| 1. |
- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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| 2. |
- Klionsky, Daniel J., et al.
(författare)
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Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
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Ingår i: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
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Forskningsöversikt (refereegranskat)abstract
- In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
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| 3. |
- Han, Jinming
(författare)
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Repopulation of a microglia-depleted central nervous system : molecular characterization during homeostasis and disease
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Microglia are predominant tissue resident macrophages within the central nervous system (CNS), and contribute to both CNS development and homeostasis. During disease conditions microglia undergo transcriptional re-programming and their dysfunction is implicated in a multitude of disorders, such as multiple sclerosis (MS). How microglia could be therapeutically targeted is a current research focus. Recent experimental microglial depletion methods using conditional genetic targeting and pharmacological therapies have broadened our perspective of these multi-tasking microglia. Newly repopulated microglia following experimental microglial ablation hold great promise for reducing neuroinflammation and treating a variety of neurological disorders. In Study 1 our results indicated that microglia could be ablated (approximately 95%) by systemic use of tamoxifen in Cx3cr1CreER/+Rosa26DTA/+ mice. Microglial repopulation ensued through both the proliferation of surviving microglia in the CNS, and from the infiltration of Ly6Chi monocytes. Under this condition infiltrating monocytes could be shaped into microglia-like cells by the CNS microenvironment. Furthermore, isolated newly repopulated resident microglia and infiltrating microglia-like cells following experimental depletion exhibited differential functionality in vitro, such as phagocytic capacity and cytokine production. In Study 2 we used the microglial depletion and repopulation model mentioned above and demonstrated that the presence of infiltrating microglia-like cells following ablation could exacerbate experimental autoimmune encephalomyelitis (EAE) symptoms in Cx3cr1CreER/+Rosa26DTA/+ female mice. This was not evident in male mice, indicating a potential sex effect. Under this condition there was a higher expression of major histocompatibility complex class II and a greater secretion of proinflammatory cytokines during the acute period in the female mice. In Study 3 we discovered a novel subpopulation of microglia that escape the genetic modification of Cx3cr1 in Cx3cr1CreER-EYFP/+Rosa26DTA/+ mice. Following microglial depletion using tamoxifen, newly repopulated Cx3cr1highEYFP– microglia had an advantage over Cx3cr1CreER-EYFP/+ and Cx3cr1lowEYFP+ microglia. We also found that microglial repopulation was tightly regulated by the CX3CL1-CX3CR1 signaling. The numbers of repopulated CNS-resident microglia were significantly decreased, while the numbers of infiltrating microglia-like cells were increased during repopulation in mice devoid of Cx3cr1. In Study 4 we demonstrated that experimentally removing microglia using both Cx3cr1CreER/+Rosa26DTA/+ mice and PLX3397 treatment had crucial effects on circulating monocytes and splenic macrophages, a finding that had previously received little attention. We therefore proposed that clinical translation of preclinical studies using microglial depletion should take peripheral effects into consideration.
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| 4. |
- Hering, Alessa, et al.
(författare)
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Learn2Reg: comprehensive multi-task medical image registration challenge, dataset and evaluation in the era of deep learning
- 2023
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Ingår i: IEEE Transactions on Medical Imaging. - : Institute of Electrical and Electronics Engineers (IEEE). - 0278-0062 .- 1558-254X. ; 42:3, s. 697-712
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Tidskriftsartikel (refereegranskat)abstract
- Image registration is a fundamental medical image analysis task, and a wide variety of approaches have been proposed. However, only a few studies have comprehensively compared medical image registration approaches on a wide range of clinically relevant tasks. This limits the development of registration methods, the adoption of research advances into practice, and a fair benchmark across competing approaches. The Learn2Reg challenge addresses these limitations by providing a multi-task medical image registration data set for comprehensive characterisation of deformable registration algorithms. A continuous evaluation will be possible at https:// learn2reg.grand-challenge.org. Learn2Reg covers a wide range of anatomies (brain, abdomen, and thorax), modalities (ultrasound, CT, MR), availability of annotations, as well as intra- and inter-patient registration evaluation. We established an easily accessible framework for training and validation of 3D registration methods, which enabled the compilation of results of over 65 individual method submissions from more than 20 unique teams. We used a complementary set of metrics, including robustness, accuracy, plausibility, and runtime, enabling unique insight into the current state-of-the-art of medical image registration. This paper describes datasets, tasks, evaluation methods and results of the challenge, as well as results of further analysis of transferability to new datasets, the importance of label supervision, and resulting bias. While no single approach worked best across all tasks, many methodological aspects could be identified that push the performance of medical image registration to new state-of-the-art performance. Furthermore, we demystified the common belief that conventional registration methods have to be much slower than deep-learning-based methods.
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| 5. |
- Hu, Cheng, et al.
(författare)
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On the steady-state workpiece flow mechanism and force prediction considering piled-up effect and dead metal zone formation
- 2021
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Ingår i: Journal of Manufacturing Science and Engineering. - : ASME International. - 1087-1357 .- 1528-8935. ; 143:4
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Tidskriftsartikel (refereegranskat)abstract
- The manufacturing of miniaturized components is indispensable in modern industries, where the uncut chip thickness (UCT) inevitably falls into a comparable magnitude with the tool edge radius. Under such circumstances, the ploughing phenomenon between workpiece and tool becomes predominant, followed by the notable formation of dead metal zone (DMZ) and piled-up chip. Although extensive models have been developed, the critical material flow status in such microscale is still confusing and controversial. In this study, a novel material separation model is proposed for the demonstration of workpiece flow mechanism around the tool edge radius. First, four critical positions of workpiece material separation are determined, including three points characterizing the DMZ pattern and one inside considered as stagnation point. The normal and shear stresses as well as friction factors along the entire contact region are clarified based on slip-line theory. It is found that the friction coefficient varies symmetrically about the stagnation point inside DMZ and remains constant for the rest. Then, an analytical force prediction model is developed with Johnson-Cook constitutive model, involving calibrated functions of chip-tool contact length and cutting temperature. The assumed tribology condition and morphologies of material separation including DMZ are clearly observed and verified through various finite element (FE) simulations. Finally, comparisons of cutting forces from cutting experiments and predicted results are adopted for the validation of the predictive model.
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| 6. |
- Lund, Harald, et al.
(författare)
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CD163+ macrophages monitor enhanced permeability at the blood-dorsal root ganglion barrier
- 2024
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Ingår i: Journal of Experimental Medicine. - : Rockefeller University Press. - 0022-1007 .- 1540-9538. ; 221:2
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Tidskriftsartikel (refereegranskat)abstract
- In dorsal root ganglia (DRG), macrophages reside close to sensory neurons and have largely been explored in the context of pain, nerve injury, and repair. However, we discovered that most DRG macrophages interact with and monitor the vasculature by sampling macromolecules from the blood. Characterization of the DRG vasculature revealed a specialized endothelial bed that transformed in molecular, structural, and permeability properties along the arteriovenous axis and was covered by macrophage-interacting pericytes and fibroblasts. Macrophage phagocytosis spatially aligned with peak endothelial permeability, a process regulated by enhanced caveolar transcytosis in endothelial cells. Profiling the DRG immune landscape revealed two subsets of perivascular macrophages with distinct transcriptome, turnover, and function. CD163(+) macrophages self-maintained locally, specifically participated in vasculature monitoring, displayed distinct responses during peripheral inflammation, and were conserved in mouse and man. Our work provides a molecular explanation for the permeability of the blood-DRG barrier and identifies an unappreciated role of macrophages as integral components of the DRG-neurovascular unit.
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| 7. |
- Lund, Harald, et al.
(författare)
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Competitive repopulation of an empty microglial niche yields functionally distinct subsets of microglia-like cells
- 2018
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Ingår i: Nature Communications. - : Nature Publishing Group. - 2041-1723. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Circulating monocytes can compete for virtually any tissue macrophage niche and become long-lived replacements that are phenotypically indistinguishable from their embryonic counterparts. As the factors regulating this process are incompletely understood, we studied niche competition in the brain by depleting microglia with >95% efficiency using Cx3cr1CreER/+R26DTA/+ mice and monitored long-term repopulation. Here we show that the microglial niche is repopulated within weeks by a combination of local proliferation of CX3CR1+F4/80lowClec12a– microglia and infiltration of CX3CR1+F4/80hiClec12a+ macrophages that arise directly from Ly6Chi monocytes. This colonization is independent of blood brain barrier breakdown, paralleled by vascular activation, and regulated by type I interferon. Ly6Chi monocytes upregulate microglia gene expression and adopt microglia DNA methylation signatures, but retain a distinct gene signature from proliferating microglia, displaying altered surface marker expression, phagocytic capacity and cytokine production. Our results demonstrate that monocytes are imprinted by the CNS microenvironment but remain transcriptionally, epigenetically and functionally distinct.
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| 8. |
- Weng, Jian, et al.
(författare)
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A hybrid model for force prediction in orthogonal cutting with chamfered tools considering size and edge effect
- 2020
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Ingår i: International Journal of Advanced Manufacturing Technology. - : Springer Science and Business Media LLC. - 0268-3768 .- 1433-3015. ; 110:5-6, s. 1367-1384
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Tidskriftsartikel (refereegranskat)abstract
- Researches on the modeling of machining difficult-to-cut metals are important for optimization of the processing parameters, in which the force modeling is essential due to its significant influence on the performance of tools and the quality of parts. A semi-analytical method for force prediction in orthogonal cutting with chamfered tools considering both edge and size effect is proposed in this paper. The plastic deformation in the shear band was investigated using a parallel shear zone model and unequal division shear zone model. The influence of size effect on cutting force was discussed and a simplified expression of improvement factor is introduced to describe the sharp increase of shear stress under the condition of low feed rate. Simulations of orthogonal cutting with different chamfer lengths are conducted to analyze the variation of cutting force with respect to chamfer length, which reveals that the influence of chamfer length on cutting force is determined by the ratio of chamfer length to uncut chip thickness. A modified function considering the trend of material flow condition is proposed, which treats the total cutting force as a combination of cutting forces caused by chamfered edge and rake face. The calibration of constants in the proposed method is achieved using particle swarm optimization (PSO), a meta-heuristic algorithm for complicated non-linear models. The experiments show that the method works well on both fitting and predicting modules in orthogonal cutting of AISI 304 using cemented carbide tools with 15° chamfer angle or 25° chamfer angle.
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| 9. |
- Weng, Jian, et al.
(författare)
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A machine learning based approach for determining the stress-strain relation of grey cast iron from nanoindentation
- 2020
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Ingår i: Mechanics of Materials. - : Elsevier BV. - 0167-6636. ; 148
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Tidskriftsartikel (refereegranskat)abstract
- Apart from microhardness and elastic modulus, the stress-strain relation is another important characteristic that more and more scholars have been trying to extract from nanoindentation. With the development of artificial intelligence and computer technology, a machine learning based method is proposed in this paper to extract stress-strain curve of grey cast iron using sharp nanoindentation. Firstly, the average curve is achieved by the grid-design nanoindentation to avoid the influence of different phases on indentation results. The plastic behavior is considered as a power law function in this paper. Then, finite element method supports to generate a simulation data set, with full-factor and full-level design of constants of stress-strain relation. With the simulation data set, the support vector regression machine establishes a surrogate model to correlate the input (constants of stress-strain function) and output (the mean error between predicted and measured results). The best parameters of support vector machine are determined through grid search and cross-validation. PSO serves as the optimization algorithm to find the optimum of input related to the measured results, with an inertia factor to improve the local search ability. Finally, the simulation loading curve with the optimal constants provided by PSO perfectly fits the measured loading curve, which shows the effectiveness of the inverse method proposed in this paper.
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| 10. |
- Zhang, Kaicheng, et al.
(författare)
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SN 2014J in M82 : new insights on the spectral diversity of Type Ia supernovae
- 2018
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Ingår i: Monthly notices of the Royal Astronomical Society. - : Oxford University Press (OUP). - 0035-8711 .- 1365-2966. ; 481:1, s. 878-893
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Tidskriftsartikel (refereegranskat)abstract
- We present extensive spectroscopic observations for one of the closest Type Ia supernovae (SNe Ia), SN 2014J discovered in M82, ranging from 10.4 d before to 473.2 d after B-band maximum light. The diffuse interstellar band features detected in a high-resolution spectrum allow an estimate of line-of-sight extinction as A(v) similar to 1.9 +/- 0.6 mag. Spectroscopically, SN 2014J can be put into the high-velocity (HV) subgroup in Wang's classification with a velocity of Si II lambda 6355 at maximum light of upsilon(0) = 1.22 +/- 0.01 x 10(4) km s(-1) but has a low velocity gradient (LVG, following Benetti's classification) of (v) over bar = 41 +/- 2 km s(-1) d(-1), which is inconsistent with the trend that HV SNe Ia generally have larger velocity gradients. We find that the HV SNe Ia with LVGs tend to have relatively stronger Si III (at similar to 4400 angstrom) absorptions in early spectra, larger ratios of S II lambda 5468 to S II lambda 5640, and weaker Si II 5972 absorptions compared to their counterparts with similar velocities but high velocity gradients. This shows that the HV+ LVG subgroup of SNe Ia may have intrinsically higher photospheric temperature, which indicates that their progenitors may experience more complete burning in the explosions relative to the typical HV SNe Ia.
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