Optimizing clinical O2 saturation mapping using hyperspectral imaging and diffuse reflectance spectroscopy in the context of epinephrine injection.

Clinical determination of oxygen saturation (sO2) in patients is commonly performed via non-invasive optical techniques. However, reliance on a few wavelengths and some form of pre-determined calibration introduces limits to how these methods can be used. One example involves the assessment of sO2 after injection of local anesthetic using epinephrine, where some controversy exists around the time it takes for the epinephrine to have an effect. This is likely caused by a change in the tissue environment not accounted for by standard calibrated instruments and conventional analysis techniques. The present study aims to account for this changing environment by acquiring absorption spectra using hyperspectral imaging (HSI) and diffuse reflectance spectroscopy (DRS) before, during, and after the injection of local anesthesia containing epinephrine in human volunteers. We demonstrate the need to account for multiple absorbing species when applying linear spectral unmixing in order to obtain more clinically relevant sO2 values. In particular, we demonstrate how the inclusion of water absorption greatly affects the rate at which sO2 seemingly drops, which in turn sheds light on the current debate regarding the time required for local anesthesia with epinephrine to have an effect. In general, this work provides important insight into how spectral analysis methods need to be adapted to specific clinical scenarios to more accurately assess sO2.

Defined shapes of carotid artery calcifications on panoramic radiographs correlate with specific signs of cardiovascular disease on ultrasound examination.

OBJECTIVE: The aim was to optimize diagnostics for carotid artery calcifications (CACs) on panoramic radiographs (PRs) to identify cardiovascular disease (CVD) by investigating how 4 defined CAC shapes are associated with ultrasound (US) findings indicating CVD. STUDY DESIGN: The study included 414 participants (802 neck sides) from the Malmö Offspring Dental Study, examined with PRs. The PRs were assessed for CAC shapes stratified into 4 categories: single, scattered, vessel-width defining, and vessel-outlining. The carotid arteries were examined with US for signs of CVD: the presence of plaques, largest individual area of a plaque, number of plaques, and percentage reduction of the lumen. Associations between the different CAC categories and US characteristics were analyzed. RESULTS: All categories of CAC were significantly associated with a higher degree of US findings indicating CVD compared with no CAC (P < .001). The most significant differences were found for vessel-outlining CAC, with the mean of the largest individual plaque area of 17.9 vs 2.3 mm2, mean number of plaques 1.6 vs 0.2, and mean percentage reduction of the lumen 24.1% vs 3.5% (all P < .001). CONCLUSIONS: Independent of shape, CACs detected on PRs were associated with a higher degree of US findings of CVD. This was most pronounced for vessel-outlining CAC. With refined differential diagnostics of CACs in PRs, dentists may contribute to improved identification of patients in need of cardiovascular prevention.

Oxygen saturation mapping during reconstructive surgery of human forehead flaps with hyperspectral imaging and spectral unmixing.

BACKGROUND: Optical spectroscopy is commonly used clinically to monitor oxygen saturation in tissue. The most commonly employed technique is pulse oximetry, which provides a point measurement of the arterial oxygen saturation and is commonly used for monitoring systemic hemodynamics, e.g. during anesthesia. Hyperspectral imaging (HSI) is an emerging technology that enables spatially resolved mapping of oxygen saturation in tissue (sO2), but needs to be further developed before implemented in clinical practice. The aim of this study is to demonstrate the applicability of HSI for mapping the sO2 in reconstructive surgery and demonstrate how spectral analysis can be used to obtain clinically relevant sO2 values. METHODS: Spatial scanning HSI was performed on cutaneous forehead flaps, raised as part of a direct brow lift, in eight patients. Pixel-by-pixel spectral analysis, accounting for the absorption from multiple chromophores, was performed and compared to previous analysis techniques to assess sO2. RESULTS: Spectral unmixing using a broad spectral range, and accounting for the absorption of melanin, fat, collagen, and water, provided a more clinically relevant estimate of sO2 than conventional techniques, where typically only spectral features associated with absorption of oxygenated (HbO2) and deoxygenated (HbR) hemoglobin are considered. We demonstrate its clinical applicability by generating sO2 maps of partially excised forehead flaps showed a gradual decrease in sO2 along the length of the flap from 95 % at the flap base to 85 % at the flap tip. After being fully excised, sO2 in the entire flap decreased to 50 % within a few minutes. CONCLUSIONS: The results demonstrate the capability of sO2 mapping in reconstructive surgery in patients using HSI. Spectral unmixing, accounting for multiple chromophores, provides sO2 values that are in accordance with physiological expectations in patients with normal functioning microvascularization. Our results suggest that HSI methods that yield reliable spectra are to be preferred, so that the analysis can produce results that are of clinical relevance.

Association of high cardiovascular risk and diabetes with calcified carotid artery atheromas depicted on panoramic radiographs.

OBJECTIVE: To evaluate whether estimates of risk of future cardiovascular events and death and established or unknown diabetes are significantly associated with calcified carotid artery atheromas (CCAAs) on panoramic radiographs (PRs). The main focus was on men and women without previous myocardial infarction (MI). METHODS: The PAROKRANK (Periodontitis and its Relation to Coronary Artery Disease) study included patients with a first MI and matched control subjects. In this substudy, 738 patients (138 women) and 744 control subjects (144 women) with available PRs were assessed for CCAA. Cardiovascular risk estimates were determined according to the Framingham Risk Score (FRS) and Systematic COronary Risk Evaluation (SCORE). Established and previously unknown diabetes was also determined. RESULTS: CCAA was detected on PRs in 206 control subjects (28%) and 251 patients (34%). FRS was significantly associated with CCAA among control subjects (P = .04) and patients (P = .001). SCORE was associated with CCAA among control subjects (P < .01) but not patients (P = .07). Among men, FRS and SCORE were associated with CCAA in both control subjects and patients. Diabetes was not significantly associated with CCAA after adjustments. CONCLUSIONS: Elevated cardiovascular risk scores were associated with CCAA on PRs among control subjects. Diabetes was not independently associated with CCAA, possibly owing to selection bias.

Super-beacons: Open-source probes with spontaneous tuneable blinking compatible with live-cell super-resolution microscopy.

Localization-based super-resolution microscopy relies on the detection of individual molecules cycling between fluorescent and non-fluorescent states. These transitions are commonly regulated by high-intensity illumination, imposing constrains to imaging hardware and producing sample photodamage. Here, we propose single-molecule self-quenching as a mechanism to generate spontaneous photoswitching. To demonstrate this principle, we developed a new class of DNA-based open-source super-resolution probes named super-beacons, with photoswitching kinetics that can be tuned structurally, thermally and chemically. The potential of these probes for live-cell compatible super-resolution microscopy without high-illumination or toxic imaging buffers is revealed by imaging interferon inducible transmembrane proteins (IFITMs) at sub-100 nm resolutions.

Long-term skill improvement among general dental practitioners after a short training programme in diagnosing calcified carotid artery atheromas on panoramic radiographs.

PURPOSE: To study general dental practitioners (GDPs) ability to detect calcified carotid artery atheromas (CCAAs) in panoramic radiographs (PRs) and if their diagnostic accuracy in long term is improved after a short training programme. METHODS: Fourteen GDPs had their diagnostic accuracy regarding CCAA in PR assessed at baseline, 2 weeks and 1 year after training. Comparison was made with a reference standard based on consensus results from two experienced oral and maxillofacial radiologists. At each session, 100 radiographs were assessed individually by the GDPs. After the baseline assessment, the GDPs participated in a 2-hour training programme comprising a lecture and diagnostic training by calibration. The GDPs results before and after training were compared, as well as between follow-up sessions. RESULTS: A significant improvement in diagnostic accuracy was observed with increased sensitivity (from 41.8% to 55.7%, P = 0.02) without a significant decrease in specificity (from 87.2% to 86.7%, P = 0.87). The Kappa values also increased (from 0.66 to 0.71, P = 0.04). At 1-year follow-up, the improvement compared to baseline remained significant. There were no significant changes between the 2-week and 1-year follow-up assessment. CONCLUSION: A short training programme can significantly and sustainable improve GDPs diagnostic accuracy regarding CCAA.

NanoJ: a high-performance open-source super-resolution microscopy toolbox.

Super-resolution microscopy (SRM) has become essential for the study of nanoscale biological processes. This type of imaging often requires the use of specialised image analysis tools to process a large volume of recorded data and extract quantitative information. In recent years, our team has built an open-source image analysis framework for SRM designed to combine high performance and ease of use. We named it NanoJ-a reference to the popular ImageJ software it was developed for. In this paper, we highlight the current capabilities of NanoJ for several essential processing steps: spatio-temporal alignment of raw data (NanoJ-Core), super-resolution image reconstruction (NanoJ-SRRF), image quality assessment (NanoJ-SQUIRREL), structural modelling (NanoJ-VirusMapper) and control of the sample environment (NanoJ-Fluidics). We expect to expand NanoJ in the future through the development of new tools designed to improve quantitative data analysis and measure the reliability of fluorescent microscopy studies.

A HIDDEN MARKOV MODEL APPROACH TO CHARACTERIZING THE PHOTO-SWITCHING BEHAVIOR OF FLUOROPHORES.

Fluorescing molecules (fluorophores) that stochastically switch between photon-emitting and dark states underpin some of the most celebrated advancements in super-resolution microscopy. While this stochastic behavior has been heavily exploited, full characterization of the underlying models can potentially drive forward further imaging methodologies. Under the assumption that fluorophores move between fluorescing and dark states as continuous time Markov processes, the goal is to use a sequence of images to select a model and estimate the transition rates. We use a hidden Markov model to relate the observed discrete time signal to the hidden continuous time process. With imaging involving several repeat exposures of the fluorophore, we show the observed signal depends on both the current and past states of the hidden process, producing emission probabilities that depend on the transition rate parameters to be estimated. To tackle this unusual coupling of the transition and emission probabilities, we conceive transmission (transition-emission) matrices that capture all dependencies of the model. We provide a scheme of computing these matrices and adapt the forward-backward algorithm to compute a likelihood which is readily optimized to provide rate estimates. When confronted with several model proposals, combining this procedure with the Bayesian Information Criterion provides accurate model selection.

Calcified carotid artery atheromas in panoramic radiographs are associated with a first myocardial infarction: a case-control study.

OBJECTIVE: The aim of this case-control study was to investigate whether patients with a first myocardial infarction (MI) had a higher prevalence of calcified carotid artery atheromas (CCAAs) on panoramic radiographs (PRs) than age-, gender-, and residential area-matched controls without MI. STUDY DESIGN: Six hundred ninety-six cases with a first MI and 696 controls were included in this substudy of the Swedish multicentre PAROKRANK study. All participants underwent panoramic radiography, and the PRs were evaluated for CCAAs. RESULTS: The prevalence of CCAAs detected by PR was 33.8% (235 of 696) in cases and 27.6% (192 of 696) in controls (odds ratio [OR] 1.24; 95% confidence interval [CI] 1.04-1.44; P = .012). Among males, 32.7% of cases (184 of 562) and 26.5% of controls (149 of 562) displayed CCAAs on PRs (OR 1.24; 95% CI 1.03-1.48; P = .022). Among both genders, bilateral CCAAs were significantly more common among cases than among controls (P = .002). CONCLUSIONS: Cases with recent MIs had a significantly higher prevalence of CCAAs on PRs compared with controls without MIs. This difference between groups was more pronounced for bilateral CCAAs. These findings supported the hypothesis that CCAA detection could serve as a risk indicator for future MIs.

Fast live-cell conventional fluorophore nanoscopy with ImageJ through super-resolution radial fluctuations.

Despite significant progress, high-speed live-cell super-resolution studies remain limited to specialized optical setups, generally requiring intense phototoxic illumination. Here, we describe a new analytical approach, super-resolution radial fluctuations (SRRF), provided as a fast graphics processing unit-enabled ImageJ plugin. In the most challenging data sets for super-resolution, such as those obtained in low-illumination live-cell imaging with GFP, we show that SRRF is generally capable of achieving resolutions better than 150 nm. Meanwhile, for data sets similar to those obtained in PALM or STORM imaging, SRRF achieves resolutions approaching those of standard single-molecule localization analysis. The broad applicability of SRRF and its performance at low signal-to-noise ratios allows super-resolution using modern widefield, confocal or TIRF microscopes with illumination orders of magnitude lower than methods such as PALM, STORM or STED. We demonstrate this by super-resolution live-cell imaging over timescales ranging from minutes to hours.

EB1 accelerates two conformational transitions important for microtubule maturation and dynamics.

BACKGROUND: The dynamic properties of microtubules depend on complex nanoscale structural rearrangements in their end regions. Members of the EB1 and XMAP215 protein families interact autonomously with microtubule ends. EB1 recruits several other proteins to growing microtubule ends and has seemingly antagonistic effects on microtubule dynamics: it induces catastrophes, and it increases growth velocity, as does the polymerase XMAP215. RESULTS: Using a combination of in vitro reconstitution, time-lapse fluorescence microscopy, and subpixel-precision image analysis and convolved model fitting, we have studied the effects of EB1 on conformational transitions in growing microtubule ends and on the time course of catastrophes. EB1 density distributions at growing microtubule ends reveal two consecutive conformational transitions in the microtubule end region, which have growth-velocity-independent kinetics. EB1 binds to the microtubule after the first and before the second conformational transition has occurred, positioning it several tens of nanometers behind XMAP215, which binds to the extreme microtubule end. EB1 binding accelerates conformational maturation in the microtubule, most likely by promoting lateral protofilament interactions and by accelerating reactions of the guanosine triphosphate (GTP) hydrolysis cycle. The microtubule maturation time is directly linked to the duration of a growth pause just before microtubule depolymerization, indicating an important role of the maturation time for the control of dynamic instability. CONCLUSIONS: These activities establish EB1 as a microtubule maturation factor and provide a mechanistic explanation for its effects on microtubule growth and catastrophe frequency, which cause microtubules to be more dynamic.