Search

Subarachnoid hemorrhage (SAH) can be associated with neurological deficits and has profound consequences for mortality and morbidity. Cerebral vasospasm (CVS) and delayed cerebral ischemia affect neurological outcomes in SAH patients, but their mechanisms are not fully understood, and effective treatments are limited. Here, we report that urotensin II receptor UT plays a pivotal role in both early events and delayed mechanisms following SAH in male mice. Few days post-SAH, UT expression is triggered by blood or hemoglobin in the leptomeningeal compartment. UT contributes to perimeningeal glia limitans astrocyte reactivity, microvascular alterations and neuroinflammation independent of CNS-associated macrophages (CAMs). Later, CAM-dependent vascular inflammation and subsequent CVS develop, leading to cognitive dysfunction. In an SAH model using humanized UT h+ / h+ male mice, we show that post-SAH CVS and behavioral deficits, mediated by UT through Gq/PLC/Ca 2+ signaling, are prevented by UT antagonists. These results highlight the potential of targeting UT pathways to reduce early meningeal response and delayed cerebral ischemia in SAH patients., (© 2024. The Author(s).)

GPR56, an adhesion G-protein coupled receptor (aGPCRs) with constitutive and ligand-promoted activity, is involved in many physiological and pathological processes. Whether the receptor's constitutive or ligand-promoted activation occur through the same molecular mechanism, and whether different activation modes lead to functional selectivity between G proteins is unknown. Here we show that GPR56 constitutively activates both G12 and G13. Unlike constitutive activation and activation with 3-a-acetoxydihydrodeoxygedunin (3αDOG), stimulation with an antibody, 10C7, directed against GPR56's extracellular domain (ECD) led to an activation that favors G13 over G12. An autoproteolytically deficient mutant, GPR56-T383A, was also activated by 10C7 indicating that the tethered agonist (TA) exposed through autocatalytic cleavage, is not required for this activation modality. In contrast, this proteolysis-resistant mutant could not be activated by 3αDOG indicating different modes of activation by the two ligands. We show that an N-terminal truncated GPR56 construct (GPR56-Δ1-385) is devoid of constitutive activity but was activated by 3αDOG. Similarly to 3αDOG, 10C7 promoted the recruitment of b-arrestin-2 but GPR56 internalization was β-arrestin independent. Despite the slow activation mode of 10C7 that favors G13 over G12, it efficiently activated the downstream Rho pathway in BT-20 breast cancer cells. These data show that different GPR56 ligands have different modes of activation yielding differential G protein selectivity but converging on the activation of the Rho pathway both in heterologous expressions system and in cancer cells endogenously expressing the receptor. 10C7 is therefore an interesting tool to study both the processes underlying GPR56 activity and its role in cancer cells.

TMPRSS13, a member of the Type II Transmembrane Serine Proteases (TTSP) family, is involved in cancer progression and in cell entry of respiratory viruses. To date, no inhibitors have been specifically developed toward this protease. In this study, a chemical library of 65 ketobenzothiazole-based peptidomimetic molecules was screened against a proteolytically active form of recombinant TMPRSS13 to identify novel inhibitors. Following an initial round of screening, subsequent synthesis of additional derivatives supported by molecular modelling, uncovered important molecular determinants involved in TMPRSS13 inhibition. One inhibitor, N-0430, achieved low nanomolar affinity towards TMPRSS13 activity in a cellular context. Using a SARS-CoV-2 pseudovirus cell entry model, we further show the ability of N-0430 to block TMPRSS13-dependent entry of the pseudovirus. The identified peptidomimetic inhibitors and the molecular insights of their potency gained from this study will aid in the development of specific TMPRSS13 inhibitors.

TMPRSS6 is a potential therapeutic target for the treatment of iron overload due to its role in regulating levels of hepcidin. Although potent TMPRSS6 inhibitors have been previously developed, their lack of specificity requires optimization to avoid potential side effects before pursuing preclinical development with in vivo models. Here, using computer-aided drug design based on a TMPRSS6 homology model, we reveal that the S2 position of TMPRSS6 offers a potential avenue to achieve selectivity against other members of the TTSP family. Accordingly, we synthesized novel peptidomimetic molecules containing lipophilic amino acids at the P2 position to exploit this unexplored pocket. This enabled us to identify TMPRSS6-selective small molecules with low nanomolar affinity. Finally, pharmacokinetic parameters were determined, and a compound was found to be potent in cellulo toward its primary target while retaining TTSP-subtype selectivity and showing no signs of alteration in in vitro TEER experiments.

SARS-CoV-2 Omicron subvariants with increased transmissibility and immune evasion are spreading globally with alarming persistence. Whether the mutations and evolution of spike (S) Omicron subvariants alter the viral hijacking of human TMPRSS2 for viral entry remains to be elucidated. This is particularly important to investigate because of the large number and diversity of mutations of S Omicron subvariants reported since the emergence of BA.1. Here we report that human TMPRSS2 is a molecular determinant of viral entry for all the Omicron clinical isolates tested in human lung cells, including ancestral Omicron subvariants (BA.1, BA.2, BA.5), contemporary Omicron subvariants (BQ.1.1, XBB.1.5, EG.5.1) and currently circulating Omicron BA.2.86. First, we used a co-transfection assay to demonstrate the endoproteolytic cleavage by TMPRSS2 of spike Omicron subvariants. Second, we found that N-0385, a highly potent TMPRSS2 inhibitor, is a robust entry inhibitor of virus-like particles harbouring the S protein of Omicron subvariants. Third, we show that N-0385 exhibits nanomolar broad-spectrum antiviral activity against live Omicron subvariants in human Calu-3 lung cells and primary patient-derived bronchial epithelial cells. Interestingly, we found that N-0385 is 10-20 times more potent than the repositioned TMPRSS2 inhibitor, camostat, against BA.5, EG.5.1, and BA.2.86. We further found that N-0385 shows broad synergistic activity with clinically approved direct-acting antivirals (DAAs), i.e., remdesivir and nirmatrelvir, against Omicron subvariants, demonstrating the potential therapeutic benefits of a multi-targeted treatment based on N-0385 and DAAs., Competing Interests: Declaration of competing interest PLB and RL are inventors on patent applications (US9365853B2 and US10988505B2) that cover matriptase and other type II transmembrane serine protease inhibitors for treating and preventing viral infections, respiratory disorders, inflammatory disorders, pain disorders, tissue disorders, hyperproliferative disorders, and disorders associated with iron overload. The remaining authors declare that they have no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Critical sized bone defects in the ankle are becoming increasingly more common in patients undergoing limb reconstruction with tibiotalocalcaneal arthrodesis. Bulk allografts have not fared well over time. There have been scattered preliminary reports using custom spinal cages or 3D-printed Titanium Implants to address the critical bony defect; however, the cost of these devices is prohibitive in many clinical practice settings. The purpose of this investigation is to report the preliminary experience using a commercially available Trabecular Metal (Zimmer-Biomet) tibial metaphyseal cone combined with a retrograde locked intramedullary nail to address this challenging problem. Eight consecutive patients underwent tibiotalocalcaneal arthrodesis using a commercially available Trabecular Metal tibial metaphyseal cone combined with a retrograde locked intramedullary nail. Five developed bone loss secondary to neuropathic (Charcot) bony resorption and 3 underwent surgery for failed total ankle arthroplasty. All 8 patients eventually achieved clinical and radiographic healing and were able to ambulate with standard footwear. One patient developed a postoperative wound infection at the site of calcaneal locking screws, which resolved with debridement and parenteral antibiotic therapy. Critical bone defects about the ankle have successfully addressed with custom 3D titanium implants. This small series suggests that similar clinical outcomes can be achieved with the use of a commercially available porous tantalum metaphyseal spacer borrowed from our arthroplasty colleagues, combined with the use of a retrograde locked intramedullary nail. Levels of Evidence : Level 4: Retrospective case series., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

The carriage of Salmonella in pigs is a major concern for the agri-food industry and for global healthcare systems. Humans could develop salmonellosis when consuming contaminated pig products. On the other hand, some Salmonella serotypes could cause disease in swine, leading to economic losses on farms. The purpose of the present study was to characterize the anti- Salmonella activity of a novel Bacillus -based probiotic using a bioreactor containing a piglet-derived intestinal microbiota. Two methods of probiotic administration were tested: a single daily and a continuous dose. Salmonella enumeration was performed using selective agar at T24h, T48h, T72h, T96h and T120h. The DNA was extracted from bioreactor samples to perform microbiome profiling by targeted 16S rRNA gene sequencing on Illumina Miseq. The quantification of short-chain fatty acids (SCFAs) was also assessed at T120h. The probiotic decreased Salmonella counts at T96 for the daily dose and at T120 for the continuous one. Both probiotic doses affected the alpha and beta diversity of the piglet-derived microbiota ( p < 0.05). A decrease in acetate concentration and an increase in propionate proportion were observed in the continuous condition. In conclusion, the tested Bacillus -based product showed a potential to modulate microbiota and reduce Salmonella colonization in a piglet-derived intestinal microbiota and could therefore be used in vivo.

Objective: To identify facilitators and barriers to older adults' participation in telehealth interventions for primary prevention and health promotion., Methods: Relevant articles were searched using keywords in Embase and MEDLINE. Study characteristics, type of telehealth interventions and technology involved, as well as facilitators and barriers to their use, were extracted from selected articles. The Unified Theory of Acceptance and Use of Technology 2 (UTAUT2) model was used to organise data., Results: A total of 24 articles (pertaining to 20 studies) were included. Nine facilitators and 11 barriers influencing the participation in telehealth interventions for primary prevention and health promotion among older adults were identified. The most recurrent facilitators were related to the individual's performance expectancy and effort expectancy, as well as the presence of a social dimension associated with the intervention (i.e. having a good relationship with the other participants in the program). The two most prevalent barriers were also related to effort expectancy and performance expectancy, followed by barriers related to the inherent characteristics of the technology and older adults' health condition. Experience, age and gender were also found to moderate technology use and acceptance., Conclusions: This rapid review highlights the importance of adopting a holistic perspective when designing telehealth interventions aimed at preventive and health promotion purposes among older adults., (© 2023 The Authors. Australasian Journal on Ageing published by John Wiley & Sons Australia, Ltd on behalf of AJA Inc’.)

Total ankle arthroplasty is a topic that has recently gained increasing interest, largely due to the improved outcomes, which have been demonstrated by short- and mid-term research studies on the newer, third-generation implant designs. The purpose of this review is to provide an updated assessment of the quality of outcomes research on total ankle arthroplasty., Competing Interests: Disclosure The authors of this article have no disclosures to report., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Effective perioperative pain control following knee arthroscopy allows patients to reduce narcotic intake, avoid side effects of these medications, and recover more quickly. Adductor canal nerve blockade (ACB) and intra-articular injection of local anesthetic have been described as adjuvant treatments for postoperative pain control following surgery of the knee. This study directly compares the effect of each of these treatment modalities. Patients undergoing knee arthroscopy were blinded and randomized to receive either an ACB ( n  = 60) or intra-articular injection of local anesthetic (IAB, n  = 64). Outcome measures included patient reported visual analog scale (VAS) scores at 1, 2, 4, 8, 16, 24, 36, 48 hours and 1 week and total narcotic consumption at 12, 24, and 48 hours postoperatively. Student's t -tests were used to compare unadjusted VAS scores at each time point and use of postoperative pain medication between treatment groups. Adjusted VAS scores were estimated in a multivariable general linear model with interaction of time and treatment group and other relevant covariates. There were no statistically significant differences between the two groups in terms of gender, age, body mass index, and insurance type. ACB patients had significantly higher pain scores than IAB patients at hours 1 and 2 (hour 1: 4.02 [2.99] vs. 2.59 [3.00], p  = 0.009; hour 2: 3.12 [2.44] vs. 2.17 [2.62], p  = 0.040). ACB patients had higher pain scores than IAB patients up to hour 16, though hours 4 to 16 were not significantly different. Adjusted covariate analyses demonstrate an additional statistically significant reduction in pain score in the IAB group at hour 4. There were no differences in narcotic consumption. Intraoperative local anesthetic and regional ACB each provides adequate pain control following knee arthroscopy, and intraoperative local anesthetic may provide enhanced pain control for up to 4 hours postoperatively. LEVEL OF EVIDENCE: : Level 1 evidence, randomized control trial., Competing Interests: D.E. is a paid consultant for Stryker Corporation. No other disclosures., (Thieme. All rights reserved.)

Human influenza viruses cause acute respiratory symptoms that can lead to death. Due to the emergence of antiviral drug-resistant strains, there is an urgent requirement for novel antiviral agents and innovative therapeutic strategies. Using the peptidomimetic ketobenzothiazole protease inhibitor RQAR-Kbt (IN-1, aka N-0100) as a starting point, we report how substituting P2 and P4 positions with natural and unnatural amino acids can modulate the inhibition potency toward matriptase, a prototypical type II transmembrane serine protease (TTSP) that acts as a priming protease for influenza viruses. We also introduced modifications of the peptidomimetics N-terminal groups, leading to significant improvements (from μM to nM, 60 times more potent than IN-1) in their ability to inhibit the replication of influenza H1N1 virus in the Calu-3 cell line derived from human lungs. The selectivity towards other proteases has been evaluated and explained using molecular modeling with a crystal structure recently obtained by our group. By targeting host cell TTSPs as a therapeutic approach, it may be possible to overcome the high mutational rate of influenza viruses and consequently prevent potential drug resistance., (© 2023 The Authors. ChemMedChem published by Wiley-VCH GmbH.)

Introduction: Surgical smoke generated through the use of electrical surgical devices poses a risk to the surgeon, medical personnel in the operating room, and the patient by exposing them to environmentally hazardous particulate matter. Previous investigation has shown that surgical smoke leads to an increased risk of pulmonary conditions, circulatory disorders, and irritation of the eyes, nose, and throat. Transmission of infectious disease can occur through inhalation of viral particles, and the presence of carcinogens are also of major concern. The deleterious effects of surgical smoke are well documented in several subspecialties, namely dermatology and general surgery, but there has been little discussion on the topic amongst orthopedic surgeons., Methods: A non-systematic review of the literature was completed with the aim of identifying the major categories of adverse health effects associated with surgical smoke inhalation and offering recommendations to reduce these hazards in the orthopedic surgical community., Results: Three primary categories of risk associated with surgical smoke inhalation were identified: inflammation, viral/bacterial transmission, and carcinogenicity. In addition, strategies for mitigating risk and best practice recommendations were explored., Conclusion: Surgical smoke is an under-recognized occupational hazard within the orthopedic surgery literature. There are several strategies which can be employed to reduce risk. Further investigation is needed to understand the long-term impact of these risks, as well as what can be done to improve the practicality and compliance with protective measures., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Antivirals with broad coronavirus activity are important for treating high-risk individuals exposed to the constantly evolving SARS-CoV-2 variants of concern (VOCs) as well as emerging drug-resistant variants. We developed and characterized a novel class of active-site-directed 3-chymotrypsin-like protease (3CLpro) inhibitors ( C2-C5a ). Our lead direct-acting antiviral (DAA), C5a , is a non-covalent, non-peptide with a dissociation constant of 170 nM against recombinant SARS-CoV-2 3CLpro. The compounds C2-C5a exhibit broad-spectrum activity against Omicron subvariants (BA.5, BQ.1.1, and XBB.1.5) and seasonal human coronavirus-229E infection in human cells. Notably, C5a has median effective concentrations of 30-50 nM against BQ.1.1 and XBB.1.5 in two different human cell lines. X-ray crystallography has confirmed the unique binding modes of C2-C5a to the 3CLpro, which can limit virus cross-resistance to emerging Paxlovid-resistant variants. We tested the effect of C5a with two of our newly discovered host-directed antivirals (HDAs): N-0385, a TMPRSS2 inhibitor, and bafilomycin D (BafD), a human vacuolar H + -ATPase [V-ATPase] inhibitor. We demonstrated a synergistic action of C5a in combination with N-0385 and BafD against Omicron BA.5 infection in human Calu-3 lung cells. Our findings underscore that a SARS-CoV-2 multi-targeted treatment for circulating Omicron subvariants based on DAAs ( C5a ) and HDAs (N-0385 or BafD) can lead to therapeutic benefits by enhancing treatment efficacy. Furthermore, the high-resolution structures of SARS-CoV-2 3CLpro in complex with C2-C5a will facilitate future rational optimization of our novel broad-spectrum active-site-directed 3C-like protease inhibitors.

BACKGROUND : Cold snare polypectomy (CSP) is increasingly used for polypectomy and is recommended as the first-line modality for small (< 10 mm) polyps. This study aimed to evaluate incomplete resection rates (IRRs) when using CSP for colorectal polyps of 4-20 mm. METHODS : Adults (45-80 years) undergoing screening, surveillance, or diagnostic colonoscopy and CSP by one of nine endoscopists were included. The primary outcome was the IRR for colorectal polyps of 4-20 mm, defined as the presence of polyp tissue in marginal biopsies after resection of serrated polyps or adenomas. Secondary outcomes included the IRR for serrated polyps, ease of resection, and complications. RESULTS:  413 patients were included (mean age 63; 48 % women) and 182 polyps sized 4-20 mm were detected and removed by CSP. CSP required conversion to hot snare resection in < 1 % of polyps of < 10 mm and 44 % of polyps sized 10-20 mm. The IRRs for polyps < 10 mm and ≥ 10 mm were 18 % and 21 %. The IRR was higher for serrated polyps (26 %) compared with adenomas (16 %). The IRR was higher for flat (IIa) polyps (odds ratio [OR] 2.9, 95 %CI 1.1-7.4); and when resection was judged as difficult (OR 4.2, 95 %CI 1.5-12.1), piecemeal resection was performed (OR 6.6, 95 %CI 2.0-22.0), or visible residual polyp was present after the initial resection (OR 5.4, 95 %CI 2.0-14.9). Polyp location, use of a dedicated cold snare, and submucosal injection were not associated with incomplete resection. Intraprocedural bleeding requiring endoscopic intervention occurred in 4.7 %. CONCLUSIONS : CSP for polyps of 4-9 mm is safe and feasible; however, for lesions ≥ 10 mm, CSP failure occurs frequently, and the IRR remains high even after technical success. Incomplete resection was associated with flat polyps, visual residual polyp, piecemeal resection, and difficult polypectomies., Competing Interests: D. von Renteln is supported by a “Fonds de Recherche du Québec Santé” career development award and has received research funding from ERBE, Ventage, Pendopharm, and Pentax; he is a consultant for Boston Scientific and Pendopharm.R. Djinbachian, F. Benard, A. Barkun, M. Bouin, S. Bouchard, E. Deslandres, B. Panzini, S.Sidani, R. Leduc, and B.C. Jobse declare that they have no conflicts of interest relevant to this paper., (Thieme. All rights reserved.)

The purpose of this study was to describe the social isolation of older adults in the Côte-des-Neiges neighbourhood (Montreal, Canada) from the perspectives of older adults and community stakeholders. To do so, a descriptive qualitative study was conducted, involving community-dwelling older adults and a variety of key stakeholders from the neighbourhood. Seven focus groups were held, with a total of 37 participants. Focus group transcripts were analyzed using the approach of Miles, Huberman, and Saldaña. Participants reported that social isolation of older adults is characterized by gaps in social interactions (scarcity of social interactions, lack of social support, and unsatisfying relationships) as well as by low social participation that can be depicted in three ways: (1) exclusion by society, (2) self-restriction of participation, and (3) low eagerness to socialize. This study highlights that there is a diversity in how social isolation of older adults manifests itself. It can be the result of a deliberate choice (or not), as well as being desired (or not). These aspects of the phenomenon of social isolation of older adults are still not well described. However, they offer relevant avenues for rethinking approaches to intervention development.

Nanotechnology has enabled the development of innovative therapeutics, diagnostics, and drug delivery systems. Nanoparticles (NPs) can influence gene expression, protein synthesis, cell cycle, metabolism, and other subcellular processes. While conventional methods have limitations in characterizing responses to NPs, omics approaches can analyze complete sets of molecular entities that change upon exposure to NPs. This review discusses key omics approaches, namely transcriptomics, proteomics, metabolomics, lipidomics and multi-omics, applied to the assessment of biological responses to NPs. Fundamental concepts and analytical methods used for each approach are presented, as well as good practices for omics experiments. Bioinformatics tools are essential to analyze, interpret and visualize large omics data, and to correlate observations in different molecular layers. The authors envision that conducting interdisciplinary multi-omics analyses in future nanomedicine studies will reveal integrated cell responses to NPs at different omics levels, and the incorporation of omics into the evaluation of targeted delivery, efficacy, and safety will improve the development of nanomedicine therapies., Competing Interests: Declaration of Competing Interest Rene P. Zahedi is CEO of MRM Proteomics, Inc. The rest of authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

G protein-coupled receptors engage both G proteins and β-arrestins, and their coupling can be biased by ligands and mutations. Here, to resolve structural elements and mechanisms underlying effector coupling to the angiotensin II (AngII) type 1 receptor (AT1R), we combined alanine scanning mutagenesis of the entire sequence of the receptor with pharmacological profiling of Gα q and β-arrestin engagement to mutant receptors and molecular dynamics simulations. We showed that Gα q coupling to AT1R involved a large number of residues spread across the receptor, whereas fewer structural regions of the receptor contributed to β-arrestin coupling regulation. Residue stretches in transmembrane domain 4 conferred β-arrestin bias and represented an important structural element in AT1R for functional selectivity. Furthermore, we identified allosteric small-molecule binding sites that were enclosed by communities of residues that produced biased signaling when mutated. Last, we showed that allosteric communication within AT1R emanating from the Gα q coupling site spread beyond the orthosteric AngII-binding site and across different regions of the receptor, including currently unresolved structural regions. Our findings reveal structural elements and mechanisms within AT1R that bias Gα q and β-arrestin coupling and that could be harnessed to design biased receptors for research purposes and to develop allosteric modulators.

Diabetics are a highly comorbid population with an elevated risk profile when undergoing surgery. Proper preparation and management of modifiable risk factors can optimize outcomes in diabetics. A multidisciplinary approach to preoperative optimization, including surgeons, primary care providers, and anesthesiologists, ensures diabetic patients receive comprehensive evaluation before elective surgery. Orthopedic surgeons must understand preoperative optimization goals as they pertain to nutrition, glycemic control, and cardiovascular disease in diabetic patients., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), remains a global public health crisis. The reduced efficacy of therapeutic monoclonal antibodies against emerging SARS-CoV-2 variants of concern (VOCs), such as omicron BA.5 subvariants, has underlined the need to explore a novel spectrum of antivirals that are effective against existing and evolving SARS-CoV-2 VOCs. To address the need for novel therapeutic options, we applied cell-based high-content screening to a library of natural products (NPs) obtained from plants, fungi, bacteria, and marine sponges, which represent a considerable diversity of chemical scaffolds. The antiviral effect of 373 NPs was evaluated using the mNeonGreen (mNG) reporter SARS-CoV-2 virus in a lung epithelial cell line (Calu-3). The screening identified 26 NPs with half-maximal effective concentrations (EC 50 ) below 50 μM against mNG-SARS-CoV-2; 16 of these had EC 50 values below 10 μM and three NPs (holyrine A, alotaketal C, and bafilomycin D) had EC 50 values in the nanomolar range. We demonstrated the pan-SARS-CoV-2 activity of these three lead antivirals against SARS-CoV-2 highly transmissible Omicron subvariants (BA.5, BA.2 and BA.1) and highly pathogenic Delta VOCs in human Calu-3 lung cells. Notably, holyrine A, alotaketal C, and bafilomycin D, are potent nanomolar inhibitors of SARS-CoV-2 Omicron subvariants BA.5 and BA.2. The pan-SARS-CoV-2 activity of alotaketal C [protein kinase C (PKC) activator] and bafilomycin D (V-ATPase inhibitor) suggest that these two NPs are acting as host-directed antivirals (HDAs). Future research should explore whether PKC regulation impacts human susceptibility to and the severity of SARS-CoV-2 infection, and it should confirm the important role of human V-ATPase in the VOC lifecycle. Interestingly, we observed a synergistic action of bafilomycin D and N-0385 (a highly potent inhibitor of human TMPRSS2 protease) against Omicron subvariant BA.2 in human Calu-3 lung cells, which suggests that these two highly potent HDAs are targeting two different mechanisms of SARS-CoV-2 entry. Overall, our study provides insight into the potential of NPs with highly diverse chemical structures as valuable inspirational starting points for developing pan-SARS-CoV-2 therapeutics and for unravelling potential host factors and pathways regulating SARS-CoV-2 VOC infection including emerging omicron BA.5 subvariants., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

TMPRSS6 is a type II transmembrane serine protease involved in iron homeostasis expressed as 4 isoforms in humans. TMPRSS6 isoform 2 downregulates hepcidin production by cleaving hemojuvelin and other surface proteins of hepatocytes. The functions of catalytically impaired isoforms 3 and 4 are still unknown. Here we demonstrate that TMPRSS6 isoforms 3 and 4 reduce the proteolytic activity of isoform 2 and uncover the ability of isoforms to interact. Moreover, we identified 49 potential protein partners common to TMPRSS6 isoforms, including TfR1, known to be involved in iron regulation. By co-expressing TMPRSS6 and TfR1, we show that TfR1 is cleaved and shed from the cell surface. Further, we demonstrate that TMPRSS6 isoforms 3 and 4 behave as dominant negative., Competing Interests: The authors have declared that no competing interests exist.

Latrophilin-3 (Lphn3; also known as ADGRL3) is a member of the adhesion G Protein Coupled Receptor subfamily, which participates in the stabilization and maintenance of neuronal networks by mediating intercellular adhesion through heterophilic interactions with transmembrane ligands. Polymorphisms modifying the Lphn3 gene are associated with attention-deficit/hyperactivity disorder (ADHD) in children and its persistence into adulthood. How these genetic alterations affect receptor function remains unknown. Here, we conducted the functional validation of distinct ADHD-related Lphn3 variants bearing mutations in the receptor's adhesion motif-containing extracellular region. We found that all variants tested disrupted the ability of Lphn3 to stabilize intercellular adhesion in a manner that was distinct between ligands classes, but which did not depend on ligand-receptor interaction parameters, thus pointing to altered intrinsic receptor signaling properties. Using G protein signaling biosensors, we determined that Lphn3 couples to Gαi1, Gαi2, Gαs, Gαq, and Gα13. However, all ADHD-related receptor variants consistently lacked intrinsic as well as ligand-dependent Gα13 coupling efficiency while maintaining unaltered coupling to Gαi, Gαs, and Gαq. Consistent with these alterations, actin remodeling functions as well as actin-relevant RhoA signaling normally displayed by the constitutively active Lphn3 receptor were impeded by select receptor variants, thus supporting additional signaling defects. Taken together, our data point to Gα13 selective signaling impairments as representing a disease-relevant pathogenicity pathway that can be inherited through Lphn3 gene polymorphisms. This study highlights the intricate interplay between Lphn3 GPCR functions and the actin cytoskeleton in modulating neurodevelopmental cues related to ADHD etiology., (© 2022. The Author(s).)

The COVID-19 pandemic caused by the SARS-CoV-2 virus remains a global public health crisis. Although widespread vaccination campaigns are underway, their efficacy is reduced owing to emerging variants of concern 1,2 . Development of host-directed therapeutics and prophylactics could limit such resistance and offer urgently needed protection against variants of concern 3,4 . Attractive pharmacological targets to impede viral entry include type-II transmembrane serine proteases (TTSPs) such as TMPRSS2; these proteases cleave the viral spike protein to expose the fusion peptide for cell entry, and thus have an essential role in the virus lifecycle 5,6 . Here we identify and characterize a small-molecule compound, N-0385, which exhibits low nanomolar potency and a selectivity index of higher than 10 6 in inhibiting SARS-CoV-2 infection in human lung cells and in donor-derived colonoids 7 . In Calu-3 cells it inhibits the entry of the SARS-CoV-2 variants of concern B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma) and B.1.617.2 (Delta). Notably, in the K18-human ACE2 transgenic mouse model of severe COVID-19, we found that N-0385 affords a high level of prophylactic and therapeutic benefit after multiple administrations or even after a single administration. Together, our findings show that TTSP-mediated proteolytic maturation of the spike protein is critical for SARS-CoV-2 infection in vivo, and suggest that N-0385 provides an effective early treatment option against COVID-19 and emerging SARS-CoV-2 variants of concern., (© 2022. The Author(s).)

Transient receptor potential canonical (TRPC) channels are membrane proteins involved in regulating Ca 2+ homeostasis, and whose functions are modulated by G protein-coupled receptors (GPCR). In this study, we developed bioluminescent resonance energy transfer (BRET) biosensors to better study channel conformational changes following receptor activation. For this study, two intramolecular biosensors, GFP10-TRPC7-RLucII and RLucII-TRPC7-GFP10, were constructed and were assessed following the activation of various GPCRs. We first transiently expressed receptors and the biosensors in HEK293 cells, and BRET levels were measured following agonist stimulation of GPCRs. The activation of GPCRs that engage Gα q led to a Gα q -dependent BRET response of the functional TRPC7 biosensor. Focusing on the Angiotensin II type-1 receptor (AT 1 R), GFP10-TRPC7-RLucII was tested in rat neonatal cardiac fibroblasts, expressing endogenous AT 1 R and TRPC7. We detected similar BRET responses in these cells, thus validating the use of the biosensor in physiological conditions. Taken together, our results suggest that activation of Gα q -coupled receptors induce conformational changes in a novel and functional TRPC7 BRET biosensor.

Background: The prevalence of back pain in the pediatric population is increasing, and the workup of these patients presents a clinical challenge. Many cases are selflimited, but failure to diagnose a pathology that requires clinical intervention can carry severe repercussions. Magnetic resonance imaging (MRI) carries a high cost to the patient and health care system, and may even require procedural sedation in the pediatric population. The aim of this study was to develop a scoring system based on pediatric patient factors to help determine when an MRI will change clinical management., Methods: This is a retrospective cohort analysis of consecutive pediatric patients who presented to clinic with a chief complaint of back pain between 2010 and 2018 at single orthopaedic surgery practice. Comprehensive demographic and presentation variables were collected. A predictive model of factors that influence whether MRI results in a change in management was then generated using cross-validation least absolute shrinkage and selection operator logistic regression analysis., Results: A total of 729 patients were included, with a mean age of 15.1 years (range: 3 to 20 y). Of these, 344 (47.2%) had an MRI. A predictive model was generated, with nocturnal symptoms (5 points), neurological deficit (10 points), age (0.7 points per year), lumbar pain (2 points), sudden onset of pain (3.25 points), and leg pain (3.75 points) identified as significant predictors. A combined score of greater than 9.5 points for a given patient is highly suggestive that an MRI will result in a change in clinical management (specificity: 0.93; positive predictive value: 0.92)., Conclusions: A predictive model was generated to help determine when ordering an MRI may result in a change in clinical management for workup of back pain in the pediatric population. The main factors included the presence of a neurological deficit, nocturnal symptoms, sudden onset, leg pain, lumbar pain, and age. Care providers can use these findings to better determine if and when an MRI might be appropriate., Level of Evidence: Level III-diagnostic study., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Delayed graft function (DGF) of a kidney transplant results in increased cost and complexity of management. For clinical care or a DGF trial, it would be ideal to accurately predict individual DGF risk and provide preemptive treatment. A calculator developed by Irish et al has been useful for predicting population but not individual risk., Methods: We analyzed the Irish calculator (IC) in the DeKAF prospective cohort (incidence of DGF = 20.4%) and investigated potential improvements., Results: We found that the predictive performance of the calculator in those meeting Irish inclusion criteria was comparable with that reported by Irish et al. For cohorts excluded by Irish: (a) in pump-perfused kidneys, the IC overestimated DGF risk; (b) in simultaneous pancreas kidney transplants, the DGF risk was exceptionally low. For all 3 cohorts, there was considerable overlap in IC scores between those with and those without DGF. Using a modified definition of DGF-excluding those with single dialysis in the first 24 h posttransplant-we found that the calculator had similar performance as with the traditional DGF definition. Studying whether DGF prediction could be improved, we found that recipient cardiovascular disease was strongly associated with DGF even after accounting for IC-predicted risk., Conclusions: The IC can be a useful population guide for predicting DGF in the population for which it was intended but has limited scope in expanded populations (SPK, pump) and for individual risk prediction. DGF risk prediction can be improved by inclusion of recipient cardiovascular disease., Competing Interests: R.B.M. was supported in part by the UAB-UCSD O’Brien Core Center for Acute Kidney Injury Research (NIH P30-DK079337) (5UO1DK115997) and Department of Veterans Affairs (5-IO1-BX003272). The other authors declare no conflicts of interest., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)