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Abstract Background Ehlers-Danlos syndrome Type IV (aka Vascular Ehlers Danlos, or vEDS) is a dominantly inherited mutation in the Collagen 3A1 gene (COL3A1). The disease is characterized by tissue friability and age-related susceptibility to arterial aneurysm, dissection and rupture as well as uterine and bowl tears. These clinical manifestations result in major surgical intervention and decreased life expectancy. Understanding how mutations in COL3A1 impact the structure and function of the extracellular matrix (ECM) is important to managing the disease and finding treatments. Results Skin fibroblasts from vEDS subjects heterozygous for the p.G588S pathogenic variant in the COL3A1 gene and a normal individual were cultured and studied. Proteomics analysis identified dozens of upregulated proteins related to extracellular matrix dysregulation that is characteristic of fibrosis. Gene expression libraries from cultured primary fibroblasts were screened for messenger RNA (mRNA) markers of ECM degradation. The proteomics and targeted gene expression array results were largely consistent with dysregulation of the extracellular matrix in vEDS. The data show upregulation of multiple Collagen proteins and genes, other ECM components, and enzymes related to ECM processing and turn-over. vEDS fibroblasts expressed significantly more cross linked C-Telopeptide of Collagen III (CTXIII) than normal fibroblasts, indicative of Collagen III degradation and turn-over. Further, the expression and activity of Lysyl Oxidase (LOX), an enzyme that initiates covalent cross-linking of soluble collagen and elastin into protease resistant fibers, is elevated in vEDS fibroblasts compared to normal fibroblasts. Conclusion Together, these findings suggest dysregulated ECM deposition and processing, reminiscent of a state of fibrosis. Therapeutics that target the dysregulated ECM proteins or help replace damaged tissue may improve clinical outcomes.

Abstract Background Scaffolds for tissue engineering can be received by whole organ decellularization while maintaining the site-specific extracellular matrix and the vascular tree. One among other decellularization techniques is the perfusion-based method using specific agents e.g. SDS for the elimination of cellular components. While SDS can disrupt the composition of the extracellular matrix and impair the adherence and growth of site-specific cells there are indications that xenogeneic cell types may benefit from protein denaturation by using higher detergent concentrations. The aim of this work is to investigate the effect of two different SDS-concentrations (i.e. 0.66% and 3%) on the ability of human endothelial cells to adhere and proliferate in an acellular rat kidney scaffold. Material and methods Acellular rat kidney scaffold was obtained by perfusion-based decellularization through the renal artery using a standardized protocol including SDS at concentrations of 0.66% or 3%. Subsequently cell seeding was performed with human immortalized endothelial cells EA.hy 926 via the renal artery. Recellularized kidneys were harvested after five days of pressure-controlled dynamic culture followed sectioning, histochemical and immunohistochemical staining as well as semiquantitative analysis. Results Efficacy of decellularization was verified by absence of cellular components as well as preservation of ultrastructure and adhesive proteins of the extracellular matrix. In semiquantitative analysis of recellularization, cell count after five days of dynamic culture more than doubled when using the gentle decellularization protocol with a concentration of SDS at 0.66% compared to 3%. Detectable cells maintained their endothelial phenotype and presented proliferative behavior while only a negligible fraction underwent apoptosis. Conclusion Recellularization of acellular kidney scaffold with endothelial cells EA.hy 926 seeded through the renal artery benefits from gentle decellularization procedure. Because of that, decellularization with a SDS concentration at 0.66% should be preferred in further studies and coculture experiments.

Bioreactors with a controlled physiological environment are being developed to study various cell processes. The influences of mechanostimulation on bone cell cultures can be investigated using a compression bioreactor. The developed bioreactor system applies a cyclic compression force to the specimen via an eccentrically mounted push rod. The compression force is monitored by a force sensor to detect changes in the material properties of the specimen. Depending on the piston setting, a stroke of 0.28 - 2.50 mm can be applied to the specimen. The bioreactor system was tested with a trial run of 18 days. A sample was continuously stimulated with a loading frequency of 2 Hz and a stroke of 1.50 mm. The sterility in the cell chamber as well as the functionality of the realised bioreactor stimulation system could be successfully confirmed

Attempts to develop a therapeutic vaccine against human papillomavirus (HPV)-induced malignancies have mostly not been clinically successful to date. One reason may be the hypoxic microenvironment present in most tumors, including cervical cancer. Hypoxia dysregulates the levels of human leukocyte antigen (HLA) class I molecules in different tumor entities, impacts the function of cytotoxic T cells, and leads to decreased protein levels of the oncoproteins E6 and E7 in HPV-transformed cells. Therefore, we investigated the effect of hypoxia on the presentation of HPV16 E6- and E7-derived epitopes in cervical cancer cells and its effect on epitope-specific T cell cytotoxicity. Hypoxia induced downregulation of E7 protein levels in all analyzed cell lines, as assessed by Western blotting. However, contrary to previous reports, no perturbation of antigen processing and presentation machinery (APM) components and HLA-A2 surface expression upon hypoxia treatment was detected by mass spectrometry and flow cytometry, respectively. Cytotoxicity assays performed in hypoxic conditions showed differential effects on the specific killing of HPV16-positive cervical cancer cells by epitope-specific CD8+ T cell lines in a donor- and peptide-specific manner. Effects of hypoxia on the expression of PD-L1 were ruled out by flow cytometry analysis. Altogether, our results under hypoxia show a decreased expression of E6 and E7, but an intact APM, and epitope- and donor-dependent effects on T cell cytotoxicity towards HPV16-positive target cells. This suggests that successful immunotherapies can be developed for hypoxic HPV-induced cervical cancer, with careful choice of target epitopes, and ideally in combination with hypoxia-alleviating measures.

Abstract Background For focal cartilage defects, biological repair might be ineffective in patients over 45 years. A focal metallic implant (FMI) (Hemi-CAP Arthrosurface Inc., Franklin, MA, USA) was designed to reduce symptoms. The aim of this study was to evaluate the effects of a FMI on the opposing tibial cartilage in a biomechanical set-up. It is hypothesized that a FMI would not damage the opposing cartilage under physiological loading conditions. Methods An abrasion machine was used to test the effects of cyclic loading on osteochondral plugs. The machine applied a compressive load of 33 N and sheared the samples 10 mm in the anteroposterior direction by 1 Hz. Tibial osteochondral plugs from porcine knees were placed in opposition to a FMI and cycled for 1 or 6 h. After testing each plug was fixed, stained and evaluated for cartilage damage. Results After 1 h of loading (n = 6), none of the osteochondral plugs showed histologic signs of degradation. After 6 h of loading (n = 6) three samples had histologic signs of injury in the tangential zone (grade 1) and one had signs of injury in the transitional and deep zones (grade 2). Exploration for 6 h resulted in significant more cartilage damage compared to the shorter exploration time (p = 0.06). However, no significant difference between saline and hyaluronic acid was evident (p = 0.55). Conclusion Under physiologic loading conditions, contact with a FMI leads to cartilage damage in the opposing articular cartilage in six hours. In clinical practice, a thorough analysis of pre-existing defects on the opposing cartilage is recommended when FMI is considered.

American youth are awash in media. They have television sets in their bedrooms, personal computers in their family rooms, and digital music players and cell phones in their backpacks. They spend more time with media than any single activity other than sleeping, with the average American eight- to eighteen-year-old reporting more than six hours of daily media use. The growing phenomenon of "media multitasking"--using several media concurrently--multiplies that figure to eight and a half hours of media exposure daily. Donald Roberts and Ulla Foehr examine how both media use and media exposure vary with demographic factors such as age, race and ethnicity, and household socioeconomic status, and with psychosocial variables such as academic performance and personal adjustment. They note that media exposure begins early, increases until children begin school, drops off briefly, then climbs again to peak at almost eight hours daily among eleven- and twelve-year-olds. Television and video exposure is particularly high among African American youth. Media exposure is negatively related to indicators of socioeconomic status, but that relationship may be diminishing. Media exposure is positively related to risk-taking behaviors and is negatively related to personal adjustment and school performance. Roberts and Foehr also review evidence pointing to the existence of a digital divide--variations in access to personal computers and allied technologies by socioeconomic status and by race and ethnicity. The authors also examine how the recent emergence of digital media such as personal computers, video game consoles, and portable music players, as well as the media multitasking phenomenon they facilitate, has increased young people's exposure to media messages while leaving media use time largely unchanged. Newer media, they point out, are not displacing older media but are being used in concert with them. The authors note which young people are more or less likely to use several media concurrently and which media are more or less likely to be paired with various other media. They argue that one implication of such media multitasking is the need to reconceptualize "media exposure." (Contains 4 tables, 4 figures and 50 endnotes.)

In this article, the existence, stability and bifurcation structure of time-periodic solutions (including ones that also have the property of spatial localization, i.e., breathers) are studied in an array of cantilevers that have magnetic tips. The repelling magnetic tips are responsible for the intersite nonlinearity of the system, whereas the cantilevers are responsible for the onsite (potentially nonlinear) force. The relevant model is of the mixed Fermi-Pasta-Ulam-Tsingou and Klein-Gordon type with both damping and driving. In the case of base excitation, we provide experimental results to validate the model. In particular, we identify regions of bistability in the model and in the experiment, which agree with minimal tuning of the system parameters. We carry out additional numerical explorations in order to contrast the base excitation problem with the boundary excitation problem and the problem with a single mass defect. We find that the base excitation problem is more stable than the boundary excitation problem and that breathers are possible in the defect system. The effect of an onsite nonlinearity is also considered, where it is shown that bistability is possible for both softening and hardening cubic nonlinearities.

In this study, topology optimized, patient specific osteosynthesis plates (TOPOS-implants) are evaluated for the mandibular reconstruction using fibula segments. These shape optimized implants are compared to a standard treatment with miniplates (thickness: 1.0 mm, titanium grade 4) in biomechanical testing using human cadaveric specimen. Mandible and fibula of 21 body donors were used. Geometrical models were created based on automated segmentation of CT-scans of all specimens. All reconstructions, including cutting guides for osteotomy as well as TOPOS-implants, were planned using a custom-made software tool. The TOPOS-implants were produced by electron beam melting (thickness: 1.0 mm, titanium grade 5). The fibula-reconstructed mandibles were tested in static and dynamic testing in a multi-axial test system, which can adapt to the donor anatomy and apply side-specific loads. Static testing was used to confirm mechanical similarity between the reconstruction groups. Force-controlled dynamic testing was performed with a sinusoidal loading between 60 and 240 N (reconstructed side: 30% reduction to consider resected muscles) at 5 Hz for up to 5 · 105 cycles. There was a significant difference between the groups for dynamic testing: All TOPOS-implants stayed intact during all cycles, while miniplate failure occurred after 26.4% of the planned loading (1.32 · 105 ± 1.46 · 105 cycles). Bone fracture occurred in both groups (miniplates: n = 3, TOPOS-implants: n = 2). A correlation between bone failure and cortical bone thickness in mandible angle as well as the number of bicortical screws used was demonstrated. For both groups no screw failure was detected. In conclusion, the topology optimized, patient specific implants showed superior fatigue properties compared to miniplates in mandibular reconstruction. Additionally, the patient specific shape comes with intrinsic guiding properties to support the reconstruction process during surgery. This demonstrates that the combination of additive manufacturing and topology optimization can be beneficial for future maxillofacial surgery.

BackgroundA substantial proportion of echinococcosis transmission to humans via contamination of food has been assumed. However, the relative importance of food as a transmission vehicle has previously been estimated through expert opinion rather than empirical data.ObjectiveTo find and evaluate empirical data that could be used to estimate the source attribution of echinococcosis, in particular the proportion that is transmitted through contaminated food.MethodsA systematic review was undertaken to identify reports on the risk factors for human cystic (CE) and alveolar (AE) echinococcosis. Data bases searched included PubMed, Scopus, Web of Knowledge, Cab Direct, Science Direct, Google Scholar, eLIBRARY.RU, CyberLeninka, CNKI and VIP. Search terms included Echinococc*, hydatid, epidemiology, logistic regression, risk factors, odds ratio, relative risk, risk factors. Reports, including grey literature where available, that had suitable data were selected and data were extracted. The main pathways of transmission were hypothesised to be contact with the definitive host, contaminated water, contaminated food and contaminated environment (other than food). For each study the attributable fraction for these potential sources of infection was calculated from the data presented. A meta-analysis was then undertaken to obtain pooled estimates for the relative contribution of these transmission pathways.ResultsData from 28 cross-sectional studies and 14 case-control studies were extracted. There was strong evidence for transmission by direct contact with dogs for both CE and AE. The estimated attributable fractions were 26.1% (CI 13.8%-39.6%) and 34.4% (CI 20.7% -48.2%) respectively. Transmission through contaminated water was estimated to be responsible for approximately 29.4% (CI 12.1%-51.7%) for CE and 24.8% (CI 10.6% to 42.6%) for AE. Contaminated food may be responsible for approximately 23.4% of CE cases (CI 2.1%-47.3%). Globally, there was insufficient evidence to conclude AE can be transmitted by food, although case control studies from low human incidence areas suggested that possibly 32.5% (CI 10.0%-53.2%) could be transmitted by food. There was also insufficient evidence that direct contact with foxes was a significant source of human disease. There were no suitable studies with a risk of environmental contact reported, but the residual attributable fraction thatwould likely include this pathway was approximately 21.1% for CE and 11.1% for AE.ConclusionsThe results support the hypothesis that dog contact and drinking contaminated water are major pathways of transmission of both CE and AE. For contaminated food, the results are less consistent, but suggest that it is an important transmission pathway and provide better evidence than expert elicitations as previously used.

Abstract Background Animal fracture models, primarily performed in rats, are crucial to investigate normal and pathological bone healing. However, results of biomechanical testing representing a major outcome measure show high standard deviations often precluding statistical significance. Therefore, the aim of our study was a systematical examination of biomechanical characteristics of rat femurs during three-point bending. Furthermore, we tried to reduce variation of results by individually adapting the span of bearing and loading areas to the bone’s length. Methods We examined 40 paired femurs of male Wistar-rats by DXA (BMD and BMC of the whole femur) and pQCT-scans at the levels of bearing and loading areas of the subsequent biomechanical three-point bending test. Individual adjustment of bearing and loading bars was done respecting the length of each specimen. Subgroups of light ( 400 g, n = 18) animals were formed and analysed separately. We furthermore compared the results of the individualised bending-setting to 20 femurs tested with a fix span of 15 mm. Results Femurs showed a length range of 34 to 46 mm. The failure loads ranged from 116 to 251 N (mean 175.4 ± 45.2 N; heavy animals mean 221 ± 18.9 N; light animals mean 138.1 ± 16.4 N) and stiffness ranged from 185 N/mm to 426 N/mm (mean 315.6 ± 63 N/mm; heavy animals mean 358.1 ± 34.64 N/mm; light animals mean 280.8 ± 59.85 N/mm). The correlation of densitometric techniques and failure loads was high (DXA R2 = 0.89 and pQCT R2 = 0.88). In comparison to femurs tested with a fix span, individual adaptation of biomechanical testing homogenized our data significantly. Most notably, the standard deviation of failure loads (221 ± 18.95 N individualized setting vs. 205.5 ± 30.36 N fixed) and stiffness (358.1 ± 34.64 N/mm individualized setting vs. 498.5 ± 104.8 N/mm fixed) was reduced by at least one third. Conclusions Total variation observed in any trait reflects biological and methodological variation. Precision of the method hence affects the statistical power of the study. By simply adapting the setting of the biomechanical testing, interindividual variation could be reduced, which improves the precision of the method significantly.

Abstract Background Capsular contracture is one of the most common complications in surgical interventions for aesthetic breast augmentation or post-mastectomy breast reconstruction involving the use of silicone prostheses. Although the precise cause of capsular contracture is yet unknown, the leading hypothesis is that it is caused by long-term unresolved foreign body reaction towards the silicone breast implant. To authors’ best knowledge, this is the first study that elucidates the presence of lysyl oxidase (LOX)—an enzyme that is involved in collagen and elastin crosslinking within fibrous capsules harvested from patients with severe capsular contracture. It was hypothesized that over-expression of LOX plays a role in the irreversible crosslinking of collagen and elastin which, in turn, stabilizes the fibrous proteins and contributes to the progression of capsular contracture. Methods Eight fibrous capsules were collected from patients undergoing capsulectomy procedure, biomechanical testing was performed for compressive Young’s moduli and evaluated for Type I and II collagen, elastin and LOX by means of non-linear optical microscopy and immunohistology techniques. Results Observations revealed the heterogeneity of tissue structure within and among the collected fibrous capsules. Regardless of the tissue structure, it has been shown that LOX expression was intensified at the implant-to-tissue interface. Conclusion Our results indicate the involvement of LOX in the initiation of fibrous capsule formation which ultimately contributes towards the progression of capsular contracture.

Abstract Objectives Rat fracture models are extensively used to characterize normal and pathological bone healing. Despite, systematic research on inter- and intra-individual differences of common rat bones examined is surprisingly not available. Thus, we studied the biomechanical behaviour and radiological characteristics of the humerus, the tibia and the femur of the male Wistar rat—all of which are potentially available in the experimental situation—to identify useful or detrimental biomechanical properties of each bone and to facilitate sample size calculations. Methods 40 paired femura, tibiae and humeri of male Wistar rats (10–38 weeks, weight between 240 and 720 g) were analysed by DXA, pQCT scan and three-point-bending. Bearing and loading bars of the biomechanical setup were adapted percentually to the bone’s length. Subgroups of light (skeletal immature) rats under 400 g (N = 11, 22 specimens of each bone) and heavy (mature) rats over 400 g (N = 9, 18 specimens of each bone) were formed and evaluated separately. Results Radiologically, neither significant differences between left and right bones, nor a specific side preference was evident. Mean side differences of the BMC were relatively small (1–3% measured by DXA and 2.5–5% by pQCT). Over all, bone mineral content (BMC) assessed by DXA and pQCT (TOT CNT, CORT CNT) showed high correlations between each other (BMC vs. TOT and CORT CNT: R 2 = 0.94–0.99). The load–displacement diagram showed a typical, reproducible curve for each type of bone. Tibiae were the longest bones (mean 41.8 ± 4.12 mm) followed by femurs (mean 38.9 ± 4.12 mm) and humeri (mean 29.88 ± 3.33 mm). Failure loads and stiffness ranged from 175.4 ± 45.23 N / 315.6 ± 63.00 N/mm for the femurs, 124.6 ± 41.13 N / 260.5 ± 59.97 N/mm for the humeri to 117.1 ± 33.94 N / 143.8 ± 36.99 N/mm for the tibiae. Smallest interindividual differences were observed in failure loads of the femurs (CV% 8.6) and tibiae (CV% 10.7) of heavy animals, light animals showed good consistency in failure loads of the humeri (CV% 7.7). Most consistent results of both sides (left vs. right) in failure loads were provided by the femurs of light animals (mean difference 4.0 ± 2.8%); concerning stiffness, humeri of heavy animals were most consistent (mean difference of 6.2 ± 5%). In general, the failure loads showed strong correlations to the BMC (R 2 = 0.85–0.88) whereas stiffness correlated only moderate, except for the humerus (BMC vs. stiffness: R 2 = 0.79). Discussion Altogether, the rat’s femur of mature specimens showed the most accurate and consistent radiological and biomechanical results. In synopsis with the common experimental use enabling comparison among different studies, this bone offers ideal biomechanical conditions for three point bending experiments. This can be explained by the combination of a superior aspect ratio and a round and long, straight morphology, which satisfies the beam criteria more than other bones tested.

ObjectiveAim of this study was, to demonstrate the feasibility of high-resolution grating-based X-ray phase-contrast computed tomography (PCCT) for quantitative assessment of cartilage.Materials and methodsIn an experimental setup, 12 osteochondral samples were harvested from n = 6 bovine knees (n = 2 each). From each knee, one cartilage sample was degraded using 2.5% Trypsin. In addition to PCCT and biomechanical cartilage stiffness measurements, 3T and 7T MRI was performed including MSME SE T2 and ME GE T2* mapping sequences for relaxationtime measurements. Paired t-tests and receiver operating characteristics (ROC) curves were used for statistical analyses.ResultsPCCT provided high-resolution images for improved morphological cartilage evaluation as compared to 3T and 7T MRI. Quantitative analyses revealed significant differences between the superficial and the deep cartilage layer for T2 mapping as well as for PCCT (P0.05). MRI and stiffness measurements showed significant differences between healthy and degraded osteochondral samples. Accuracy in the prediction of cartilage degradation was excellent for MRI and biomechanical analyses.ConclusionIn conclusion, high-resolution grating-based X-ray PCCT cartilage imaging is feasible. In addition to MRI and biomechanical analyses it provides complementary, water content independent, information for improved morphological and quantitative characterization of articular cartilage ultrastructure.

In this study we characterized the wear behavior of a tribological system with material combinations of the total intervertebral disc prosthesis InDisc (Institute of Medical and Polymer Engineering) – consisting of two laser sintered titanium alloy plates with an elastomer inbetween. For the tests a translatory oscillation test bench was used which simulates the kinematic and the physiological environment of the lumbar spine. In the interface between the elastomer (Silopren LSR 2670) and the laser sintered titanium alloy (Ti6Al4V) abrasion cords out of silicone rubber with a maximal area of 500 × 700 μm2 were generated as well as individual particles with a size of 10 × 20 to 40 × 50 μm2. In the second prosthesis interface of Ti6Al4V and bone, bone particles with an area of 0.5 × 0.5 to 1 × 1 μm2 were detected. The wear particles and the amount of wear volume influence the biological response of a total intervertebral disc prosthesis in the body.