromagnetic Fields Improve Osteogenic Differentiation of Human Adipose-Derived Mesenchymal Stem Cells.

Human adipose-derived mesenchymal stem cells (Ad-MSCs) have been proposed as suitable option for cell-based therapies to support bone regeneration. In the bone environment, Ad-MSCs will receive stimuli from resident cells that may favor their osteogenic differentiation. There is recent evidence that this process can be further improved by extremely low frequency pulsed electromagnetic fields (ELF-PEMFs). Thus, the project aimed at (i) investigating whether co-culture conditions of human osteoblasts (OBs) and Ad-MSCs have an impact on their proliferation and osteogenic differentiation; (ii) whether this effect can be further improved by repetitive exposure to two specific ELF-PEMFs (16 and 26 Hz); (iii) and the effect of these ELF-PEMFs on human osteoclasts (OCs). Osteogenic differentiation was improved by co-culturing OBs and Ad-MSCs when compared to the individual mono-cultures. An OB to Ad-MSC ratio of 3:1 had best effects on total protein content, alkaline phosphatase (AP) activity, and matrix mineralization. Osteogenic differentiation was further improved by both ELF-PEMFs investigated. Interestingly, only repetitive exposure to 26 Hz ELF-PEMF increased Trap5B activity in OCs. Considering this result, a treatment with gradually increasing frequency might be of interest, as the lower frequency (16 Hz) could enhance bone formation, while the higher frequency (26 Hz) could enhance bone remodeling.

Dual inhibition of thrombin and activated factor X attenuates disseminated intravascular coagulation and protects organ function in a baboon model of severe Gram-negative sepsis.

BACKGROUND: Inhibition of procoagulant pathways may improve outcome in sepsis. We examined whether a dual short-acting thrombin (factor II) and factor X (FX)a inhibitor (SATI) ameliorates sepsis-induced disseminated intravascular coagulation (DIC) and is organ-protective. METHODS: Escherichia coli were infused for 2 h in 22 anesthetized baboons. The control (CO) group (n = 8) received sterile isotonic solution only. In the treatment groups, SATI was administered starting 15 minutes after the end of the bacterial exposure. In the low-dose group (LD-SATI, n = 8), SATI was infused with 75 µg/kg/h for the first hour, followed by 23 µg/kg/h until the end of the study. In the high-dose SATI group (HD-SATI, n = 6), 225 µg/kg/h was administered for the first hour followed by continuous infusion of 69 µg/kg/h until termination of the study. RESULTS: Sepsis-induced DIC was attenuated, as reflected by lower peak thrombin-antithrombin complexes (threefold) and D-dimer levels (twofold) in both SATI groups compared to the CO. This coincided with strongly improved cell/organ protection assessed by decreased levels of lactate dehydrogenase (threefold), creatinine (twofold), aspartate aminotransferase (threefold), and amylase (twofold) compared to the CO group. Anuria, which started at 8 h in the CO group, was prevented in both SATI groups. Peak interleukin-6 release at 12 h was prevented in the treatment groups. In both SATI groups, fewer catecholamines were necessary and no bleeding complications were observed. CONCLUSIONS: Dual inhibition of thrombin and FXa preserved activation of coagulation, protected organ function and ameliorated inflammation in severe Gram-negative sepsis in baboons. SATI could be a novel therapeutic agent against sepsis-induced DIC.