Bone material contains a hierarchical network of micro- and nano-cavities and channels, known as the lacuna-canalicular network (LCN), that is thought to play an important role in mechanobiology and turnover. The LCN comprises micrometer-sized lacunae, voids that house osteocytes, and submicrometer-sized canaliculi that connect bone cells. Characterization of this network in three dimensions is crucial for many bone studies. To quantify X-ray Zernike phase-contrast nanotomography data, deep learning is used to isolate and assess porosity in artifact-laden tomographies of zebrafish bones. A technical solution is proposed to overcome the halo and shade-off domains in order to reliably obtain the distribution and morphology of the LCN in the tomographic data. Convolutional neural network (CNN) models are utilized with increasing numbers of images, repeatedly validated by 'error loss' and 'accuracy' metrics. U-Net and Sensor3D CNN models were trained on data obtained from two different synchrotron Zernike phase-contrast transmission X-ray microscopes, the ANATOMIX beamline at SOLEIL (Paris, France) and the P05 beamline at PETRA III (Hamburg, Germany). The Sensor3D CNN model with a smaller batch size of 32 and a training data size of 70 images showed the best performance (accuracy 0.983 and error loss 0.032). The analysis procedures, validated by comparison with human-identified ground-truth images, correctly identified the voids within the bone matrix. This proposed approach may have further application to classify structures in volumetric images that contain non-linear artifacts that degrade image quality and hinder feature identification.

Background Obesity is a known cardiovascular risk factor and associated with higher postoperative complication rates in patients undergoing cardiac surgery. In heart failure (HF), conflicting evidence in terms of survival has been reported, whereas sarcopenia is associated with poor prognosis. An increasing number of HF patients require left ventricular assist device (LVAD) implantations. The postoperative mortality has improved in recent years but is still relatively high. The impact of body composition on outcome in this population remains unclear. The aim of this investigation was to examine the preoperative computed tomography (CT) body composition as a predictor of the postoperative outcome in advanced HF patients, who receive LVAD implantations.

Methods Preoperative CT scans of 137 patients who received LVADs between 2015 and 2020 were retrospectively analysed using an artificial intelligence (AI)-powered automated software tool based on a convolutional neural network, U-net, developed for image segmentation (Visage Version 7.1, Visage Imaging GmbH, Berlin, Germany). Assessment of body composition included visceral and subcutaneous adipose tissue areas (VAT and SAT), psoas and total abdominal muscle areas and sarcopenia (defined by lumbar skeletal muscle indexes). The body composition parameters were correlated with postoperative major complication rates, survival and postoperative 6-min walk distance (6MWD) and quality of life (QoL).

Results The mean age of patients was 58.21 ± 11.9 years; 122 (89.1%) were male. Most patients had severe HF requiring inotropes (Interagency Registry for Mechanically Assisted Circulatory Support [INTERMACS] profile I–III, 71.9%) secondary to coronary artery diseases or dilated cardiomyopathy (96.4%). Forty-four (32.1%) patients were obese (body mass index ≥ 30 kg/m2), 96 (70.1%) were sarcopene and 19 (13.9%) were sarcopene obese. Adipose tissue was associated with a significantly higher risk of postoperative infections (VAT 172.23 cm2 [54.96, 288.32 cm2] vs. 124.04 cm2 [56.57, 186.25 cm2], P = 0.022) and in-hospital mortality (VAT 168.11 cm2 [134.19, 285.27 cm2] vs. 135.42 cm2 [49.44, 227.91 cm2], P = 0.033; SAT 227.28 cm2 [139.38, 304.35 cm2] vs. 173.81 cm2 [97.65, 254.16 cm2], P = 0.009). Obese patients showed no improvement of 6MWD and QoL within 6 months postoperatively (obese: +0.94 ± 161.44 months, P = 0.982; non-obese: +166.90 ± 139.00 months, P < 0.000; obese: +0.088 ± 0.421, P = 0.376; non-obese: +0.199 ± 0.324, P = 0.002, respectively). Sarcopenia did not influence the postoperative outcome and survival within 1 year after LVAD implantation.

Conclusions Preoperative AI-based CT body composition identifies patients with poor outcome after LVAD implantation. Greater adipose tissue areas are associated with an increased risk for postoperative infections, in-hospital mortality and impaired 6MWD and QoL within 6 months postoperatively.

Introduction. Cervical dystonia (CD) is viewed as a circumscribed movement disorder. However, beyond motor symptoms, it can imply subtle cognitive change, among others with respect to theory of mind (ToM) capacities. Here, affective and cognitive ToM performances and potential relations to other mental functions were investigated to refine the concept of social cognitive dysfunction in CD. Methods. 20 persons with CD were clinically assessed, together with 20 healthy controls engaged in overview cognitive testing, executive function (EF) tasks, and the Faux Pas Recognition Test (FPRT) as well as the Reading the Mind in the Eyes Tests (RMET) addressing cognitive and affective ToM functions, respectively. Results. Persons with CD showed lower cognitive, but not affective ToM performance than controls. Further, they had abnormally low word production in verbal fluency (VF) tasks, imposing high EF demands. Generally, ToM performance correlated with VF and, further, with the decreased quality of life score in persons with CD. Conclusion. Cognitive ToM deficits seem to occur in the context of executive dysfunction in CD. They belong to an underrecognized spectrum of nonmotor symptoms of likely clinical relevance.

Background Subcutaneous allergen immunotherapy (SCIT) with depigmented, polymerized (DPP) birch pollen extract has been marketed at doses of up to 1000 DPP units/mL since 2001. We sought to determine the dose-dependent efficacy of a DPP birch pollen extract formulation in patients suffering from birch-pollen-induced allergic rhinitis or rhinoconjunctivitis with or without intermittent asthma.

Methods A titrated conjunctival provocation test (CPT) was applied as a surrogate marker. This Phase II randomized, double-blind, parallel-group, dose-ranging clinical trial was performed at 39 centres in Germany, Lithuania and Poland. After randomization to four dose-level groups (100, 1000, 5000 and 10,000 DPP units/mL) and up-dosing, participants received maintenance SCIT with five monthly subcutaneous injections. The primary endpoint was the proportion of patients in whom a higher concentration of birch pollen (vs. baseline) was needed to elicit a positive CPT.

Results Three hundred forty-three patients were included (mean (range) age: 42.6 (19–70)). The highest CPT responder rates were seen in the higher dose-level groups. In the intention-to-treat analysis, the difference between the 100 and 10,000 groups was statistically significant (p = 0.0118). Although the proportion of patients with ≥1 treatment-emergent adverse events increased with the dose, almost all these events were mild (65.6%) or moderate (18.5%).

Conclusion Judging by the results of a CPT, the efficacy/safety ratio in SCIT appears to be favourable for a high-dose-level preparation of a DPP birch pollen extract.

Background

Vasoregulatory autoantibodies including autoantibodies targeting G‐protein–coupled receptors might play a functional role in vascular diseases. We investigated the impact of vasoregulatory autoantibodies on clinical outcome after ischemic stroke.

Methods and Results

Data were used from the PROSCIS‐B (Prospective Cohort With Incident Stroke–Berlin). Autoantibody‐targeting receptors such as angiotensin II type 1 receptor (AT1R), endothelin‐1 type A receptor, complement factor‐3 and ‐5 receptors, vascular endothelial growth factor receptor‐1 and ‐2, vascular endothelial growth factor A and factor B were measured. We explored associations of high antibody levels with (1) poor functional outcome defined as modified Rankin Scale >2 or Barthel Index <60 at 1 year after stroke, (2) Barthel Index scores over time using general estimating equations, and (3) secondary vascular events (recurrent stroke, myocardial infarction) or death up to 3 years using Cox proportional hazard models. We included 491 patients with ischemic stroke with data on autoantibody levels and outcome. In models adjusted for demographics and vascular risk factors, high autoantibody concentrations (quartile 4) targeting complement factor C3a receptor, vascular endothelial growth factor receptor‐2, and vascular endothelial growth factor B were associated with poor functional outcome at 1 year: (odds ratio, 2.0 [95% CI, 1.1–3.6]; odds ratio, 1.8 [95% CI, 1.1–3.2]; and odds ratio, 2.1 [95% CI, 1.2–3.6], respectively) and with lower Barthel Index scores over 3 years (complement factor C3a receptor: adjusted β=−3.3 [95% CI, −5.7 to −0.5]; VEGF‐B: adjusted β=−2.4 [95% CI, −4.8 to −0.06]). Patients with high autoantibody levels were not at higher risk for secondary vascular events or death.

Conclusions

High levels of autoantibodies against vascular endothelial growth factor receptor‐2, vascular endothelial growth factor B, and complement factor C3a receptor measured are associated with poor functional outcome after stroke but not with recurrent vascular events or death.

Registration

URL: https://www.clinicaltrials.gov; Unique identifier: NCT01363856.

Objective Retrograde trans-synaptic neuroaxonal degeneration is considered a key pathological factor of subclinical retinal neuroaxonal damage in multiple sclerosis (MS). We aim to evaluate the longitudinal association of optic radiation (OR) lesion activity with retinal neuroaxonal damage and its role in correlations between retinal and brain atrophy in people with clinically isolated syndrome and early MS (pweMS).

Methods Eighty-five pweMS were retrospectively screened from a prospective cohort (Berlin CIS cohort). Participants underwent 3T magnetic resonance imaging (MRI) for OR lesion volume and brain atrophy measurements and optical coherence tomography (OCT) for retinal layer thickness measurements. All pweMS were followed with serial OCT and MRI over a median follow-up of 2.9 (interquartile range: 2.6–3.4) years. Eyes with a history of optic neuritis prior to study enrollment were excluded. Linear mixed models were used to analyze the association of retinal layer thinning with changes in OR lesion volume and brain atrophy.

Results Macular ganglion cell-inner plexiform layer (GCIPL) thinning was more pronounced in pweMS with OR lesion volume increase during follow-up compared to those without (Difference: −0.82 μm [95% CI:-1.49 to −0.15], p = 0.018). Furthermore, GCIPL thinning correlated with both OR lesion volume increase (β [95% CI] = −0.27 [−0.50 to −0.03], p = 0.028) and brain atrophy (β [95% CI] = 0.47 [0.25 to 0.70], p < 0.001). Correlations of GCIPL changes with brain atrophy did not differ between pweMS with or without OR lesion increase (  = 5.92e−7, p = 0.762).

Interpretation Faster GCIPL thinning rate is associated with increased OR lesion load. Our results support the value of GCIPL as a sensitive biomarker reflecting both posterior visual pathway pathology and global brain neurodegeneration.

The DNA damage response (DDR) acts as a barrier to malignant transformation and is often impaired during tumorigenesis. Exploiting the impaired DDR can be a promising therapeutic strategy; however, the mechanisms of inactivation and corresponding biomarkers are incompletely understood. Starting from an unbiased screening approach, we identified the SMC5-SMC6 Complex Localization Factor 2 (SLF2) as a regulator of the DDR and biomarker for a B-cell lymphoma (BCL) patient subgroup with an adverse prognosis. SLF2-deficiency leads to loss of DDR factors including Claspin (CLSPN) and consequently impairs CHK1 activation. In line with this mechanism, genetic deletion of Slf2 drives lymphomagenesis in vivo. Tumor cells lacking SLF2 are characterized by a high level of DNA damage, which leads to alterations of the post-translational SUMOylation pathway as a safeguard. The resulting co-dependency confers synthetic lethality to a clinically applicable SUMOylation inhibitor (SUMOi), and inhibitors of the DDR pathway act highly synergistic with SUMOi. Together, our results identify SLF2 as a DDR regulator and reveal co-targeting of the DDR and SUMOylation as a promising strategy for treating aggressive lymphoma.

Bone has the fascinating ability to self-regenerate. However, under certain conditions, such as type 2 diabetes mellitus (T2DM), this ability is impaired. T2DM is a chronic metabolic disease known by the presence of elevated blood glucose levels that is associated with reduced bone regeneration capability, high fracture risk, and eventual non-union risk after a fracture. Several mechanical and biological factors relevant to bone regeneration have been shown to be affected in a diabetic environment. However, whether impaired bone regeneration in T2DM can be explained due to mechanical or biological alterations remains unknown. To elucidate the relevance of either one, the aim of this study was to investigate the relative contribution of T2DM-related alterations on either cellular activity or mechanical stimuli driving bone regeneration. A previously validated in silico computer modeling approach that was capable of explaining bone regeneration in uneventful conditions of healing was further developed to investigate bone regeneration in T2DM. Aspects analyzed included the presence of mesenchymal stromal cells (MSCs), cellular migration, proliferation, differentiation, apoptosis, and cellular mechanosensitivity. To further verify the computer model findings against in vivo data, an experimental setup was replicated, in which regeneration was compared in healthy and diabetic after a rat femur bone osteotomy stabilized with plate fixation. We found that mechanical alterations had little effect on the reduced bone regeneration in T2DM and that alterations in MSC proliferation, MSC migration, and osteoblast differentiation had the highest effect. In silico predictions of regenerated bone in T2DM matched qualitatively and quantitatively those from ex vivo & mu;CT at 12 weeks post-surgery when reduced cellular activities reported in previous in vitro and in vivo studies were included in the model. The presented findings here could have clinical implications in the treatment of bone fractures in patients with T2DM.

Alternative medicine methods and treatments enjoy great popularity. However, many users fail to recognise the dangers associated with these methods. One of these is called Black Ointment or Black Salve. Oncologists and attending physicians should be aware of alternative medical methods such as these and be in a position to provide appropriate advice, intervention, if necessary, at any time when counselling patients in difficult situations.

Background Body composition alterations are frequent in patients with cancer or chronic liver disease, but their prognostic value remains unclear in many cancer entities.

Objective We investigated the impact of disease aetiology and body composition after surgery for intrahepatic cholangiocarcinoma (iCCA), a rare and understudied cancer entity in European and North American cohorts.

Methods Computer tomography-based assessment of body composition at the level of the third lumbar vertebra was performed in 173 patients undergoing curative-intent liver resection for iCCA at the Department of Surgery, Charité – Universitätsmedizin Berlin. Muscle mass and -composition as well as subcutaneous and visceral adipose tissue quantity were determined semi-automatically. (Secondary) sarcopenia, sarcopenic obesity, myosteatosis, visceral and subcutaneous obesity were correlated to clinicopathological data.

Results Sarcopenia was associated with post-operative morbidity (intraoperative transfusions [p = 0.027], Clavien–Dindo ≥ IIIb complications [p = 0.030], post-operative comprehensive complication index, CCI [p < 0.001]). Inferior overall survival was noted in patients with myosteatosis (33 vs. 23 months, p = 0.020).

Fifty-eight patients (34%) had metabolic (dysfunction)-associated fatty liver disease (MAFLD) and had a significantly higher incidence of sarcopenic (p = 0.006), visceral (p < 0.001) and subcutaneous obesity (p < 0.001). Patients with MAFLD had longer time-to-recurrence (median: 38 vs. 12 months, p = 0.025, log-rank test). Multivariable cox regression analysis confirmed only clinical, and not body, composition parameters (age > 65, fresh frozen plasma transfusions) as independently prognostic for overall survival.

Conclusion This study evidenced a high prevalence of MAFLD in iCCA, suggesting its potential contribution to disease aetiology. Alterations of muscle mass and adipose tissue were more frequent in patients with MAFLD.

Background The coronavirus disease pandemic and its containing measures have caused concerns for patients with hereditary angioedema (HAE) and their treating physicians. Both faced challenges surrounding interaction, and communication had to adapt to facilitate appropriate management. Specifically, the pandemic resulted in reduced in-person contact in clinics. Where possible, telemedicine appointments were offered and treatment outside the hospital setting was encouraged.

Body The pandemic markedly affected patient-physician communication, which is essential to maintain partnerships and optimize care. Although patients with HAE are often experts in their condition, guidance by their physicians is essential, especially with the recent shift toward patient-centered management for rare diseases and shared decision-making (SDM). SDM enables patients to take control of their disease and allows the risks and benefits of treatment to be discussed with their physicians. This review explores perspectives from patients and physicians in the HAE clinical setting, particularly regarding their experiences with communication throughout the pandemic. We discuss the importance of SDM in rare diseases such as HAE, factors that impact effective communication, and potential solutions.

Conclusion Since patient-centered care and SDM have particular relevance in rare diseases in general, we believe our findings could be transferrable and applicable in the management of other rare diseases.

Introduction The integrated care pathways for atopic dermatitis (AD-ICPs) aim to bridge the gap between existing AD treatment evidence-based guidelines and expert opinion based on daily practice by offering a structured multidisciplinary plan for patient management of AD. ICPs have the potential to enhance guideline recommendations by combining interventions and aspects from different guidelines, integrating quality assurance, and describing co-ordination of care. Most importantly, patients can enter the ICPs at any level depending on AD severity, resources available in their country, and economic factors such as differences in insurance reimbursement systems.

Methods The GA2LEN ADCARE network and partners as well as all stakeholders, abbreviated as the AD-ICPs working group, were involved in the discussion and preparation of the AD ICPs during a series of subgroup workshops and meetings in years 2020 and 2021, after which the document was circulated within all GAL2EN ADCARE centres.

Results The AD-ICPs outline the diagnostic procedures, possible co-morbidities, different available treatment options including differential approaches for the pediatric population, and the role of the pharmacists and other stakeholders, as well as remaining unmet needs in the management of AD.

Conclusion The AD-ICPs provide a multidisciplinary plan for improved diagnosis, treatment, and patient feedback in AD management, as well as addressing critical unmet needs, including improved access to care, training specialists, implementation of educational programs, assessment on the impact of climate change, and fostering a personalised treatment approach. By focusing on these key areas, the initiative aims to pave the way for a brighter future in the management of AD.

Background The Angioedema Control Test (AECT) is a patient-reported outcome measure developed and validated for the assessment of disease control in patients with recurrent angioedema. Its sensitivity to change and minimal clinically important difference (MCID) have hitherto not been established.

Methods Patients with recurrent angioedema due to chronic spontaneous urticaria, hereditary angioedema, or acquired C1-inhibitor deficiency were repeatedly asked to complete the AECT along with the Angioedema Quality of Life Questionnaire (AE-QoL), Dermatology Life Quality Index (DLQI), and anchors for disease control and whether treatment was sufficient during routine care visits. The sensitivity to the change of the AECT was determined by correlating changes in its scores over time with changes in the applied anchors. The MCID was determined using anchor-based and distributional criterion-based approaches.

Results Eighty-six cases were used for this analysis. Changes in AECT scores correlated well with AE-QoL changes (but less with changes in the DLQI) as well as other applied anchors, demonstrating its sensitivity to change. The MCID was found to be three points for improvement of angioedema control. The available number of cases with meaningful deterioration in our dataset was too low to reach a definite conclusion on the MCID for deterioration of angioedema control.

Conclusion The AECT is a valuable tool to assess changes in disease control in patients with recurrent angioedema over time. The lowest AECT score change that reflects a meaningful improvement of disease control to patients (MCID) is three points.

As AI becomes increasingly relevant, especially in high-stakes domains such as healthcare, it is important to investigate which approaches can improve human-AI collaboration and, if so, why. Current research focuses primarily on technically available approaches, such as explainable AI (XAI), often overlooking human needs. This study bridges this gap by adopting a well-established technical approach - model uncertainty representations - by considering users' familiarity with the format and numeracy skills. Despite being provided with uncertainty representations, users may still struggle to handle uncertain decisions. Thus, we introduce an educational approach that communicates the capabilities of humans and the AI system to users, supplementing the uncertainty representations. We conducted a pre-registered, between-subjects user study to determine whether these approaches resulted in improved human-AI team performance, mediated by the user's mental model of the AI. Our findings indicate that solely providing uncertainty representations does not improve team performance or the user's mental model in comparison to only providing AI recommendations. However, incorporating capability-focused guidance alongside uncertainty representations significantly enhances correct self-reliance and, to some extent, overall team performance. Our additional exploratory analyses suggest that factors such as task uncertainty, case difficulty, and case type, rather than numeracy skills, the need for cognition or familiarity, can influence team performance. We discuss these factors in detail, provide practical implications, and suggest directions for further research. This work contributes to the CSCW discourse by demonstrating how technical approaches can be augmented with educational approaches to enhance human-AI collaboration in decision-making tasks.

It is demonstrated that the longitudes of the planetary exaltations mentioned in numerous Greco-Roman and Late Antique astrological sources derive from Babylonian Normal-Star longitudes. This is achieved through a comparison between both sets of longitudes. Supporting evidence is found in the Late Babylonian astral compendium BM 36609+. The Babylonian longitudes were transferred to exaltations without significant changes, but their association with stars was discarded. The sun’s exaltation constitutes an exception, since it does not correspond to a Normal Star, but appears to have been defined in relation to the longitude of the Pleiades, which became the moon’s exaltation.

Social housing has been experiencing a global trend toward residualization, primarilytargeting vulnerable groups through non-market methods. Meanwhile, it is undergoinga parallel trend of financialization, where it is increasingly treated as a financial assetfor trading and speculation. In contrast, social rented housing in China is transitioningto a mass model and embracing financialization, exemplified by the government'sproactive promotion of the Affordable Rental Housing (ARH) program and theintroduction of Real Estate Investment Trusts (REITs) for social housing investment.However, the combination of seemingly non-profit social housing with profit-drivenREITs remains an understudied phenomenon. This paper addresses this gap by drawingon theories of overaccumulation and state entrepreneurialism. We explore the rationalesdriving the promotion of ARH (-REITs) and investigate the state's role in facilitatingthis process. We argue that China's development and financialization of ARH isincreasingly adopted as a national strategy to mitigate the overaccumulation crisis inthe homeownership sector through a 'spatial fix' wherein capital circulates within thesecondary circuit rather than switches between circuits of capital. This strategy involvesestablishing legal frameworks and institutions to make ARH profitable, stimulatingdemand by expanding target groups to include middle- to high-income “talents” withadditional benefits, stimulating supply by reducing costs and political directives, andpromoting financial innovation and credit enhancement. We conclude with concernsabout the potential long-term ‘crisis-magnifying’ effects of this spatial fix and itsimplications for vulnerable households.

Predictive Coding (PC) is a neuroscientific theory that has inspired a variety of training algorithms for biologically inspired deep neural networks (DNN). However, many of these models have only been assessed in terms of their learning performance, without evaluating whether they accurately reflect the underlying mechanisms of neural learning in the brain. This study explores whether predictive coding inspired Deep Neural Networks can serve as biologically plausible neural network models of the brain. We compared two PC-inspired training objectives, a predictive and a contrastive approach, to a supervised baseline in a simple Recurrent Neural Network (RNN) architecture. We evaluated the models on key signatures of PC, including mismatch responses, formation of priors, and learning of semantic information. Our results show that the PC-inspired models, especially a locally trained predictive model, exhibited these PC-like behaviors better than a Supervised or an Untrained RNN. Further, we found that activity regularization evokes mismatch response-like effects across all models, suggesting it may serve as a proxy for the energy-saving principles of PC. Finally, we find that Gain Control (an important mechanism in the PC framework) can be implemented using weight regularization. Overall, our findings indicate that PC-inspired models are able to capture important computational principles of predictive processing in the brain, and can serve as a promising foundation for building biologically plausible artificial neural networks. This work contributes to our understanding of the relationship between artificial and biological neural networks such as the brain, and highlights the potential of PC-inspired algorithms for advancing brain modelling as well as brain-inspired machine learning.

Irregular or deviant burials, i. e. cases of sepulture divergent from the usual burial rite, are well known from many cultures. Based on a review of numerous published burial sites, the existence of similar practices can now be documented for ancient Mesopotamia of the 3rd to 1st millennium BCE as well. Unusual features include absence or dislocation of skull or limb bones, partial burning, burial in prone position, and complete disarticulation. While a part of these may go back to disturbance or secondary reburial, perimortal violence, or ritual removal of bones, certain body manipulations such as skeletal mutilation, fettering, and covering with stones may be explained as arrangements against the return of the dead, comparable to similar practices against alleged revenants and vampires in other cultures.

Protein folding stability can be probed using urea, a chaotropic agent that disrupts non-covalent interactions at molar concentrations. The denaturation process is typically monitored via optical spectroscopy, which provides ensemble-averaged measurements and may struggle to resolve folding intermediates. In contrast, electrospray ionization mass spectrometry (ESI-MS) captures a non-averaged snapshot of all populated assembly and folding states within a protein conformational ensemble. However, high urea concentrations have traditionally been considered incompatible with ESI. Here, we leverage recent advancements in nano ESI emitter design, utilizing well-defined small-diameter emitters which enables protein charge states to be resolved from solutions containing up to 8 M urea. This approach allows us to directly detect the disruption of native tertiary and quaternary structures and to monitor stability changes in response to solution pH and ligand binding. We demonstrate this using single-domain proteins that follow simple two-state unfolding pathways, as well as more complex multidomain proteins and multimeric protein complexes. Our results show strong agreement with conventional urea–denaturation curves obtained via optical spectroscopy, while also providing enhanced resolution of intermediate folding and assembly states that are challenging to capture using traditional methods.