Image Analysis Protocol Research Articles

Caspase 3 Expression Profiles in Meningioma Subtypes Based on Tissue Microarray Analysis.

Concerning primary central nervous system neoplasms, meningiomas demonstrate the most common type in adults worldwide. Deregulation of apoptotic pathways in malignancies, including meningiomas, is correlated with chemoresistance and poor prognosis. Caspases represent crucial proteins that induce cell apoptosis. This study aimed to correlate caspase 3 protein expression levels to meningioma clinic-pathological features. A set of fifty (n=50) meningioma lesions was included in the current analysis including a broad spectrum of histopathological subtypes (meningotheliomatous, psammomatus, transitional, fibrous, angiomatous, microcystic, atypical and anaplastic). Immunohistochemistry was implemented on tissue microarray cores of selected paraffin blocks by applying an anti-caspase 3 antibody. Additionally, an image analysis protocol was also performed in the corresponding immunostained slides. Caspase 3 protein over-expression was detected in 17/50 (34%) cases, whereas the remaining 33 cases (66%) were characterized by medium to low levels of the molecule. Caspase 3 expression was statistically significantly associated with the grade of the analyzed tumors and the mitotic index (p=0.002, p=0.001, respectively). Caspase 3 expression status was also correlated with the histotype of the selected meningiomas (p=0.016). Caspase 3 demonstrated low expression levels in a significant subset of the examined meningiomas correlated with differentiation grade, mitotic activity, and partially with specific histotypes. Agents that could enhance caspase 3 expression - inducing its apoptotic activity - represent a very promising area in oncology for developing novel treatment regimens.

Doppler Sonography Evaluation of Median Nerve Intraneural Blood Flow: A Systematic Review

Objective: This systematic review aims to determine the extent, scope, and nature of research using the sonographic measurement of intraneural blood flow within the median nerve and to identify, characterize, and compare image acquisition and analysis protocols that have been reported as potential candidate techniques for standardizing research and clinical applications. Materials and Methods: This systematic review summarizes image acquisition protocols and analysis methodologies from 52 current research studies using Doppler ultrasound to examine median nerve intraneural blood flow. Results: Four types of Doppler technologies were identified: power Doppler, color Doppler, spectral Doppler, and Superb Microvascular Imaging, but there were inconsistencies in how images were acquired and analyzed. Intraneural blood flow measurements were categorized into four types based on a combination of measurement level (e.g., binary, ordinal, continuous) and type of flow indicator (e.g., pixel count, intensity, velocity). Conclusion: Standardized imaging protocols and reporting guidelines are needed to improve consistency. Future studies should evaluate the accuracy and reliability of different image acquisition methods and analysis measurements.

Multiscale chromatin dynamics and high entropy in plant iPSC ancestors.

Plant protoplasts provide starting material for of inducing pluripotent cell masses that are competent for tissue regeneration in vitro, analogous to animal induced pluripotent stem cells (iPSCs). Dedifferentiation is associated with large-scale chromatin reorganisation and massive transcriptome reprogramming, characterised by stochastic gene expression. How this cellular variability reflects on chromatin organisation in individual cells and what factors influence chromatin transitions during culturing are largely unknown. Here, we used high-throughput imaging and a custom supervised image analysis protocol extracting over 100 chromatin features of cultured protoplasts. The analysis revealed rapid, multiscale dynamics of chromatin patterns with a trajectory that strongly depended on nutrient availability. Decreased abundance in H1 (linker histones) is hallmark of chromatin transitions. We measured a high heterogeneity of chromatin patterns indicating intrinsic entropy as a hallmark of the initial cultures. We further measured an entropy decline over time, and an antagonistic influence by external and intrinsic factors, such as phytohormones and epigenetic modifiers, respectively. Collectively, our study benchmarks an approach to understand the variability and evolution of chromatin patterns underlying plant cell reprogramming in vitro.

Optimizing SUV Analysis: A Multicenter Study on Preclinical FDG-PET/CT Highlights the Impact of Standardization

PurposePreclinical imaging, with translational potential, lacks a standardized method for defining volumes of interest (VOIs), impacting data reproducibility. The aim of this study was to determine the interobserver variability of VOI sizes and standard uptake values (SUVmean and SUVmax) of different organs using the same [18F]FDG-PET and PET/CT datasets analyzed by multiple observers. In addition, the effect of a standardized analysis approach was evaluated.ProceduresIn total, 12 observers (4 beginners and 8 experts) analyzed identical preclinical [18F]FDG-PET-only and PET/CT datasets according to their local default image analysis protocols for multiple organs. Furthermore, a standardized protocol was defined, including detailed information on the respective VOI size and position for multiple organs, and all observers reanalyzed the PET/CT datasets following this protocol.ResultsWithout standardization, significant differences in the SUVmean and SUVmax were found among the observers. Coregistering CT images with PET images improved the comparability to a limited extent. The introduction of a standardized protocol that details the VOI size and position for multiple organs reduced interobserver variability and enhanced comparability.ConclusionsThe protocol offered clear guidelines and was particularly beneficial for beginners, resulting in improved comparability of SUVmean and SUVmax values for various organs. The study suggested that incorporating an additional VOI template could further enhance the comparability of the findings in preclinical imaging analyses.

Microstructural Hierarchy Descriptor Enabling Interpretative AI for Microelectronic Failure Analysis

Abstract This article proposes the MicroStructural Hierarchy Descriptor (µSHD) as a systematic and quantitative approach to spectra and image data in microelectronics failure analysis. It discusses concrete routes for employing µSHD directly as the quantitative descriptor for supervised and unsupervised machine learning. The authors propose that µSHD tools can be used to automate and improve characterization techniques and image processing and analysis protocols.

Quantitative analysis of trabecular bone tissue cryosections via a fully automated neural network-based approach.

Cryosectioning is known as a common and well-established histological method, due to its easy accessibility, speed, and cost efficiency. However, the creation of bone cryosections is especially difficult. In this study, a cryosectioning protocol for trabecular bone that offers a relatively cheap and undemanding alternative to paraffin or resin embedded sectioning was developed. Sections are stainable with common histological dying methods while maintaining sufficient quality to answer a variety of scientific questions. Furthermore, this study introduces an automated protocol for analysing such sections, enabling users to rapidly access a wide range of different stainings. Therefore, an automated 'QuPath' neural network-based image analysis protocol for histochemical analysis of trabecular bone samples was established, and compared to other automated approaches as well as manual analysis regarding scattering, quality, and reliability. This highly automated protocol can handle enormous amounts of image data with no significant differences in its results when compared with a manual method. Even though this method was applied specifically for bone tissue, it works for a wide variety of different tissues and scientific questions.

Studying Microtubule Dynamics in Human Neurons: Two-Dimensional Microtubule Tracing and Kymographs in iPSC- and SH-SY5Y-Derived Neurons for Tau Research.

The study of microtubule (MT) dynamics is essential for the understanding of cellular transport, cell polarity, axon formation, and other neurodevelopmental mechanisms. All these processes rely on the constant transition between assembly and disassembly of tubulin polymers to/from MTs, known as dynamic instability. This process is well-regulated, among others, by phosphorylation of microtubule-associated proteins (MAP), including the Tau protein. Protein kinases, in particular the microtubule affinity regulating kinase (MARK), regulate the MT-Tau interaction, inducingTau dissociation by phosphorylation. Phosphorylated Tau dissociates from microtubules forming insoluble aggregates known as neurofibrillary tangles. These accumulations of hyperphosphorylated Tau in the neurons disrupt the physiological MT-based transport machinery within the cell and can potentially lead to the development of neurodegenerative disorders, such as Alzheimer's disease (AD) and related tauopathies. Further investigations on the MT cytoskeleton dynamics are essential as they may elucidate pathomechanisms of neurodegenerative diseases - particularly tauopathies - as well as fundamental neurodevelopmental processes.The study of thedynamic assembly and disassembly of the MT network requires live-cell imaging rather than conventional immunocytochemistry based on fixed samples. To investigate MT dynamics, we perform live-cell imaging of neurons transfected with a fluorescently tagged version of the microtubule plus-end tracking protein (+TIP) EB3. This protein associates with the growing ends of MTs and thus visualizes MT growth in real time. Our imaging analysis protocol allows the determination of quantity, orientation, and velocity of MT growth in the soma and neurites of transfected neurons,using ImageJ-based tracking software and kymographs. Furthermore, functional effects of Tau and MARK kinases on the MT cytoskeleton can be assessed by overexpression or downregulation experiments of the respective protein prior to the live imaging assay. We use two different human neuronal cell models, naive and differentiated SH-SY5Y neuroblastoma cells, and neurons derived from induced pluripotent stem cells (iPSCs), both of which have shown success as models to study Tau-related pathologies.This protocol describes an optimized method for analysis of microtubule dynamics using fluorescent tagged EB3 protein as microtubule plus end marker. In this chapter, we outline the process of neuronal transfection, live-cell imaging, and necessary time-lapse image analysis based on ImageJ in two human-derived neuronal systems, which are suitable for the analysis of Tau trafficking and sorting studies.

Rationale and design of the RAPID-WATER-FLOW trial: Radiolabeled perfusion to identify coronary artery disease using water to evaluate responses of myocardial FLOW

ObjectivesThe objective of this study was to determine the diagnostic performance of 15O-water positron emission tomography (PET) myocardial perfusion imaging to detect coronary artery disease (CAD) using the truth-standard of invasive coronary angiography (ICA) with fractional flow reserve (FFR) or instantaneous wave-Free Ratio (iFR) or coronary computed tomography angiogram (CCTA). Background15O-water has a very high first-pass extraction that allows accurate quantification of myocardial blood flow and detection of flow-limiting CAD. However, the need for an on-site cyclotron and lack of automated production at the point of care and relatively complex image analysis protocol has limited its clinical use to date. MethodsThe RAPID WATER FLOW study is an open-label, multicenter, prospective investigation of the accuracy of 15O-water PET to detect obstructive angiographic and physiologically significant stenosis in patients with suspected CAD. The study will include the use of an automated system for producing, dosing, and injecting 15O-water and enrolling approximately 215 individuals with suspected CAD at approximately 10 study sites in North America and Europe. The primary endpoint of the study is the diagnostic sensitivity and specificity of the 15O-water PET study using the truth-standard of ICA with FFR or iFR to determine flow-limiting stenosis, or CCTA to rule out CAD and incorporating a quantitative analytic platform developed for the 15O-water PET acquisitions. Sensitivity and specificity are to be considered positive if the lower bound of the 95% confidence interval is superior to the threshold of 60% for both, consistent with prior registration studies.Subgroup analyses include assessments of diagnostic sensitivity, specificity, and accuracy in female, obese, and diabetic individuals, as well as in those with multivessel disease. All enrolled individuals will be followed for adverse and serious adverse events for up to 32 hours after the index PET scan.The study will have >90% power (one-sided test, α = 0.025) to test the hypothesis that sensitivity and specificity of 15O-water PET are both >60%. ConclusionsThe RAPID WATER FLOW study is a prospective, multicenter study to determine the diagnostic sensitivity and specificity of 15O-water PET as compared to ICA with FFR/iFR or CCTA. This study will introduce several novel aspects to imaging registration studies, including a more relevant truth standard incorporating invasive physiologic indexes, coronary CTA to qualify normal individuals for eligibility, and a more quantitative approach to image analysis than has been done in prior pivotal studies. Clinical trial registration informationClinical-Trials.gov (#NCT05134012).

Novel and pragmatic exploration of variation in glottic parameters in non-parallel versus parallel vocal cord CT planes with potential reporting pitfalls.

Oblique orientation of vocal cord demands strict compliance, by technicians and clinicians, to the recommended parallel plane CT scan of larynx. Repercussions of non-compliance has never been investigated before. We aimed to observe influence of non-parallel vocal cord plane CT scan on qualitative and quantitative glottic parameters, keeping parallel plane CT as a standard for comparison. Simultaneous identification of potential suboptimal imaging sequelae as a result of unformatted CT plane was also identified. In this study we included 95 normal adult glottides and retrospectively analyzed their anatomy in two axial planes, non-parallel plane ① and parallel to vocal cord plane ②. Qualitative (shape, structures at glottic level) and quantitative (anterior commissure ACom, vocal cord width VCw, anteroposterior AP, transverse Tr, cross-sectional area CSA) glottic variables were recorded. Multivariate statistical analysis was used to predict pattern and their impact on glottic anatomy. Plane ① displayed supraglottic features in glottis; adipose (90.5%) and split thyroid laminae (70.6%). Other categorical variables: atypical shape, submental structures and multilevel vertebral crossing were also in majority. All glottic dimensions varied significantly between two planes with most in ACom (-5.8mm) and CSA (-15.0 mm2). In contrast, plane ② manifested higher VCw (>73%), Tr (66.3%), CSA (64.2%) and AP (44.2%) measurements. On correlation analysis, variation in ACom, CSA, Tr was positively associated with VC or plane obliquity (p<0.05). This variability was more in obese and short necked subjects. Change in one parameter also modified other significantly i.e., ACom versus AP and CSA versus Tr. Results indicated statistically significant change in subjective and objective anatomical parameters of glottis on non-application of appropriate CT larynx protocol for image analysis hence highlighting importance of image reformation.

Pearls and Pitfalls of Adaptive Optics Ophthalmoscopy in Inherited Retinal Diseases.

Adaptive optics (AO) retinal imaging enables individual photoreceptors to be visualized in the clinical setting. AO imaging can be a powerful clinical tool for detecting photoreceptor degeneration at a cellular level that might be overlooked through conventional structural assessments, such as spectral-domain optical coherence tomography (SD-OCT). Therefore, AO imaging has gained significant interest in the study of photoreceptor degeneration, one of the most common causes of inherited blindness. Growing evidence supports that AO imaging may be useful for diagnosing early-stage retinal dystrophy before it becomes apparent on fundus examination or conventional retinal imaging. In addition, serial AO imaging may detect structural disease progression in early-stage disease over a shorter period compared to SD-OCT. Although AO imaging is gaining popularity as a structural endpoint in clinical trials, the results should be interpreted with caution due to several pitfalls, including the lack of standardized imaging and image analysis protocols, frequent ocular comorbidities that affect image quality, and significant interindividual variation of normal values. Herein, we summarize the current state-of-the-art AO imaging and review its potential applications, limitations, and pitfalls in patients with inherited retinal diseases.

Clodronate Liposome-Mediated Phagocytic Hemocyte Depletion Affects the Regeneration of the Cephalic Tentacle of the Invasive Snail, Pomacea canaliculata.

After amputation, granular hemocytes infiltrate the blastema of regenerating cephalic tentacles of the freshwater snail Pomacea canaliculata. Here, the circulating phagocytic hemocytes were chemically depleted by injecting the snails with clodronate liposomes, and the effects on the cephalic tentacle regeneration onset and on Pc-Hemocyanin, Pc-transglutaminase (Pc-TG) and Pc-Allograft Inflammatory Factor-1 (Pc-AIF-1) gene expressions were investigated. Flow cytometry analysis demonstrated that clodronate liposomes targeted large circulating hemocytes, resulting in a transient decrease in their number. Corresponding with the phagocyte depletion, tentacle regeneration onset was halted, and it resumed at the expected pace when clodronate liposome effects were no longer visible. In addition to the regeneration progress, the expressions of Pc-Hemocyanin, Pc-TG, and Pc-AIF-1, which are markers of hemocyte-mediated functions like oxygen transport and immunity, clotting, and inflammation, were modified. After the injection of clodronate liposomes, a specific computer-assisted image analysis protocol still evidenced the presence of granular hemocytes in the tentacle blastema. This is consistent with reports indicating the large and agranular hemocyte population as the most represented among the professional phagocytes of P. canaliculata and with the hypothesis that different hemocyte morphologies could exert diverse biological functions, as it has been observed in other invertebrates.

Machine learning for the generation of personalized image analysis protocol in echocardiography: a pilot study in arterial hypertension

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Agence National de la Recherche Europian Union-NextGenerationEU. Background Current workflow in cardiac imaging is based on protocols meant to capture a set of predefined images for a certain condition [1]. Although standardized, such approaches are not personalized, and are often time- and resource-ineffective. Reinforcement learning (RL), a subfield of machine learning that addresses decision making [2], can be used to incorporate complex, whole cardiac cycle imaging data, while optimizing costs associated to data acquisition and processing – including economy, patient safety/comfort and examination time. Our aim is to demonstrate the viability of RL to create a personalized, resource-effective data analysis routine from echocardiography to separate remodelling in arterial hypertension from normal cardiac function. Methods An echo study was performed in 189 clinically managed hypertensive (HTN) patients, and 60 non-hypertensive healthy control subjects [3]. Speckle-tracking analysis of the left ventricle was performed and deformation curves extracted. Aortic and mitral flow pulsed-wave (PW) Doppler and septal basal tissue PW Doppler velocity profiles were obtained. These whole cardiac cycle deformation and velocity curves were used as input data to the RL algorithm. RL optimized the required image analysis sequence to maximize the probability of obtaining a correct diagnosis while reducing the number of analysed images. Results The RL-generated protocol identified three separate image analysis groups to determine cardiac remodelling (Figure 1). The proposed protocol starts with the analysis of the mitral PW, and based on its velocity pattern, assigns each individual to one of the three groups. In Group 1 (G1, n=158) analysing the mitral PW acquisition was sufficient to determine disease status. HTN patients were separated based on a pattern of an inversed E/A ratio and EA fusion reflecting signs of diastolic dysfunction. In G2 (n=56), mitral flow showed a slightly decreased E/A ratio and longer E deacceleration time, and adding aortic PW was required to determine remodelling: HTN had higher aortic peak velocity and shorter peak acceleration time due to higher systemic afterload. Finally, G3 (n=19) required global longitudinal strain, which showed pathological early systolic stretching, reflecting the change in the timing of cardiac events in arterial hypertension. Fifteen (6%) participants did not match any of the groups, requiring the acquisition of remaining sequences (TDI, regional strain) to secure a correct diagnosis. In average, the protocol analysed 1.44 images per patient, out of the four available. Conclusion Through a pilot study utilizing a limited set of imaging sequences, we demonstrate a template for automated generation of efficient, tailored image analysis protocols, that minimimize the number of analyzed images to determinine patient phenotypes. RL is a promising tool to reduce analysis time, potentially saving costs in preventive screening programs.

The Application of Ultrasound to Quantify Hyoid Motion During Drug-Induced Sleep Endoscopy.

The significance of hyoid dynamics in OSA pathophysiology remains unclear. Drug-induced sleep endoscopy (DISE) is often used for evaluating patients intolerant to positive airway pressure (PAP) therapy. We performed DISE with concurrent hyoid-focused ultrasonography to quantify hyoid dynamics during obstructive and non-obstructive breathing. A cross-sectional analysis from a prospective cohort of patients undergoing DISE with PAP titration (DISE-PAP) and hyoid-focused ultrasound was conducted. Hyoid ultrasound was performed during obstructive breathing, and non-obstructive breathing after PAP administration. Motion was quantified by generating displacement curves based on echo-tracking hyoid movement. The image analysis protocol for quantifying hyoid displacement was performed independently by two researchers, and reliability of measures was assessed. Univariate and multivariate regressions were performed for various clinical data and hyoid displacement during obstructive breathing. Twenty patients met inclusion criteria. On average, the cohort was male (75%), elderly (65.9 ± 10 years), overweight (29.3 ± 3.99 kg/m2 ), and with moderate-to-severe OSA (29.3 ± 12.5 events/h). Mean hyoid displacement during obstructive breathing was 5.81 mm (±3.48). In all patients, hyoid displacement decreased after PAP administration (-3.94 mm [95% CI: -5.10, -2.78]; p < 0.0001). Inter-rater reliability for measures of hyoid displacement was excellent. After multivariate regression, hyoid displacement at baseline was associated with higher AHI (β [95% CI] = 0.18 [0.03, 0.33], p = 0.020). During DISE, hyoid displacement is greater during obstructive breathing with significant variability amongst patients. Further, these ultrasonographic measurements had excellent intra- and inter-rater reliability. Additional, larger studies are needed to understand contributors to hyoid mobility. 4 Laryngoscope, 133:3221-3227, 2023.

Correcting bias in cardiac geometries derived from multimodal images using spatiotemporal mapping

Cardiovascular imaging studies provide a multitude of structural and functional data to better understand disease mechanisms. While pooling data across studies enables more powerful and broader applications, performing quantitative comparisons across datasets with varying acquisition or analysis methods is problematic due to inherent measurement biases specific to each protocol. We show how dynamic time warping and partial least squares regression can be applied to effectively map between left ventricular geometries derived from different imaging modalities and analysis protocols to account for such differences. To demonstrate this method, paired real-time 3D echocardiography (3DE) and cardiac magnetic resonance (CMR) sequences from 138 subjects were used to construct a mapping function between the two modalities to correct for biases in left ventricular clinical cardiac indices, as well as regional shape. Leave-one-out cross-validation revealed a significant reduction in mean bias, narrower limits of agreement, and higher intraclass correlation coefficients for all functional indices between CMR and 3DE geometries after spatiotemporal mapping. Meanwhile, average root mean squared errors between surface coordinates of 3DE and CMR geometries across the cardiac cycle decreased from 7 ± 1 to 4 ± 1 mm for the total study population. Our generalised method for mapping between time-varying cardiac geometries obtained using different acquisition and analysis protocols enables the pooling of data between modalities and the potential for smaller studies to leverage large population databases for quantitative comparisons.

Method development for the evaluation of emergency decontamination protocol effectiveness using an ultraviolet fluorescent aerosol.

After hazardous material incidents, it is important to perform emergency decontamination procedures to remove contamination from the body. As these emergency decontamination procedures are developed, it is important to understand the efficacy of a given protocol. This study discusses a method that was developed to evaluate the efficacy of decontamination procedures by using an ultraviolet fluorescent aerosol and an image analysis protocol. This method involves imaging a mannequin while both unclothed and clothed prior to exposure to the fluorescent aerosol. After exposure, it was imaged again, disrobed, and decontaminated following an unconscious patient wet decontamination method. This work describes in detail the materials and methods used to develop the final methodology. Two clothing types (black cotton and Tyvek) were used to simulate civilian and first responder casualties. Image analysis was used to measure the extent of contamination on the mannequin at each stage of the procedure. These measurements were then compared to determine decontamination efficacy for each step (disrobing, wet decontamination, and total removal). The exposure protocol was shown to provide repeatable deposition of aerosol onto the mannequin. Decontamination was also shown to be repeatable, with no trends toward efficacy changing over time.

Large-scale analysis of structural brain asymmetries in schizophrenia via the ENIGMA consortium

Left-right asymmetry is an important organizing feature of the healthy brain that may be altered in schizophrenia, but most studies have used relatively small samples and heterogeneous approaches, resulting in equivocal findings. We carried out the largest case-control study of structural brain asymmetries in schizophrenia, with MRI data from 5,080 affected individuals and 6,015 controls across 46 datasets, using a single image analysis protocol. Asymmetry indexes were calculated for global and regional cortical thickness, surface area, and subcortical volume measures. Differences of asymmetry were calculated between affected individuals and controls per dataset, and effect sizes were meta-analyzed across datasets. Small average case-control differences were observed for thickness asymmetries of the rostral anterior cingulate and the middle temporal gyrus, both driven by thinner left-hemispheric cortices in schizophrenia. Analyses of these asymmetries with respect to the use of antipsychotic medication and other clinical variables did not show any significant associations. Assessment of age- and sex-specific effects revealed a stronger average leftward asymmetry of pallidum volume between older cases and controls. Case-control differences in a multivariate context were assessed in a subset of the data (N = 2,029), which revealed that 7% of the variance across all structural asymmetries was explained by case-control status. Subtle case-control differences of brain macrostructural asymmetry may reflect differences at the molecular, cytoarchitectonic, or circuit levels that have functional relevance for the disorder. Reduced left middle temporal cortical thickness is consistent with altered left-hemisphere language network organization in schizophrenia.

Effects of fitting methods, high b-values and image quality on diffusion and perfusion quantification and reproducibility in the calf

PurposesThe study aimed to optimize diffusion-weighted imaging (DWI) image acquisition and analysis protocols in calf muscles by investigating the effects of different model-fitting methods, image quality, and use of high b-value and constraints on parameters of interest (POIs). The optimized modeling methods were used to select the optimal combinations of b-values, which will allow shorter acquisition time while achieving the same reliability as that obtained using 16 b-values. MethodsTest-retest baseline and high-quality DWI images of ten healthy volunteers were acquired on a 3T MR scanner, using 16 b-values, including a high b-value of 1200 s/mm2, and structural T1-weighted images for calf muscle delineation. Three and six different fitting methods were used to derive ADC from monoexponential (ME) model and Dd, fp, and Dp from intravoxel incoherent motion (IVIM) model, with or without the high b-value. The optimized ME and IVIM models were then used to determine the optimal combinations of b-values, obtainable with the least number of b-values, using the selection criteria of coefficient of variance (CV) ≤10% for all POIs. ResultsThe find minimum multivariate algorithm was more flexible and yielded smaller fitting errors. The 2-steps fitting method, with fixed Dd, performed the best for IVIM model. The inclusion of high b-value reduced outliers, while constraints improved 2-steps fitting only. ConclusionsThe optimal numbers of b-values for ME and IVIM models were nine and six b-values respectively. Test-retest reliability analyses showed that only ADC and Dd were reliable for calf diffusion evaluation, with CVs of 7.22% and 4.09%.

Microscopy-based phenotypic monitoring of MDA-MB-231 spheroids allows the evaluation of phenotype-directed therapy

Breast cancer (BC) is the most commonly diagnosed cancer among women. Prognosis has improved over the years, to a large extent, owing to personalized therapy informed by molecular profiling of hormone receptors. However, there is a need for new therapeutic approaches for a subgroup of BCs lacking molecular markers, the Triple Negative Breast Cancer (TNBC) subgroup. TNBC is the most aggressive type of BC, lacks an effective standard of care, shows high levels of resistance and relapse is often inevitable. High resistance to therapy has been hypothesized to be associated with high intratumoral phenotypic heterogeneity. To characterize and treat this phenotypic heterogeneity, we optimized a whole-mount staining and image analysis protocol for three-dimensions (3D) spheroids.Applying this protocol to TNBC spheroids located in the outer region of the spheroid the cells with selected phenotypes: dividing, migrating, and high mitochondrial mass phenotypes. To evaluate the relevance of phenotype-based targeting these cell populations were targeted with Paclitaxel, Trametinib, and Everolimus, respectively, in a dose-dependent manner. Single agents cannot specifically target all phenotypes at the same time. Therefore, we combined drugs that should target independent phenotype. With this rationale we observed that combining Trametinib and Everolimus achieves the highest cytotoxicity at lower doses from all the tested combinations. These findings suggest a rational approach to design treatments can be evaluated in spheroids prior to pre-clinical models and potentially reduce adverse effects.

Impact of CD34-dependent Micro Vessel Density on Periapical Odontogenic Cysts.

Odontogenic cysts belong to a type of lesions with endodontic origin that in some cases mimic even aggressive odontogenic tumors sharing with them similar radiographic features. Periapical cysts (PCs) belong to the inflammatory odontogenic cysts sub-category and rarely squamous cell carcinoma arises from their hyperplastic/ dysplastic epithelia. This study aimed to explore the impact of cluster differentiation 34 (CD34) protein expression combined with micro vessel density (MVD) on PCs. Forty-eight (n=48) archival, formalin-fixed, and paraffin-embedded PC tissue specimens were included in the study. Immunohistochemistry (IHC) was performed in the corresponding tissue sections using an anti- CD34 antibody. CD34 expression levels and also MVD in the examined cases were measured by implementing a digital image analysis protocol. CD34 over-expression (moderate to high staining intensity levels) were detected in 29/48 (60.4%) cases, whereas the rest of them (19/48-39.6%) were characterized by low levels of expression. Extended MVD was identified in 26/48 (50.1%) cases correlated with CD34 over-expression, epithelial hyperplasia (p-value=0.001), and marginally with inflammatory infiltration level in the examined lesions (p-value=0.056). CD34 over-expression combined with increased MVD is associated with a neoplastic-like (hyperplastic) phenotype in PCs as a result of increased neo-angiogenic activity. These histopathological characteristics rarely form an eligible substrate for squamous cell carcinoma onset in untended cases.

Quantitative morphometric analysis in tibiofemoral joint osteoarthritis imaging: A literature review

ObjectiveTo highlight published quantitative morphometric analysis (QMA) methods for assessing tibiofemoral joint osteoarthritis in magnetic resonance imaging and computed tomography and underline the need for open and interoperable protocols for quantitative image analysis. DesignA literature search on PubMed/MEDLINE and Web of Science of keywords relating to QMA in magnetic resonance imaging and computed tomography in the context of tibiofemoral osteoarthritis in the past 23 years (2000–2022). The search was based on, but not limited to: “quantitative morphometric analysis” or “quantitative image analysis” in combination with “osteoarthritis”, “articular cartilage”, “subchondral bone”, “tibiofemoral joint”, “joint”, “CT”, and “MRI”. The search found 73 relevant publications that were manually screened and sorted for QMA methods. A further search to extract key functions of the underlying algorithms was performed. ResultMRI is generally used for QMA of articular cartilage and joint contact area, while CT is generally used for QMA of subchondral bone and joint space width. Studies have shown that QMA algorithms can be adapted to new tissues and modalities. However, many methods are not easily accessible, and there is fragmentation of computational tools and platforms in the research field. ConclusionQMA is an active research area, and many techniques from one modality can be readily extended to another. Adoption of open-source practices can allow algorithms developed for other imaging modalities to be shared, making it possible to bridge the knowledge gap for structures and pathological features for which QMA has not yet been investigated and to increase research output overall.