Proteins Marking the Sequence of Genotoxic Signaling from Irradiated Mesenchymal Stromal Cells to CD34+ Cells.

Non-targeted effects (NTE) of ionizing radiation may initiate myeloid neoplasms (MN). Here, protein mediators (I) in irradiated human mesenchymal stromal cells (MSC) as the NTE source, (II) in MSC conditioned supernatant and (III) in human bone marrow CD34+ cells undergoing genotoxic NTE were investigated. Healthy sublethal irradiated MSC showed significantly increased levels of reactive oxygen species. These cells responded by increasing intracellular abundance of proteins involved in proteasomal degradation, protein translation, cytoskeleton dynamics, nucleocytoplasmic shuttling, and those with antioxidant activity. Among the increased proteins were THY1 and GNA11/14, which are signaling proteins with hitherto unknown functions in the radiation response and NTE. In the corresponding MSC conditioned medium, the three chaperones GRP78, CALR, and PDIA3 were increased. Together with GPI, these were the only four altered proteins, which were associated with the observed genotoxic NTE. Healthy CD34+ cells cultured in MSC conditioned medium suffered from more than a six-fold increase in γH2AX focal staining, indicative for DNA double-strand breaks, as well as numerical and structural chromosomal aberrations within three days. At this stage, five proteins were altered, among them IQGAP1, HMGB1, and PA2G4, which are involved in malign development. In summary, our data provide novel insights into three sequential steps of genotoxic signaling from irradiated MSC to CD34+ cells, implicating that induced NTE might initiate the development of MN.

Evaluating Usability of a Touchless Image Viewer in the Operating Room.

BACKGROUND: Availability of patient-specific image data, gathered from preoperatively conducted studies, like computed tomography scans and magnetic resonance imaging studies, during a surgical procedure is a key factor for surgical success and patient safety. Several alternative input methods, including recognition of hand gestures, have been proposed for surgeons to interact with medical image viewers during an operation. Previous studies pointed out the need for usability evaluation of these systems. OBJECTIVES: We describe the accuracy and usability of a novel software system, which integrates gesture recognition via machine learning into an established image viewer. METHODS: This pilot study is a prospective, observational trial, which asked surgeons to interact with software to perform two standardized tasks in a sterile environment, modeled closely to a real-life situation in an operating room. To assess usability, the validated "System Usability Scale" (SUS) was used. On a technical level, we also evaluated the accuracy of the underlying neural network. RESULTS: The neural network reached 98.94% accuracy while predicting the gestures during validation. Eight surgeons with an average of 6.5 years of experience participated in the usability study. The system was rated on average with 80.25 points on the SUS. CONCLUSION: The system showed good overall usability; however, additional areas of potential improvement were identified and further usability studies are needed. Because the system uses standard PC hardware, it made for easy integration into the operating room.

Usefulness of slice encoding for metal artifact correction (SEMAC) technique for reducing metal artifacts after total knee arthroplasty.

PURPOSE: To evaluate the usefulness of a novel MRI sequence strategy in the assessment of the periprosthetic anatomical structures after primary total knee arthroplasty. METHODS: Two MR sequences were retrospectively compared for the imaging of 15 patients with implanted cruciate-retaining/fixed-bearing TKAs (DePuy, PFC Sigma): a slice encoding sequence for metal artifact correction (SEMAC) and a standard sequence. Images were acquired on a 1.5-T system. The degree of artifact reduction was assessed using several qualitative (Likert-type scale) (artifact size, distorsion, blur, image quality, periprosthetic bone, posterior cruciate ligament, lateral collateral ligament, medial collateral ligament, patella tendon, popliteal vessels) and quantitative (artifact volume, Insall-Salvati index, length of patella/tendon, prosthesis dimensions) parameters by blinded reads performed by four investigators. The SEMAC sequences were statistically compared with the standard sequence using Wilcoxon test. Additionally, the intraclass correlation coefficient (ICC) for interobserver agreement was calculated. RESULTS: Higher levels of blurring were found with SEMAC compared to standard sequences (p < 0.001). All other qualitative parameters improved significantly with the application of SEMAC. In comparison with conventional sequences, the artifact volume was reduced by 59% utilizing SEMAC. Thus, the artifact reduction improved the precision of measurements such as Insall-Salvati index and length of patella/tendon (p < 0.001). The dimension of the tibial component (Ti alloy/polyethylene) revealed accurate values with both MRI sequences. A sufficient interobserver agreement among all readers was found with SEMAC, qualitatively ICC 0.9 (range 0.8-1) as well as quantitatively ICC 0.95 (range 0.92-0.98). CONCLUSIONS: SEMAC effectively reduces artifacts caused by metallic implants after total knee arthroplasty relative to standard imaging. This allows for an improved assessment of periprosthetic anatomical structures. This might enable an improved detectability of postoperative complications in the future. LEVEL OF EVIDENCE: Diagnostic Study Level III.

Tourniquet-induced ischaemia during total knee arthroplasty results in higher proteolytic activities within vastus medialis cells: a randomized clinical trial.

PURPOSE: Recent data suggest diminished post-operative quadriceps muscle strength after tourniquet application during total knee arthroplasty (TKA). The metabolic effects of the commonly utilized intraoperative tourniquet with consecutive ischaemia on the skeletal muscle cells were unknown. Ubiquitin proteasome system represents one of the main pathways involved in muscle protein breakdown contributing to muscle atrophy. Therefore, the purpose of the present study was to quantify the acute effects of the tourniquet application during TKA on the (1) concentrations of free/conjugated ubiquitin, (2) total ubiquitin-protein ligase activity, (3) proteasome-dependent and (4) proteasome-independent peptidase activities in the cells of vastus medialis. METHODS: The randomized, controlled, monocentric trial included 34 patients scheduled to undergo primary TKA. Each patient was randomly assigned to the tourniquet (n = 17) or non-tourniquet group (n = 17) after receiving a written consent. Muscle biopsies of (5 × 5 × 5 mm) 125 mm3 were obtained from vastus medialis immediately after performing the surgical approach and exactly 60 min later. After preparation of the muscle tissue specimen, the concentrations of the free/conjugated ubiquitin (Ub) were measured by western blot analyses. The ubiquitination was determined as biotinylated Ub incorporated into the sum of the cytosolic proteins and expressed as total ubiquitin-protein ligase activity (tUbPL). The quantification of the proteasome-dependent and proteasome-independent peptidase activities was performed with peptidase assays. RESULTS: Tourniquet application did not influence the concentration of the free/conjugated Ub. There were no differences in tUbPL activities between groups and time points. Tourniquet-induced ischaemia resulted in statistically significant higher proteasome-dependent (caspase-like p = 0.0034; chymotryptic-like p = 0.0013; tryptic-like p = 0.0036) and proteasome-independent (caspase-like p = 0.03; chymotryptic-like p = 0.0001; tryptic-like p = 0.0062) peptidase activities. CONCLUSION: Tourniquet application did not affect the free/conjugated Ub as well as tUbPL significantly, emphasizing the sophisticated regulation of ubiquitination. The proteasome-dependent peptidase activities were significantly upregulated during tourniquet application, suggesting an increase in protein degradation, which in turn might explain the skeletal muscle atrophy occurring after TKA. These findings add further knowledge and should raise the awareness of surgeons about the effects of tourniquet-induced ischaemia at the molecular level. Additional high-quality research may be warranted to examine the short- and long-term clinical significance of the present data. LEVEL OF EVIDENCE: I.

Myelodysplastic cells in patients reprogram mesenchymal stromal cells to establish a transplantable stem cell niche disease unit.

Myelodysplastic syndromes (MDSs) are a heterogeneous group of myeloid neoplasms with defects in hematopoietic stem and progenitor cells (HSPCs) and possibly the HSPC niche. Here, we show that patient-derived mesenchymal stromal cells (MDS MSCs) display a disturbed differentiation program and are essential for the propagation of MDS-initiating Lin(-)CD34(+)CD38(-) stem cells in orthotopic xenografts. Overproduction of niche factors such as CDH2 (N-Cadherin), IGFBP2, VEGFA, and LIF is associated with the ability of MDS MSCs to enhance MDS expansion. These factors represent putative therapeutic targets in order to disrupt critical hematopoietic-stromal interactions in MDS. Finally, healthy MSCs adopt MDS MSC-like molecular features when exposed to hematopoietic MDS cells, indicative of an instructive remodeling of the microenvironment. Therefore, this patient-derived xenograft model provides functional and molecular evidence that MDS is a complex disease that involves both the hematopoietic and stromal compartments. The resulting deregulated expression of niche factors may well also be a feature of other hematopoietic malignancies.

Fibula head is a useful landmark to predict the location of posterior cruciate ligament footprint prior to total knee arthroplasty.

PURPOSE: The hypothesis of our study is that a routine tibial cut during cruciate retaining TKA may result in a partial or a total removal of the PCL footprint. Therefore providing a reliable landmark is essential to estimate the probability of PCL damage with a tibial cut and to enable the surgeon to decide pre-operatively whether a cruciate retaining implant design is suitable. METHODS: In a case series of 175 cruciate retaining TKA, the routinely made standing postoperative AP-view radiographs were evaluated to determine the distance between fibula head and tibial cutting plane. In a second case series knee MRI of 223 subjects were consecutively used to measure the vertical distance between tibial attachment of PCL and fibula head. The probability of partial or total PCL damage was calculated for different vertical distances between tibial cut and fibula head. RESULTS: The vertical distance between the tibial cut and the most proximal point of the fibula head averaged 6.1 mm ±4.8 mm. The mean vertical distance from fibula head to proximal and to distal PCL footprint revealed to be 11.4 mm ±3.7 mm and 5.4 mm ±2.9 mm, respectively. The location of the insertion was not significantly different between subgroups such as age (<50 or >50 years), gender and side. Based on our results 11 (7%) knees were considered at high risk of an entire PCL removal after implantation of a cruciate retaining TKA design. CONCLUSIONS: Currently available routine tibial preparation techniques result in partial or total posterior cruciate ligament detachment. Fibula head as a landmark aids to predict the PCL location and to estimate its disruption pre- and postoperatively on AP-view radiographs.