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Human brain undergoes rapid growth during the first few years of life. While previous research has employed graph theory to study early brain development, it has mostly focused on the topological attributes of the whole brain. However, examining regional graph-theory features may provide unique insights into the development of cognitive abilities. Utilizing a large and longitudinal rsfMRI dataset from the UNC/UMN Baby Connectome Project, we investigated the developmental trajectories of regional efficiency and evaluated the relationships between these changes and cognitive abilities using Mullen Scales of Early Learning during the first twenty-eight months of life. Our results revealed a complex and spatiotemporally heterogeneous development pattern of regional global and local efficiency during this age period. Furthermore, we found that the trajectories of the regional global efficiency at the left temporal occipital fusiform and bilateral occipital fusiform gyri were positively associated with cognitive abilities, including visual reception, expressive language, receptive language, and early learning composite scores (P

Presenilin 1 (PS1) is a central component of γ-secretase, an enzymatic complex involved in the generation of the amyloid-β (Aβ) peptide that deposits as plaques in the Alzheimer’s disease (AD) brain. The M146L mutation in the PS1 gene (PSEN1) leads to an autosomal dominant form of early-onset AD by promoting a relative increase in the generation of the more aggregation-prone Aβ42. This change is evident not only in the brain but also in peripheral cells of mutation carriers. In this study we used the CRISPR-Cas9 system from Streptococcus pyogenes to selectively disrupt the PSEN1M146L allele in human fibroblasts. A disruption of more than 50% of mutant alleles was observed in all CRISPR-Cas9-treated samples, resulting in reduced extracellular Aβ42/40 ratios. Fluorescence resonance energy transfer-based conformation and western blot analyses indicated that CRISPR-Cas9 treatment also affects the overall PS1 conformation and reduces PS1 levels. Moreover, our guide RNA did not lead to any detectable editing at the highest-ranking candidate off-target sites identified by ONE-seq and CIRCLE-seq. Overall, our data support the effectiveness of CRISPR-Cas9 in selectively targeting the PSEN1M146L allele and counteracting the AD-associated phenotype. We believe that this system could be developed into a therapeutic strategy for patients with this and other dominant mutations leading to early-onset AD.

Childhood neuroblastomas exhibit plasticity between an undifferentiated neural crest–like mesenchymal cell state and a more differentiated sympathetic adrenergic cell state. These cell states are governed by autoregulatory transcriptional loops called core regulatory circuitries (CRCs), which drive the early development of sympathetic neuronal progenitors from migratory neural crest cells during embryogenesis. The adrenergic cell identity of neuroblastoma requires LMO1 as a transcriptional cofactor. Both LMO1 expression levels and the risk of developing neuroblastoma in children are associated with a single nucleotide polymorphism, G/T, that affects a GATA motif in the first intron of LMO1. Here, we showed that WT zebrafish with the GATA genotype developed adrenergic neuroblastoma, while knock-in of the protective TATA allele at this locus reduced the penetrance of MYCN-driven tumors, which were restricted to the mesenchymal cell state. Whole genome sequencing of childhood neuroblastomas demonstrated that TATA/TATA tumors also exhibited a mesenchymal cell state and were low risk at diagnosis. Thus, conversion of the regulatory GATA to a TATA allele in the first intron of LMO1 reduced the neuroblastoma-initiation rate by preventing formation of the adrenergic cell state. This mechanism was conserved over 400 million years of evolution, separating zebrafish and humans.

BackgroundAdult primary retroperitoneal sarcomas (RPSs) are a group of heterogeneous tumors with different histological subtypes. Comprehensive genomic profiling (CGP) analyses have recently provided significant insights into the biology of sarcomas by identifying genomic alterations (GAs) which could benefit from targeted therapies. MethodsRPS were evaluated by CGP using next-generation sequencing of up to 406 cancer-related genes. Tumor mutational burden (TMB) was determined on 0.83 to 1.14 mut/Mb of sequenced DNA. Finally, PD-L1 expression was determined. ResultsOverall, 296 cases of primary RPS were analyzed. Liposarcoma (LPS) subtype had more GA/tumor than leiomyosarcoma (LMS) subtypes, with follicular dendritic cell sarcomas harboring the highest and synovial sarcomas the lowest. TP53 and Rb1 alterations were the highest in LMS, and CDK4/6 and MDM2 in LPS. However, both the TMB and targetable GA rates were low across subtypes. PD-L1 immunostaining was low positive in 21% and high positive in 5% of patients, respectively. ConclusionsCGP analysis revealed that potentially actionable genomic targets were rare in our cohort of RPS. Moreover, RPSs seem less likely to respond to immune checkpoint inhibitors based on putative biomarkers status. Nevertheless, genomic stratification according to histological subtypes led to description of GAs that can inform future clinical trials design.

Abstract Background While chiropractic care is most commonly provided within a private practice context, the ‘traditional’ solo practice is now uncommon. Chiropractors, manual therapists and related health professionals commonly work within the same practice bringing obvious advantages to both the practitioners and their patients. However, multi-practitioner, multi-disciplinary clinics also carry often unrecognized liabilities. We refer here to vicarious liability and non-delegable duties. Vicarious liability refers to the strict liability imposed on one person for the negligent acts of another person. The typical example is an employer being held vicariously liable to the negligent acts of an employee. However, vicarious liability can arise outside of the employer-employee relationship. For example, under non-delegable duty provisions, an entity owing a non-delegable duty can be liable for an independent contractor’s wrongdoing. After a plain English explanation of this complex area of law, we provide seven scenarios to demonstrate how vicarious liability can envelop practice principals when things go wrong. We also make suggestions for risk mitigation. Conclusion Practice owners may unexpectedly find themselves legally liable for another’s actions with dire consequences. A knowledge of vicarious liability along with implementing risk mitigation strategies has the potential to minimize the likelihood of this unwanted event. Recommendations are made to this end.

Abstract Optical manipulation of colloidal systems is of high interest for both fundamental studies and practical applications. It has been shown that optically induced thermophoresis and nonlinear interactions can significantly affect the properties of dense colloidal media. However, macroscopic scale phenomena can also be generated at thermal equilibrium. Here, we demonstrate that steady-state variations of particle density can be created over large, three-dimensional regions by appropriately structured external optical fields. We prove analytically and experimentally that an optical vortex beam can dynamically control the spatial density of microscopic particles along the direction of its propagation. We show that these artificial steady-states can be generated at will and can be maintained indefinitely, which can be beneficial for applications such as path clearing and mass transportation.

Prime editing guide RNA design is more complex than for standard CRISPR-based nucleases or base editors. Here the authors present PrimeDesign and PrimeVar for the rapid and simplified design of pegRNA and ngRNA combinations.

Abstract Background The COVID-19 pandemic has seen the emergence of unsubstantiated claims by vertebral subluxation-based chiropractors that spinal manipulative therapy has a role to play in prevention by enhancing the body’s immune function. We contend that these claims are unprofessional and demonstrate a disturbing lack of insight into the doctrine of informed consent. As such it is timely to review how informed consent has evolved and continues to do so and also to discuss the attendant implications for contemporary health practitioner practice. We review the origins of informed consent and trace the duty of disclosure and materiality through landmark medical consent cases in four common law (case law) jurisdictions. The duty of disclosure has evolved from a patriarchal exercise to one in which patient autonomy in clinical decision making is paramount. Passing time has seen the duty of disclosure evolve to include non-medical aspects that may influence the delivery of care. We argue that a patient cannot provide valid informed consent for the removal of vertebral subluxation. Further, vertebral subluxation care cannot meet code of conduct standards because it lacks an evidence base and is practitioner-centered. The uptake of the expanded duty of disclosure has been slow and incomplete by practitioners and regulators. The expanded duty of disclosure has implications, both educative and punitive for regulators, chiropractic educators and professional associations. We discuss how practitioners and regulators can be informed by other sources such as consumer law. For regulators, reviewing and updating informed consent requirements is required. For practitioners it may necessitate disclosure of health status, conflict of interest when recommending “inhouse” products, recency of training after attending continuing professional development, practice patterns, personal interests and disciplinary findings. Conclusion Ultimately such matters are informed by the deliberations of the courts. It is our opinion that the duty of a mature profession to critically self-evaluate and respond in the best interests of the patient before these matters arrive in court.

Most individuals affected with DYT1 dystonia have a heterozygous 3-bp deletion in the TOR1A gene (c.907_909delGAG). The mutation appears to act through a dominant-negative mechanism compromising normal torsinA function, and it is proposed that reducing mutant torsinA may normalize torsinA activity. In this study, we used an engineered Cas9 variant from Streptococcus pyogenes (SpCas9-VRQR) to target the mutation in the TOR1A gene in order to disrupt mutant torsinA in DYT1 patient fibroblasts. Selective targeting of the DYT1 allele was highly efficient with most common non-homologous end joining (NHEJ) edits, leading to a predicted premature stop codon with loss of the torsinA C terminus (delta 302–332 aa). Structural analysis predicted a functionally inactive status of this truncated torsinA due to the loss of residues associated with ATPase activity and binding to LULL1. Immunoblotting showed a reduction of the torsinA protein level in Cas9-edited DYT1 fibroblasts, and a functional assay using HSV infection indicated a phenotypic recovery toward that observed in control fibroblasts. These findings suggest that the selective disruption of the mutant TOR1A allele using CRISPR-Cas9 inactivates mutant torsinA, allowing the remaining wild-type torsinA to exert normal function.

Abstract Background Chiropractic emerged in 1895 and was promoted as a viable health care substitute in direct competition with the medical profession. This was an era when there was a belief that one cause and one cure for all disease would be discovered. The chiropractic version was a theory that most diseases were caused by subluxated (slightly displaced) vertebrae interfering with “nerve vibrations” (a supernatural, vital force) and could be cured by adjusting (repositioning) vertebrae, thereby removing the interference with the body’s inherent capacity to heal. DD Palmer, the originator of chiropractic, established chiropractic based on vitalistic principles. Anecdotally, the authors have observed that many chiropractors who overtly claim to be “vitalists” cannot define the term. Therefore, we sought the origins of vitalism and to examine its effects on chiropractic today. Discussion Vitalism arose out of human curiosity around the biggest questions: Where do we come from? What is life? For some, life was derived from an unknown and unknowable vital force. For others, a vital force was a placeholder, a piece of knowledge not yet grasped but attainable. Developments in science have demonstrated there is no longer a need to invoke vitalistic entities as either explanations or hypotheses for biological phenomena. Nevertheless, vitalism remains within chiropractic. In this examination of vitalism within chiropractic we explore the history of vitalism, vitalism within chiropractic and whether a vitalistic ideology is compatible with the legal and ethical requirements for registered health care professionals such as chiropractors. Conclusion Vitalism has had many meanings throughout the centuries of recorded history. Though only vaguely defined by chiropractors, vitalism, as a representation of supernatural force and therefore an untestable hypothesis, sits at the heart of the divisions within chiropractic and acts as an impediment to chiropractic legitimacy, cultural authority and integration into mainstream health care.

Abstract Background Internet analytics are increasingly being integrated into public health regulation. One specific application is to monitor compliance of website and social media activity with respect to jurisdictional regulations. These data may then identify breaches of compliance and inform disciplinary actions. Our study aimed to evaluate the novel use of internet analytics by a Canadian chiropractic regulator to determine their registrants compliance with three regulations related to specific health conditions, pregnancy conditions and most recently, claims of improved immunity during the COVID-19 crisis. Methods A customized internet search tool (Market Review Tool, MRT) was used by the College of Chiropractors of British Columbia (CCBC), Canada to audit registrants websites and social media activity. The audits extracted words whose use within specific contexts is not permitted under CCBC guidelines. The MRT was first used in October of 2018 to identify words related to specific health conditions. The MRT was again used in December 2019 for words related to pregnancy and most recently in March 2020 for words related to COVID-19. In these three MRT applications, potential cases of word misuse were evaluated by the regulator who then notified the practitioner to comply with existing regulations by a specific date. The MRT was then used on that date to determine compliance. Those found to be non-compliant were referred to the regulator’s inquiry committee. We mapped this process and reported the outcomes with permission of the regulator. Results In September 2018, 250 inappropriate mentions of specific health conditions were detected from approximately 1250 registrants with 2 failing to comply. The second scan for pregnancy related terms of approximately1350 practitioners revealed 83 inappropriate mentions. Following notification, all 83 cases were compliant within the specified timeframe. Regarding COVID-19 related words, 97 inappropriate mentions of the word “immune” were detected from 1350 registrants with 7 cases of non-compliance. Conclusion Internet analytics are an effective way for regulators to monitor internet activity to protect the public from misleading statements. The processes described were effective at bringing about rapid practitioner compliance. Given the increasing volume of internet activity by healthcare professionals, internet analytics are an important addition for health care regulators to protect the public they serve.

Summary: Repeat elements can be dysregulated at a genome-wide scale in human diseases. For example, in Ewing sarcoma, hundreds of inert GGAA repeats can be converted into active enhancers when bound by EWS-FLI1. Here we show that fusions between EWS and GGAA-repeat-targeted engineered zinc finger arrays (ZFAs) can function at least as efficiently as EWS-FLI1 for converting hundreds of GGAA repeats into active enhancers in a Ewing sarcoma precursor cell model. Furthermore, a fusion of a KRAB domain to a ZFA can silence GGAA microsatellite enhancers genome wide in Ewing sarcoma cells, thereby reducing expression of EWS-FLI1-activated genes. Remarkably, this KRAB-ZFA fusion showed selective toxicity against Ewing sarcoma cells compared with non-Ewing cancer cells, consistent with its Ewing sarcoma-specific impact on the transcriptome. These findings demonstrate the value of ZFAs for functional annotation of repeats and illustrate how aberrant microsatellite activities might be regulated for potential therapeutic applications.

Germline mutations within the Krebs cycle enzyme genes fumarate hydratase (FH) or succinate dehydrogenase (SDHB, SDHC, SDHD) are associated with an increased risk of aggressive and early metastasizing variants of renal cell carcinoma (RCC). These RCCs express significantly increased levels of intracellular fumarate or succinate that inhibit 2-oxoglutarate-dependent dioxygenases, such as the TET enzymes that regulate DNA methylation. This study evaluated the genome-wide methylation profiles of 34 RCCs from patients with RCC susceptibility syndromes and 11 associated normal samples using the Illumina HumanMethylation450 BeadChip. All the HLRCC (FH mutated) and SDHB-RCC (SDHB mutated) tumors demonstrated a distinct CpG island methylator phenotype (CIMP). HLRCC tumors demonstrated an extensive and relatively uniform level of hypermethylation that showed some correlation with tumor size. SDHB-RCC demonstrated a lesser and more varied pattern of hypermethylation that overlapped in part with the HLRCC hypermethylation. Combined methylation and mRNA expression analysis of the HLRCC tumors demonstrated hypermethylation and transcription downregulation of genes associated with the HIF pathway, HIF3A and CITED4, the WNT pathway, SFRP1, and epithelial-to-mesenchymal transition and MYC expression, OVOL1. These observations were confirmed in the TCGA CIMP-RCC tumors. A selected panel of probes could identify the CIMP tumors and differentiate between HLRCC and SDHB-RCC tumors. This panel accurately detected all CIMP-RCC tumors within the TCGA RCC cohort, identifying them as HLRCC -like, and could potentially be used to create a liquid biopsy-based screening tool. The CIMP signature in these aggressive tumors could provide both a useful biomarker for diagnosis and a target for novel therapies.

Childhood lead exposure remains a key health concern for officials worldwide, contributing some 600,000 new cases of intellectually disabled children annually. Most children affected by high exposure to lead live in low- and middle-income countries. The leaded gasoline phase out in India was completed in 2000. Yet, in 2020, an estimated 275 million children aged 0 to 9 years had blood lead levels (BLLs) ≥ 5 μg/dL known to adversely affect intelligence and behavior. Lead sources reported in India include spices, cookware, paint, traditional medicines and cosmetics, and lead-acid battery recycling and repair. However, their relative contribution has not been characterized. More than 200 lead pollution sites related to battery recycling and repair activities were identified in Bihar and Jharkhand, India. Ninety percent of the recycling sites had soil lead concentrations exceeding the US Environmental Protection Agency’s standards. We compared blood and environmental lead levels in two groups of children in Patna, Bihar. Households in proximity to battery recycling operations (Proximal n = 67) versus households distal to these operations (Distal n = 68). The average age of children was 40 months; 46% were female. Overall, the geometric mean (GM) BLL was 11.6 μg/dL. GM BLLs of children in Proximal and Distal households were not significantly different (10.2 μg/dL vs. 13.1 μg/dL respectively; p≤0.07). About 87% children, 56 Proximal and 62 Distal had BLLs ≥5 μg/dl. Lead concentrations in environmental samples were significantly higher in Proximal households (soil mean 9.8 vs. 1.6 μg/ft2; dust mean 52.9 vs. 29.9 μg/ft2 p

In-depth characterization of adeno-associated virus (AAV)-mediated CRISPR delivery is still lacking. Here, the authors show high levels of integration into Cas9-induced double-strand breaks (DSBs) in therapeutically relevant genes in vivo.

Abstract Background Society expects professionals to promote their businesses in an ethical manner, refraining from misleading or deceptive marketing due to the potential to harm members of the community. In Australia this expectation resides in the Australian registration board advertising guidelines or the Health Practitioner Regulation National Law. Registration board data indicate there are many health care professionals failing to meet these expectations. The aims of this research were to determine the frequency, type and nature of at-risk advertising by Australian chiropractors and physiotherapists and whether there is a correlation between professional association membership and advertising guideline compliance. Method A cross sectional audit examining practitioner advertising was performed on representative samples of Australian chiropractors and physiotherapists. Two auditors examined advertising by 380 physiotherapists and 359 chiropractors for material potentially in breach of the regulatory authorities’ advertising guidelines. The advertising appeared on practitioner websites and linked Facebook pages. Results Two-hundred and fifty-eight (72%) audited chiropractors and 231 (61%) audited physiotherapists had breaches of the Advertising Guidelines on their websites and linked Facebook pages. The frequency of breaches by chiropractors was higher. The type and nature of the breaches by chiropractors was potentially more harmful. Membership in a professional association influenced neither the frequency nor the severity of breaches with chiropractors. Discussion Advertising breaches were common in both samples even though regulators and professional associations provide practitioners with explicit information on how to comply with advertising guidelines. Breaches by chiropractors were more numerous and more serious due to their greater potential to lead consumers to make inappropriate and potentially harmful healthcare decisions. Stronger enforcement strategies may have a positive effect on compliance.

Metal dissolution from atmospheric aerosol deposition plays an important role in enhancing and inhibiting phytoplankton growth and community structure. Here, the authors review the impacts of trace metal leaching from natural and anthropogenic aerosols on marine microorganisms over short and long timescales.

The APPswe (Swedish) mutation in the amyloid precursor protein (APP) gene causes dominantly inherited Alzheimer’s disease (AD) as a result of increased β-secretase cleavage of the amyloid-β (Aβ) precursor protein. This leads to abnormally high Aβ levels, not only in brain but also in peripheral tissues of mutation carriers. Here, we selectively disrupted the human mutant APPSW allele using CRISPR. By applying CRISPR/Cas9 from Streptococcus pyogenes, we generated allele-specific deletions of either APPSW or APPWT. As measured by ELISA, conditioned media of targeted patient-derived fibroblasts displayed an approximate 60% reduction in secreted Aβ. Next, coding sequences for the APPSW-specific guide RNA (gRNA) and Cas9 were packaged into separate adeno-associated viral (AAV) vectors. Site-specific indel formation was achieved both in primary neurons isolated from APPSW transgenic mouse embryos (Tg2576) and after co-injection of these vectors into hippocampus of adult mice. Taken together, we here present proof-of-concept data that CRISPR/Cas9 can selectively disrupt the APPSW allele both ex vivo and in vivo—and thereby decrease pathogenic Aβ. Hence, this system may have the potential to be developed as a tool for gene therapy against AD caused by APPswe and other point mutations associated with increased Aβ.

Abstract Since its inception, the chiropractic profession has been divided along ideological fault lines. These divisions have led to a profession wide schism, which has limited mainstream acceptance, utilisation, social authority and integration. The authors explore the historical origins of this schism, taking time to consider historical context, religiosity, perpetuating factors, logical fallacies and siege mentality. Evidence is then provided for a way forward, based on the positioning of chiropractors as mainstream partners in health care.

Abstract Aim This paper examines appeal to fear in general: its perceived positive aspects, its negative characteristics, its appropriate as well as its fallacious use. Background Appeal to fear is a commonly used marketing method that attempts to change behaviour by creating anxiety in those receiving a fearful message. It is regularly used in public health initiatives such as anti-smoking, anti-drunk driving campaigns as well as in hypertension awareness campaigns. Some chiropractors appear to use appeal to fear to promote subluxation awareness and thereby encourage the use of chiropractic treatment. Research supporting its use is equivocal; nevertheless, when used judiciously, appeal to fear probably has sufficient strengths to warrant its continued conditional use. When used to promote care for which there is no supporting evidence, its use is fallacious. Discussion Appeal to fear has been used in health promotion campaigns for sixty years or more with the intent of modifying behaviours. While there is evidence to suggest that appeal to fear may motivate some individuals to modify offending behaviour or adopt recommended behaviour there is growing resistance to the use of appeal to fear on ethical and psychological grounds. Using appeal to fear as a tool of persuasion can be valid or fallacious depending on the truth of the premises within the argument. When used to raise awareness about genuine health concerns such as smoking, drunk driving and hypertension appeal to fear is considered to be a valid approach with certain caveats. However, when appeal to fear, not based on evidence or reason, is used as motivator to get others to accept unnecessary interventions for unproven disorders, the use of appeal to fear is fallacious. Conclusion In spite of the evidence against its use, it seems likely that appeal to fear will continue to be used in conjunction with other public awareness initiatives to modify recognized detrimental behaviours such as smoking and drunk driving as well as silent killers such as hypertension. However, when used to promote a treatment that has no evidentiary basis such as subluxation based practice in chiropractic the appeal to fear is a fallacy and must be stopped.

Endophyte-infected tall fescue (E+) produces ergovaline and ergovalinine, which are mycotoxins that act as dopamine agonists to suppress prolactin and induce vasoconstriction. The experiment was designed as a 3 × 2 × 2 factorial with DRD2 genotype (AA, AG, GG), fescue seed (endophyte-free, E− or endophyte-infected, E+), stage of gestation (MID, d (day) 35–85; LATE, d 86–parturition) and all interactions in the model. Pregnant Suffolk ewes (n = 60) were stratified by genotype and fed E+ or E− seed in a total mixed ration according to treatment assignment. Serum prolactin concentrations were lower (p < 0.05) in ewes fed E+ seed but did not differ by maternal DRD2 genotype or two-way interaction. Lamb birth weight was lower (p < 0.05) in ewes fed E+ seed in last trimester. Pre-weaning growth rate, milk production and total weaning weight was reduced (p < 0.05) in ewes fed E+ fescue seed during MID and LATE gestation. Ingestion of ergovaline/ergovalinine in last trimester reduces lamb birth weight; however, lamb growth rate, milk production and total weaning weight are reduced in all ewes fed E+ during mid and last trimester.

Weaned lambs (n = 82), born to ewes fed endophyte-free (E−) or endophyte-infected (E+; 1.77 mg hd−1 d−1 ergovaline + ergovalinine) tall fescue seed from d 35 to 85 of gestation (MID) and/or d 86 of gestation to parturition (LATE), were used to examine how ergot alkaloid exposure during fetal development altered subsequent puberty attainment or carcass quality. Lambs were weaned at 75 d of age and separated by sex to assess puberty in ewe lambs (n = 39) and to evaluate growth, carcass and meat quality in wethers (n = 43). Data were analyzed with maternal fescue treatment, stage of gestation, and two-way interaction in the model. Age at puberty tended (P = 0.06) to be longer for ewe lambs born to dams fed E+ fescue during LATE gestation versus those fed E−. Post-weaning average daily gain tended to be higher (P = 0.07) for wether lambs born to dams fed E+ fescue seed during MID gestation compared to E−. Exposure to ergot alkaloids during fetal growth altered (P < 0.10) longissimus muscle weight and color, lipid deposition, fatty acid composition, and shear force values of semimembranosus muscle in wether lambs. These results indicate that exposure to ergot alkaloids in utero does alter subsequent post-weaning puberty attainment and body composition in offspring.

Numerous studies have been conducted to delineate the early development of different functional networks, based on measuring the temporal synchronization of spontaneous blood oxygenation level-dependent (BOLD) signals acquired using resting state functional MRI (rsfMRI). However, little attention has been paid to the change of the frequency properties of these signals during early brain development. Such frequency properties may reflect important physiological changes and potentially have significant cognitive consequences. In this study, leveraging a large (N = 86 subjects), longitudinal sample of human infants scanned during the first two years of life, we aimed to specifically delineate the developmental changes of the frequency characteristics of spontaneous BOLD signals. Both whole-brain and network-level examinations were carried out and the frequency–behavior relationship was explored. Our results revealed a clear right-ward shift of BOLD signal frequency during the first year of life. Moreover, the power at the peak-frequency for sensorimotor and lateral visual networks correlates with domain-specific Mullen Scales in 1-year-olds, suggesting the behavioral significance of the BOLD signal frequency during infancy. Findings from this study shed light into early functional brain development and provide a new perspective for future searches for functional developmental abnormalities.

Peripheral blood as a surrogate tissue for transcriptome profiling holds great promise for the discovery of diagnostic and prognostic disease biomarkers, particularly when target tissues of disease are not readily available. To maximize the reliability of gene expression data generated from clinical blood samples, both the sample collection and the microarray probe generation methods should be optimized to provide stabilized, reproducible and representative gene expression profiles faithfully representing the transcriptional profiles of the constituent blood cell types present in the circulation. Given the increasing innovation in this field in recent years, we investigated a combination of methodological advances in both RNA stabilisation and microarray probe generation with the goal of achieving robust, reliable and representative transcriptional profiles from whole blood. To assess the whole blood profiles, the transcriptomes of purified blood cell types were measured and compared with the global transcriptomes measured in whole blood. The results demonstrate that a combination of PAXgeneTM RNA stabilising technology and single-stranded cDNA probe generation afforded by the NuGEN Ovation RNA amplification system V2TM enables an approach that yields faithful representation of specific hematopoietic cell lineage transcriptomes in whole blood without the necessity for prior sample fractionation, cell enrichment or globin reduction. Storage stability assessments of the PAXgeneTM blood samples also advocate a short, fixed room temperature storage time for all PAXgeneTM blood samples collected for the purposes of global transcriptional profiling in clinical studies.

Counter-selectable markers can be used in two-hybrid systems to search libraries for a protein or compound that interferes with a macromolecular interaction or to identify macromolecules from a population that cannot mediate a particular interaction. In this report, we describe the adaptation of the yeast URA3/5-FOA counter-selection system for use in bacterial interaction trap experiments. Two different URA3 reporter systems were developed that allow robust counter-selection: (i) a single copy F′ episome reporter and (ii) a co-cistronic HIS3-URA3 reporter vector. The HIS3-URA3 reporter can be used for either positive or negative selections in appropriate bacterial strains. These reagents extend the utility of the bacterial two-hybrid system as an alternative to its yeast-based counterpart.

Purpose: Limited information is available comparing toxicity of accelerated radiotherapy (RT) to those of standard fractionation RT for early stage breast cancer, after breast conserving surgery. We report early and late toxicities of two prone regimens of accelerated intensity modulated radiation therapy (IMRT) with a concomitant boost (CB) to the tumor bed delivered over 3 or 5 weeks as compared to standard 6 week RT with a sequential electron boost.Methods: From 2/2003 to 12/2007, 169 consecutive patients with Stage I-II breast cancer were offered the choice to undergo prone RT with either: a 6 weeks standard RT regimen of 46 Gy/23 fractions (fx) to the whole breast (WB), followed by a14 Gy sequential boost (SB) to the tumor bed (6wSB) ; a 5 week regimen of 50 Gy to WB a with a IMRT CB of 6.25 Gy in 25 fx (5wCB); or a 3 week protocol of 40.5 Gy to WB with an IMRT CB of 7.5 Gy in 15 fx (3wCB). These regimens were estimated as biologically equivalent, based on alpha/beta=4 for tumor control. Toxicities were reported using RTOG and LENT/SOMA scoring.Results: 51/169 patients chose standard 6wSB, 28 selected 5wCB, and 90 enrolled in 3wCB protocol. Maximum acute toxicity was grade 3 dermatitis in 4% of the patients in the 6wSB compared 1% in 3wCB. In general, acute complications (breast pain, fatigue, and dermatitis) were significantly less in the 3wCB than in the other schedules (P

Background Mediastinal gray zone lymphoma is a newly recognized entity with transitional morphological and immunophenotypic features between the nodular sclerosis subtype of Hodgkin’s lymphoma and primary mediastinal large B-cell lymphoma. Diagnostic criteria for mediastinal gray zone lymphoma are still challenging, and the optimal therapy is as yet undetermined. Epigenetic changes have been implicated in the loss of the B-cell program in classical Hodgkin’s lymphoma, and might provide a basis for the immunophenotypic alterations seen in mediastinal gray zone lymphoma.Design and Methods We performed a large-scale DNA methylation analysis of microdissected tumor cells to investigate the biological underpinnings of mediastinal gray zone lymphoma and its association with the related entities classical Hodgkin’s lymphoma and primary mediastinal large B-cell lymphoma, making comparisons with the presumptively less related diffuse large B-cell lymphoma.Results Principal component analysis demonstrated that mediastinal gray zone lymphoma has a distinct epigenetic profile intermediate between classical Hodgkin’s lymphoma and primary mediastinal large B-cell lymphoma but remarkably different from that of diffuse large B-cell lymphoma. Analysis of common hypo- and hypermethylated CpG targets in mediastinal gray zone lymphoma, classical Hodgkin’s lymphoma, primary mediastinal large B-cell lymphoma and diffuse large B-cell lymphoma was performed and confirmed the findings of the principal component analysis. Based on the epigenetic profiles we were able to establish class prediction models utilizing genes such as HOXA5, MMP9, EPHA7 and DAPK1 which could distinguish between mediastinal gray zone lymphoma, classical Hodgkin’s lymphoma and primary mediastinal large B-cell lymphoma with a final combined prediction of 100%.Conclusions Our data confirm a close relationship between mediastinal gray zone lymphoma and both classical Hodgkin’s lymphoma and primary mediastinal large B-cell lymphoma. However, important differences were observed as well, allowing a clear distinction from both parent entities. Thus, mediastinal gray zone lymphoma cannot be assigned to either classical Hodgkin’s lymphoma or primary mediastinal large B-cell lymphoma, validating the decision to create an intermediate category in the World Health Organization classification.

In 2020, lung cancer was the tumor type with the highest mortality, with 1.8 million deaths worldwide. (1) The most common type of lung cancer is non-small cell lung carcinoma [...], Context.--Multiple procedural techniques can be used to obtain tissue to create a formalin-fixed, paraffin-embedded specimen for comprehensive genomic profiling (CGP) in lung cancer. The literature is mixed on whether the procedure affects CGP success. Objective.--To examine whether biopsy procedure affects lung cancer CGP success. Design.--This was a cross-sectional study of all patients with lung cancer whose specimens were submitted for CGP between January and February 2020. Multiple quality control metrics were used to determine whether cases were successfully profiled. Results.--In all, 3312 samples were identified. Overall, 67.5% (2236 of 3312) of samples were obtained from biopsies, 13.0% (432 of 3312) from fine-needle aspirations (FNAs), 9.7% (321 of 3312) from resections, 5.3% (174 of 3312) from fluid cytology cell blocks, and 4.5% (149 of 3312) from bone biopsies. Overall, 70.1% (2321 of 3312) of cases passed CGP, 15.4% (510 of 3312) of cases were released as qualified reports, and 14.5% (481 of 3312) of cases failed CGP. Resection samples were the most likely to be successfully sequenced, failing in only 2.8% (9 of 321) of instances, while fluid cytology specimens were the least likely, failing in 23.0% (40 of 174) of instances. Biopsy (14.5% [324 of 2236]), FNA (18.5% [80 of 432]), and bone biopsy (18.8% [28 of 149]) specimens failed at intermediate frequencies. On multivariate logistic regression analysis of CGP success on specimen type, fluid cytology (odds ratio [OR], 0.08; 95% CI, 0.03-0.19), biopsy (OR, 0.25; 95% CI, 0.11-0.52), FNA (OR, 0.14; 95% CI, 0.06-0.32), and bone biopsy (OR, 0.07; 95% CI, 0.03-0.17) specimens had decreased odds of CGP success relative to resection samples. Among patients with successfully sequenced samples, 48.0% were eligible for at least 1 therapy, based on a companion diagnostic or National Comprehensive Cancer Network biomarker. Conclusions.--The method of tissue acquisition was an important preanalytic factor that determined whether a sample would be successfully sequenced and whether a clinically actionable genomic alteration would be detected. doi: 10.5858/arpa.2021-0313-OA

Climate-driven depletion of ocean oxygen strongly impacts the global cycles of carbon and nutrients as well as the survival of many animal species. One of the main uncertainties in predicting changes to marine oxygen levels is the regulation of the biological respiration demand associated with the biological pump. Derived from the Redfield ratio, the molar ratio of oxygen to organic carbon consumed during respiration (i.e., the respiration quotient, [Formula: see text]) is consistently assumed constant but rarely, if ever, measured. Using a prognostic Earth system model, we show that a 0.1 increase in the respiration quotient from 1.0 leads to a 2.3% decline in global oxygen, a large expansion of low-oxygen zones, additional water column denitrification of 38 Tg N/y, and the loss of fixed nitrogen and carbon production in the ocean. We then present direct chemical measurements of [Formula: see text] using a Pacific Ocean meridional transect crossing all major surface biome types. The observed [Formula: see text] has a positive correlation with temperature, and regional mean values differ significantly from Redfield proportions. Finally, an independent global inverse model analysis constrained with nutrients, oxygen, and carbon concentrations supports a positive temperature dependence of [Formula: see text] in exported organic matter. We provide evidence against the common assumption of a static biological link between the respiration of organic carbon and the consumption of oxygen. Furthermore, the model simulations suggest that a changing respiration quotient will impact multiple biogeochemical cycles and that future warming can lead to more intense deoxygenation than previously anticipated.