N = 67, Mage = 9 years) and adolescents . Parents completed an online survey that asked about child and parental mental health, infection experience, and changes to education arrangements, family life, housing and finances during the pandemic. Pre-existing measures of mental health, autism and adaptive functioning were also utilised. More engagement and enjoyment in education provision and going outside was associated with better child and parental mental health. In multivariate multiple linear regression models, more pre-existing attention deficit hyperactivity disorder symptoms were associated with more behavioural/attention deficit hyperactivity disorder symptoms during the pandemic in the pre-adolescent cohort, and with greater emotional symptoms in the adolescent cohort. More pre-existing parental mental health problems were associated with more parental mental health symptoms during the pandemic in both cohorts. Knowledge of pre-existing mental health and pandemic-related stressors may help care planning. Encouraging engagement and enjoyment in education and promoting physical exercise are key intervention targets. Ensuring access to attention deficit hyperactivity disorder medication and support is important, especially if this is managed jointly across school and home.The current study explored the role of pre-existing and pandemic-time child, family or environmental factors in the presentation of mental health symptoms of autistic youth and their parents during the pandemic. Participants were parents/carers of autistic children (Autism Spectrum Treatment and Resilience Cohort, What is already known about the topic: The COVID-19 pandemic and the associated restrictions impacted all of society. There is emerging evidence showing a range of impacts on autistic children and young people and their families. Further research that looks at how individuals coped during the pandemic while considering how they were doing before the pandemic is needed.What this paper adds: This article explores whether how well autistic youth were doing before the pandemic influenced how they coped during the pandemic. It also looked at how well their parents were doing during the pandemic and whether any pre-pandemic factors influenced how they coped. Samples of both primary-school-aged autistic children and autistic teenagers and their parents were surveyed to answer these questions. More engagement and enjoyment in education provision during the pandemic and getting outside more were linked with better child and parental mental health during the pandemic. More attention deficit hyperactivity disorder before the pandemic was linked with more attention deficit hyperactivity disorder and behavioural problems during the pandemic in primary-school-aged autistic children, and more emotional problems during the pandemic in autistic teenagers. Parents with more mental health problems during the pandemic had more mental health problems before the pandemic.Implications for practice, research or policy: Encouraging engagement and enjoyment in education and promoting physical exercise are key intervention targets. Ensuring access to attention deficit hyperactivity disorder medication and support is important, especially if this is managed jointly across school and home. The COVID-19 pandemic had a widespread impact on society worldwide. Three key vulnerable groups were: children and young people (CYP) and families affected by school closures; those with pre-existing mental health symptoms, broadly known as a latent trait of psychopathology  e.g. em. A systeAutistic CYP were particularly at risk for negative impacts of the pandemic for several reasons . First, decrease in parent-reported emotional, behavioural and ADHD symptoms in SEN CYP over the first UK lockdown; whereas a slight increase in these symptoms was observed among CYP without SEN. In However, findings from studies exploring the mental health of CYP with neurodevelopmental disorders during the pandemic present a varied picture. For example, the Co-SPACE study showed children with special educational needs (SENs) or neurodevelopmental disorders to have markedly elevated levels of emotional symptoms, behavioural symptoms and ADHD during the pandemic, compared to those without SEN or neurodevelopmental problems . Since tThe findings from studies exploring the mental health of parents during the pandemic also are mixed. A systematic review of literature reporting mental health symptoms in parents/carers of CYP reported that 52% had anxiety and 27% had depression during the pandemic . In Co-SCOVID-19-related research conducted with CYP with neurodevelopmental disorders has mainly surveyed or tracked CYP since the pandemic began. While this provides essential information about the current mental health of this population and how it has changed over the course of the pandemic, the lack of systematically selected samples means that there is limited understanding of participant representativeness, and the lack of pre-pandemic measures of mental health symptoms does not allow for comparisons to symptoms during the pandemic or identification of risk and protective factors. Further research examining who is most at risk of negative impacts of the pandemic based on pre-existing factors and lockdown circumstances is needed. Such information is essential to understand whether there are sub-groups who may be more adversely affected and may in turn be used to guide future care planning and pandemic response.The current study aims to address the following research questions:Research Question 1. How did the pandemic impact the lives of families of autistic children?Research Question 2. Were the mental health symptoms of autistic CYP during the pandemic associated with pre-existing mental health symptoms, over and above autism characteristics, adaptive functioning and other contextual factors related to the pandemic?Research Question 3. Were the mental health symptoms of parents/carers of autistic CYP during the pandemic associated with their own/their child’s pre-existing mental health symptoms, over and above their child’s autism characteristics, adaptive functioning and other contextual factors related to the pandemic?We included two samples of well-characterised pre-adolescent and adolescent CYP to examine impact across the developmental period and allow for internal replication of results. Both samples were originally recruited from a similar geographical area and tracked over time as part of the wider Improving Autism Mental Health  programme and hence policies around education and social restrictions will have been similar for both cohorts. Comparing the cohorts in this way enabled us to look at the commonalities and differences that apply to autistic CYP at two important developmental stages during the pandemic.We hypothesised that greater autism characteristics, pre-existing mental health and behavioural symptoms and environmental stressors would be associated with increased mental health symptoms during the pandemic among both pre-adolescent and adolescent CYP. For parents, we hypothesised that those with CYP with more pre-existing mental health symptoms, those who had more pre-existing parental mental health symptoms and those with pandemic-related environmental stressors would report more parental mental health symptoms during the pandemic.Supplementary Materials.The current study consists of two samples of autistic CYP drawn from the South London area. Separate public involvement panels of autistic adults and parents of autistic children advised on the research conducted with both cohorts. This involvement consisted of seeking guidance on all aspects of the research, from objectives to design and interpretation of findings via regular group meetings and electronic feedback. Data collection for the current study was conducted early in the pandemic. Pre-existing measures were taken pre-pandemic. N = 82/85, 96%) were invited to participate in the current study via email  cohort consisted of 85 parents of young autistic children who consented to participate in a feasibility study or pilot randomised controlled trial of novel group-based parent-mediated interventions (ISRCTN91411078), as part of IAMHealth . All parThe sample participating in the current study consisted of 67 parents (92.5% mothers) of autistic children. Most children were male (83.6%) and were on average 8 years 10 months (range 6:0–11:10) during the pandemic-time data collection. Thirty-five (52.2%) children were minimally verbal and 32 (47.8%) were verbal, as verbal ability was stratified for in the original study design. The children were split across specialist (42.4%) and mainstream (57.6%) education provision when originally assessed.N = 211) were invited to participate in the current study. In previous waves of data collection for the QUEST cohort, children were split into an ‘intensively studied’ (hereafter intensive) and ‘extensively studied’ group (hereafter extensive). The intensive group completed in-depth assessments (including direct observations of the CYP) whereas the extensive group completed shorter online parent-report questionnaires only. For the current study, families were approached and data was collected in the same manner as described above for the ASTAR cohort. All parents/carers completed the same measures, with data collection taking place between July and October 2020  parents/carers (95.5% mothers) participated. 79% of their CYP were male; 37% had an IQ <70 when assessed pre-pandemic; and 49% were in specialist education at the last point of data collection approximately 18 months prior to the pandemic.The European Child & Adolescent Psychopharmacology Network (ECNP) Child and Adolescent Mental Health Services (CAMHS) COVID survey was designed to tap into a broad range of mental health symptoms experienced during the pandemic and contextual factors particularly pertinent to periods of social restrictions. Questions from the Coronavirus Health and Impact Survey mental health section  were useSupplementary Materials for loadings for the current sample). The questions on the child’s emotional and behavioural symptoms formed two factors: (1) emotional symptoms  and (2) behavioural/ADHD symptoms . The questions on parental emotional and behavioural symptoms formed two factors: (1) emotional symptoms (measured using the same six items) and (2) irritable behaviour .Factor analysis of this measure using a larger dataset collected from CYP in South London and Maudsley (SLaM) CAMHS (manuscript in preparation) led us to adopt a two-factor solution for both CYP and parent mental health symptoms, which performed well .The Adaptive Behaviour Assessment System (ABAS) was used as a parent-report measure of adaptive functioning in ASTAR  Preschool Anxiety Scale–Revised PASR, , a 28-itIn the QUEST cohort, pre-existing CYP mental health and behavioural symptoms were measured using the Strengths and Difficulties Questionnaire SDQ, . The SDQThe Autism Parenting Stress Index APSI,  was usedData analysis was conducted in Stata MP 16/17 . All varSupplementary Table 4). Variables that were significant at p < .05 in either cohort were entered into a multivariate multiple regression model, using structural equation modelling with full information maximum likelihood. This analysis was chosen to generate standardised coefficients for comparison across the samples and deal with missing data. In the models examining CYP symptoms, the dependent variables were CYP emotional symptoms or CYP behavioural/ADHD symptoms (as measured by the COVID survey). Independent variables included: CYP age, level of adaptive functioning, autism characteristics, pre-existing CYP mental health problems, pre-existing parental distress and the family’s current financial situation, home accommodation, access to garden and CYP physical attendance at school/college. In addition to these, level of adaptive functioning and autism characteristics were retained as co-variates to account for CYP ability levels and autism characteristics, even if they weren’t significant predictors at the bivariate level.The bivariate associations between the independent and dependent variables were first examined . The independent variables were the same as those specified in the CYP outcome models above.In the models examining parent symptoms, again any variables with a significant bi-variate association with any pandemic-time outcome at Supplementary Material for further information).For both cohorts, there were no significant differences between participants and non-participants with regard to child and parent characteristics, with exception of parental unemployment being more common among QUEST non-participants versus participants (see Supplementary Table 4). However, less engagement and enjoyment in education was associated with more CYP emotional symptoms in ASTAR, more behavioural/ADHD symptoms in the CYP in both samples and more parental emotional symptoms in QUEST . Consistent across cohorts, lower pre-existing observed restricted and repetitive behaviour was associated with more behavioural/ADHD symptoms during the pandemic. However, lower pre-existing parent-reported social communication impairment scores were associated with more CYP emotional symptoms in ASTAR during the pandemic consistent with the observed restricted and repetitive behaviour association; whereas the opposite pattern was found in QUEST with higher parent-reported social communication impairment scores being associated with CYP emotional symptoms. Furthermore, in ASTAR, lower pre-pandemic adaptive functioning and increased pre-existing CYP ADHD behaviours were also associated with greater CYP behavioural/ADHD symptoms during the pandemic. In QUEST, higher levels of pre-existing parental distress were additionally associated with more pandemic-time behavioural/ADHD symptoms in the CYP. Pre-existing CYP emotional symptoms was not associated with pandemic-time emotional symptoms in neither sample, although more pre-existing ADHD was associated with greater emotional symptoms during the pandemic in QUEST.Supplementary Table 6 for details). Consistent across both ASTAR and QUEST, greater pre-existing parental distress was associated with increased parent emotional symptoms and increased parent irritable behaviour during the pandemic. For QUEST only, increased current financial concerns were also associated with increased parent emotional symptoms and irritable behaviour during the pandemic. In ASTAR, lower pre-existing child-observed SA was associated with greater pandemic-time parental emotional symptoms. However, no child factors were associated with parental mental health symptoms during the pandemic in QUEST.The current study explored whether mental health symptoms in autistic CYP and their parents during the pandemic were associated with pre-existing mental health symptoms, over and above pre-existing measures of autism characteristics and adaptive functioning, and other contextual factors related to the pandemic. We utilised previously studied cohorts of both pre-adolescent and adolescent autistic CYP to examine the impact of pre-existing and current vulnerabilities across childhood.For almost all families, the experience of the pandemic during data collection had been one of the severe restrictions. A large proportion of both pre-adolescent and adolescent CYP did not access education as they usually would, even though it was UK policy that CYP with SEN  should have access to their usual educational placement . We founFor pre-adolescent autistic CYP, higher levels of pre-existing ADHD symptoms were associated with higher levels of behavioural/ADHD symptoms during the pandemic, perhaps simply highlighting the stability of symptoms within this domain over time . This waIn contrast to our expectation, less observed restricted and repetitive behaviours were associated with greater pandemic-time behavioural/ADHD symptoms. This was consistent across both pre-adolescent and adolescent CYP. Given the inconsistencies between this finding and the relationship between social communication and interaction impairments and CYP’s mental health during the pandemic mentioned below, caution should be taken when interpreting this result. Different from the pattern of relationships between restricted and repetitive behaviours and CYP mental health, fewer parent-reported social communication and interaction impairments were associated with more emotional symptoms in the pre-adolescent children, but fewer emotional symptoms in the adolescent children during the pandemic. This could be due to the different ages of the two cohorts; the differential impact of the pandemic on emotional well-being for pre-adolescents versus adolescents; the role of school as a potential stressor for adolescents versus pre-adolescents; the use of different versions of the measure tapping into social communication and interaction impairments which asked about different time periods for the two cohorts (lifetime impairment for ASTAR versus current impairment for QUEST). Furthermore, the association between social communication impairment scores and CYP emotional symptoms in the pre-adolescent sample was non-significant at the bivariate level. Future research should be undertaken to replicate these effects.In both cohorts, few familial or environmental stressors were associated with pandemic CYP mental health symptoms, except for pre-existing parental distress being associated with pandemic-time behavioural/ADHD symptoms among adolescents. This suggests that pre-existing mental health symptoms in both the child and parent are more pertinent to CYP mental health symptoms experienced during the pandemic than other environmental factors. This finding is consistent with pre-pandemic literature .For parents of autistic CYP in our samples, more pre-existing parental distress was associated with increased pandemic-time mental health symptoms in parents of both pre-adolescent and adolescent children indicating the stability of these difficulties and vulnerability to pandemic impacts. More pandemic-time parental mental health symptoms were associated with increased financial concerns during the pandemic in parents of autistic adolescents; this relationship was replicated in the pre-adolescent cohort but only at the bivariate level when other factors weren’t controlled for. This finding is in line with the previous literature that demonstrated the impact of the pandemic on parents . Such imThe combined findings from the two cohorts indicate that for autistic CYP and their parents, those more vulnerable to the impacts of the COVID-19 pandemic restrictions may have had heightened mental health symptoms pre-pandemic. This highlights the need for services to consider autistic CYP and their families’ pre-existing mental health symptoms when, for example, prioritising access . Two types of continuity in mental health symptoms have been suggested, heterotypic and homotypic continuity . The finThe survey used to assess the impact of the COVID-19 pandemic on CYP and their parents is not a standardised measure. This means that we cannot compare the levels of mental health symptoms among our samples with other samples or the population. In addition, measures of pre-existing mental health symptoms varied across the cohorts and over time. We therefore cannot make definite conclusions about whether mental health symptoms changed as a result of the pandemic and associated restrictions. In addition, the survey only provides a snapshot of experience as it asks about the 2 weeks prior to survey completion, and data were collected early in the pandemic. We do not know the prolonged impact of the restrictions or impacts when restrictions were re-imposed in the UK. However, there is growing recognition that referrals to CAMHS and evidence of increased CYP distress became apparent after the period of this survey completion . FurtherThis study used two well-characterised pre-existing cohorts to examine the effects of the UK pandemic on autistic children and parental mental health. The inclusion of these two cohorts, one a community sample and the other a sample involved in a feasibility pilot randomised controlled trial with few eligibility criteria drawn from similar geographical areas enabled us to understand what factors are consistently associated with mental health symptoms during the pandemic across pre-adolescent and adolescent CYP and their families. In summary, autistic CYP and their parents with greater pre-pandemic mental health symptoms are likely to be more at risk of having more pandemic-time mental health symptoms. This information should be used to guide care planning for autistic CYP and their families.Click here for additional data file.Supplemental material, sj-pdf-1-aut-10.1177_13623613231153694 for Factors associated with mental health symptoms among UK autistic children and young people and their parents during the COVID-19 pandemic by Melanie Palmer, Susie Chandler, Virginia Carter Leno, Farah Mgaieth, Isabel Yorke, Matthew Hollocks, Andrew Pickles, Vicky Slonims, Stephen Scott, Tony Charman and Emily Simonoff in Autism


Cardiovascular disease (CVD) can begin early and progress over one’s lifetime. Chronic disease limits the ability of therapies to reverse pathophysiology once symptoms present. An estimated 17.9 million lives are lost annually to CVD, making it the leading cause of death globally. Current treatments are mostly reactive and used primarily when a patient becomes ill. So, how are the medical and public health communities tackling CVD, and what will disease prevention look like in the coming decades?Recent results from the highly anticipated DELIVER trial exemplify the limitations of contemporary cures as opposed to preventative strategies. Dapagliflozin, a sodium-glucose cotransporter-2 inhibitor (SGLT2i), was compared to placebo in heart failure patients with preserved ejection fraction (HFpEF). Positive results were expected, as SGLT2 inhibition was previously shown effective in patients with type 2 diabetes, kidney disease, or reducing heart failure. Dapagliflozin met its primary outcome: a reduction in the composite score of worsening heart failure or cardiovascular death. However, just like the EMPEROR-Preserved trial with another SGLT2i empagliflozin, the positive effect of dapagliflozin was mainly driven by a reduction in heart failure hospitalizations with no effect on cardiovascular death. Given that heart failure hospitalizations represent only a low number of all hospitalizations in patients with HFpEF, this high-cost drug class may be of low value in treating difficult-to-manage HFpEF. As baseline therapies have improved over time, it is harder to demonstrate big gains with newer expensive drugs. It is becoming less reasonable to warrant the spending of precious healthcare funds, unless significant improvements to health are clearly proven. Hence, there is an urgent need for preventative strategies to lower the incidences of CVD before patients become symptomatic and require costly care.Prevention is better than cure was a key theme at this year’s European Society of Cardiology’s (ESC) Congress. Population-based screening has been viewed as one method to detect subclinical CVD. The DANCAVAS randomized trial assessed the efficacy of a comprehensive cardiovascular screen versus standard care in > 45,000 Danish men. Investigators found that screening was associated with a significant 11% reduction in mortality in 65- to 69-year-olds. Over a 5-year follow-up, prescriptions of antiplatelet and lipid-lowering agents significantly increased in the intervention arm. No differences in the outcomes in men aged > 70 years were observed, thus implying that the target age for screening should be < 70 years old. The eBRAVE trial utilized conventional smartphones to double the detection of treatment-relevant atrial fibrillation in older adults versus traditional care. Digital screening with a certified app more than doubled the rate of atrial fibrillation detection in both the randomization and cross-over phases of this trial. Both studies confirm that screening identified CVD earlier than traditional methods and may lead to better treatment outcomes.Artificial intelligence (AI) offers the potential to enable better CVD diagnosis and management. Research findings have been promising, although questions remain regarding limited clinical implementation to date. During a keynote ESC presentation, Professor Brian Ference  presented the novel CAUSAL AI algorithm. CAUSAL AI combined deep and machine learning to accurately estimate the lifetime risk of major cardiac events among assessed participants when embedded into the JBS3 risk score, a standard tool used to estimate CVD risk. This algorithm more precisely predicted the benefit of lowering low-density lipoprotein (LDL) cholesterol, systolic blood pressure (SBP), or both on atherosclerotic CVD events in multiple clinical cohorts. These benefits started at any age and extended for any duration, with further analyses validating these original findings. CAUSAL AI’s findings agree with evidence from previous trials and Mendelian randomization studies, which stipulate that lowering LDL and SBP levels reduces the risk of atherosclerotic CVD. This new algorithm can improve existing risk prediction scores, which currently do not account for the causal effect of LDL and SBP in humans. Neither have risk scores accurately estimated baseline cardiovascular risk or the benefit of lowering LDL or SBP in atherosclerotic CVD.While CAUSAL AI must be peer-reviewed and subsequently investigated in real-world populations, algorithms that learn the effects of modifiable disease causes have the potential to benefit many in society. It opens the door to transformative changes in primary prevention. That is “providing the essential information needed to inform individual treatment decisions about the optimal timing, duration, and intensity of lowering LDL and SBP to optimally prevent cardiovascular events for each person,” predicted Prof. Ference in his presentation. Extrapolating such AI into personalized medicine programs promises to provide actionable information to guide clinical decision-making. These precision algorithms could be targeted to at-risk individuals in early and mid-adulthood to minimize their likelihood of chronic disease progression. In the near future, these groups might hypothetically be prescribed specific lifestyle changes via smartphone apps to reduce their risk profile and likelihood of developing CVD.Knowledge of likely successful preventative actions is a powerful tool to prevent CVD. But how does one then translate this knowledge into actionable and efficacious strategies? Well-established knowledge should be applied to elicit meaningful benefits, such as clinicians prescribing exercise as medicine for patients. Measures known to assist healthy living must be promoted and not scrapped, such as the UK Soft Drinks Industry Levy, currently under governmental review. Moreover, encouraging innovations like screening and predictive algorithms should be funded and trialed to determine if they can benefit whole populations. Only then could societies imagine alleviating disease burden and healthcare costs.


Photoacoustic remote sensing has been recently developed as an all-optical imaging modality capable of imaging a variety of endogenous contrast agents label-free. Initially predicted laser pulse-induced refractive index perturbation-based interrogation beam reflectivity modulations have been found to be orders of magnitude smaller than those typically observed experimentally. In this report we utilize a 10 million frames-per-second camera to further investigate these predicted reflectivity modulations, while also exploring other potential mechanisms of laser pulse-induced reflectivity modulations. Laser-induced motion is demonstrated both laterally for gold wires suspended and submerged in air and water, respectively, and carbon fibers submerged in water, and axial motion is observed in gold wires submerged in a depth gradient of intralipid solution. This laser-induced sample motion is anticipated to cause reflectivity modulations local to the interrogation beam profile in microscopy set-ups. Non-motion-based maximum intensity modulations of 3% are also observed in gold wires submerged in water, indicating the presence of the originally predicted reflectivity modulations. Overall, these observations are important as they provide a widefield view of laser-pulse interactions unavailable in previous point scanning-based photoacoustic remote sensing microscopy configurations, where observed mechanisms occur on time-scales orders of magnitude faster than equivalent field of view point scanning capabilities. Recently, we developed photoacoustic remote sensing (PARS) microscopy, an all-optical non-contact version of PAM, demonstrating high sensitivity to a wide variety of endogenous chromophores5 and ready integration into existing technologies due to its all-optical architecture10. In comparison to PAM, PARS microscopy utilizes an additional interrogation beam to probe reflectivity modulations rather than directly detecting generated acoustic waves using an ultrasound transducer. Without a priori knowledge of the underlying mechanism, early work strove to explain the intensity modulations observed when pulsed laser light was absorbed. Initially, a model was described in which reflectivity modulations were attributed to elasto-optic refractive index perturbations arising due to photoacoustic initial pressure transients following excitation pulse absorption2. However, in practice predicted reflectivity modulations have been found to be orders of magnitude smaller than those typically observed experimentally, and other recent work has suggested the presence of laser-induced cavitation, which may affect reflectivity11. In this report, a 10 million frames-per-second (fps) camera is utilized to investigate other potential mechanisms of laser pulse-induced reflectivity modulations, with the hypothesis that these modulations may be additionally attributed to laser-induced sample motion or surrounding media displacement local to the detection beam profile.Photoacoustic microscopy (PAM) generates absorption-contrast imagery by tuning an excitation wavelength to match chromophore absorption peaks and using an ultrasound transducer to detect acoustic waves generated via the photoacoustic effect following excitation pulse absorption13. Nanosecond pulsed laser-induced bubble interactions with micro-particles have also been observed using high-speed cameras at up to 540,000 fps, describing particle dynamics in relation to proximity to the bubble center. In closest proximity, rapid ejection of the particle occurred along with the formation of a cavity behind the particle. Lower proximity resulted in particle repulsion that accompanied bubble expansion, followed by attraction of the particle during bubble collapse15. Since laser pulse absorption can result in pressure wave generation via the photoacoustic effect in the form of ultrasound and/or shockwaves, induced cavitation16 may also interact with particles, with observations made at up to 1 Mfps18. Additionally, microjets may form at boundaries19 or even following shockwave-induced bubble collapse20, exerting forces on nearby targets. These small volume liquid jets have even been utilized in microfluidic cell sorting applications to move cells between channels21. Laser-induced ablation recoil momentum has been demonstrated to cause sample motion through ejection of ablated material opposite to the laser irradiation direction22. Using cantilever dynamics and nanosecond pulsed laser fluences of up to 1400 J/cm23. For immobile samples, 70-ns pulsed Er:YAG laser irradiation with up to 5.4 J/cm24. Optical tweezers can be used in the manipulation of small dielectric particles exploiting photon gradient forces, with radiation pressure also exerting a force in the direction of beam propagation. These forces however are very small, typically being on the order of piconewtons25. Alternatively, acoustic radiation force, describing the force exerted on targets by acoustic waves, has been shown to induce micron order displacements in tissues26. This effect has also been explored in the form of photoacoustic radiation force on microbubbles27. Photoacoustic/photothermal tweezers have recently demonstrated target manipulation within absorbing medium with theoretical forces on the order of nanonewtons, with the caveat that the technique relies on the generation of acoustic and thermal gradients which may induce many physical phenomena responsible for observed motion, requiring further exploration28. This example in particular is very compelling, displaying the complexity and range of mechanisms that may follow laser pulse absorption. Together with refractive index perturbations, one or more of these mechanisms may also contribute to observed reflectivity modulations in PARS microscopy explaining the magnitude of reflectivity modulation measurements.A literature review finds several different mechanisms in which laser-induced sample motion is generated. Phase-sensitive optical coherence tomography (OCT) has recently shown that thermoelastic expansion following 5-ns pulsed excitation absorption can generate deformations with axial displacements on the order of several hundred nanometers for a 29, the orders-of-magnitude higher frame rate in this work enables observation of motion dynamics on the sub-microsecond scale. Additionally, we demonstrate that the magnitude of observed intensity modulations arising potentially due to refractive index-based perturbations are significantly weaker than the magnitude of observed motion in samples, providing further evidence for the hypothesis of motion-based reflectivity modulations. The manuscript is laid out as follows: details are provided pertaining to system characterization; results of non-motion-based intensity modulations are described; lastly, observations of laser-induced sample motion in air and in liquid are shown with discussion as to the potential underlying mechanisms responsible.To investigate the presence of sample motion in PARS microscopy, a detection scheme utilizing widefield parallelized 10 Mfps readout was combined with laser pulse excitation fluence levels comparable to those reported in previous PARS microscopy set-ups. Compared to previous experiments which were only capable of observing a singular post pulse excitation frame at a synchronized 10 fps readoutFirst we characterized the lateral resolution of our system. A 1951 USAF resolution test target was imaged with our system and the denoted line-pair widths were used to determine an effective pixel size of 5 2.The system was next utilized to explore widefield parallelized camera-based detection of intensity modulations following laser pulse excitation. Despite best efforts, laser pulse excitation leakage in the form of diffuse scattering from the target into the camera sensor was unavoidable due to the strong angle of incidence dependence of the notch filter used. To address this leakage, three acquisitions were taken sequentially of 25 To investigate potential laser-induced sample motion in air, a vertical 25 We next imaged a target consisting of a 7 10. It should be noted that when submerged in water there appeared to be an absence of target ablation possibly due to better stress propagation in water compared to air, this likely explains the reliance on this type of sample preparation in PARS microscopy experiments. In the presence of water, laser-induced bubble formation was observed with bubble dissipation occurring as early as 10 Next, to explore potential laser-induced sample motion in liquid media, 25 Lastly, in an attempt to observe axial motion within the system’s large depth of field, we imaged a target consisting of a 25 In this work, we report the observation of laser-induced motion in a variety of phantoms using widefield parallelized 10 Mfps readout. Targets suspended in tension in air indicated the potential emission of ablated material, which may suggest a laser-induced ablation recoil momentum source to explain the motion. In carbon fibers, restoring forces were also observed in the displacement trajectory demonstrating a similar response to that of an underdamped oscillator. We hypothesize that this tension-induced restoring force may be more prevalent in focused laser pulse experiments where local structural forces should be stronger relative to induced localized motion. Targets submerged in water indicated motion due to laser-induced bubble formation, with the magnitude and velocity of displacement being dependent on proximity to the bubbles or whether bubbles were pre-existing. In closer proximity to the bubble nucleation site, these results also indicated the presence of restoring forces on the phantoms. Nonlinear displacement trajectories observed suggest that we should expect nonlinear reflectivity modulation responses in a point-based PARS microscopy set-up for dominant motion-based mechanisms. Additionally, axial motion was explored using scattering media to generate a depth gradient across the target, with results indicating an intensity change along the target following pulse excitation, which suggests axial motion of the sample or displacement of the surrounding scattering media. Targets submerged in water showing no bubble formation post pulse-excitation were not observed to exhibit any in-plane lateral motion. The absence of apparent laser-induced ablation recoil momentum in these cases may be due to better stress propagation in water compared to air. Weak intensity modulations observed following pulse leakage removal are believed to be due to the refractive index-based reflectivity perturbation mechanism previously reported.30 and laser Doppler vibrometry31, although it should be noted that these methods would only be able to measure axial motion. Moreover, work should also explore the presence of other potential mechanisms such as photoacoustic radiation force, thermal expansion, and cavitation. In particular, in immersion environments, even if bubble nucleation is absent, motion may result from photoacoustic radiation pressure being created on one side of the target where absorption is strong, but not on the other side of the target where there is minimal fluence. Better understanding the underlying mechanisms in PARS may allow more effective detection of laser-induced changes, better informing system design characteristics and sample preparation for improved PARS microscopy imaging. Additionally, the potential for decoupling signals according to their respective source mechanisms may provide further information not currently available in PARS microscopy images.In conclusion, we hypothesize that these observed motion responses, both lateral and axial, may additionally contribute to reflectivity modulations observed in PARS microscopy, with induced sample motion local to the detection beam profile generating a localized reflectivity change. These high-speed camera-based observations are important as they provide a widefield view of excitation pulse interactions unavailable in previous point scanning-based PARS microscopy configurations, where observed mechanisms occur on time-scales orders of magnitude faster than equivalent field of view point scanning capabilities. Future work should investigate the presence of this observed motion in a point scanning-based PARS microscopy system using methods capable of detecting localized sample displacement with high sensitivity, such as phase-contrast OCTFigure In order to ensure that we were able to capture a signal immediately following the excitation event, the following synchronization set-up was used, with a timing diagram shown in Fig. 2, resulting in a fluence of 177.6 mJ/cmIn comparison to our previously reported point-based scanning PARS microscopy, the excitation beam in this set-up is both unfocused and has oblique incidence on the sample. With point-based scanning PARS, in order to image 7 2. Additionally, taking into consideration the Nyquist limit, 10 Mfps camera capture should provide a detection bandwidth of 5 MHz. Compared to the typical photodiode bandwidth of 75 MHz used in previous point-based PARS microscopy systems32, we expect a loss in detectable frequency content which could result in aliasing in the case of detecting high-frequency modulations greater than 5 MHz. However, for the purposes of capturing motion, these high-speed cameras have been used without issue to image frames of high-speed events involving projectiles moving at 33.The high-speed camera uses a FTCMOS2 image sensor with 10-bit monochrome output and is capable of recording at 10 Mfps, with 50 ns exposure time, and 5 Mfps with 110 ns exposure time. When operating at 5 Mfps, the full 100,000 pixels can be used, however, when operated at 10 Mfps, half the pixels are used in a zigzag lattice pixel array. Based on previous PARS microscopy modulation observations, a 10-bit dynamic range is anticipated to be sufficient to detect intensity modulations, previously predicted to be on the order of Supplementary Information.Supplementary Video S1.Supplementary Video S2.Supplementary Video S3.Supplementary Video S4.Supplementary Video S5.


Ribosomally-synthesized cyclic peptides are widely found in plants and exhibit useful bioactivities for humans. The identification of cyclic peptide sequences and their precursor proteins is facilitated by the growing number of sequenced genomes. While previous research largely focused on the chemical diversity of these peptides across various species, there is little attention to a broader range of potential peptides that are not chemically identified.Linum usitatissimum). Phylogenetic analysis clustered the 5 known linusorb precursor proteins into two clades and one singleton. Preliminary tBLASTn search of the published flax genome using the whole protein sequence as query could only retrieve its homologues within the same clade. This limitation was overcome using a profile-based mining strategy. After genome reannotation, a hidden Markov Model (HMM)-based approach identified 58 repeats homologous to the linusorb-embedded repeats in 8 novel proteins, implying that they share common ancestry with the linusorb-embedded repeats. Subsequently, we developed a customized profile composed of a random linusorb-like domain (LLD) flanked by 5 conserved sites and used it for string search of the proteome, which extracted 281 LLD-containing repeats (LLDRs) in 25 proteins. Comparative analysis of different repeat categories suggested that the 5 conserved flanking sites among the non-homologous repeats have undergone convergent evolution driven by functional selection.A pioneering study was initiated to explore the genetic diversity of linusorbs, a group of cyclic peptides uniquely occurring in cultivated flax (The profile-based mining approach is suitable for analyzing repetitive sequences. The 25 LLDR proteins identified herein represent the potential diversity of cyclic peptides within the flax genome and lay a foundation for further studies on the functions and evolution of these protein tandem repeats.The online version contains supplementary material available at 10.1186/s12864-022-08735-x. Plant secondary metabolites play important roles in plant development, mediating communication with other organisms, responding to biotic stresses, and acclimating to the abiotic environment . While pOldenlandia affinis (Violaceae) and Lycium barbarum (Solanaceae). A wide range of biological activities have been reported in cyclic peptides, including enzyme (trypsin) inhibition, immunosuppression, anti-inflammation, antibacterial activity and antiviral effects [One typical form of RiPPs are cyclic peptides, which are derived from linear precursor proteins which are cyclized by specialized proteases Fig. . Well ch effects –10. It i effects –13.Fig. Lycium barbarum) [Since the isolation of the first cyclic peptide, cyclolinopeptide A, from the sediment of flaxseed oil in 1959 , more tharbarum) . SelectiComparative genomics analyses identified potential orthologues of individual precursor proteins, although it has been shown that precursor proteins embedded with cyclic peptides of the same structural class (cyclotides) can have different (i.e. non-homologous) biosynthetic origins . These fViola tricolor using a set of published cyclotide sequences as queries, which revealed 108 transcripts with homology to the cyclotide sequences [Viola species [Another potential limitation lies in the chemical verification of predicted peptides. Hellinger et al. 2015) conducted a tBLASTn search in the transcriptome of equences . Of thes15 conducLinum usitatissimum L., 2n = 2x = 30). Flax is one of the most ancient crops widely grown as a source of fiber, oil, and health products. More than 20 orbitides have been isolated from flaxseed and are collectively named linusorbs [In this study, we developed proteome mining methods to explore the genetic diversity of orbitides in flax  an iterative search for homologous sequences of linusorb-embedded repeats using algorithm-derived profiles, and (2) a string search for sequences matching profiles composed of conserved flanking sites. Both strategies are based on different assumptions and provide unique perspectives for exploring linusorb-like sequence diversity.Iterative searches can efficiently identify sequences with varying homology. Such searches begin with the generation of profiles from the set of linusorb-embedded repeats which are then used as queries to search the proteome for similar sequences. Significant matches are combined to update the profile which is then used in the next round of search. The search is iterated until no more new sequences are found within the significance threshold. Two models were used to describe profiles. The Position Specific Score Matrix (PSSM) is based on the frequencies of each residue in a specific position of a multiple sequence alignment and is used as query in the Position Specific Iterative BLAST (PSI-BLAST) search. The second is a probabilistic “profile hidden Markov model” (profile HMM) trained from a multiple alignment and used as query to search a sequence database using the HMMERsuite. The mining results of both iterative search methods are as follows:− 10 and 1e− 4, respectively. The third and fourth matches are G3-449N (g53356) and G4-136N (g33422) with E-values of 0.006 and 0.007, respectively. The reason for this order of matches is that the first sequence in the multiple alignment  within one site and across adjacent sites in the alignment, while PSSM assumes all sites are independent of one another . A profiats Fig.  was usedns Data S. Of the Within the 8 protein matches there are 58 motifs significantly matching the profile HMM, as the E-values are lower than both per-sequence and per-domain inclusion thresholds Data S. HMMER chmmsearch are members of the identified repeats, and thus are potential homologues of the 12 linusorb-embedded repeats in Fig. All 58 motifs retrieved by As revealed in the above two algorithm-based searches, potential homologues of the linusorb-embedded repeats are characterized of (1) repetitiveness, (2) variable LLDs and (3) conserved flanking residues. The conservation of flanking sites has been reported to play a vital role in the recognition by modification enzymes during the biosynthesis of cyclic peptides in other species , 33–38. We thus developed a mining method focused on flanking site conservation by generating 26 profiles to characterize an LLDR Table S. These 22 = 0.83) in the proteome and exponentially (R2 = 0.70) in the ORF library  of the matching strings. The extracted proteins containing no fewer than 3 matches were parsed by repeat finder RADAR to determine if the matching strings exhibit repeat patterns. As a result, in addition to sequences that matched the 4 known repeat-containing linusorb precursor proteins, 25 new proteins were detected to contain a total of 281 profile-matching repeats and thus were identified as LLDR proteins. For easier tracking the locations of LLDR encoding genes in the genome, the 25 LLDR proteins were denoted the format “Lu (chromosome number)-(accession number of predicted gene)”, e.g. Lu1–17,106. Names of the 4 linusorb precursor proteins remain unchanged for consistency with published literature. Among them only 10 were identified by the previous genome annotation . This protein was neglected in the profile-based proteome mining because it contains fewer than 3 repeats. Nonetheless, the fact that proteome mining based on leader peptide homology found potential homologues of repeats in the core peptide region suggests strong correlation between the leader peptide and the core peptide region in these LLDR proteins. Conservation of both the leader peptide and flanking signatures further suggests that these LLDR proteins may undergo similar post-translational modification to the linusorb precursor proteins.There are 4 hits with larger E-values, among which Lu1–18,055 (g18055) and Lu3–56,299 (g56299) are members of the 25 LLDR proteins. The other 2 hits, g39692 and g11605, were analysed by RADAR for the presence of any repeats. Only g39692 contains repeats, and importantly, the 2 repeats match our customized profile: Since the advent of next-generation sequencing, omics database mining has been driving new approaches to the discovery of natural products, especially RiPPs because of their direct link between the gene sequences and the peptide products. However, the diversity of RiPPs poses a challenge for mining to predict RiPPs-encoding sequences. Previous mining practices have largely focused on identifying potential homologues of cyclic peptide precursor proteins in closely related species. Numerous peptides have been identified by MS analysis, but a larger majority of mining candidates have not been validated through chemical identification of products. Little attention has been paid to candidates identified by omics with no corresponding product or the relationship between these sequences and the detected ones. Most mining candidates have been retrieved by employing search tools like BLAST based on their high similarity with a known protein sequence, and thus contain genetic information related to the biosynthetic mechanisms and evolution of cyclic peptides. Here we employed bioinformatics tools to explore the genetic diversity of linusorbs, the flaxseed orbitides. Each search tool identified patterns from different regions of the linusorb precursor proteins, which resulted in the detection of novel linusorb-like sequences. The collection of linusorb-like sequences provides a diverse pool for subsequent studies on the evolutionary relationship to the known linusorb sequences and expands the candidate pool of potential linusorbs that can be synthesized in engineered organisms for practical applications.Theoretically, genetic diversity and chemical diversity of RiPPs are related to each other in that every RiPP a plant produces is encoded by a gene sequence; however, the link between the gene and the peptide has proven to be weak. Several factors, both biological and technical, limit the diversity of peptides that can be identified chemically. From the biological perspective, peptide biosynthesis involves expression of a protein as a precursor for post-translational modification by proteolytic enzymes. The production of peptides depends on regulation of the precursor protein expression , the activity of modification enzymes and many other factors associated with downstream metabolic pathways. Consequently, only those minimally modified peptides resemble the primary sequences the most and are readily detectable, representing a subset of all possible peptide sequences, as the discovery of peptides and their variants largely relies on prior knowledge of their physicochemical properties.To achieve unambiguous identification, analytical processes which involve extraction of crude peptides, separation by liquid chromatography (LC) and determination by mass spectrometry (MS) need to be customized for a specific class of peptides based on their molecular weight, polarity, linkage moiety and other structural information, most of which is obtained from empirical studies of known peptides. This is a bottleneck for identifying novel peptides with unknown modifications which might require special treatment for successful analysis. Undetected peptides might share the same precursor proteins or their homologues with the detected peptides, underlying useful clues about cyclic peptide evolution. Elucidating biological mechanisms solely from those chemically validated peptides may result in ascertainment bias. Therefore, while chemical diversity based on product validation provides solid evidence, the conserved patterns from the known peptide sequences are more important for exploring a greater diversity of genetically relevant sequences to reveal a bigger picture at the omics level.The currently identified > 20 linusorb structural variants are derived from 11 linusorb domains in 5 precursor proteins . The disThe limitation of sequence similarity searches for local sequence alignment with the whole precursor protein query calls for a more specific mining strategy. Repeat detection and analysis of the core peptide region with respect to multiple orbitide-embedded repeats provides an informative approach for resolving the relationships of the 5 linusorb precursor proteins. Alignment of repeats revealed the hypervariability of linusorb domains flanked by several highly conserved sites. Such a uniform pattern of linusorb-embedded repeat units across different precursor proteins has fostered a profile-based mining strategy. We compared 3 search methods that employ different algorithms of profile recognition to perform proteome mining for profile-matching sequences. While different methods retrieved different matches, all linusorb precursor proteins present in the predicted proteome were recovered by all 3 methods, highlighting the power of a profile-based mining strategy to identify distantly related proteins. Among the 2 algorithm-based search methods, HMMER extracted 8 new proteins in which 58 repeats significantly matched the profile HMM built from linusorb-embedded repeats, while PSI-BLAST found only 2 of the 8 proteins as matches with large E-values. The recovered sequences contrasted due to differences in profile construction and match scoring between both tools. Although non-significant matches do not meet the significance threshold, many of them contain sites matching the conserved sites in the input alignment. Additionally, using aligned repeats as query achieved higher E-values compared to using the whole protein as query due to the much shorter length than protein. A previous study on the repeat-containing kinetochore protein scaffold KNL1 revealed that classical homology searches using BLAST failed to detect sufficient homology for KNL1 genes, and demonstrated the power of iterative homology search using HMMER that found 110 diverged homologues in a variety of eukaryotic species . The KNLBoth PSI-BLAST and HMMER build profiles from the alignment of linusorb-embedded repeats, meaning that any conserved sites in the linusorb domain will contribute to the profile. To maximize the variability of the linusorb domain, we developed customized profiles using the 5 most conserved sites flanking the linusorb domain represented by 4–13 random residues and wrote Python scripts to search the proteome for strings matching the profile. Screening the primary matches for those containing multiple repeats identified 25 LLDR proteins with 281 profile-matching repeats. The 8 proteins with 58 potential homologues of the linusorb-embedded repeats are a subset of the total collection. This profile-based string search method is one step further from HMMER by extracting a wider range of repeats that are distantly related to the linusorb-embedded repeats. The randomization of the linusorb domain region expands the collection of orbitides flax can potentially produce, from which we can gain more insights into the genetic relationship among these sequences.et. al. (2017) employed two search tools, BLAST and PROSITE, to search the transcriptomes of various Viola species using known cyclotide sequences as queries [Viola genus, this mining process could extract sequences like the known cyclotides in other Viola species, enabling the reconstruction of the evolutionary relationships of orthologues among different species at the cyclotide level. However, this approach cannot identify cyclotide-like domains with low similarity to known cyclotides but anchored by the 6 conserved cysteines. This contradicts their assumption that the sequence is a cyclotide precursor if the sequence contains the 6 conserved cysteines aligned with previously known cyclotides. Therefore, it is imperative to identify a specific goal and design a cohesive mining scheme towards the goal.Some of the published mining practices also appear to be profile-based, especially for cyclotides, for which the structures have been well studied. Park et. al. 17 employOne of the structural features shared by many cyclic peptide precursor proteins is the repeats that constitute the core peptide region. These repeats have high sequence similarity and thus are identified as low-complexity sequences like those non-coding repeats. As it is a routine treatment to mask the repetitive elements in many genome assembly practices , a consiAnother treatment specific for repeats is to dissect them into individual repeat units and align them across different precursor proteins, due to their varying copy number and length from protein to protein. However, few studies have performed this treatment before their mining experiments. The potential relationships among the repeats could not be resolved until they were identified by the repeat finder RADAR and properly aligned. In terms of mining, using the entire protein as query failed to differentiate the hypervariable linusorb domain from the conserved flanking sites. The short length and discrete conserved sites interspersed by variable lengths of hypervariable domains hinder BLAST from accurately finding homologues with different numbers of repeat units , 44. TheOur profile-based mining approach identified 25 LLDR proteins that contain 281 LLD repeats. These repeats share a structural profile with the linusorb-embedded repeats where a variable domain is flanked by 5 conserved sites. From an evolutionary perspective, it would be interesting to know whether these conserved sites originate from shared ancestry or are resulted from convergent evolution . The HMM search result has shed light on this question. Among the 281 LLD repeats, 58 exhibit significant sequence homology to the linusorb-embedded repeats, implying that they share common ancestry. However, the 8 LLDR proteins containing these 58 LLD repeats are unrelated to the 5 linusorb precursor proteins, as tBLASTn found no additional matches in the genome with similarity comparable to the linusorb precursor proteins to themselves  tree was constructed with 1000 bootstrap replications using MEGA7 . MidpoinL. usitatissimum cv. CDC Bethune (GenBank accession: GCA_000224295.2) with tBLASTn. The substitution matrix was BLOSUM62. The word size was set to 2 and the expect value  threshold was 10 by default. Low-complexity filtration was disabled to ensure the whole protein sequence was queried.The 5 known linusorb precursor proteins were used as queries to search the genome assembly of The genome annotation conducted by Wang et al.  removed The eukaryotic genome annotation pipeline, BRAKER2 , was utiAn iterative search was performed with the tool PSI-BLAST to mine the proteome generated from genome reannotation for potential homologues of the linusorb-embedded repeats. First, a position specific scoring matrix (PSSM) was generated from the multiple alignment of linusorb-embedded repeats in the original 5 precursor proteins. Then a PSI-BLAST search was conducted to identify sequences matching the PSSM input. Matches with E-values below 0.01 were accepted and combined with the original alignment for the next round of search until no new matches were identified.hmmbuild tool with default parameters in the HMMER suite [hmmsearch was conducted to search the BRAKER2-predicted proteome with the input profile HMM. Statistically significant protein matches were taken above the default inclusion threshold . Hits marked with “!” were determined as significant by HMMER, and the subject segments of these hits were considered as “true” homologues. These homologous subject segments were then extracted from the output file and aligned to the original alignment using hmmalign based on the input profile. The combined alignment was used to build a new profile HMM for the next round of search. Such searching process was iterated until no additional proteins were identified that met the inclusion threshold.Proteins containing repeat sequences that are potential homologues of the linusorb-embedded repeats were identified based on the profile HMM match. First, a profile HMM was built from the alignment of original linusorb-embedded repeats using the ER suite . Next, hBased on conserved sites in the alignment of linusorb-embedded repeats, sequence profiles were created in the form of regular expressions that can be interpreted by Python, whereby a script was developed to automate proteome mining. Accessions containing sequence strings that matched the specified regular expression were extracted to a primary candidate pool. The primary candidate protein pool was refined using a Python script to enumerate candidate protein matches and screen for those with multiple (≥ 3) matches. The refined candidate pool was then filtered by detecting matching strings that exhibit repeat pattern using RADAR . FinallyWebLogo3 was employed to create sequence logos from the multiple sequence alignments of the following 3 datasets : (a) theAdditional file 1: Fig. S1. Phylogenetic trees of (a) 11 linusorb (LO) domains and (b) 5 linusorb precursor proteins constructed with the Maximum Likelihood (ML) method.Additional file 2: Data S1. BLAST search for potential homologues of 5 precursor proteins.Additional file 3: Data S2. RADAR-detected repetitive motifs in linusorb precursor proteins.Additional file 4: Data S3. Repeat PSI-BLAST output.Additional file 5: Data S4. Repeat HMM search output.Additional file 6: Data S5. Leader peptide HMM search outputAdditional file 7: Table S1. Statistics of 26 profiles


Autoantibodies are increasingly recognized for their pathogenic potential in a growing number of neurological diseases. While myasthenia gravis represents the prototypic antibody (Ab)-mediated neurological disease, many more disorders characterized by Abs targeting neuronal or glial antigens have been identified over the past two decades. Depletion of humoral immune components including immunoglobulin G (IgG) through plasma exchange or immunoadsorption is a successful therapeutic strategy in most of these disease conditions. The neonatal Fc receptor (FcRn), primarily expressed by endothelial and myeloid cells, facilitates IgG recycling and extends the half-life of IgG molecules. FcRn blockade prevents binding of endogenous IgG to FcRn, which forces these antibodies into lysosomal degradation, leading to IgG depletion. Enhancing the degradation of endogenous IgG by FcRn-targeted therapies proved to be a powerful therapeutic approach in patients with generalized MG and is currently being tested in clinical trials for several other neurological diseases including autoimmune encephalopathies, neuromyelitis optica spectrum disorders, and inflammatory neuropathies. This review illustrates mechanisms of FcRn-targeted therapies and appraises their potential to treat neurological diseases.The online version contains supplementary material available at 10.1007/s13311-021-01175-7. A growing spectrum of neurological disorders is characterized by disease-associated immunoglobulin G (IgG) autoantibodies targeting structures of the central or peripheral nervous system as well as the neuro-muscular junction Table . These dThe neonatal Fc-receptor (FcRn) is a major histocompatibility class I–related receptor responsible for the transfer of humoral immunity from the mother to the newborn . ThroughFcRn was first cloned in 1994 . StructuFunctionally, β2m knockout mice with impaired FcRn had reduced IgG levels after birth . FurtherIgG and albumin bind to FcRn at distinct sites under acidic but not neutral pH conditions , 13. FcRExpression of FcRn was recorded for numerous cells and tissues. In human tissues, expression was detected including but not limited to the placenta , spleen + T cell stimulation by APC, providing a novel mechanism by which FcRn controls immune processes ) induce myasthenia gravis, i.e. a chronic autoimmune disease characterized by weakness of skeletal muscles, which can be generalized or involve only few muscular units, often including the extraocular muscles, with diplopia and ptosis. The weakness typically increases with muscle use (fatigability) and fluctuates during the day and from day to day. Although the disease can initially be localized, for instance in the purely ocular forms, within the first 2 years, it becomes generalized in more than 80% of the patients . OverallThe main therapeutic strategies for myasthenia gravis include (1) symptomatic treatment, which aims to potentiate the neuromuscular transmission through the use of acetylcholinesterase inhibitors, and (2) immunosuppressive therapy  to induce a durable remission of symptoms over time. Given that the thymus is considered to play a key role in inducing AChR Ab production, total thymectomy is recommended not only in patients with myasthenia gravis and thymoma, but it has been shown to be beneficial even in those without thymoma , 73.Based on the established efficacy of plasmapheresis (PLEX) or IVIg for amelioration of several Ab-mediated diseases, harnessing FcRn might allow to elegantly achieve comparable efficacy while avoiding adverse effects and reducing supply issues. Indeed, therapeutic Abs targeting FcRn, commonly referred to as Abdegs (ab that enhance IgG degradation), are effective at reducing serum IgG levels . Abdegs Abdegs are under investigation for a number of autoimmune conditions, most notably MG, neuromyelitis optica spectrum disorder (NMOSD) or chronic inflammatory demyelinating polyneuropathy (CIDP).We will first dissect the clinical landscape of FcRn targeted therapies in the context of MG as a model disease for Ab-mediated autoimmunity. MG constitutes the major autoimmune disorder affecting the neuromuscular junction. The pathophysiological hallmark of MG are pathogenic IgG Ab directed against structures of the postsynaptic membrane . In receIndeed, Abdegs are under investigation in preclinical as well as in clinical studies for treatment of MG. An overview over current FcRn therapies is given in Table Given the promising effects observed for MG, indications for Abdegs are likely to evolve. Besides MG, the phase II ADHERE trial (NCT04281472) investigates Efgartigimod for treatment of CIDP, including 400 participants in a randomized, placebo-controlled setting. Similarly, Rozanolixizumab (NCT03861481) and the fully human anti-FcRn mAb Batoclimab are under investigation in clinical trials for treatment of CIDP. Abdegs might provide a novel mechanism capable of ameliorating Ab-mediated disease while curtailing costs and risks associated with standard therapies. Nonetheless, long-term outcomes of IgG depletion remain unknown and will likely shape our understanding of FcRn modulation in the future.A potential drawback of Abdeg technologies is the unselective depletion of IgG, including those mediating host defence. To overcome this caveat, research teams engineered a novel class of agents that selectively clear antigen-specific Abs by exploiting the FcRn mechanism . IndicatThe FcRn mechanism could be used therapeutically in two ways: (i) to reduce the half-life of pathological antibodies or (ii) to increase the half-life of therapeutic antibodies. Abdeg and Seldeg technologies target the first mechanism. Many therapeutic mAbs have proven efficacious for a number of neurological indications, i.e. Eculizumab inhibits complement activation in MG by inhibiting C5 , while TApproximately two-thirds of NMSOD patients have detectable IgG Abs against aquaporin-4 (AQP-4) . IL-6 leFor MG, Eculizumab has proven effective for treatment-refractory disease . HoweverRemoval of serum proteins by PLEX and even by immunoadsorption is unselective, while FcRn-targeted therapeutics are confined to reduction of IgG and albumin, possibly providing fewer off-target effects. As of now, safety concerns of Abdegs are focused on the consequences of IgG depletion . ClinicaNeisseria meningitidis or Streptococcus pneumoniae, are resolved by the innate immune response [However, it is important to note that there is a lack of long-term studies investigating effects of IgG depletion. Given the mechanism of action, infectious complications arising from IgG deficiency remain the primary concern for treatment. To better understand long-term outcomes, we may consider IgG depletion as a type of secondary immunodeficiency (SID) . As opporesponse , with Abresponse . In contresponse , 104. ThBesides, FcRn function is evolving across human tissues. As such, FcRn is expressed in the microvascular epithelium constituting the blood–brain-barrier (BBB) , 106. ThTargeting the FcRn is a novel and promising approach for the treatment of a number of Ab-mediated neurological diseases due to selective IgG depletion and can additionally be used to extend the half-life and efficacy of therapeutic mAbs. FcRn blockade is particularly intriguing as it does not result in general immune suppression, in contrast to many conventional therapies in routine clinical use, while the long-term safety of recurrent IgG depletion cycles remains to be addressed. As IgG molecules are the preeminent effector proteins of the immune system and recruit and activate leukocytes through Fc interactions with Fc receptors (FcRs) expressed by innate immune cells and B cells, the removal of pathogenic IgG molecules by FcRn targeted therapies likely affects both humoral and cellular immunity. Along these lines, a third of patients with gMG positive for AChR Abs who responded to efgartigimod in experienced clinical improvement that lasted more than 12 weeks [Supplementary file1 (DOCX 34 KB)Supplementary file2 (DOCX 34 KB)Supplementary file3 (DOCX 34 KB)Supplementary file4 (DOCX 35 KB)Below is the link to the electronic supplementary material.


Sensory loss leads to widespread adaptation of neural circuits to mediate cross-modal plasticity, which allows the organism to better utilize the remaining senses to guide behavior. While cross-modal interactions are often thought to engage multisensory areas, cross-modal plasticity is often prominently observed at the level of the primary sensory cortices. One dramatic example is from functional imaging studies in humans where cross-modal recruitment of the deprived primary sensory cortex has been observed during the processing of the spared senses. In addition, loss of a sensory modality can lead to enhancement and refinement of the spared senses, some of which have been attributed to compensatory plasticity of the spared sensory cortices. Cross-modal plasticity is not restricted to early sensory loss but is also observed in adults, which suggests that it engages or enables plasticity mechanisms available in the adult cortical circuit. Because adult cross-modal plasticity is observed without gross anatomical connectivity changes, it is thought to occur mainly through functional plasticity of pre-existing circuits. The underlying cellular and molecular mechanisms involve activity-dependent homeostatic and Hebbian mechanisms. A particularly attractive mechanism is the sliding threshold metaplasticity model because it innately allows neurons to dynamically optimize their feature selectivity. In this mini review, I will summarize the cellular and molecular mechanisms that mediate cross-modal plasticity in the adult primary sensory cortices and evaluate the metaplasticity model as an effective framework to understand the underlying mechanisms. Cross-modal plasticity refers to changes in brain function following a sensory loss that allows the spared senses to be used more effectively to guide behavior. There are two main changes: cross-modal recruitment of the brain areas that serve the lost sensory modality and compensatory plasticity of the brain areas that process the spared senses . Cross-mIn animal models, early sensory loss results in cross-modal anatomical changes to the primary sensory cortices, especially of thalamocortical inputs , but in via LTD, but in the opposite direction that ultimately leads to inactivity. Such bistable property of Hebbian plasticity on neural network activity suggests a need for homeostatic control to provide stability. The sliding threshold model allows such homeostatic control by postulating that the induction threshold for LTP and LTD, referred to as the synaptic modification threshold (θm), slides as a function of the integrated past activity of a neuron  has been shown to occur with neuromodulators acting on LTP/LTD expression mechanisms  are likely a major substrate for adult cross-modal plasticity. L2/3 synapses retain their ability to undergo LTP and LTD as well as homeostatic synaptic plasticity into adulthood . This coEven a short duration of sensory loss impacts normal development , but theKey distinguishing features of synaptic scaling and sliding threshold model are dependence on activity, especially of NMDARs, and input-specific nature of plasticity. Inactivity-driven synaptic scaling was first demonstrated in neuronal cultures upon pharmacological blockade of activity using tetrodotoxin (TTX) or AMPAR blockers . Subsequvia LTP. This allows V1 neurons, devoid of visual inputs, to adopt these newly potentiated inputs as their main driver. Inputs with activity below the new θm will still undergo LTD, which permits V1 neurons to maintain selectivity to the newly adopted inputs. Therefore, the sliding threshold model enables neurons to dynamically adopt new features based on the changes in the landscape of sensory experience. Particularly for cross-modal plasticity, the metaplasticity model allows V1 neurons to respond to non-visual activity carried by the newly potentiated inputs. This could serve as a substrate for cross-modal recruitment observed in blind subjects  receptor; NR2A, NMDAR 2A subunit; NR2B, NMDAR 2B subunit; PLC, phospholipase C; TRN, thalamic reticular nucleus; TTX, tetrodotoxin; V1, primary visual cortex; θm, synaptic modification threshold.H-KL synthesized and conceptualized the ideas and wrote the manuscript.


Furthermore, tensile properties were shown to withstand the physiological mechanical stresses and strains. In vitro characterization, using L929 mouse fibroblasts, demonstrated that the multilayered scaffold is a suitable platform for cell infiltration and proliferation from the innermost to the outermost layer as is needed for vascular wall regeneration. Our work holds promise as a strategy for the low-cost manufacture of next-generation polymer-based hierarchical scaffolds with high bioactivity and resemblance of ECM’s microstructure to accurately guide cell attachment and proliferation.Fabrication of scaffolds with hierarchical structures exhibiting the blood vessel topological and biochemical features of the native extracellular matrix that maintain long-term patency remains a major challenge. Within this context, scaffold assembly using biodegradable synthetic polymers (BSPs) via electrospinning had led to soft-tissue-resembling microstructures that allow cell infiltration. However, BSPs fail to exhibit the sufficient surface reactivity, limiting protein adsorption and/or cell adhesion and jeopardizing the overall graft performance. Here, we present a methodology for the fabrication of three-layered polycaprolactone (PCL)-based tubular structures with biochemical cues to improve protein adsorption and cell adhesion. For this purpose, PCL was backbone-oxidized (O-PCL) and cast over a photolithography-manufactured microgrooved mold to obtain a bioactive surface as demonstrated using a protein adsorption assay (BSA), Fourier transform infrared spectroscopy (FTIR) and calorimetric analyses. Then, two layers of PCL:gelatin (75:25 and 95:5  Cardiovascular diseases represent one of the main causes of death in the world. About 8.5 million people worldwide have diseased blood vessels that must be replaced with vascular grafts ,2. CurreTo overcome the limitations of the currently available alternatives, the tissue-engineering field is a strategy for the implantation of a hemodynamic-responsive conduit able to maintain long-term patency . TherefoNevertheless, the lack of surface bioactivity of most BSPs limits their further applicability to promote cell adhesion, growth, and proliferation . For ins2 and NO2 plasma to improve hydrophilicity and protein adsorption as well as conjugate bioactive molecules ..−1 , (111), and (200) of the crystalline phase of PCL. Furthermore, a halo centered near 21° in the XRD profile indicates the presence of amorphous structures in the samples as expected from the semicrystalline structure of PCL. Observable differences can be identified in the deconvoluted peak intensities and peak widths among the orthorhombic unit cell diffractions when oxidation takes place, indicating variations on the lamellar organization of the polymer.2 plasma treatment, indicating that the oxidation introduced here is a cost-effective alternative for polyester oxidation  when compared with PCL. Vero cells remained metabolically active with no major changes in cell membrane integrity after 24 and 72 h as demonstrated by cell viability levels above 95% for both MTT and LDH assays , α-helices (1650 cm−1), parallel β-sheets (1630 cm−1), and intermolecular interactions between secondary structures (1610 cm−1) .,34.26,34The circumferentially aligned mesh demonstrated via the FFT analysis  revealedOne of the major challenges of manufacturing vascular grafts is to assure that they not only have geometries that are like those of native vessels but that they mimic their tissue organization and chemistries. This is critical to promote the appropriate biochemical and mechanotransduction signals that lead to controlled cell-surface interactions. The present study was dedicated to addressing this challenge by tuning the topological and bioactive cues of hierarchical tubular structures using a layer-by-layer rational design to assure control over protein adsorption capacity and the subsequent cell adhesion events. This was achieved by incorporating a radial gelatin gradient into the manufactured multilayer scaffold, which was composed of electrospun polycaprolactone (PCL) microfibrils deposited on a microgrooved oxidized-PCL (O-PCL) film. The obtained scaffold showed interconnected pores, which were formed after particle leaching conducted on the PCL microfibril structure. The achieved 3D topology mimicked very closely that of the media and adventitia layers of the blood vessels’ ECM as evidenced by the preferential cell attachment and protein adsorption observed along the intermediate layer. Our PCL-based multilayer scaffold also proved to be a suitable platform for fibroblast cell attachment and proliferation where cells exhibited elongated and spindle-like morphologies, which are typically observed under native conditions. Additionally, the scaffold maintains the basal intracellular ROS production levels of immune cells (THP-1).All in all, our work highlights a relatively simple and cost-effective route to engineer multilayered tubular scaffolds with surface and bulk features that resemble native ECMs. We are certain that this is one step forward to approaching hierarchical structures that provide the physicochemical and mechanical cues to overcome early failure of chemically inert BSP. Future work will focus on studying the interplay of cell phenotypes at the engineered vascular wall to identify possible design improvements to elicit a desired cellular response, attachment and ROS signaling profile.Injerto vascular regenerativo multicapa basado en colágeno, con recubrimiento luminal bioactivo). Patent application number in Colombia: NC2021/0017700, submitted on 22 December 2021. Collagen based multilayered regenerative vascular graft with bioactive luminal coating. (Original in Spanish: