Heterotrimeric collagen helix with high specificity of assembly results in a rapid rate of folding.

The most abundant natural collagens form heterotrimeric triple helices. Synthetic mimics of collagen heterotrimers have been found to fold slowly, even compared to the already slow rates of homotrimeric helices. These prolonged folding rates are not understood. Here we compare the stabilities, specificities and folding rates of three heterotrimeric collagen mimics designed through a computationally assisted approach. The crystal structure of one ABC-type heterotrimer verified a well-controlled composition and register and elucidated the geometry of pairwise cation-π and axial and lateral salt bridges in the assembly. This collagen heterotrimer folds much faster (hours versus days) than comparable, well-designed systems. Circular dichroism and NMR data suggest the folding is frustrated by unproductive, competing heterotrimer species and these species must unwind before refolding into the thermodynamically favoured assembly. The heterotrimeric collagen folding rate is inhibited by the introduction of preformed competing triple-helical assemblies, which suggests that slow heterotrimer folding kinetics are dominated by the frustration of the energy landscape caused by competing triple helices.

Impact of low-load resistance exercise with and without blood flow restriction on muscle strength, endurance, and oxidative capacity: A pilot study.

Low-load resistance exercise (LLRE) to failure can increase muscle mass, strength, endurance, and mitochondrial oxidative capacity (OXPHOS). However, the impact of adding blood flow restriction to low-load resistance exercise (LLBFR) when matched for volume on these outcomes is incompletely understood. This pilot study examined the impact of 6 weeks of single-legged LLBFR and volume-matched LLRE on thigh bone-free lean mass, strength, endurance, and mitochondrial OXPHOS. Twenty (12 males and 8 females) untrained young adults (mean ± SD; 21 ± 2 years, 168 ± 11 cm, 68 ± 12 kg) completed 6 weeks of either single-legged LLBFR or volume-matched LLRE. Participants performed four sets of 30, 15, 15, and 15 repetitions at 25% 1-RM of leg press and knee extension with or without BFR three times per week. LLBFR increased knee extension 1-RM, knee extension endurance, and thigh bone-free lean mass relative to control (all p < 0.05). LLRE increased leg press and knee extension 1-RM relative to control (p = 0.012 and p = 0.054, respectively). LLRE also increased mitochondrial OXPHOS (p = 0.047 (nonparametric)). Our study showed that LLBFR increased muscle strength, muscle endurance, and thigh bone-free lean mass in the absence of improvements in mitochondrial OXPHOS. LLRE improved muscle strength and mitochondrial OXPHOS in the absence of improvements in thigh bone-free lean mass or muscle endurance.

Enhanced dynamic covalent chemistry for the controlled release of small molecules and biologics from a nanofibrous peptide hydrogel platform.

Maintaining safe and potent pharmaceutical drug levels is often challenging. Multidomain peptides (MDPs) assemble into supramolecular hydrogels with a well-defined, highly porous nanostructure that makes them attractive for drug delivery, yet their ability to extend release is typically limited by rapid drug diffusion. To overcome this challenge, we developed self-assembling boronate ester release (SABER) MDPs capable of engaging in dynamic covalent bonding with payloads containing boronic acids (BAs). As examples, we demonstrate that SABER hydrogels can prolong the release of five BA-containing small-molecule drugs as well as BA-modified insulin and antibodies. Pharmacokinetic studies revealed that SABER hydrogels extended the therapeutic effect of ganfeborole from days to weeks, preventing Mycobacterium tuberculosis growth better than repeated oral administration in an infection model. Similarly, SABER hydrogels extended insulin activity, maintaining normoglycemia for six days in diabetic mice after a single injection. These results suggest that SABER hydrogels present broad potential for clinical translation.

"Mourning and Melancholia" Meets The Babadook: Emptiness and its Relation to Absence.

This paper explores how the film The Babadook illuminates psychoanalytic understandings of melancholia and mourning. The author attempts to unwind the complicated character of melancholia, using Freud as an initial point of orientation, then relying on a few ideas from Klein and later writers. The paper attempts to refine our understanding of the difference between absence and emptiness, especially the difference between being captured in the nothing or deadness of melancholic emptiness, on the one hand, and being alive enough to suffer the absence of a lost object, which bears a potential for mourning, on the other. The possibility of psychic tension between these states is explored. Some implications of the relationship between absence and emptiness for the mourning process are considered. The author uses the film as a resource throughout.

Topical pravibismane as adjunctive therapy for moderate or severe diabetic foot infections: A phase 1b randomized, multicenter, double-blind, placebo-controlled trial.

This Phase 1b study was designed to evaluate the safety and efficacy of pravibismane, a novel broad-spectrum topical anti-infective, in managing moderate or severe chronic diabetic foot ulcer (DFU) infections. This randomized, double-blind, placebo-controlled, multicenter study consisted of 39 individuals undergoing pravibismane treatment and 13 individuals in the placebo group. Assessment of safety parameters included clinical observations of tolerability and pharmacokinetics from whole blood samples. Pravibismane was well-tolerated and exhibited minimal systemic absorption, as confirmed by blood concentrations that were below the lower limit of quantitation (0.5 ng/mL) or in the low nanomolar range, which is orders of magnitude below the threshold of pharmacological relevance for pravibismane. Pravibismane treated subjects showed approximately 3-fold decrease in ulcer size compared to the placebo group (85% vs. 30%, p = 0.27). Furthermore, the incidence of ulcer-related lower limb amputations was approximately 6-fold lower (2.6%) in the pooled pravibismane group versus 15.4% in the placebo group (p = 0.15). There were no treatment emergent or serious adverse events related to study drug. The initial findings indicate that topical pravibismane was safe and potentially effective treatment for improving recovery from infected chronic ulcers by reducing ulcer size and facilitating wound healing in infected DFUs (ClinicalTrials.gov Identifier NCT02723539).

A TLR7 Agonist Conjugated to a Nanofibrous Peptide Hydrogel as a Potent Vaccine Adjuvant.

Toll-like receptors (TLRs) recognize pathogen- and damage-associated molecular patterns and, in turn, trigger the release of cytokines and other immunostimulatory molecules. As a result, TLR agonists are increasingly being investigated as vaccine adjuvants, though many of these agonists are small molecules that quickly diffuse away from the vaccination site, limiting their co-localization with antigens and, thus, their effect. Here, the small-molecule TLR7 agonist 1V209 is conjugated to a positively-charged multidomain peptide (MDP) hydrogel, K 2 , which was previously shown to act as an adjuvant promoting humoral immunity. Mixing the 1V209-conjugated K 2 50:50 with the unfunctionalized K 2 produces hydrogels that retain the shear-thinning and self-healing physical properties of the original MDP, while improving the solubility of 1V209 more than 200-fold compared to the unconjugated molecule. When co-delivered with ovalbumin as a model antigen, 1V209-functionalized K 2 produces antigen-specific IgG titers that were statistically similar to alum, the gold standard adjuvant, and a significantly lower ratio of Th2-associated IgG1 to Th1-associated IgG2a than alum, suggesting a more balanced Th1 and Th2 response. Together, these results suggest that K 2 MDP hydrogels functionalized with 1V209 are a promising adjuvant for vaccines against infectious diseases, especially those benefiting from a combined Th1 and Th2 immune response. Table of Contents: Activation of toll-like receptors (TLRs) stimulates a signaling cascade to induce an immune response. A TLR7 agonist was conjugated to an injectable peptide hydrogel, which was then used to deliver a model vaccine antigen. This platform produced antibody titers similar to the gold standard adjuvant alum and demonstrated an improved balance between Th1- and Th2-mediated immunity over alum.

Successful Sequential Liver and Isolated Intestine Transplantation for Mitochondrial Neurogastrointestinal Encephalopathy Syndrome: A Case Report.

BACKGROUND Mitochondrial neurogastrointestinal encephalopathy syndrome (MNGIE) is an autosomal recessive disease caused by thymidine phosphorylase deficiency leading to progressive gastrointestinal dysmotility, cachexia, ptosis, ophthalmoparesis, peripheral neuropathy and leukoencephalopathy. Although liver transplantation corrects thymidine phosphorylase deficiency, intestinal deficiency of the enzyme persists. Retrospective chart review was carried out to obtain clinical, biochemical, and pathological details. CASE REPORT We present a case of liver and subsequent intestine transplant in a 28-year-old man with MNGIE syndrome with gastrointestinal dysmotility, inability to walk, leukoencephalopathy, ptosis, cachexia, and elevated serum thymidine. To halt progression of neurologic deficit, he first received a left-lobe partial liver transplantation. Although his motor deficit improved, gastrointestinal dysmotility persisted, requiring total parenteral nutrition. After exhaustive intestinal rehabilitation, he was listed for intestine transplantation. Two-and-half years after liver transplantation, he received an intestine transplant. At 4 years after LT and 20 months after the intestine transplant, he remains off parenteral nutrition and is slowly gaining weight. CONCLUSIONS This is the first reported case of mitochondrial neurogastrointestinal encephalomyopathy to undergo successful sequential liver and intestine transplantation.

Effect of ammonium sulfate combined with aqueous bio-chelator on Cd uptake by Cd-hyperaccumulator Solanum nigrum L.

The efficacy of using plants to phytoremediate heavy metal (HM) contaminated soils can be improved using soil amendments. These amendments may both increase plant biomasses and HMs uptake. We aimed to determine the composite effect of ammonium sulfate ((NH4)2SO4) combined with the application of an aqueous stem-extracted bio-chelator (Bidens tripartita L) on the plant biomasses and cadmium (Cd) phytoextraction by Solanum nigrum L. The constant (NH4)2SO4 application mode plus bio-chelator additives collectively enhanced the shoot Cd extraction ability owing to the increased plant biomass and shoot Cd concentration by S. nigrum. The shoot Cd extraction and the soil Cd decreased concentration confirmed the optimal Cd phytoextraction pattern in K8 and K9 treatments (co-application of (NH4)2SO4 and twofold/threefold bio-chelators). Accordingly, Cd contamination risk in the soil (2 mg kg-1) could be completely eradicated (<0.2 mg kg-1) after three rounds of phytoremediation by S.nigrum based on K8 and K9 treatments through calculating soil Cd depletion. The microorganism counts and enzyme activities in rhizosphere soils at treatments with the combined soil additives apparently advanced. In general, co-application mode of (NH4)2SO4 and aqueous bio-chelator was likely to be a perfect substitute for conventional scavenger agents on account of its environmental friendliness and cost saving for field Cd contamination phytoremediation by S. nigrum.

Survey of genetic counselors identifies a knowledge gap discerning properly regulated cell and gene therapy trials.

BACKGROUND AND AIMS: As cell and gene therapy (CGT) has grown in availability and scope, more unapproved regenerative medicine is being marketed to the public. It is essential that health care providers have sufficient knowledge and comfort to determine whether treatments are properly regulated and address these topics with patients. Due to the applicability of CGT to genetic disease, genetic counselors could be key in providing education and answering patients' questions about these topics. However, previous studies have focused only on physicians' knowledge and comfort with CGT and unapproved regenerative medicine. The purpose of this study was to assess genetic counselors' self-reported knowledge and comfort discussing these topics with patients and to explore what factors predict increased knowledge and comfort. METHODS: The authors designed an online survey distributed to genetic counselors who were part of the National Society of Genetic Counselors Student Research Program e-mail list. The survey addressed genetic counselors' demographics, practice experience with CGT, education about CGT, knowledge and comfort. RESULTS: The survey was completed by 144 genetic counselors. The best predictor of increased knowledge and comfort was experience discussing CGT in practice. In addition, those who worked at an institution at which CGT trials were offered had greater knowledge and comfort. However, most genetic counselors reported their knowledge was not sufficient to address questions from patients, and most had little-to-no knowledge or comfort determining whether a trial was properly regulated. There was no correlation between education and either knowledge or comfort; however, most participants desired more education about these topics. CONCLUSIONS: This study suggests that genetic counselors who (i) have experience with CGT in practice or (ii) work at institutions at which CGT trials are offered may have better knowledge regarding CGT. These results may help identify individuals and/or institutions in whom increasing knowledge regarding CGT could be beneficial. This is crucial as CGT becomes mainstream, leading to more widely marketed unapproved regenerative medicine. Several gaps in knowledge and comfort were identified, including participants' ability to determine whether a treatment is properly regulated. Further research is needed to better characterize the educational needs of genetic counselors surrounding these topics to address these gaps.

Supervisor integrity empowers employees to advocate for diversity in problematic climates.

Supervisors struggle to encourage employees to engage in diversity advocacy-key behaviors that help promote more equitable workplaces. Research hints that one reason for this struggle may be that employees lack the empowerment to engage in such behaviors. Drawing on perspectives that conceptualize diversity advocacy as a moral and virtuous behavior, we integrate research on leadership and empowerment to suggest that supervisor integrity can empower observers to engage in diversity advocacy. In exploring boundary conditions, we draw on performance models to counterintuitively suggest that this effect is strongest when employees perceive a negative diversity climate, as employees see the greatest need for change in these contexts. We test our theory in three complementary studies: A field sample with employees, a preregistered experimental vignette study, and an additional preregistered immersive experiment with a behavioral dependent variable. Our results contribute to theory on diversity, empowerment, and organizational climate. Additionally, we make an empirical contribution by developing and validating a four-item diversity advocacy scale. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

Kids gone wild - Alcohol use and patient characteristics in pediatric trauma during the coronavirus disease 2019 pandemic.

Importance: Reported coronavirus disease 2019 (COVID-19) pandemic effects on pediatric trauma have been variable. Objective: We investigated the characteristics of pediatric trauma including alcohol use during the pandemic at our urban trauma center. Methods: The trauma database of our adult level 1 trauma center was queried for all pediatric (age ≤ 18 years) patients presenting between March 1, 2020, and October 30, 2020. Data from 2017 to 2019 served as a control. Variables analyzed included demographics, mechanisms, injury severity, hospitalization characteristics, and positive blood alcohol. Results: Pandemic pediatric trauma volumes increased by 67.5% (330/year vs. 197/year). Pandemic patients were younger (median age 13 vs. 14 years, P = 0.011), but similar in gender, ethnicity, severity, hospital length of stay, mortality, and rates of penetrating injury. Falls doubled (79/year vs. 34/year) and shifted away from high falls >6 meters (0% vs. 7.9%) to moderate falls 1-6 meters (58.2% vs. 51.5%) (P = 0.028). Transportation injury rates were similar however mechanisms shifted from motor vehicle crashes (-13.5%) towards recreational vehicles including motorcycles (+2.1%), all-terrain vehicles (+8.6%), and bicycles (+3.8%) (P = 0.018). Pediatric-positive blood alcohol was significantly higher (11.2% vs. 5.1%, P < 0.001), especially for ages 14-18 years (21.7% vs. 9.5%, P < 0.001). Interpretation: Pediatric trauma volumes during the COVID-19 pandemic increased. Pandemic patients had more recreational vehicle injuries and higher rates of positive blood alcohol. This suggests an increased need for alcohol assessment and targeted interventions in the pediatric population during pandemics or periods of school closures.

Loss of succinyl-CoA synthetase in mouse forebrain results in hypersuccinylation with perturbed neuronal transcription and metabolism.

Lysine succinylation is a subtype of protein acylation associated with metabolic regulation of succinyl-CoA in the tricarboxylic acid cycle. Deficiency of succinyl-CoA synthetase (SCS), the tricarboxylic acid cycle enzyme catalyzing the interconversion of succinyl-CoA to succinate, results in mitochondrial encephalomyopathy in humans. This report presents a conditional forebrain-specific knockout (KO) mouse model of Sucla2, the gene encoding the ATP-specific beta isoform of SCS, resulting in postnatal deficiency of the entire SCS complex. Results demonstrate that accumulation of succinyl-CoA in the absence of SCS leads to hypersuccinylation within the murine cerebral cortex. Specifically, increased succinylation is associated with functionally significant reduced activity of respiratory chain complex I and widescale alterations in chromatin landscape and gene expression. Integrative analysis of the transcriptomic data also reveals perturbations in regulatory networks of neuronal transcription in the KO forebrain. Together, these findings provide evidence that protein succinylation plays a significant role in the pathogenesis of SCS deficiency.

Specifications of the ACMG/AMP guidelines for ACADVL variant interpretation.

Very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency (VLCADD) is a relatively common inborn error of metabolism, but due to difficulty in accurately predicting affected status through newborn screening, molecular confirmation of the causative variants by sequencing of the ACADVL gene is necessary. Although the ACMG/AMP guidelines have helped standardize variant classification, ACADVL variant classification remains disparate due to a phenotype that can be nonspecific, the possibility of variants that produce late-onset disease, and relatively high carrier frequency, amongst other challenges. Therefore, an ACADVL-specific variant curation expert panel (VCEP) was created to facilitate the specification of the ACMG/AMP guidelines for VLCADD. We expect these guidelines to help streamline, increase concordance, and expedite the classification of ACADVL variants.

Elucidating the clinical and molecular spectrum of SMARCC2-associated NDD in a cohort of 65 affected individuals.

PURPOSE: Coffin-Siris and Nicolaides-Baraitser syndromes are recognizable neurodevelopmental disorders caused by germline variants in BAF complex subunits. The SMARCC2 BAFopathy was recently reported. Herein, we present clinical and molecular data on a large cohort. METHODS: Clinical symptoms for 41 novel and 24 previously published affected individuals were analyzed using the Human Phenotype Ontology. For genotype-phenotype correlations, molecular data were standardized and grouped into non-truncating and likely gene-disrupting (LGD) variants. Missense variant protein expression and BAF-subunit interactions were examined using 3D protein modeling, co-immunoprecipitation, and proximity-ligation assays. RESULTS: Neurodevelopmental delay with intellectual disability, muscular hypotonia, and behavioral disorders were the major manifestations. Clinical hallmarks of BAFopathies were rare. Clinical presentation differed significantly, with LGD variants being predominantly inherited and associated with mildly reduced or normal cognitive development, whereas non-truncating variants were mostly de novo and presented with severe developmental delay. These distinct manifestations and non-truncating variant clustering in functional domains suggest different pathomechanisms. In vitro testing showed decreased protein expression for N-terminal missense variants similar to LGD. CONCLUSION: This study improved SMARCC2 variant classification and identified discernible SMARCC2-associated phenotypes for LGD and non-truncating variants, which were distinct from other BAFopathies. The pathomechanism of most non-truncating variants has yet to be investigated.

Succinyl-CoA Synthetase Dysfunction as a Mechanism of Mitochondrial Encephalomyopathy: More than Just an Oxidative Energy Deficit.

Biallelic pathogenic variants in subunits of succinyl-CoA synthetase (SCS), a tricarboxylic acid (TCA) cycle enzyme, are associated with mitochondrial encephalomyopathy in humans. SCS catalyzes the interconversion of succinyl-CoA to succinate, coupled to substrate-level phosphorylation of either ADP or GDP, within the TCA cycle. SCS-deficient encephalomyopathy typically presents in infancy and early childhood, with many patients succumbing to the disease during childhood. Common symptoms include abnormal brain MRI, basal ganglia lesions and cerebral atrophy, severe hypotonia, dystonia, progressive psychomotor regression, and growth deficits. Although subunits of SCS were first identified as causal genes for progressive metabolic encephalomyopathy in the early 2000s, recent investigations are now beginning to unravel the pathomechanisms underlying this metabolic disorder. This article reviews the current understanding of SCS function within and outside the TCA cycle as it relates to the complex and multifactorial mechanisms underlying SCS-related mitochondrial encephalomyopathy.

Visual Misrepresentations: The Lack of Skin Tone and Sex Equity in General Surgical Textbooks.

OBJECTIVE: Lack of racial and ethnic diversity in educational material contributes to health disparities. This study sought to determine if images of skin color and sex in general surgery textbooks were reflective of the U.S. DESIGN: All human figures with discernable sex characteristics and/or skin tone were evaluated independently by 4 coders. Each image was categorized as male or female. Skin tone in each image was categorized using the Massey- Martin skin color scale. This data was compared to 2020 U.S. Census Data. SETTING: U.S. Medical School. PARTICIPANTS: Not applicable. RESULTS: A total of 1179 images were evaluated for skin tone alone; 293 images for sex alone. 650 images depicted characteristics of both sex and skin tone. Interrater reliability was 0.78 for skin tone and 0.91 for sex. While the U.S. population is 59.3% white, 29.5% non-black persons of color and 13.6% black, in surgical textbooks, 90.7% of images were white, 6.5% were non-black persons of color, and 2.8% were black. Distribution of skin tone for all textbooks were significantly different. (p < 0.001) compared to the U.S. POPULATION: The U.S. population is 49.5% male and 50.5% female. When images of sex specific genitalia and breasts are excluded, surgical textbook images are 62.9% male and 37.1% female. Only 1 textbook had a distribution of sex that was similar to the U.S. CONCLUSIONS: Despite increasing diversity in the U.S. population there is a lack of skin tone and sex diversity in traditional surgical textbooks.

Tumor-associated fibrosis impairs immune surveillance and response to immune checkpoint blockade in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is a leading cause of cancer-related deaths. Immune checkpoint blockade has improved survival for many patients with NSCLC, but most fail to obtain long-term benefit. Understanding the factors leading to reduced immune surveillance in NSCLC is critical in improving patient outcomes. Here, we show that human NSCLC harbors large amounts of fibrosis that correlates with reduced T cell infiltration. In murine NSCLC models, the induction of fibrosis led to increased lung cancer progression, impaired T cell immune surveillance, and failure of immune checkpoint blockade efficacy. Associated with these changes, we observed that fibrosis leads to numerically and functionally impaired dendritic cells and altered macrophage phenotypes that likely contribute to immunosuppression. Within cancer-associated fibroblasts, distinct changes within the Col13a1-expressing population suggest that these cells produce chemokines to recruit macrophages and regulatory T cells while limiting recruitment of dendritic cells and T cells. Targeting fibrosis through transforming growth factor-ß receptor signaling overcame the effects of fibrosis to enhance T cell responses and improved the efficacy of immune checkpoint blockade but only in the context of chemotherapy. Together, these data suggest that fibrosis in NSCLC leads to reduced immune surveillance and poor responsiveness to checkpoint blockade and highlight antifibrotic therapies as a candidate strategy to overcome immunotherapeutic resistance.