SnapShot: Psychedelics and serotonin receptor signaling.

Serotonin (5-hydroxytryptamine; 5HT) signaling regulates processes in every major organ system, but it is most widely known for its role as a neurotransmitter in modulating a plethora of human behaviors. Psychedelics target the 5HT2A receptor and represent potentially transformative therapeutics for neuropsychiatric disorders. To view this SnapShot, open or download the PDF.

A Compendium of Genetic Modifiers of Mitochondrial Dysfunction Reveals Intra-organelle Buffering.

Mitochondrial dysfunction is associated with a spectrum of human conditions, ranging from rare, inborn errors of metabolism to the aging process. To identify pathways that modify mitochondrial dysfunction, we performed genome-wide CRISPR screens in the presence of small-molecule mitochondrial inhibitors. We report a compendium of chemical-genetic interactions involving 191 distinct genetic modifiers, including 38 that are synthetic sick/lethal and 63 that are suppressors. Genes involved in glycolysis (PFKP), pentose phosphate pathway (G6PD), and defense against lipid peroxidation (GPX4) scored high as synthetic sick/lethal. A surprisingly large fraction of suppressors are pathway intrinsic and encode mitochondrial proteins. A striking example of such "intra-organelle" buffering is the alleviation of a chemical defect in complex V by simultaneous inhibition of complex I, which benefits cells by rebalancing redox cofactors, increasing reductive carboxylation, and promoting glycolysis. Perhaps paradoxically, certain forms of mitochondrial dysfunction may best be buffered with "second site" inhibitors to the organelle.

Maintenance of CD4 T cell fitness through regulation of Foxo1.

Foxo transcription factors play an essential role in regulating specialized lymphocyte functions and in maintaining T cell quiescence. Here, we used a system in which Foxo1 transcription-factor activity, which is normally terminated upon cell activation, cannot be silenced, and we show that enforcing Foxo1 activity disrupts homeostasis of CD4 conventional and regulatory T cells. Despite limiting cell metabolism, continued Foxo1 activity is associated with increased activation of the kinase Akt and a cell-intrinsic proliferative advantage; however, survival and cell division are decreased in a competitive setting or growth-factor-limiting conditions. Via control of expression of the transcription factor Myc and the IL-2 receptor ß-chain, termination of Foxo1 signaling couples the increase in cellular cholesterol to biomass accumulation after activation, thereby facilitating immunological synapse formation and mTORC1 activity. These data reveal that Foxo1 regulates the integration of metabolic and mitogenic signals essential for T cell competitive fitness and the coordination of cell growth with cell division.

Bitter taste receptor activation by cholesterol and an intracellular tastant.

Bitter taste sensing is mediated by type 2 taste receptors (TAS2Rs (also known as T2Rs)), which represent a distinct class of G-protein-coupled receptors1. Among the 26 members of the TAS2Rs, TAS2R14 is highly expressed in extraoral tissues and mediates the responses to more than 100 structurally diverse tastants2-6, although the molecular mechanisms for recognizing diverse chemicals and initiating cellular signalling are still poorly understood. Here we report two cryo-electron microscopy structures for TAS2R14 complexed with Ggust (also known as gustducin) and Gi1. Both structures have an orthosteric binding pocket occupied by endogenous cholesterol as well as an intracellular allosteric site bound by the bitter tastant cmpd28.1, including a direct interaction with the α5 helix of Ggust and Gi1. Computational and biochemical studies validate both ligand interactions. Our functional analysis identified cholesterol as an orthosteric agonist and the bitter tastant cmpd28.1 as a positive allosteric modulator with direct agonist activity at TAS2R14. Moreover, the orthosteric pocket is connected to the allosteric site via an elongated cavity, which has a hydrophobic core rich in aromatic residues. Our findings provide insights into the ligand recognition of bitter taste receptors and suggest activities of TAS2R14 beyond bitter taste perception via intracellular allosteric tastants.

Stem-loop recognition by DDX17 facilitates miRNA processing and antiviral defense.

DEAD-box helicases play essential roles in RNA metabolism across species, but emerging data suggest that they have additional functions in immunity. Through RNAi screening, we identify an evolutionarily conserved and interferon-independent role for the DEAD-box helicase DDX17 in restricting Rift Valley fever virus (RVFV), a mosquito-transmitted virus in the bunyavirus family that causes severe morbidity and mortality in humans and livestock. Loss of Drosophila DDX17 (Rm62) in cells and flies enhanced RVFV infection. Similarly, depletion of DDX17 but not the related helicase DDX5 increased RVFV replication in human cells. Using crosslinking immunoprecipitation high-throughput sequencing (CLIP-seq), we show that DDX17 binds the stem loops of host pri-miRNA to facilitate their processing and also an essential stem loop in bunyaviral RNA to restrict infection. Thus, DDX17 has dual roles in the recognition of stem loops: in the nucleus for endogenous microRNA (miRNA) biogenesis and in the cytoplasm for surveillance against structured non-self-elements.

Cancer-associated PTEN mutants act in a dominant-negative manner to suppress PTEN protein function.

PTEN dysfunction plays a crucial role in the pathogenesis of hereditary and sporadic cancers. Here, we show that PTEN homodimerizes and, in this active conformation, exerts lipid phosphatase activity on PtdIns(3,4,5)P3. We demonstrate that catalytically inactive cancer-associated PTEN mutants heterodimerize with wild-type PTEN and constrain its phosphatase activity in a dominant-negative manner. To study the consequences of homo- and heterodimerization of wild-type and mutant PTEN in vivo, we generated Pten knockin mice harboring two cancer-associated PTEN mutations (PtenC124S and PtenG129E). Heterozygous Pten(C124S/+) and Pten(G129E/+) cells and tissues exhibit increased sensitivity to PI3-K/Akt activation compared to wild-type and Pten(+/-) counterparts, whereas this difference is no longer apparent between Pten(C124S/-) and Pten(-/-) cells. Notably, Pten KI mice are more tumor prone and display features reminiscent of complete Pten loss. Our findings reveal that PTEN loss and PTEN mutations are not synonymous and define a working model for the function and regulation of PTEN.

Diverse clonal fates emerge upon drug treatment of homogeneous cancer cells.

Even among genetically identical cancer cells, resistance to therapy frequently emerges from a small subset of those cells1-7. Molecular differences in rare individual cells in the initial population enable certain cells to become resistant to therapy7-9; however, comparatively little is known about the variability in the resistance outcomes. Here we develop and apply FateMap, a framework that combines DNA barcoding with single-cell RNA sequencing, to reveal the fates of hundreds of thousands of clones exposed to anti-cancer therapies. We show that resistant clones emerging from single-cell-derived cancer cells adopt molecularly, morphologically and functionally distinct resistant types. These resistant types are largely predetermined by molecular differences between cells before drug addition and not by extrinsic factors. Changes in the dose and type of drug can switch the resistant type of an initial cell, resulting in the generation and elimination of certain resistant types. Samples from patients show evidence for the existence of these resistant types in a clinical context. We observed diversity in resistant types across several single-cell-derived cancer cell lines and cell types treated with a variety of drugs. The diversity of resistant types as a result of the variability in intrinsic cell states may be a generic feature of responses to external cues.

Molecular basis for selective activation of DREADD-based chemogenetics.

Designer receptors exclusively activated by designer drugs (DREADDs) represent a powerful chemogenetic technology for the remote control of neuronal activity and cellular signalling1-4. The muscarinic receptor-based DREADDs are the most widely used chemogenetic tools in neuroscience research. The Gq-coupled DREADD (hM3Dq) is used to enhance neuronal activity, whereas the Gi/o-coupled DREADD (hM4Di) is utilized to inhibit neuronal activity5. Here we report four DREADD-related cryogenic electron microscopy high-resolution structures: a hM3Dq-miniGq complex and a hM4Di-miniGo complex bound to deschloroclozapine; a hM3Dq-miniGq complex bound to clozapine-N-oxide; and a hM3R-miniGq complex bound to iperoxo. Complemented with mutagenesis, functional and computational simulation data, our structures reveal key details of the recognition of DREADD chemogenetic actuators and the molecular basis for activation. These findings should accelerate the structure-guided discovery of next-generation chemogenetic tools.

Structure, function and pharmacology of human itch GPCRs.

The MRGPRX family of receptors (MRGPRX1-4) is a family of mas-related G-protein-coupled receptors that have evolved relatively recently1. Of these, MRGPRX2 and MRGPRX4 are key physiological and pathological mediators of itch and related mast cell-mediated hypersensitivity reactions2-5. MRGPRX2 couples to both Gi and Gq in mast cells6. Here we describe agonist-stabilized structures of MRGPRX2 coupled to Gi1 and Gq in ternary complexes with the endogenous peptide cortistatin-14 and with a synthetic agonist probe, respectively, and the development of potent antagonist probes for MRGPRX2. We also describe a specific MRGPRX4 agonist and the structure of this agonist in a complex with MRGPRX4 and Gq. Together, these findings should accelerate the structure-guided discovery of therapeutic agents for pain, itch and mast cell-mediated hypersensitivity.

Ligand recognition and allosteric modulation of the human MRGPRX1 receptor.

The human MAS-related G protein-coupled receptor X1 (MRGPRX1) is preferentially expressed in the small-diameter primary sensory neurons and involved in the mediation of nociception and pruritus. Central activation of MRGPRX1 by the endogenous opioid peptide fragment BAM8-22 and its positive allosteric modulator ML382 has been shown to effectively inhibit persistent pain, making MRGPRX1 a promising target for non-opioid pain treatment. However, the activation mechanism of MRGPRX1 is still largely unknown. Here we report three high-resolution cryogenic electron microscopy structures of MRGPRX1-Gαq in complex with BAM8-22 alone, with BAM8-22 and ML382 simultaneously as well as with a synthetic agonist compound-16. These structures reveal the agonist binding mode for MRGPRX1 and illuminate the structural requirements for positive allosteric modulation. Collectively, our findings provide a molecular understanding of the activation and allosteric modulation of the MRGPRX1 receptor, which could facilitate the structure-based design of non-opioid pain-relieving drugs.

Neighborhood-level deprivation and survival in lung cancer.

BACKGROUND: Despite recent advances in lung cancer therapeutics and improving overall survival, disparities persist among socially disadvantaged populations. This study aims to determine the effects of neighborhood deprivation indices (NDI) on lung cancer mortality. This is a multicenter retrospective cohort study assessing the relationship between NDI and overall survival adjusted for age, disease stage, and DNA methylation among biopsy-proven lung cancer patients. State-specific NDI for each year of sample collection were computed at the U.S. census tract level and dichotomized into low- and high-deprivation. RESULTS: A total of 173 non small lung cancer patients were included, with n = 85 (49%) and n = 88 (51%) in the low and high-deprivation groups, respectively. NDI was significantly higher among Black patients when compared with White patients (p = 0.003). There was a significant correlation between DNA methylation and stage for HOXA7, SOX17, ZFP42, HOXA9, CDO1 and TAC1. Only HOXA7 DNA methylation was positively correlated with NDI. The high-deprivation group had a statistically significant shorter survival than the low-deprivation group (p = 0.02). After adjusting for age, race, stage, and DNA methylation status, belonging to the high-deprivation group was associated with higher mortality with a hazard ratio of 1.81 (95%CI: 1.03-3.19). CONCLUSIONS: Increased neighborhood-level deprivation may be associated with liquid biopsy DNA methylation, shorter survival, and increased mortality. Changes in health care policies that consider neighborhood-level indices of socioeconomic deprivation may enable a more equitable increase in lung cancer survival.

Non-suicidal self-injury among individuals with an eating disorder: A systematic review and prevalence meta-analysis.

OBJECTIVE: This study aimed to quantify the prevalence of non-suicidal self-injury across eating disorders (EDs) and within diagnostic categories through systematic review and proportional, or so-called prevalence, meta-analysis. METHOD: Included studies had to contain individuals with a verified diagnosis of an ED. The last literature search was conducted on September 11, 2023, for studies published on or before September 2023 without a restriction on earliest publication year. Results were synthesized and analyzed using the "metaprop" package in R and presented using forest plots. Bias was assessed by a Peters' regression test and funnel plot. RESULTS: 79 studies published between 1985 and 2023 were included encompassing 32,334 individuals with an ED. Importantly, 42 studies were not included in any other meta-analyses on self-injury in EDs to date. Overall prevalence of non-suicidal self-injury was 34.59% (95%CI = 30.49-38.81). Prevalence in anorexia nervosa restrictive type, binge/purge type, bulimia nervosa, binge eating disorder and other specified feeding/eating disorder were 23.19% (95%CI = 16.96-30.03%), 41.98% (95%CI = 32.35-51.91%), 36.97% (95%CI = 30.69-43.46%), 21.21% (95%CI = 14.93-28.12%) and 37.65% (95%CI = 28.59-47.09%), respectively. Prevalence estimations could not be estimated for other ED categories due to lack of a sufficient number of studies. DISCUSSION: Non-suicidal self-injury is prevalent across both binge/purge and restrictive EDs. Considering the transdiagnostic nature of self-injurious behaviors in ED, the results highlight the importance of assessment and monitoring of self-injury in people with ED, irrespective of specific diagnoses. The method of determining self-injury varied across studies and may limit this study. PUBLIC SIGNIFICANCE: This study highlights the prevalence of self-injury across eating disorders irrespective of diagnosis and within specific EDs. While diagnoses known to exhibit self-injurious behaviors (e.g., bulimia nervosa, anorexia nervosa binge/purge subtype) demonstrated the highest prevalence of self-injury, all diagnoses were found to have a prevalence greater than 20%. These findings suggest the importance of assessing and monitoring all individuals with an eating disorder for the presence of self-injury.

OBJETIVO: Este estudio tuvo como objetivo cuantificar la prevalencia de la autolesión no suicida en los trastornos de la conducta alimentaria (TCA) y dentro de las categorías diagnósticas mediante una revisión sistemática y un metaanálisis proporcional, también llamado metaanálisis de prevalencia. MÉTODO: Los estudios incluidos debían contener individuos con un diagnóstico verificado de un TCA. La última búsqueda bibliográfica se realizó el 11 de septiembre de 2023, para estudios publicados en o antes de septiembre de 2023 sin restricción en el año de publicación más temprano. Los resultados fueron sintetizados y analizados utilizando el paquete "metaprop" en R y presentados mediante gráficos de bosque. El sesgo se evaluó mediante una prueba de regresión de Peters y un gráfico de embudo. RESULTADOS: Se incluyeron 79 estudios publicados entre 1985 y 2023 que abarcaron a 32,334 individuos que padecían un TCA. Es importante destacar que 42 estudios no se incluyeron en ningún otro metaanálisis sobre autolesión en TCA hasta la fecha. La prevalencia general de la autolesión no suicida fue del 34.59% (IC del 95% = 30.49-38.81). La prevalencia en la anorexia nerviosa subtipo restrictivo, subtipo atracones/purga, bulimia nerviosa, trastorno de atracones y otros trastornos especificados de la conducta alimentaria y de la alimentación fue del 23.19% (IC del 95% = 16.96-30.03%), 41.98% (IC del 95% = 32.35-51.91%), 36.97% (IC del 95% = 30.69-43.46%), 21.21% (IC del 95% = 14.93-28.12%) y 37.65% (IC del 95% = 28.59-47.09%), respectivamente. No se pudieron estimar las estimaciones de prevalencia para otras categorías de TCA debido a la falta de un número suficiente de estudios. DISCUSIÓN: La autolesión no suicida es prevalente tanto en los TCA subtipo de atracón/purgación como en los restrictivos. Dada la naturaleza transdiagnóstica de los comportamientos autolesivos en los TCA, los resultados resaltan la importancia de la evaluación y el monitoreo de la autolesión en personas que padecen TCA, independientemente de los diagnósticos específicos. El método para determinar la autolesión varió entre los estudios y puede limitar este estudio.

Is Suffering a Useless Concept?

"Suffering" is a central concept within bioethics and often a crucial consideration in medical decision making. As used in practice, however, the concept risks being uninformative, ambiguous, or even misleading. In this paper, we consider a series of cases in which "suffering" is invoked and analyze them in light of prominent theories of suffering. We then outline ethical hazards that arise as a result of imprecise usage of the concept and offer practical recommendations for avoiding them. Appeals to suffering are often getting at something ethically important. But this is where the work of ethics begins, not where it ends.

Evaluation and application of American College of Radiology Thyroid Imaging Reporting and Data System for improved malignancy detection in paediatric thyroid nodules.

OBJECTIVE: The American College of Radiology Thyroid Imaging Reporting and Data System (TI-RADS) is a widely used method for the management of adult thyroid nodules. However, its use in paediatric patients is controversial because adult fine needle aspiration biopsy (FNAB) recommendations may lead to delayed diagnoses of cancer in children. The objectives of this study were to evaluate the performance of TI-RADS in paediatric thyroid nodules and to tailor FNAB recommendations for children. METHODS: Consecutive surgically resected paediatric thyroid nodules from two tertiary care centres between 2003 and 2021 were reviewed. Ultrasounds were blindly scored by radiologists according to TI-RADS. Management recommendations based on TI-RADS were evaluated. Various modelling methodologies were used to determine the optimal cutoff for FNAB in children. RESULTS: Of the 96 patients, 79 (82%) were female and the median age at surgery was 16.1 years. Fifty (52%) nodules were malignant on surgical pathology. The area under the receiver operating characteristic curve of TI-RADS for predicting malignancy was 0.78. Adult TI-RADS recommendations would have resulted in 4% of cancerous nodules being lost to follow-up. Modifications to TI-RADS (FNAB of all TR3 nodules ≥1.5 cm, FNAB of TR4 and TR5 nodules ≥0.5 cm, surveillance of nodules ≥1 cm, consider surgery for nodules >4 cm) reduced this missed malignancy rate to 0%. CONCLUSIONS: TI-RADS can risk-stratify paediatric thyroid nodules. However, the system requires modifications to reduce the missed malignancy rate in paediatric thyroid nodules. Our data suggest that lower size thresholds for FNAB are warranted in children.

Spindle Cell Carcinoma of the Ocular Adnexa: A Case Report and Literature Review.

A 68-year-old woman was seen in oculoplastic consultation with a medial canthal lesion initially diagnosed as an atypical fibroxanthoma. On excisional biopsy, she was found to have a spindle cell carcinoma, which is a rare and reportedly more aggressive form of squamous cell carcinoma. We highlight the surgical technique of biopsy and reconstruction, the detailed histologic and immunohistochemical analysis required for accurate diagnosis, considerations for adjuvant treatment, and suggestions for systemic workup and surveillance. This case adds to the small body of available literature on primary spindle cell carcinoma of the ocular surface and ocular adnexa, which we have summarized. We hope that as more data becomes available, there will be clearer diagnostic and treatment algorithms for this uncommon presentation.

Inadvertent Orbital Mitomycin C Injection as a Cause of Ptosis and Ophthalmoplegia from Orbital Necrosis.

Mitomycin C is an alkylating agent with the ability to suppress fibroblast proliferation and activity, making it a powerful antifibrotic. It has therefore become popular in glaucoma filtering surgeries, used both intraoperatively during bleb formation and postoperatively as an adjunct to bleb needling. This report presents a rare but serious risk of bleb needling with Mitomycin C at the slit lamp, where inadvertent movement of the patient resulted in an orbital injection. The patient quickly developed focal tissue inflammation and necrosis, presenting one day after the procedure with complete ptosis, ophthalmoplegia, and a palpable orbital mass. After appropriate imaging and an excisional biopsy to exclude infectious, infiltrative, and neoplastic causes, this was managed with close observation and continued improvement, and resolution of most orbital sequelae.

Patella and Trochlea Osteochondritis Dissecans: Demographics and Treatment Paradigms.

OBJECTIVE: Previous research on patellar and trochlear groove osteochondritis dissecans (OCD) is limited by small sample sizes. This study aims to describe the presentation of patients with OCD lesions of the patella and trochlea and characterize the outcomes of operative and nonoperative treatments. METHODS: This retrospective cohort study identified all patients from a single institution from 2008 to 2021 with patellar and/or trochlear OCD lesions. Patients were excluded from the study if surgical records were unavailable or if the patient had knee surgery for a different injury at index surgery or in the 12 months postoperative. Minimum follow-up was 12 months. Outcomes included a return to sports (RTS), pain resolution, radiographic healing, and treatment "success" (defined as full RTS, complete pain resolution, and full healing on imaging). RESULTS: A total of 68 patients (75 knees) were included-45 (60%) with patellar OCD and 30 (40%) with trochlear. Of the patients, 69% were males. The median age at knee OCD diagnosis was 14 years. At the final follow-up, 62% of knees (n = 44) recovered sufficiently to allow a full RTS and 54% of knees (n = 39) had full pain resolution. Of the 46 knees with radiographic imaging at least 1 year apart, 63% had full healing of the lesion. There was no significant difference in RTS, pain resolution, radiographic healing, or overall success when comparing treatments. CONCLUSIONS: This study provides valuable epidemiologic demographic and outcome data regarding the scarcely reported patellar and trochlear OCD. While over half of patients fully returned to sports and reported full pain resolution, a large proportion continued to experience symptoms over a year after presentation. Future research should aim to better define the treatment algorithms for these OCD subtypes. LEVEL OF EVIDENCE: Level III.

A conserved virus-induced cytoplasmic TRAMP-like complex recruits the exosome to target viral RNA for degradation.

RNA degradation is tightly regulated to selectively target aberrant RNAs, including viral RNA, but this regulation is incompletely understood. Through RNAi screening in Drosophila cells, we identified the 3'-to-5' RNA exosome and two components of the exosome cofactor TRAMP (Trf4/5-Air1/2-Mtr4 polyadenylation) complex, dMtr4 and dZcchc7, as antiviral against a panel of RNA viruses. We extended our studies to human orthologs and found that the exosome as well as TRAMP components hMTR4 and hZCCHC7 are antiviral. While hMTR4 and hZCCHC7 are normally nuclear, infection by cytoplasmic RNA viruses induces their export, forming a cytoplasmic complex that specifically recognizes and induces degradation of viral mRNAs. Furthermore, the 3' untranslated region (UTR) of bunyaviral mRNA is sufficient to confer virus-induced exosomal degradation. Altogether, our results reveal that signals from viral infection repurpose TRAMP components to a cytoplasmic surveillance role where they selectively engage viral RNAs for degradation to restrict a broad range of viruses.

HbA1c and risk factors' prevalence in patients with stroke: a retrospective study in a tertiary care hospital in Saudi Arabia.

OBJECTIVES: To determine the relationship between hemoglobin A1c (HbA1c) and different types of stroke, and how different comorbidities and risk factors are related to the occurrence of stroke in a Saudi Arabian tertiary care hospital. METHODS: This retrospective study was conducted at King Abdulaziz Medical City, Riyadh, Kingdom of Saudi Arabia. The study included patients who experienced either hemorrhagic stroke (HS), ischemic stroke (IS), or transient ischemic attack (TIA) between 2015 and 2020. RESULTS: In total, 976 patients were included, of whom 670 were males (68.6%). The incidence of HS was significantly higher in males compared to females (14.2% vs. 6.9%), whereas the incidence of IS was higher in females (76.8% vs. 74.6%) (p=0.001). Ischemic stroke was significantly higher in the 65 years or older age group, whereas HS was comparatively higher among those aged <65 years. The means HbA1c levels in all three types of stroke were abnormally high. However, HbA1c levels were significantly higher in IS than in the other 2 stroke types (p=0.017). The HbA1c levels showed statistically significant differences between the different types of stroke, where the estimated marginal means were higher in patients with IS with a small effect size. Heart disease was also more prevalent in the IS group. Stroke-related mortality was reported in 16 patients and was significantly higher in the IS group than in the HS group. CONCLUSION: The HbA1c levels were elevated in all types of stroke, significantly in IS. Controlling patients' HbA1c and other modifiable risk factors could significantly reduce the risk of stroke.