Pooled multicolour tagging for visualizing subcellular protein dynamics.

Imaging-based methods are widely used for studying the subcellular localization of proteins in living cells. While routine for individual proteins, global monitoring of protein dynamics following perturbation typically relies on arrayed panels of fluorescently tagged cell lines, limiting throughput and scalability. Here, we describe a strategy that combines high-throughput microscopy, computer vision and machine learning to detect perturbation-induced changes in multicolour tagged visual proteomics cell (vpCell) pools. We use genome-wide and cancer-focused intron-targeting sgRNA libraries to generate vpCell pools and a large, arrayed collection of clones each expressing two different endogenously tagged fluorescent proteins. Individual clones can be identified in vpCell pools by image analysis using the localization patterns and expression level of the tagged proteins as visual barcodes, enabling simultaneous live-cell monitoring of large sets of proteins. To demonstrate broad applicability and scale, we test the effects of antiproliferative compounds on a pool with cancer-related proteins, on which we identify widespread protein localization changes and new inhibitors of the nuclear import/export machinery. The time-resolved characterization of changes in subcellular localization and abundance of proteins upon perturbation in a pooled format highlights the power of the vpCell approach for drug discovery and mechanism-of-action studies.

Impact of 3D surface scanning protocols on the Os coxae digital data: Implications for sex and age-at-death assessment.

The 3D imaging technologies have become of paramount importance for example in disciplines such as forensic anthropology and bioarchaeology, where they are being used more and more frequently. There are several new possibilities that they offer; for instance, the easier and faster sharing of data among institutions, the possibility of permanent documentation, or new opportunities of data analysis. An important requirement, however, is whether the data obtained from different scanning devices are comparable and whether the possible varying outputs could affect further analyses, such as the estimation of the biological profile. Therefore, we aimed to investigate two important questions: (1) whether 3D models acquired by two different scanning technologies (structured light and laser) are comparable and (2) whether the scanning equipment has an effect on the anthropological analyses, such as age-at-death estimation and sex assessment. 3D models of ossa coxa (n = 29) were acquired by laser (NextEngine) and structured light (HP 3D Structured Light Scanner PRO 2) scanners. The resulting 3D models from both scanners were subjected to age-at-death analyses (via the quantitative method of Stoyanova et al., 2017) and sex analyses (via Diagnose Sexuelle Probabiliste 2 of Bruzek et al., 2017). Furthermore, high quality scans of a small sample (n = 5) of pubic symphyseal surfaces with the RedLux Profiler device were acquired as reference surfaces to which the outputs from both scanners were compared. Small deviations between surfaces were more evident in more rugged surfaces (in areas of depression and protrusion). Even though small differences from the reference surfaces were found, they did not have a significant effect on the age and sex estimates. It never resulted in the opposite sex assignment, and no significant differences were observed between age estimates (with the exception of those with the TPS/BE model).

Magnification of digital hip radiographs differs between clinical workplaces.

Preoperative planning for total hip arthroplasty includes templating on anteroposterior radiographs. It is necessary to consider radiographic magnification in order to scale templates accurately. Studies dealing with hip templating report different values of radiographic magnification. It is not clear if the observed difference in magnification between the studies is caused by variability in studied groups, methodology or instrumentation. We hypothesize that there is a difference in magnification between clinical workplaces. Within this study, radiographic magnification was estimated on 337 radiographs of patients after total hip surgery from five orthopaedic departments in the Czech Republic. Magnification was determined for each patient as a ratio between diameter of implanted femoral head measured on radiogram and its true size. One-way ANOVA revealed significant differences in magnification between workplaces (F(4,332) = 132, p≤0.001). These results suggest that radiographic magnification depends on the workplace where it is taken or more precisely on radiographic device. It indicates potential limits in generalizability of results of studies dealing with preoperative planning accuracy to other institutions.

Design, analysis and verification of a knee joint oncological prosthesis finite element model.

BACKGROUND: The aim of this paper was to design a finite element model for a hinged PROSPON oncological knee endoprosthesis and to verify the model by comparison with ankle flexion angle using knee-bending experimental data obtained previously. METHOD: Visible Human Project CT scans were used to create a general lower extremity bones model and to compose a 3D CAD knee joint model to which muscles and ligaments were added. Into the assembly the designed finite element PROSPON prosthesis model was integrated and an analysis focused on the PEEK-OPTIMA hinge pin bushing stress state was carried out. To confirm the stress state analysis results, contact pressure was investigated. The analysis was performed in the knee-bending position within 15.4-69.4° hip joint flexion range. RESULTS: The results showed that the maximum stress achieved during the analysis (46.6 MPa) did not exceed the yield strength of the material (90 MPa); the condition of plastic stability was therefore met. The stress state analysis results were confirmed by the distribution of contact pressure during knee-bending. CONCLUSION: The applicability of our designed finite element model for the real implant behaviour prediction was proven on the basis of good correlation of the analytical and experimental ankle flexion angle data.

Knee joint endoprosthesis loading in full extension.

This paper deals with a knee joint endoprosthesis finite element analysis. Based on a three dimensional geometric model of a lower extremity, a mechanical axis of the limb was designed. This axis is important for several reasons. Firstly, the endoprosthesis was positioned due to its direction, secondly, boundary conditions was defined on its proximal and distal end and finally, the axis enabled reasonable simplification of the model which led to the time saving analysis while preserving principal features of the model like the natural boundary conditions or knee joint's degrees of freedom . Having this, one leg stance was simulated. Results of the analysis were encouraging for future models. Especially the choice of the mechanical axis was suitable and enabled a better distribution of contact pressures and stress on both femoral and tibial component compared to our former models. Also their magnitudes correspond better the manufacturer's experience and our findings. The stresses did not exceeded 30MPa for the UHMWPE tibial plateau and 100MPa for the femoral component. The contact pressures were lower than 40MPa.

MRI-based registration of pelvic alignment affected by altered pelvic floor muscle characteristics.

BACKGROUND: Pelvic floor muscles have potential to influence relative pelvic alignment. Side asymmetry in pelvic floor muscle tension is claimed to induce pelvic malalignment. However, its nature and amplitude are not clear. There is a need for non-invasive and reliable assessment method. An intervention experiment of unilateral pelvic floor muscle activation on healthy females was performed using image data for intra-subject comparison of normal and altered configuration of bony pelvis. METHODS: Sequent magnetic resonance imaging of 14 females in supine position was performed with 1.5 T static body coil in coronal orientation. The intervention, surface functional electrostimulation, was applied to activate pelvic floor muscles on the right side. Spatial coordinates of 23 pelvic landmarks were localized in each subject and registered by specially designed magnetic resonance image data processing tool (MPT2006), where individual error calculation; data registration, analysis and 3D visualization were interfaced. FINDINGS: The effect of intervention was large (Cohen's d=1.34). We found significant differences in quantity (P<0.01) and quality (P=0.02) of normal and induced pelvic displacements. After pelvic floor muscle activation on the right side, pelvic structures shifted most frequently to the right side in ventro-caudal direction. The right femoral head, the right innominate and the coccyx showed the largest displacements. INTERPRETATION: The consequences arising from the capacity of pelvic floor muscles to displace pelvic bony structures are important to consider not only in management of malalignment syndrome but also in treatment of incontinence. The study has demonstrated benefits associated with processing of magnetic resonance image data within pelvic region with high localization and registration reliability.

Shape characteristics of the foot arch: dynamics in the pregnancy period.

OBJECTIVES: The aim is data detection and finding some load consequences generated by various mechanical or physiological changes in the interaction of the end segment of the body--the foot--and the environment. DESIGN: Shape instability of the foot caused by e.g. loading of the foot by long-term frequency loads--walking, by extreme loads--sport, by hormonal changes--pregnancy, by aging, by pathologies, etc. The footprint surface was numerically described in 3D by means of stereo-photo-gram-metrical method--DMR digital relief model. Density of discrete points--250-400 per one print. SETTING: Detailed DMR was constructed by means of triangular web including contour picture with the use of Atlas program. The specified generated web is characterized by triangles with a cca 1 mm side in the number of up to 4,500 elements per one footprint model. RESULTS: The results enable us to deduce shape characteristics of DMR--the shape of the interactive boundary of the foot--the rest surface, to solve foot arch straining, to solve issues of discomfort and distribution of the pressure at the boundary of the foot--the rest surface, the shoe, etc. The gained findings can be interpreted in the field of prevention, therapy, orthopedics, podology, and enable us to come up with recommendations for the orthopedic practice and industrial use in the footwear production, etc. THE MAIN FINDINGS: The difference between volume reductions of the space under the foot arch characterizes the level of "fall" of the arch. This criterion is independent of the foot size, and is in 3D. CONCLUSIONS: Shape characteristics of footprints in pregnant women and in the period after childbirth were calculated on the basis of the defined criterion. The results of the group of four women tested in three periods suggest that there is no clear tendency towards the foot arch falling/increasing of the foot arch "fall" during the pregnancy period.