Microbes are routinely engineered to synthesize high-value chemicals from renewable materials through synthetic biology and metabolic engineering. Microbial biosynthesis often relies on expression of heterologous biosynthetic pathways, i.e., enzymes transplanted from foreign organisms. Metallocluster enzymes are one of the most ubiquitous family of enzymes involved in natural product biosynthesis and are of great biotechnological importance. However, the functional expression of recombinant metallocluster enzymes in live cells is often challenging and represents a major bottleneck. The activity of metallocluster enzymes requires essential supporting pathways, involved in protein maturation, electron supply, and/or enzyme stability. Proper function of these supporting pathways involves specific protein–protein interactions that remain poorly characterized and are often overlooked by traditional synthetic biology approaches. Consequently, engineering approaches that focus on enzymatic expression and carbon flux alone often overlook the particular needs of metallocluster enzymes. This review highlights the biotechnological relevance of metallocluster enzymes and discusses novel synthetic biology strategies to advance their industrial application, with a particular focus on iron-sulfur cluster enzymes. Strategies to enable functional heterologous expression and enhance recombinant metallocluster enzyme activity in industrial hosts include: (1) optimizing specific maturation pathways; (2) improving catalytic stability; and (3) enhancing electron transfer. In addition, we suggest future directions for developing microbial cell factories that rely on metallocluster enzyme catalysis. Escherichia coli  cluster; (ii) a catalytic MoFe protein (the α2β2-tetramer NifDK) containing four complex metalloclusters: two M-clusters (or FeMo cofactor), and two P-clusters   hydrogenase (MBH) from Ralstonia eutropha holds a special [4Fe-3S] cluster that enables the enzyme to reduce O2 into harmless water  hydrogenases -hydrogenase-3 (Hyd-3) from its natural electron donor formate. An engineered Hyd-3 covalently attached to a ferredoxin from Thermotoga maritima accepts electron from pyruvate instead, and sustains in vivo hydrogen production when co-expressed with a pyruvate-ferredoxin oxidoreductase (PFOR) [The physical linkage of metalloenzymes with electron carriers can also be employed to engineer alternative electron transfer pathways with improved efficiency. For instance, alternative electron donors have been engineered in e (PFOR) .E. coli. Attachment of a hydrogenase and a ferredoxin on a synthetic protein scaffold improved hydrogen production by three-fold [Since the physical interaction between partner proteins is critical for electron transfer, their spatial organization can be optimized to enhance electron flux through desired metabolic pathways. Optimized spatial organization of electron donor and acceptor proteins increases their chance of physical contact, and reduces cross-talk with other cellular pathways . Moleculree-fold . The useree-fold . Other sree-fold .Eukaryotic organisms, such as yeast or microalgae, are essential hosts in the development of microbial cell factories. As many industrial bioprocesses use eukaryotic microbial hosts, implementing biotechnologically relevant pathways in these organisms is crucial to the large-scale production of chemicals. Unfortunately, attempts of transferring pathways that rely on the activity of bacterial metallocluster enzymes into eukaryotes have encountered limited success. For instance, many bacterial FeS enzymes involved in industrially relevant pathways display little-to-no activity when expressed in yeast, and common synthetic biology interventions have failed to improve their activity . AttemptThe barriers to the functional expression of recombinant metallocluster enzymes in eukaryotic organisms are analogous to those that exist between bacterial species: (i) inefficient maturation and/or electron supply due to incompatibilities between phyla-specific systems; and (ii) oxidative stress. The co-expression of bacterial FeS maturation and reducing pathways in yeast is unfortunately insufficient to enhance enzymatic activity, as demonstrated for the enzymes IspG and IspH . Other sE. coli cell chassis to determine that electron transfer pathways from chloroplasts, but not mitochondria, are capable of supporting bacterial nitrogenase systems [Engineering nitrogen fixation in eukaryotes by expressing bacterial nitrogenases has been a long-sought goal due to the cost of synthetic nitrogen fertilizers; however, nitrogen fixation in plants for self-fertilization has not been achieved due to difficulties in expressing functional nitrogenase systems . Neverth systems . More re systems . This woThe aim of making biology easier to engineer is the central goal of synthetic biology. To this purpose, tools and standardized principles for manipulating DNA have been developed over the past two decades. Genetic engineering technologies have enabled the engineering of a wide range of organisms with novel biological functions, including biotechnologically relevant microbial cell factories. However standard synthetic biology tools focused on genetic manipulation have faced challenges in engineering pathways that rely on many catalytically powerful metalloenzymes. Here, we presented an outline of the obstacles to the functional expression and optimization of heterologous metallocluster enzymes in biosynthetic pathways. We also described novel synthetic biology strategies and investigations that could enable harnessing their full biotechnological potential. Such efforts are focused on overcoming obstacles specific to the functional requirements of catalytic metallocluster enzymes: (i) engineering essential supporting pathways involved in enzyme stability and maturation; and (ii) engineering pathways for electron supply that sustain redox active catalysis. The combination and further development of these strategies could be used to optimize novel microbial cell factories with economically viable yields.However, major challenges to the full exploitation of these engineering approaches remain, as our fundamental knowledge on the biochemistry and barriers to the heterologous expression of metallocluster enzymes is still very limited. Indeed, synthetic biologists cannot easily predict the transferability of a given metallocluster enzyme, nor anticipate what will be required for their activation in heterologous hosts. More studies of phylogenetic compatibility and functional requirements will help develop intuitive rules for harnessing heterologous metallocluster enzymes. To this purpose, original research at the intersection between biochemistry, metabolomics, functional genomics, and synthetic biology is necessary. For instance, large-scale cellular assays could be developed to screen the in vivo functionality of relevant metallocluster enzymes in foreign hosts, and to address the barriers to their transferability and catalytic activity . FurtherMoreover, recent advances in synthetic biology have enabled de novo engineering of artificial metallocluster enzymes by incorporating catalytic metallocofactors (including abiotic cofactors) into designed protein scaffolds. These artificial enzymes catalyze a remarkable range of natural and synthetic reactions , and theBesides their potential for microbial biosynthesis, metallocluster enzymes participate in a wide range of key biological processes across kingdoms, including some of technological importance, such as antiviral  or antib


Dear readers,The Journal of Innovations in Cardiac Rhythm Management, Alqam et al.1 assess the discontinuation of anticoagulation after typical flutter ablation. Although retrospective and nonrandomized, their study offers important information that can help physicians managing patients with atrial flutter.In this issue of 2 recommended adherence to AF anticoagulation in patients with a history of AF ablation, regardless of procedural success or failure. Continuous or frequent electrocardiographic monitoring to screen for AF recurrence should also be considered in patients in whom anticoagulation discontinuation is being weighed based on patient values and preferences.2 Some well-designed nonrandomized studies8 have found that catheter ablation for AF reduces the risk of stroke; stopping anticoagulation after successful ablation can also be safely accomplished in some patients if careful rhythm monitoring is performed.Meanwhile, no randomized studies have investigated the continuation versus cessation of oral anticoagulation after catheter ablation for atrial fibrillation (AF). A 2017 expert consensus statement on catheter and surgical ablation of AFThe field of cardiac rhythm monitoring has rapidly expanded in recent years. I believe the discontinuation of anticoagulation after AF ablation can be safely accomplished in selected patients when paired with careful rhythm monitoring; however, randomized clinical studies that consider different monitoring practices and groups of patients remain critical.Sincerely,md, fhrs, faccMoussa Mansour, Editor in ChiefThe Journal of Innovations in Cardiac Rhythm ManagementMMansour@InnovationsInCRM.comDirector, Atrial Fibrillation ProgramJeremy Ruskin and Dan Starks Endowed Chair in CardiologyMassachusetts General HospitalBoston, MA 02114


Community-based participatory research (CBPR), a bottom-up approach that community stakeholders and academics are involved equitably, is an effective approach for enhancing relevance and value in public health research and has gained popularity in recent decades. However, little is known about how CBPR can be used in mental health studies with older adults. This systematic review examined the current state of knowledge about how CBPR approach has been adopted in mental health research among older adults in different societies. According to the PRISMA guidelines, we searched five major databases and screened the literature using these criteria: 1) journal articles reporting use of CBPR in mental health research among older adults, 2) articles published in English language, 3) studies conducted in any settings with any mental health research. Initial search found 3,227 articles and preliminary screening identified 23 eligible articles. We found that around 90% of studies were conducted in the West. Most studies adopted CBPR to develop community-based mental health interventions or to revise current interventions or models while addressing the cultural needs of their studied population. Few studies adopted CBPR to evaluate existing mental health workshops or programmes. The extent of involvement of older adults in the CBPR approach varied across studies, from questionnaire design to programme evaluation. Our review uncovered ways of CBPR implementation across different societies and elements of successful implementation in CBPR practices in mental health research among older adults.


Pathology education conventional methods have been disrupted by the Corona-Virus Disease 2019 (COVID-19) outbreak, compelling a re-evaluation of the means of educational interactions from the undergraduate to the postgraduate level. This commentary explores how the COVID-19 outbreak has challenged pathology education.We reviewed the current challenges and determined the potential implications of virtual technologies on modern pathology education for the future of pathology competency learning and assessment.The challenges are partly due to transferring from in-person teaching to a virtual education. Other reasons are shifting away from discipline-based teaching to organ-system based in medical curriculum and additional pressures on pathology faculties, such as increased demand for pathology services, lack of time, and learning resources. Keeping the national standards in pathology education even in the constant disruptions from pandemic outbreaks are current challenges. Pathology expertise will need to use emergent technologies in providing educational material to ensure quality pathology education. However, virtualization of pathology education produces a value of digital pathology and web-based pathology training materials. Medical students could review clinical cases remotely with their supervisors and gain the pathology competencies necessary for clinical practice. We need new innovative strategies, and we suggested the following steps to take advantage of the current opportunity to meet the challenges: evaluating the available digital training materials for formal pathology education, investing in the virtual infrastructure for competency-based pathology education, expanding student/residents exposure to pathology educational cases through virtual platforms; applying digital pathology solutions for virtual pathology education.  Pathology education conventional methods have been disrupted by the COVID-19 outbreak, compelling a re-evaluation of the means of educational interactions from the undergraduate to the postgraduate level , 3. ThisWorldwide, the health systems confront with a shortage of pathologists .Pathology Education in Modern Medical CurriculaPathology is the vital component of diagnostic strategies and central to medical education , 5. PathThe most medical curriculum has already reformed toward horizontal integration of courses, case presentation in the preclinical phase. The undergraduate medical education shifted away from discipline-based teaching to organ-system based in the medical curriculum. These changes affect pathology education at the undergraduate level. Moreover, forming medical educator teams composed of various medical specialists increasing the role that interdisciplinary teams play in medical education. Integration of disciplines has eliminated formal pathology classes in many medical schools , 6. HoweWith current educational policies to reduce transmission of COVID-19, Pathology departments and multidisciplinary networks should evaluate where the virtual educational technologies fit into their new educational plans in an integrated medical curriculum.Keeping the Competency-based Pathology Education The pathology competencies represent the mini-mum requirements of what the national pathology education board has agreed upon to educate undergraduate medical students for practice and for entry into any residency program.Pathology competency-based education focuses on applying pathology knowledge into practice, clinical reasoning, and pathology diagnostic skills.Keeping the competency-based standards in pathology education even in constant disruptions from pandemic outbreaks are current challenges of pathologists. The ability to gain a broad knowledge of the physiological and pathological processes of each organ system, the competency to apply disease mechanisms to explore the pathobiology, and the competency to improve the pathology diagnostic and therapeutic skills through lifelong learning is essential in becoming a competent physician .Pathology Education Challenges in Shifting to Virtual Learning Pathology education's main general challenges are deciding how much to change the courses, adapting to online education, determining the efficiency and effectiveness of virtual learning, and maintaining student engagement . The othWe Need new Innovative Strategies, and We Suggested the Following Steps to Take Advantage of the Current Opportunity to Meet the Challenges. However, contextual priorities may lead medical education experts to make individualized decisions based on unique circumstances.Evaluating the free available web-based pathology training materials for formal pathology education1) The studies indicate that medical students adopted the online resources as a learning tool, and a combina-tion of supervised small group discussions and the self-directed web-based (online) format improved path-ology teaching , 9. Web-The educational impact-not availability- of pathology web resources is a key for a successful transition to effective virtual pathology education. Therefore, pathology educational experts should evaluate the online videos according to national standards in pathology education whether they contribute to pathology education at the undergraduate and postgraduate levels. Some publicly available websites illustrate pathological principles, present rare disorders that are less relevant to undergraduate education. Some studies indicate that YouTube as a popular online video sharing website is an informative and accurate media of histopathology (cellular pathology) learning for both undergraduates and postgraduates. Moreover, autopsies were present on the site .Besides, Case X is an online platform that provides case-based learning materials and video-based medical cases. Online Case X helps student’s clinical reasoning through self-directed learning . Self-diInvesting in the Virtual Infrastructure and Products for Competency-based Pathology Education and Assessment2) The innovative learning technologies will change the future of pathology education and can facilitate training pathology with interactive simulation learn-ing . The intThe virtual educational products need massive infrastructures, including management software and hardware for clinical simulation, case designing tools, and cloud-based eLearning. Pathology content experts should support the virtual system for developing specific learning objectives, pathologic cases, providing updated educational material, virtual supervision, and support .The virtual reality (VR) platform and interactive virtual learning tools can respond to specific issues for pathology discipline that is practical in training and competency assessment . The Advancements in VR technology make its use as a training and assessment tool in online objective structured clinical examination (OSCEs) and Virtual OSCEs (VOSCEs) possible , 13. The formative ‘‘online OSCE’’ for pathology education could assess the disease recognition skills in pathology residents. A sequence of stations presents high-quality specimen images, and the residents should answer the questions and determine the organ and diagnosis within a time limit. After the deadline, they can find the correct answers and case histories of the specimens .Moreover, this can include standardized virtual patient resembling a real patient encounter in virtual stations. Variety of tools are applied in the health care system such as medical history taking, examination, diagnosis, laboratory/ imaging tests, and treating the VP. The system helps medical students master main pathology competencies, including disease mechanisms, organ system pathology, and diagnostic and therapeutic pathology skills. Besides, automated scoring is complementary to assess their progress. Virtual simulation provides the necessary tools to provide benchmarking, better preparing them for real-world medical practice.Expanding Pathology residents’ Exposure to Educational Cases Through Virtual Platforms3) Previous studies showed that some pathology educational programs should increase the residents’ exposure to pathology cases and practice because pathology subspecialty training programs require residents to apply for fellowships as early as the end of their specialty training. Also, studies indicate that changes in pathology specialty curricula and educa-tional activities have affected resident choices of fellowships , 15.. Keeping the national standards in pathology education are current challenges. Pathology expertise will need to use emergent technologies in providing educational material to ensure quality pathology educationThe clinical pathology cases presented through a patient’s laboratory data or images by detailed explanations help the learner to understand and apply diagnostic principles and incorporate morphologic or laboratory results for accurate diagnosis and treatment. Virtual discussion forums provide the context for sharing the core educational points to promote clinical reasoning.Applying Digital Pathology Solutions for Virtual Pathology Education4) Virtualization of pathology education produces a value of digital pathology; it also assists medical universities to support the continuing education of pathologists in the COVID-19 crisis.Digital pathology (DP) becomes the necessary components in clinical pathology service and cellular pathology training. Pathologists require innovative DP tools and artificial intelligence for digital-enabled care. Residents need these tools to acquire their pathology competencies necessary for the practice of pathology during their early years and discover which subspecialty areas most interest them .DP provides real-time access to clinical/ pathology cases and virtual simulation-based education technologies provide the necessary tools to review clinical cases remotely with supervisors and discuss cases with a histopathologists consultant through virtual conferencing during this pandemic.Some DP technology products are also ready for deployment on the virtual medical education platforms. England’s national health systems (NHS) partners from Oxford, Belfast, Nottingham Universities, and PathLAKE Features University are creating fully digital cellular pathology laboratories with a database of anonymous scanned slide images. These university diagnostic labs are developing AI algorithms for their generated slides , 17.Pathology education is witnessing a paradigm shift where scientific progress and technological advances are growing faster than pathologists’ workforces. Digital pathology and AI algorithm for image analysis would promote pathologic diagnosis. Such technologies and innovations are still in their early stages, but the future is promising. Applying digital pathology solutions for virtual pathology education will increase the efficiency of individual pathologists.We have described the challenges of keeping pathology education on track during the pandemic, and how virtual education platforms and digital technologies in pathology can provide real-time solutions to pathology competencies development. Pathology aca-demic departments should evaluate new technologies and encourage routine adoption of virtual education for digital native medical students. Moreover, the virtual platforms allowed the pathology educators to use DP with providing a safer and more efficient education during the COVID-19 crisis.The following four steps were suggested for utilizing the formal pathology education (from the undergraduate to the postgraduate) to support uninterrupted learning and assessment: 1) Evaluating the free available web-based patho-logy training materials for formal pathology education. 2) Investing in the virtual infrastructure and pro-ducts for competency-based pathology education and assessment.3) Expanding pathology residents’ exposure to educational cases through virtual platforms. 4) Applying digital pathology solutions for virtual pathology education.The Advancements in DP make it a pathology competencies training and assessment tool. The technologies still need to progress before virtual OSCEs become a comprehensive assessment plat-form. Despite this, the future is promising.


Hypertensive disorders of pregnancy (HDP) contribute to adverse gene-environment interactions prior to conception and continue throughout pregnancy. Embryonic/fetal brain disorders occur from interactions between genetic susceptibilities interacting with acquired diseases or conditions affecting the maternal/placental fetal (MPF) triad. Trimester-specific pathophysiological mechanisms, such as maternal immune activation and ischemic placental syndrome, contribute to adverse peripartum, neonatal and childhood outcomes. Two diagnostic approaches provide timelier diagnoses over the first 1000 days from conception until two years of age. Horizontal analyses assess the maturation of the triad, neonate and child. Vertical analyses consider systems-biology from genetic, molecular, cellular, tissue through organ networks during each developmental niche. Disease expressions associated with HDP have cumulative adverse effects across the lifespan when subjected to subsequent adverse events. Critical/sensitive periods of developmental neuroplasticity over the first 1000 days are more likely to result in permanent sequelae. Novel diagnostic approaches, beginning during pre-conception, will facilitate the development of effective preventive, rescue and reparative neurotherapeutic strategies in response to HDP-related trimester-specific disease pathways. Public health policies require the inclusion of women’s health advocacy during and beyond their reproductive years to reduce sequelae experienced by mothers and their offspring. A lower global burden of neurologic disease from HDP will benefit future generations.  Two diagnostic approaches assist in the prediction of brain health or disease throughout the life-span.  Over theBoth diagnostic approaches allow the fetal/neonatal neurologist (FNN) to anticipate adverse outcomes from any disease process that may potentially affect the developing nervous system across time-periods. This analytic process can be applied to pathophysiological mechanisms associated with specific categories of disease affecting the MPF triad, such as from hypertensive disorders of pregnancy (HDP). Brain health or disease experienced after HDP during critical/sensitive developmental time-periods over the first 1000 days will more likely remain permanent across the lifespan. Novel diagnostic approaches, starting during pre-conception are required that potentially can be applied to all levels of maternal care across the three trimesters. These advances will lead to effective and timelier neurotherapeutic interventions that combine preventive, rescue and reparative strategies for the developing brain in response to the diseases and adversities  associatThis analytic approach requires a working knowledge by an interdisciplinary team of practitioners regarding the maturing embryonic/fetal brain within the MPF triad across three trimesters in association with HDP. Such an approach will better select the appropriate diagnostic tests and treatment interventions that are patient-centric from a developmental perspective regarding a vulnerable MPF triad.The practitioner is confronted with a spectrum of HDP disorders in women, including hypertensive diseases before conception, gestational hypertension, early and late preeclampsia, HELLP syndrome and eclampsia. For this discussion of potential sequelae after HDP, no single diagnostic test or therapeutic intervention will be effective for all women at all times before and during pregnancy. New obstetrical research will offer the clinician with different options and timing that can be standardized for maximal effectiveness, beginning prior to conception. This will be discussed in HDP affects 5–7% of the world population of women and their offspring. In one recent study, as many as 7–15% of the world’s population of women will experience HDP when assessed on a per-woman basis . Given sImproved diagnostic algorithms of HDP will help reduce the global burden of neurologic disease. As discussed at the conclusion of this review, this should be a public health priority for women and their offspring given the high prevalence of HDP. International efforts, such as the sustainable development goals proposed by the WHO ,9, must Transgenerational followed by early pregnancy G × E influences subject the MPF triad to HDP, with different expressions of neurologic sequelae by the offspring. Pre-conceptional genetic susceptibilities have been identified through investigations such as high-throughput genome-wide association studies (GWAS) and epigenetic studies . ImproveCandidate genes and pathways responsible for sequelae are more accurately identified by integrated bioinformational analysis . DifferiCo-morbidities associated with maternal diseases and adversities further augment genetic/epigenetic effects  beginninCross-species expression of the MPF triad structure and function suggests preserved evolutionary biological processes, with either beneficial and adverse outcomes in humans . This trStructural markers detected by fetal surveillance using sonography and more detailed fetal neuroimaging with magnetic resonance imaging include a wider range of detectable brain lesions, including major malformations with or without lethality to region-specific lesions represented by focal developmental anomalies and destructive lesions. The form and severity of these anatomical biomarkers depend on the trimester-specific timing and specific pathophysiological pathways that were activated by G × E interactions associated with HDP as well as other diseases entities with similar harmful effects from hypoxia/ischemia, inflammation and coagulopathy. Early pregnancy anomalies range in severity from neural tube defects, holoprosencephaly and schizencephaly to focal cortical dysplasias. During the latter half or pregnancy, destructive lesions predominate and may accompany earlier anomalous lesions associated with HDP-related effects, such as the maturing MPF triad experiences in on-going disease.G × E interactions result in different neurologic sequalae specific to pre-conception and the trimester-specific time-periods when the MPF triad is affected by HDP. Primary prenatal neurologic structural/functional disorders without systemic involvement may occur. Secondary brain lesions alternatively result from multi-systemic conditions that also contribute to adverse neurologic outcomes. Disease processes, such as maternal immune activation  and ischG × E interactions associated with HDP continue to be expressed after birth as the great neonatal neurological syndromes (GNNS) of encephalopathy (NE), seizures (NS), stroke (NSK) or encephalopathy of prematurity (EP) . The sevPediatric infections, trauma and multi-systemic diseases later in childhood may further worsen sequelae after HDP, stressing the continuity of risk from G × E interactions during postnatal maturation. Both communicable and noncommunicable childhood diseases contribute to adverse outcomes, involving hypoxic/ischemic, inflammatory and hemostatic disease pathways. Biosocial adversities affecting the child within the family and community contribute to the extent and complexity of sequelae. Developmental disorders, such as intellectual disability, autism and cerebral palsy, as well as behavioral/cognitive deficits and epilepsies, are potential sequalae expressed throughout childhood and adolescence. Each disorder may either present independently or as co-morbidities. Associations with HDP need to be considered.Sequalae later present with maturation into adulthood. The aging brain’s pathophysiological mechanisms will result in long-term responses to the prenatal conditions of HDP, as well as to other adverse diseases or conditions affecting the MPF triad. Diverse sequalae include cerebrovascular diseases, dementias, adult-onset epilepsy and neurodegenerative disorders. G × E interactions at the end of the lifespan are significantly influenced by permanent maladaptive developmental neuroplasticity that occurred during the first 1000 days.Complex MPF triad G × E interactions from HDP early during pregnancy profoundly impairs neuronal and glial progenitor cells within transitory brain structures . DisruptMulti-potential precursor populations undergo proliferation, differentiation and migration during the first twenty weeks within multiple transient structures , such asAltered progenitor cell populations within the neuroectoderm or yolk sac later promote abnormal connectivity during the second half of pregnancy with postnatal neurologic sequelae A, B. AltImpaired brain development during the first half of pregnancy associated with HDP affects the interactions of multiple neuronal, glial, and angiogenic precursors, particularly within the developing neurovascular unit . Deficient pro-angiogenic growth factors, such as placental growth factor  and vascImmune intolerance between the mother and embryo/fetus from abnormal placental implantation and development result in maternal immune activation (MIA), which later influences the expression of multiple psychiatric and neurological disorders over the life-span . SpecifiConsideration of G × E interactions over the first 1000 days begins with an understanding of the vulnerability from MIA effects after infectious and non-infectious inflammatory conditions, particularly during early pregnancy. A broad range of diseases and conditions result in MIA, including HDP, and affect the maturing components of the MPF triad, with altered fetal brain connectivity. While prevalence estimates of MIA remain imprecise, a significant percentage of children and adults later express early and long-term neurologic disorders from maladaptive developmental neuroplasticity as a result of MIA associated with diseases such as those from HDP. Improved neurotherapeutic interventions need to be developed in order to apply an understanding of MIA  in relatDisordered cellular migration, dendritic arborization, synaptogenesis and myelination within the developing fetal brain is predominant during the latter half of pregnancy. These developmental fetal brain stages are particularly active within the maturing cortex and subplate zone . AnomaloDi Renzo introduced the term “the great obstetrical syndromes” (GOS) 2009), referring to abnormal maternal, placental and fetal outcomes as a result of abnormal placental angiogenesis. A growing list of adverse outcomes from GOS can affect the MPF triad. Phenotypes specific to each of the three triad components include preeclampsia, diabetes, morbidly adherent placenta, abruptio placenta, premature rupture of membranes, fetal growth restriction, fetal demise, and prematurity. All these conditions adversely affect MPF triad health, with impairment to fetal brain development specific to the developmental stage. Abnormal placental angiogenesis during the second half of pregnancy consists of incomplete spiral artery remodeling and shallow embedding A,B. This09, referHDP are expressions of abnormal placental angiogenesis associated with IPS . PathophAdverse trimester-specific conditions from HDP can negatively impact peripartum events close to and including labor and delivery . ObstetrAdverse intrapartum events associated with HDP may only later be identified during neonatal or childhood time-periods. Whether sentinel or gradual in onset, presumed fetal distress detected during labor and delivery by surveillance testing is also a poor proxy that cannot reliably identify the presence or timing of brain injury. The peripheral chemoreflex remains primarily an adaptive response, activated during uterine contractions to protect against primarily brain, cardiac and adrenal injuries in most situations as the child descends the vaginal tract. Specific clinical scenarios can result in intrapartum fetal brain injury , particuComplex disease conditions represented by HDP are best interpreted by applying horizontal and vertical analytic perspectives to anticipate risk or occurrence of brain injuries prior to, as well as during, labor and delivery. Interdisciplinary consensus reports periodically update the evolving scientific bases for these analyses ,49. CompComplex phenotypes after birth are expressed collectively as the great neonatal neurological syndromes (GNNS). Similar to the GOS, trimester-specific G × E interactions affect the MPF triad during postnatal life, expressed as the GNNS. Differences from GOS include: 1) clinical presentations include events specific to the peripartum and neonatal time-periods; (2) phenotypic expressions of GNNS are not reliably detected across trimesters for all maternal levels of care, particularly with seemingly low-risk pregnancies, (3) early and later pregnancy factors cumulatively affect MPF triad components closer to delivery expressed as the GNNS. NE, NS, NSK, and EP are the four major GNNS categories [ clinicalNegative outcomes associated with the GNNS occur despite adjustments in obstetrical management closer to delivery to conditions associated with HDP. Introduction of resuscitative interventions and neurocritical care may not significantly alter antepartum adverse outcomes ,3. FNN aAcute infections, metabolic-toxic and traumatic etiologies may best explain peripartum or neonatal-acquired encephalopathies and/or brain injuries expressed by the GNNS. Alternatively, trimester-specific MPF triad G × E interactions associated with HDP have a varying clinical expression during peripartum and neonatal time-periods, based on the timing and etiologies associated with brain anomalies or injuries superimposed on acute disease. Childhood neurologic disorders may later present, despite neonatal neurocritical care interventions for the GNNS as well as in the absence of early neonatal disease expression. Neonatal evaluations specific to each GNNS have been discussed in more detail elsewhere and are applicable to clinical assessment after HDP . NE, NS,Paradoxically, lower mortalities and adverse neurological outcomes such as periventricular leukomalacia, intraventricular hemorrhage and cerebral palsy have been retrospectively reported in specific studies of extremely and very preterm infant populations when associated with HDP, compared with cohorts without HDP . HDP mayDiagnostic strategies to assess the GNNS requires a time-sensitive interpretation of serial examinations, neuroimaging, electroencephalography and blood/serum studies throughout the neonatal hospitalization . Multi-sOne important diagnostic test regarding the timing and etiology of brain injuries should include gross and microscopic examinations of the placenta, cord and uterus. While the completed pathology report may not be available until after the first week of life, findings often more accurately estimate an antepartum time-period to diseases that precede events closer to delivery. Trimester-specific pathophysiological mechanisms may better explain different disease processes associated with HDP after pathological analyses of the placenta, cord and uterus. This explanation will better assist the family for both diagnostic accuracies, as well as prognosis and anticipatory care. Revised pathological classifications have strengthened these chronic associations . HoweverDuring the first two years of life, the expression of developmental disorders and epilepsies ,58 may bChildren who experienced the GOS and GNNS after diseases such as HDP more likely require earlier medical interventions at older ages. Hospitalizations, may require pediatric neurointensive care  for a moPediatric neurology consultations in the clinic or hospital settings may originate from primary-care practitioner or pediatric subspecialist referrals when communicable and noncommunicable diseases worsen neurologic function associated with organ-specific systemic diseases. Congenital heart disease (CHD) is the most common anomalous organ system disease of childhood that can be associated with HDP, with sequelae including cognitive/psychiatric disorders . ContinuInvolvement during the first 1000 days facilitates continued pediatric neurology consultative input throughout childhood into adolescence. This continuity of care more effectively addresses ongoing or new conditions affecting brain health when clinical disorders are expressed in association with HDP. Clinical expression requires brain maturation.Epileptic, behavioral/cognitive and mental health disorders  are assoNeurologic and mental health disorders associated with HDP may only become clinically apparent with maturation into adulthood . These sCombined developmental origins and life-course approaches to diagnosis and prognosis therefore stress the relevance of the first 1000 days to neurologic/psychiatric diseases presenting across the life-span. Cerebrovascular, cognitive, epileptic and neurodegenerative disorders  into oldCurrent practice guidelines and stress conventional management strategies to control HDP . PregnanAdverse effects by HDP on embryonic and early fetal brain development within the MPF triad requires neurotherapeutic interventions, targeted to immature neuronal populations and connectivities that may be impaired during the first half of pregnancy. Pre-conception testing will target inherited genomic biomarkers associated with hypertensive disorders affecting a future pregnancy . The nexReduced burden of neurologic disorders across the lifespan from conditions associated with HDP should constitute one of multiple sustainable goals required for maternal and pediatric health initiatives . SignifiImprovements in both person-centric and population-based healthcare define a nation’s medical, economic and social well-being. Cooperation between government, industry, and nonprofit entities collectively strengthen healthcare policy for populations specific to resource-rich and poor nations. This advocacy must include medical deserts within resource-rich nations. Health disparity research augments efforts to address prenatal and early childhood brain disorders in association with HDP required by public health programs. This is exemplified by the WHO Millennium sustainable goals , which can more effectively reduce disease burden and financial costs, as well as elevate the quality of life . The sigInterventions need to be offered that are gender-specific and sensitive to sexual-orientation, specifically recognizing the unique aspects of healthcare relevant to women and men . Outcome


Stem-cell derived in vitro cardiac models have provided profound insights into mechanisms in cardiac development and disease. Efficient differentiation of specific cardiac cell types from human pluripotent stem cells using a three-step Wnt signaling modulation has been one of the major discoveries that has enabled personalized cardiovascular disease modeling approaches. Generation of cardiac cell types follow key development stages during embryogenesis, they intuitively are excellent models to study cardiac tissue patterning in primitive cardiac structures. Here, we provide a brief overview of protocols that have laid the foundation for derivation of stem-cell derived three-dimensional cardiac models. Further this article highlights features and utility of the models to distinguish the advantages and trade-offs in modeling embryonic development and disease processes. Finally, we discuss the challenges in improving robustness in the current models and utilizing developmental principles to bring higher physiological relevance. In vitro human cardiac models are complimentary tools that allow mechanistic interrogation in a reductionist way. The unique advantage of utilizing patient specific stem cells and continued improvements in generating reliable organoid mimics of the heart will boost predictive power of these tools in basic and translational research. Over the past decade, several strides made in stem cell research has led to unparallel opportunities to study human diseases in vitro. Cardiovascular diseases (CVD) comprising of congenital heart disease, heart failure, stroke and hypertension contribute toward nearly one-third of all deaths worldwide , 2. CardThe heart forms from the mesodermal embryonic tissue from anterior primitive streak. The cardiac specification and differentiation take place after the lateral movement of the anterior region into splanchnic mesoderm. This tissue further self-organizes into three distinct regions where the cardiac crescent or first heart field (FHF) is ‘sandwiched’ between head folds and second heart field (SHF). During the morphogenetic movements, the heart regions lie in close contact with pharyngeal or foregut endoderm . TranscrFrom a molecular point-of-view, orchestration of T-box containing transcription factors such as TBX20 and TBX5 trigger chamber and septa formation while BMP driven TBX2 and TBX3 driven activation leads to atrioventricular canal and outflow tract. Cardiac neural crest influx into the outflow tract and complete septation is completed around week 12 in humans where it resembles the heart structure morphologically . Heart bIn vitro differentiation of cardiomyocytes (CMs) from iPSCs follow sequential stages observed during embryonic development, wherein heart forming progenitor cells arise from the mesodermal cell layer sandwiched between the ectoderm and endoderm in the primitive streak. Cardiomyogenesis in vertebrates is driven by classes of growth factor proteins such as wingless/INT (WNTs), bone morphogenetic proteins (BMPs) and fibroblast growth factors (FGFs) , 10. ConMost recently, there has been a surge in cardiac ‘organoid’ models which are self-organized, spatially restricted clusters of cardiac-specific cell types derived from pluripotent stem cells. Some organoid models have been touted as new tools to study several congenital and developmental disorders such as hypoplastic left heart syndrome (HLHS) where the left ventricle fails to form due to mutation in NOTCH1 gene . SeveralDevelopmental cardiac organoid-like models do not serve as an attractive tool to perform investigations in a reductionist manner due to lack of high fidelity in cellular composition and niche. Diseases where the pathophysiology is unclear in vivo due to complex microenvironment could benefit from engineered tissue-like structures that can be manipulated in a modular fashion. Several protocols have been developed in recent years to generate major non-myocyte cell types of the heart from iPSCs such as endothelial cells , cardiacBoth cardiac organoid and heart-on-a chip models are excellent surrogates to understand several important aspects of heart function during development and disease. However, it is important to acknowledge that utility in predicting in vivo functions will remain limited due to stochastic self-patterning variability between every batch of stem cell differentiation. In addition, several factors that currently rely on self-assembly time need to be coupled with controlled delivery of cues. For example, in addition to progenitor-cell types, cardiogenesis is driven by a combination of long- and short-range paracrine interactions, gene regulatory networks and ECM-guided geometrical cues. Efforts toward obtaining blueprints of stage-wise development will enable us to isolate and embed temporal cues in these models to influence multi-axial patterning and cellular organization, rather than focusing on autonomous self-assembly under different conditions. Another important aspect in cardiac models is obtaining innervation and vascular anastomoses which provide feedback on carbon dioxide, dissolved oxygen and fluid pressure. Technical challenges that prevent widespread use of cardiac models can be addressed through standardized characterization of stem cell sources, using well-defined platforms and most importantly choosing the right model in the context of use . With inCurrently, both ECM and ECM-free cardiac models are proving useful as an obligate substitute for pre-clinical animal models, to validate clinical observations and as a screening tool for therapeutics or toxicology. With success through continued efforts in addressing some of the discussed challenges, we must revisit our expectations and utility of the model based on evidence and reproducibility. For example, current 2D and 3D tissue engineered models with limited heart-like features would be useful from a reductionist approach to understand cell specific mechanism or direct pharmacological response. Future availability of morphologically and spatially defined organoid mimics will further assist in mapping direct and indirect interactions due to concerted cellular response and fluidic stress. Such systematic approach toward building complexity will drive utility-based research in cardiovascular organoid biology as a complimentary tool and not a panacea for studying multifactorial cardiovascular diseases.


To the Editors,Endovascular treatment of carotid artery disease has gone through fast-paced evolution since the advent of balloon angioplasty and stents three decades ago. Nevertheless, it was not until significant improvements in neuroprotection were achieved that carotid artery stenting became a solid contender to the gold standard carotid artery endarterectomy. Embolic protection devices, such as proximal common and/or external carotid artery balloon occlusion devices and internal carotid artery filters paved the way toward a safer and more definitive strategy. The concept of flow reversal in the early 2000s,Carotid artery calcific disease remains a challenge in patients undergoing carotid artery stenting regardless of whether it is done via a transfemoral or transcarotid approach. By far, the most challenging scenario is circumferentially calcified internal carotid artery plaques that can cause inadequate expansion of stents and limited luminal gain despite vigorous pre- and post-dilation of the lesion.The two main strategies use either atherectomy devices or the new intravascular lithotripsy (IVL) balloon. We have reported both use of orbital atherectomy


Although loneliness and social isolation are often discussed together, they are mainly examined separately. The few studies examining both concepts simultaneously focus usually on the wider category of older people (65+), with no or little attention to very old age. Our main aim was to investigate loneliness and social isolation in combination among near-centenarians and centenarians. Analyzing data from the Fordham Centenarian Study , we found no or very weak associations between loneliness and social isolation. Combining measures of loneliness  and social isolation  we built a typology with four different groups . The factors that most strongly predicted the distribution among these four groups were gender, widowhood, education, and self-rated health. Findings highlight the importance of jointly studying both concepts to better understand social risks in very old age.


Fusarium, continue to be a major threat to agricultural crops worldwide. In this issue, Tracanna and coworkers  implement a targeted amplicon sequencing approach to identify conserved domains and specific metabolic pathways shared among soil samples with antagonistic activities against Fusarium culmorum. They also introduce dom2BGC, an open-source annotation platform that builds co-occurrence networks of natural product-associated domains across samples and aids in putative gene cluster reconstruction. When coupled with metagenomics, functional amplicon sequencing and the dom2BGC pipeline can aid in identifying mechanisms and potential metabolites associated with particular microbiome-associated phenotypes.Soil-dwelling microorganisms associated with plant roots carry out essential processes that promote plant growth and productivity. In addition to these beneficial functions, the rhizosphere microbiome also serves as the first line of defense against many plant pathogens. While many rhizobacteria are capable of producing antifungal natural products, fungal pathogens, such as those belonging to the genus  Soils are biologically diverse habitats and represent some of the most complex ecosystems found in nature . As sustInvestigations into the rhizosphere have shown that this is a complex and dynamic environment . Plants Fusarium species being among the most devastating. While a previous study identified soil samples with antagonistic activity against Fusarium culmorum  are a large class of bioactive natural products and are typically biosynthesized in an assembly line-like fashion by nonribosomal peptide synthetases (NRPSs). Adenylation domains are essential components of the NRPS assembly line and are responsible for selecting the amino acid building blocks that get incorporated into the natural product. Using degenerate primers, adenylation domains were PCR amplified from rhizosphere DNA collected from four of the authors’ soil samples possessing activity against programs . Furtherin silico amplicons extracted from the antiSMASH and MIBiG databases. Taxonomic diversity and community structure relationships are calculated across sample sets, and domain co-occurrence networks establish biosynthetic distribution patterns that can be used to explain microbiome-associated phenotypes. While a similar PCR-based targeting approach was used in the discovery of the malacidins, the bioinformatics program eSNaPD (With the adenylation sequences from the rhizosphere microbiomes in hand, the authors developed an automated annotation platform to assist with comparative analyses across the sample sets. Briefly, dom2BGC annotates biosynthetic domains based on similarities with m eSNaPD  that iniF. culmorum.Using the dom2BGC platform, the authors identified community structure overlap between adenylation domain profiles from suppressive rhizobacteria and were able to map the sequences to multiple taxonomic groups. From the co-occurence patterns, dom2BGC was also successful at reconstructing multiple biosynthetic gene clusters. Results from the amplicon clustering were validated using 10× metagenome assembly and demonstrated that dom2BGC is a powerful platform for identifying functional elements in complex microbial communities. Detailed analysis of the amplified adenylation domains suggested an enrichment of cyclic and branched lipopeptide-producing biosynthetic clusters in the soil samples possessing antifungal activity, and bioinformatic analysis of the metagenomic data revealed several siderophore, lipopeptide, and 2,4-diacetylphloroglucinol gene clusters. It was speculated that the agents associated with the respective gene clusters could be responsible for the antagonistic activity against Tracanna and coworkers  demonstr


Palliative care is included within the universal health coverage goal of the sustainable development goals as an essential health service and is considered a human right.COVID-19 patients and their families report multidimensional symptoms, ranging from those that are physical, to psychosocial, spiritual and existential distress from the threat to survival.In our recent review of African COVID-19 case management guidelines, we found few palliative care approaches and a focus on clinical management to the neglect of important psychosocial and spiritual stressors affecting morbidity and outcomes.The supply of medical products, including palliative medicines, has also been disrupted by the pandemic, even more so in countries still experiencing increased cases, including Kenya, South Africa and Uganda.Before the pandemic strained African health systems, only 5–11% of people requiring palliative care had access. The negative impact on services has heightened the risk of unnecessary suffering in patients with COVID-19 and other serious illnesses. Therefore, policy-makers in Africa must try to lessen the impact of the pandemic on palliative care services by integrating palliative care within the COVID-19 response. This integration involves training family caregivers and community health-care workers caring for COVID-19 patients and other seriously ill patients whose home care by specialist teams has been disrupted by pandemic restrictions, and ensuring ongoing funding for palliative care services.African institutions must rapidly expand palliative care training to health-care professionals outside palliative care services and support the training with clear, comprehensive case management guidelines focused on palliative care. Service providers must also urgently collaborate with researchers in adapting palliative care delivery approaches and testing new technology-based service delivery models, such as mobile health applications.


Knowledge of school attendance problems (SAPs) is needed to inform treatments targeting SAPs and protecting youths from negative outcomes associated with SAPs.This study examined the school absence, absence categories , sociodemographic characteristics, and mental health problems among youths seeking psychological treatment for SAPs.The study used a cross-sectional design. Sociodemographic and clinical characteristics of 152 help-seeking youths with SAPs  and their parents were examined. The data were derived from the baseline assessment conducted before treatment start.Older youths, youths with mental health problems, and youths whose parents had mental health problems exhibited higher levels of absence. Lower levels of non-excused absence were found among youths with highly educated fathers, and youths living with both parents. Many youths had clinical levels of anxiety, depression, or ‘emotional and behavioral difficulties’.The study highlights the need for early intervention, addressing a broad range of mental health problems.NCT03459677.ClinicalTrials.gov:  School attendance problem (SAP) refers to difficulty attending school or absence from school that is problematic because of its frequency and/or duration  while studies addressing truancy have focused on non-excused absence . Sociodemographic characteristic and mental health problems are often described for distinct subgroups of youths with SAPs  are more likely to be effective for youths presenting high levels of school absence together with a complex clinical presentation including anxiety, depression, and/or behavioral disorders.S1 Table(PDF)Click here for additional data file.


Previous research indicates volunteering promotes well-being of individuals and communities. Volunteering in later-life may buffer some of the negative health effects experienced during retirement, facilitating opportunities for older adults to engage in meaningful activities and stay active. The current study examined characteristics of older adults who volunteered outside of participation in a regular cognitive monitoring study. All 124 members  of a regular cognitive monitoring study, requiring completion of a 15-minute cognitive online test once a month, with complete data on personal characteristics, volunteer activities, as well as study adherence and dropout rates were included. ANCOVA and logistic regression analyses adjusted for sociodemographic characteristics were used to assess differences between volunteers and non-volunteers. Results indicated that volunteers were less educated (p<.05), and slightly more likely to be younger and women compared to non-volunteers. There were no differences in cognitive performance (ps>.05). Volunteers had lower scores for neuroticism (p=.02) and were marginally higher agreeable and extraverted (ps<.09). Volunteers needed more reminders to complete the monthly test (ps<.01) but had lower dropout rates (p=.001). The most frequent type of volunteer activity reported was religious. Volunteers were motivated mainly by altruism, although most reported multiple reasons such as building social relationships and feeling important. Findings provide information about characteristics that can help identify older adults who are likely to volunteer. Results regarding study adherence may have implications for promoting recruitment and retention among older adult volunteers.


Inflammatory biomarkers and sex hormones have been investigated as independent risk and resilience factors for cognitive decline in older adults. Many sex hormones are anti-inflammatory and there is emerging evidence that sex hormones may buffer the risk for cognitive decline associated with higher inflammation. However, few studies have included concurrent examination of inflammation and sex hormones in studies of cognitive performance and cognitive aging. A diverse sample of older adults  had blood drawn before and after a two-week measurement burst that included three cognitive tests (6x per day) assessing working spatial memory, perceptual speed, and feature binding. Testosterone, estradiol, estrone, and six basal cytokine concentrations were quantified. Composite scores of basal inflammation were calculated. Multilevel modeling indicated that heightened inflammation related to poorer spatial working memory performance . In addition, sex hormones moderated the association of cytokine concentration with perceptual speed . Decomposition these interactions revealed that heightened inflammation predicted poorer performance, but only among individuals with lower sex-hormone concentrations. This study provides evidence of immune and hormonal-by-immune associations with performance in two cognitive domains in older adults. Examining the functional crosstalk between immune and sex hormone functioning will improve understanding of risk and resilience factors related to cognitive performance and help predict cognitive decline in older adults.


ABSTRACT IMPACT: Our data identify a novel candidate for combination strategy in melanoma treatment, and can inform clinicians in their decision-making process regarding therapeutic intervention for melanoma patients. OBJECTIVES/GOALS: Soluble adenylyl cyclase (sAC) is a novel source of cyclic AMP (cAMP). In melanoma, nuclear sAC localization has an established diagnostic utility and we newly found that nuclear sAC functions as a tumor suppressor by inhibiting Hippo pathway, which affects treatment response. Here, we examine the effect of nuclear sAC on melanoma treatment response. METHODS/STUDY POPULATION: We developed a doxycycline inducible system for increasing sAC activity only in the nucleus. We assessed whether nuclear sAC activity affects treatment response, using BRAFV600 human melanoma cell lines. Using a clonogenic assay, we examined how nuclear sAC activity affects growth inhibition in the presence of a BRAF inhibitor, vemurafenib. Our findings will be confirmed in vivo using tumor xenografts. After tumor formation in NSG mice, mice will be randomized to be fed normal or doxycycline chow for nuclear sAC induction, then subdivided to receive vehicle or vemurafenib to examine the effect of nuclear sAC activity on treatment response in vivo. We will also compare melanoma biopsies collected before and after treatment with BRAF inhibitors to assess how nuclear sAC staining affects tumor morphology in vivo. RESULTS/ANTICIPATED RESULTS: So far, nuclear sAC activity has rendered SkMel178 and M263 cell lines more susceptible to vemurafenib. Cell viability was inversely correlated both with vemurafenib and with doxycycline concentration. Cell viability after vemurafenib treatment was dramatically reduced when nuclear sAC was activated. It appears that nuclear sAC enhances the sensitivity of BRAF mutant melanomas to vemurafenib in vitro. We anticipate that xenografts of these cells in mice will be more susceptible to vemurafenib when nuclear sAC is activated. We also anticipate that positive nuclear sAC staining will correlate with a favorable response to therapy. DISCUSSION/SIGNIFICANCE OF FINDINGS: Targeted therapy with BRAF inhibitors is used in late-stage melanomas, but its use is limited as patients invariably acquire resistance. Here, we identified nuclear sAC activation as a novel candidate for combination strategy. Our data will also inform clinicians how best to integrate this biomarker into their decision-making regarding therapy.


There remains a lack of knowledge on marital satisfaction of African Americans generally, but particularly older African Americans. In addition, only a handful of studies investigate satisfaction among couples who are unmarried. With data from the National Survey of American Life, this study examined the correlates of romantic and marital satisfaction among older African Americans. Findings reveal that married older African Americans were slightly more satisfied with their relationship than individuals who were either remarried or unmarried but in a romantic relationship. Among older African American married adults, older age was associated with higher marital satisfaction, and men had higher levels of marital satisfaction than women. Also, married older African Americans with lower family incomes reported higher marital satisfaction. Given the limited research on older African Americans couples, either married or unmarried, this study offers valuable implications for individuals and professionals engaging these couples in practical settings.


In Japan, care managers engage frail older adults to support their assisted living in long term care insurance system. However, due to the lack of some or all supervision, many care managers face problems such as low work engagement and high turnover rate. This study aims to examine what types of supervision have positive effects on work engagement and turnover intensions of care managers in Japan. The sample of 241 care managers were asked whether they have received individual supervision in the workplace (ISVW), individual supervision in the community (ISVC), group supervision in the workplace (GSVW), or group supervision in the community (GSVC). Independent samples t-tests and one-way ANOVAs were conducted to examine the effectiveness of each types of supervision on work engagement and turnover intension. T-tests showed that only GSVW was significantly related to work engagement . Whereas, only ISVW had a significant effect on turnover intensions . One-way ANOVAs revealed that 28 care managers receiving GSV had significantly higher work engagement than 92 care managers who did not receive any SV . 40 care managers receiving both ISV and GSV showed significantly lower turnover intentions than 92 care managers who received neither ISV nor GSV . Since the results have implications for the importance of supervisions to enhance work engagement or to reduce turnover intension of care managers, a larger sample will need to confirm these effects.


One of the well reported but difficult to manage symptoms of spinal cordinjury (SCI) is neurogenic lower urinary tract dysfunction (NLUTD). The type ofNLUTD is variable based on location and extent of injury. SCI affects more malesand NLUTD is especially debilitating for men with incomplete injury. This reviewsummarizes the anatomical basis of NLUTD in SCI and discusses current diagnosticand management strategies that are being utilized clinically. The last twosections address new innovations and emerging discoveries with the goal ofincreasing scientific interest in improving treatment options for people withSCI. Areas warranting further investigation are pinpointed to address currentgaps in knowledge and/or appropriate technology. These pThe spinal level of SCI causes distinct neuroanatomical disruption patternsresulting in different treatment strategies. Suprasacral lesions produce neurogenicdetrusor overactivity (NDO) and a noncentralized sacral spinal reflex. Sacral lesions directly involve the spinalmicturition center and cause mixed NLUTD symptoms and detrusor underactivity . Pudenda2.Prompt evaluation of lower urinary tract function is important consideringits cause of increased morbidity and mortality and high prevalence in SCI(70%–84%) male victims. Clinical patterns can help determine the presence ofNLUTD and help localize the lesion to guide conservative, pharmacologic, andsurgical therapies. Several guidelines exist for evaluating NLUTD including EuropeanUrological Association, Joint Societe Internationale D’Urologie-InternationalConsultation on Urologic Diseases (SIU-ICUD), and Japanese clinical guidelines–12. ConsWorkup begins with a thorough history and physical exam especially with focuson male urologic history regarding benign prostate hypertrophy, urinary infrequency,and any urologic malignancies. Review of the initial trauma and severity of the SCIwith a validated system like the ASIA score is first performed. It is important toconsider factors in a male’s medical history, like severe hypertension orhistory of acute angle closure glaucoma, that may contraindicate the use ofmedications in the treatment in NLUTD . AbladdBladder function can be objectively evaluated using urodynamic studies atleast once within the months following injury and can measure bladder sensitivity.For incomplete SCI, this can help determine return of function. Video urodynamicsserve as the gold standard for those with NLUTD. It is generally recommended toperform the first urodynamic study 3–6 month following injury.Electromyography may evaluate external urethral sphincter function. The resultingmorphologies based on the information gained regarding intravesicular pressure,detrusor activity, bladder compliance, bladder sensitivity, and bladder storagecapacity can help diagnose NLUTD, differentiate between the different subtypes, suchas those that involve detrusor overactivity or DSD, localize the level of SCI,identify the extent of injury on ASIA scale, give prognosis, and monitor treatmenteffect.Post-void residuals can be measured using bladder ultrasound or cleanintermittent catheterization. Other non-neurogenic conditions can be ruled out usingrenal and pelvic ultrasonography. MRI or CT of the brain can rule out otherneurogenic causes, particularly in the setting of polytrauma where the brain may beaffected and cause disruption of the pontine micturition center. MRI of the spinecan reveal spinal cord compromise that may be amenable to surgical intervention withpossible subsequent symptom improvement .This isDilation of the ureters, calyxes, and hydronephrosis should be considered inthose with NLUTD. The upper urinary tract can be evaluated using ultrasound andmagnetic resonance urography . Several3.3.1SCI is unique from other NLUTD etiologies in that it is oftenaccompanied by traumatic injuries and spinal shock syndrome. These factorscreate distinct phases of treatment for urologic issues affecting males: theacute recovery phase, post-acute rehabilitation phase, and long-term managementphase . Allowin3.2SCI patients may require indwelling catheterization during the acuterecovery phase until renal and cardiovascular function is optimized andurodynamic parameters improved .Beyond Pharmacological treatments of NLUTD generally target neuromuscularjunctions in the smooth muscle of the detrusor or internal urethral sphincter.Antagonism of alpha-1 adrenergic receptors relaxes the internal sphincter andbladder neck to improve voiding. Terazosin and tamsulosin are two of the mostwell-studied α1 antagonists and both have been demonstrated to reduceautonomic dysreflexia and improve urodynamics, especially in males –22. AntaChemodenervation of the detrusor by botulinum neurotoxins (BoNT) hasalso emerged as a minimally invasive treatment for NLUTD in men with incompleteSCI. The most commonly used BoNT serotype is onabotulinumtoxin A (BoNT/A),commercially known as Botox  . Althoug3.3via implantation ofexternally-controlled electrodes at the sacral nerve roots may be considered incases where conservative management fails to adequately control symptoms orproduces unfavorable side effects. The Brindley Procedure, combined sacralanterior root stimulation (SARS) and rhizotomy of dorsal sacral rootsS1–S5 can be considered in NLUTD patients where conservative managementfails [Neurostimulation ntfails . Deafferntfails –30. The ntfails –32.4.As outlined above, bladder storage and micturition are controlled by acomplex interplay between the smooth muscle of the lower urinary tract, the striatedmuscle of the urethral sphincter, the cerebral cortex, pontine micturition center,sacral micturition center, and both the sympathetic and parasympathetic portions ofthe peripheral nervous system. In incomplete SCI, the range of damage is variable,and, therefore, may affect different elements of these cascades, which explains thedifferent etiologies and presentations of NLUTD after SCI. Despite this complexity,a simplified view of NLUTD after SCI may be described as improper sensory awarenessfor the need to void or improper stimulus delivery to cause the bladder to void.Therefore, new treatment options have focused on the use of sensors to monitorbladder pressure parameters, neuromodulation to deliver precise stimulation to causethe micturition cascade, and combined closed-loop neuromodulation systems to bothsense bladder fullness and void. Some of the key findings were highlighted above incurrent treatments section but the experimental approaches are addressed furtherbelow and may be especially valuable for males with SCI. Outline of promisingapproaches highlighted in Urodynamic testing is a known study that examines the parasympathetic andsympathetic function of the urinary system, enabling examiners to evaluate bladderand urinary sphincter function and integrity . ClinicaMany of the recent innovations have been as a result of improved targetingof electrical stimulation and noninvasive modalities . Traditi5.Technical considerations to improve outcomes of bladder dysfunction andmicturition continue to be explored in experimental animal models of SCI. Thesediscoveries encompass portable and reliable urodynamic monitoring, percutaneous orimplantable devices for specific neuromodulation of bladder control, and thecombination of these elements into biofeedback devices to approach restored bladderfunction after SCI. As described, innovations in urodynamic testing aim to guidecare management in real time for men with incomplete SCI. Several groups haverecently used porcine models of bladder physiology to investigate submucosalimplantation of sensors for localized bladder function . ImplantAmbulatory urodynamic monitoring (AUM) is used to permit specificmeasurement of functional status in clinic. However, sensitivity and portabilityremain significant challenges since the aim of AUM is to reproduce bladderdysfunction in the clinic setting and requires connection to an external powersource. For increased sensitivity, wearable telemetric devices would provide morerelevant functional assessment during the course of a patient’s dailyactivities . Novel dReal-time monitoring, notification, and subsequent intervention forincreased intravesical pressure would enable timely voiding and minimize theincidence of complications in patients with SCI . Treatme6.The management of NLUTD after SCI in males is continuing to evolve. Standarddiagnostic tools are being optimized for widespread applicability. In this review,we highlighted the underlying anatomical contribution and current diagnosticstrategies. We delved into current treatment approaches and highlighted the newinnovations and emerging discoveries. Research is proceeding regarding improvedtreatment options in this patient population both for men with incomplete andcomplete SCI.


Spatially resolved transcriptomic data demand new computational analysis methods to derive biological insights. Here, we comment on these associated computational challenges as well as highlight the opportunities for standardized benchmarking metrics and data-sharing infrastructure in spurring innovation moving forward. To preserve such spatial information, advances in imaging technologies have enabled high-throughput in situ, targeted transcriptomic profiling of pre-selected RNAs at molecular and single-cell resolution2. In addition, technologies based on spatially resolved RNA capture followed by sequencing have enabled non-targeted, genome-wide transcriptional profiling at the 10-100 µm pixel resolution3. Though the suitability of each spatial transcriptomics technology in addressing a particular biological question will currently involve balancing the need for experimental throughput versus spatial resolution, with current imaging-based technologies generally offering higher spatial resolution but lower experimental throughput and current sequencing-based technologies generally offering higher experimental throughput but lower spatial resolution, all of these resulting large-scale spatially resolved transcriptomic data demand new computational methods to take advantage of this new spatial information to derive biological insights.Advances in single-cell sequencing technologies have enabled high-throughput transcriptomic profiling for individual cells, allowing the characterization and discovery of transcriptionally distinct cell types and cell states. However, current protocols require dissociating cells from tissues, thereby losing potentially valuable spatial information that may inform how cell types and cell states are organized within tissues and how such organization may ultimately impact phenotype and function4, generalized linear models5, and spatial autocorrelation analysis6 have been developed to identify genes whose expression exhibits significant spatial variability. Some of these methods can also classify different patterns of spatial variation, such as linear or periodic gene expression4, as well as identify spatial features such as gene expression hotspots7. Such identification of spatially variable genes can lend insight into position-specific phenotypes as well as developmental and migration gradients. Spatial information can also augment the identification of putative cell–cell communication networks. With single-cell sequencing data, cell–cell communication inference has relied on identifying coordinated expression of known ligand–receptor pairs8. Computational methods that leverage the added spatial information from spatially resolved transcriptomic data using graph convolutional neural networks9, optimal transport approaches10, and spatial cross-correlation analysis6 can narrow down candidates to ligand–receptor pairs that are spatially colocalized, potentially indicative of autocrine or paracrine signaling. Furthermore, spatially resolved transcriptomic data with co-registered imaging data present additional sources of heterogeneity, such as morphological variability, which can be used for clustering, as differences in morphology can be a proxy for differences in cell states or other functional phenotypes such as cell cycle position, transformation, or invasiveness. Computational methods that incorporate spatial and morphological information in addition to gene expression information have been applied to further dissect heterogeneity in single-cell populations to identify clusters of single cells that are not only transcriptionally distinct but also morphologically and spatially distinct12. Although these aforementioned computational methods can be applied to both single-cell resolution and multi-cell pixel-resolution spatially resolved transcriptomic data, interpretation of the resulting trends with multi-cell pixel-resolution data will need to take into consideration potential confounding from pixels containing cells of different cell types.Given the nascency of such high-throughput spatially resolved transcriptomic technologies, new computational methods for analyzing the resulting data are still actively being developed. Already, computational methods leveraging Gaussian processes14. Integrating additional information such as cellular transcriptional composition and prior knowledge of cell type-specific gene expression can further enhance segmentation performance, particularly with crowded but transcriptionally distinct cells16. However, for cells with more complex morphologies such as neurons, additional computational methods for reliable cell segmentation are still needed. Beyond ensuring more accurate estimation of single-cell gene counts, reliable segmentation opens the door to additional downstream computational methods to incorporate subcellular spatial information. For example, by accurately accounting for the subcellular location of RNA counts, these downstream methods enable the prediction of future cellular transcriptional states by inferring RNA velocity in situ or the characterization of the subcellular spatial heterogeneity of RNAs and its functional impact18.Data from spatially resolved transcriptomic technologies present unique analytical challenges and opportunities. For spatially resolved transcriptomic data from in situ imaging-based technologies, individual identified RNA molecules must be aggregated into cells to achieve single-cell resolution transcriptomic profiling. Therefore, reliable cell segmentation is needed to fully dissect the heterogeneity of single cells in their spatial context, as well as to probe their morphological features and to characterize their intracellular variability. Several cell segmentation pipelines exist and work well with images of cells in culture or fluorescent labeled cells21 or by applying generative modeling approaches22. Although these deconvolution approaches infer the proportional representation of cell types within multi-cellular pixels, additional methods are needed to further dissect the spatial organization of cell types and enable the inference of sub-pixel spatial information.Likewise, spatially resolved transcriptomic data from sequencing-based, pixel-resolution, spatially resolved RNA capture technologies present a different set of unique analytical challenges. In particular for technologies with larger pixel sizes, transcripts from multiple cells may be captured in each spatially resolved pixel. As such, each resulting spatially resolved transcriptomic profile may reflect multiple cells of different cell types, thereby hindering the identification of cell-type-specific spatial organizational patterns. To overcome this challenge, several computational methods have been developed to deconvolve cell-type mixtures within each multi-cellular spatially resolved pixel, often by integrating the cell-type-specific transcriptomic profiles derived from a suitable single-cell reference23.Still, additional computational methods for analyzing spatially resolved transcriptomic data are needed. Notably, although computational methods have been developed to identify and characterize spatial gene expression patterns, we find that additional methods to systematically characterize and statistically evaluate how such patterns relate to anatomical features of tissues such as blood vessels or organ borders are still needed to understand the relationship between structure and phenotype. Furthermore, current computational methods generally limit spatial analysis to individual tissue sections or multiple contiguous sections from the same sample. To analyze samples collected from different individuals, time points, or perturbations, we anticipate that additional computational methods for aligning to a common coordinate system will be needed to compare, contrast, and characterize differences in spatial gene expression patterns and cellular organizationAs these spatially resolved transcriptomic technologies become more widely adopted, we anticipate that beyond the development of new computational methods for spatially informed data analysis, such computational methods must be implemented and made accessible as robust and usable software. This is needed to ensure that users can apply these technologies and analyze the resulting data effectively and efficiently. We believe the software developed to preprocess and analyze spatially resolved transcriptomic data should therefore adhere to best practices in open-source software development, such as providing adequate documentation of software functionality and maintaining responsive issue tracking. Further, support mechanisms need to be made available to promote and incentivize such adherence. Adherence to such best practices will be especially critical to ensure that these technologies and tools are accessible to researchers with more limited computational expertise. Moreover, as these spatially resolved transcriptomic technologies and protocols are further developed to enable data collection from larger tissue sections with more genes and cells across more samples, analytical algorithms and software implementations that are scalable with respect to runtime and memory will also be critical to ensure that these technologies and tools are accessible to researchers with more limited computational resources.24, further efforts are needed to encourage adoption by enhancing their ease of use, offering comparable features to existing in-house pipelines, while maintaining flexibility across available technological platforms, as well as demonstrating robustness and reproducibility across use cases.As current spatially resolved transcriptomic technologies continue to mature, we believe standardized metrics and benchmarks will need to be created, to enable comparisons across these technologies, in particular with regards to detection sensitivity, specificity, and capture efficiency. Such standardized metrics and benchmarks will be important for understanding which technologies may be better suited for specific biological questions such as those that demand detection of lowly expressed genes or single-nucleotide variations. Such standardized metrics and benchmarks will also facilitate the development of computational methods for harmonized analysis of data across multiple technologies. In particular for spatially resolved transcriptomic data from in situ imaging-based technologies, standardized metrics for reporting confidence in spot calling, gene identification, and gene-to-cell assignment remain to be established. We anticipate that such specific standardized metrics will be useful in mitigating error propagation to downstream analyses that may lead to incorrect biological interpretations. For example, errors in spot calling, gene identification, and cell segmentation may lead to inaccurate cellular gene expression counts that result in the misidentification of seemingly new, transcriptionally distinct cell types that are the result of propagated technical errors. One challenge towards establishing a set of standardized metrics for spatially resolved transcriptomic data from in situ imaging-based technologies is related to the current dearth of uniform preprocessing pipelines. Many of the current spatially resolved transcriptomic in situ imaging-based protocols rely on in-house image preprocessing pipelines. Although uniform preprocessing pipelines are being established25. Further, it makes readily accessible not only the processed gene counts but also raw sequences, as well as metadata on the machines and organisms used to generate the data and metrics regarding the quality of the data such as base call quality scores. Establishing a similar infrastructure for spatially resolved transcriptomic data from in situ imaging-based technologies may prove to be more complex given the range of protocols and modalities that exist and the sheer size of the raw imaging data as well. However, establishing such an accessible data-sharing infrastructure will be especially important for accelerating the development of computational methods to analyze such spatially resolved transcriptomic data, as it ensures the availability of a wide range of data for method testing and enables the characterization of method performance with respect to data quality. We envision that additional discussion and collaboration from the community will be needed to establish the form of processed data and range of standardized metrics most useful for all invested parties, from those interested in developing new computational methods to those interested in further enhancing the technologies, and those interested in probing deeper into datasets for biological insights.In addition, we find that an accessible and centralized infrastructure is currently still needed for sharing spatially resolved transcriptomic data, in particular from in situ imaging-based technologies. Such an accessible and centralized infrastructure already exists for RNA-sequencing dataIn conclusion, spatially resolved transcriptomic technologies offer an exciting new way of probing the intricate spatial mechanisms at play within tissue ecosystems. Computational methods are needed to enable the characterization of tissue heterogeneity using the high informational content data obtained from such spatially resolved transcriptomic technologies. Still, there remains a need for targeted perturbation, experimental validation, and investigation of generalizability to validate the insights gained from applying these computational methods. For example, although computational methods have been developed to integrate spatial and morphological information in single-cell clustering, further validation is needed to understand if new cell clusters identified through such integrative approaches represent meaningful functional heterogeneity. Furthermore, investigating the extent to which spatial and morphological characteristics of cells are independent of their gene expression can lend insights into other cell intrinsic and cell extrinsic factors that influence cell phenotype. Likewise, intracellular spatial heterogeneity and its functional consequences remain to be characterized. Ultimately, computational methods for analyzing spatially resolved transcriptomic data offer the potential to identify and characterize the heterogeneity of cells within their spatial contexts and contribute to important fundamental biological insights regarding how tissues are organized in both the healthy and diseased settings.


This session will provide updates on how the pandemic led to horrific situations in long-term care facilities and how the pandemic influenced major federal efforts to address elder abuse, neglect, and exploitation.


Pseudomonas aeruginosa (PSAR) is challenging to treat due to its multiple resistance mechanisms, limited anti-PSAR agents, and population pharmacokinetic (PK) variances. Beta-lactam antibiotics (BLA) are commonly used to treat PSAR infections and although they have a wide therapeutic index, suboptimal exposures may lead to treatment failure and antimicrobial resistance while high exposure may result in adverse effects. Certain patient populations may benefit from BLA therapeutic drug monitoring (TDM) due to their significant PK variability. The purpose of this study was to compare clinical outcomes in patients with PSAR pneumonia (PNA) or bloodstream infection (BSI) receiving BLA with and without the guidance of TDM.Retrospective, parallel cohort study conducted at UF Shands Gainesville and UF Health Jacksonville evaluating five years of patients with PSAR PNA or BSI. TDM group was defined for routine BLA TDM compared to nonroutine BLA TDM service (non-TDM). Patients were excluded if they died before a culture result, transferred in with a positive PSAR culture, were transplant recipients, cystic fibrosis or burn injury patients. The primary outcome was a composite of presumed clinical cure defined as the absence of the following: all-cause in-hospital mortality, escalation, and/or additional antimicrobial therapy for PSAR infection after 48 hours of treatment with primary susceptible regimen due to worsening clinical status or transfer to a higher level of care.Two-hundred patients were included . The overall primary composite outcome of presumed clinical cure occurred in 73% of patients . A post-hoc multivariate analysis was conducted to assess predictors of not attaining clinical cure.While there was no difference in the primary composite outcome of presumed clinical cure, future studies can use these data to assess TDM patient selection and whether a bundled care approach of BLA regimens with known clinical benefit, early TDM-guided dose optimization, and continued clinical assessment improves outcomes in patients with PSAR PNA or BSI compared to use of each modality individually. All Authors: No reported disclosures


Difficulties in parent-child interaction are easily observed and are a potential target for early intervention. This study aimed to assess the utility of observation of parent-child interaction in the first year of life in identifying children at risk of developing later psychopathology, using a rigorous systematic review method.EMBASE, CINAHL, PsycINFO, MIDIRS, MEDLINE and Cochrane Library databases were searched using MeSH terms and keywords, and reference lists screened. Two authors independently reviewed papers for inclusion and completed data extraction. All peer reviewed papers studying the association between an independent observation of parent-child interaction and later childhood psychopathology in community-based samples were included. Studies based on ‘high risk’ samples  were excluded. Results were synthesised qualitatively due to high heterogeneity.18,226 papers were identified, nine were included in this study. Childhood psychopathology was associated with fewer positive parent-infant interactions, lower parent vocalisation frequency and lower levels of adult speech and activity. Maternal sensitivity was inversely related to separation anxiety and oppositional defiant/conduct disorders were associated with lower shared look rates. Disruptive behaviour disorders were associated with higher frequency of child vocalisation. Pervasive developmental disorders were associated with ‘abnormal’ maternal infant interactions, as assessed by community health nurses using a standardised measure.Included studies reported small samples, and several of these samples overlapped. Some studies were of poor quality, but were included due to a paucity of available data. The findings may therefore have limited generalisability. Difficulties in parent-child interaction are easily observed and assessments could be made by non-specialists such as health visitors or general practitioners. Such difficulties may be an early indicator of later childhood psychopathology. Childhood psychiatric diagnoses (with the exception of Autistic Spectrum Disorders) appear associated with level of maternal activity . Assessments may identify at-risk families for early intervention, but further work is required to develop and validate reliable methods for risk stratification in community-based practice.