Search

Abstract Background Ligament tension balance is a major determinant for the success of total knee replacement (TKR). The present study aimed at determining the inter-rater and intra-rater reliability in performing ligament tension assessment using an imageless robotic-assisted TKR. Methods Twenty-four knees in 21 patients who received robotic-assisted TKR for end-stage varus osteoarthritis were examined. Three orthopedic specialists and six orthopedic trainees participated in the operations. Data from the ligament tension assessment were collected during the operations. Results For the inter-rater reliability, “extension medial” and “flexion medial” had excellent reliability whilst “extension lateral” and “flexion lateral” had good-to-excellent reliability. For the intra-rater reliability, “extension medial” showed excellent reliability, “extension lateral” and “flexion medial” showed good-to-excellent reliability, and “flexion lateral” showed moderate-to-excellent reliability. Conclusions Robotic-assisted technology provides a reliable solution to improve ligament tension assessment. All ligament tension assessments with the use of the technology could demonstrate at least good-to-excellent reliability except for the intra-rater reliability of “flexion lateral”.

Abstract Whole virion inactivated vaccine CoronaVac (C) and Spike (S) mRNA BNT162b2 (B) vaccines differ greatly in their ability to elicit neutralizing antibodies but have somewhat comparable effectiveness in protecting from severe COVID-19. We conducted further analyses for a randomized trial (Cobovax study, NCT05057169) of third dose homologous and heterologous booster vaccination, i.e. four interventions CC-C, CC-B, BB-C and BB-B. Here, we assess vaccine immunogenicity beyond neutralizing function, including S and non-S antibodies with Fc receptor (FcR) binding, antibody avidity and T cell specificity to 6 months post-vaccination. Ancestral and Omicron S-specific IgG and FcR binding are significantly higher by BNT162b2 booster than CoronaVac, regardless of first doses. Nucleocapsid (N) antibodies are only increased in homologous boosted CoronaVac participants (CC-C). CoronaVac primed participants have lower baseline S-specific CD4+ IFNγ+ cells, but are significantly increased by either CoronaVac or BNT162b2 boosters. Priming vaccine content defined T cell peptide specificity preference, with S-specific T cells dominating B primed groups and non-S structural peptides contributing more in C primed groups, regardless of booster type. S-specific CD4+ T cell responses, N-specific antibodies, and antibody effector functions via Fc receptor binding may contribute to protection and compensate for less potent neutralizing responses in CoronaVac recipients.

Abstract Background Rapid point-of-care tests for malaria are now widely used in many countries to guide the initial clinical management of patients presenting with febrile illness. With China having recently achieved malaria elimination, better understanding regarding the identity and distribution of major non-malarial causes of febrile illnesses is of particular importance to inform evidence-based empirical treatment policy. Methods A systematic review of published literature was undertaken to characterise the spectrum of pathogens causing non-malaria febrile illness in China (1980–2015). Literature searches were conducted in English and Chinese languages in six databases: Ovid MEDLINE, Global Health, EMBASE, Web of Science™ – Chinese Science Citation Database SM, The China National Knowledge Infrastructure (CNKI), and WanFang Med Online. Selection criteria included reporting on an infection or infections with a confirmed diagnosis, defined as pathogens detected in or cultured from samples from normally sterile sites, or serological evidence of current or past infection. The number of published articles, reporting a given pathogen were presented, rather than incidence or prevalence of infection. Results A total of 57,181 records from 13 provinces of China where malaria used to be endemic were screened, of which 392 met selection criteria and were included in this review. The review includes 60 (15.3%) records published from 1980 to 2000, 211 (53.8%) from 2001 to 2010 and 121 (30.9%) from 2011 to 2015;. Of the 392 records, 166 (42.3%) were from the eastern region of China, 120 (30.6%) were from the south-west, 102 (26.0%) from south-central, and four (1.0%) were multi-regional studies. Bacterial infections were reported in 154 (39.3%) records, viral infections in 219 (55.9%), parasitic infections in four (1.0%), fungal infections in one (0.3%), and 14 (3.6%) publications reported more than one pathogen group. Participants of all ages were included in 136 (34.7%) studies, only adults in 75 (19.1%), only children in 17 (4.3%), only neonates in two (0.5%) and the age distribution was not specified in 162 (41.3%) records. The most commonly reported bacterial pathogens included Typhoidal Salmonella (n = 30), Orientia/ Rickettsia tsutsugamushi (n = 31), Coxiella burnetii (n = 17), Leptospira spp. (n = 15) and Brucella spp. (n = 15). The most commonly reported viral pathogens included Hantavirus/Hantaan virus (n = 89), dengue virus (DENV) (n = 76 including those with unknown serovars), Japanese encephalitis virus (n = 21), and measles virus (n = 15). The relative lack of data in the western region of the country, as well as in in neonates and children, represented major gaps in the understanding of the aetiology of fever in China. Conclusions This review presents a landscape of non-malaria pathogens causing febrile illness in China over 36 years as the country progressed toward malaria elimination. These findings can inform guidelines for clinical management of fever cases and infection surveillance and prevention, and highlight the need to standardize operational and reporting protocols for better understanding of fever aetiology in the country.

AbstractPersonalized educational interventions have been shown to facilitate successful and inclusive statistics, mathematics, and data science (SMDS) in higher education through timely and targeted reduction of heterogeneous training disparities caused by years of cumulative, structural challenges in contemporary educational systems. However, the burden on the institutions and instructors to provide personalized training resources to large groups of students is also formidable, and often unsustainable. We present Individualized Pathways and Resources to Adaptive Control Theory-Inspired Scientific Education (iPRACTISE), a free, publicly available web app that serves as a tool to facilitate personalized trainings on SMDS and related topics through provision of personalized training recommendations as informed by computerized assessments and individuals’ training preferences. We describe the resources available in iPRACTISE, and some proof-of-concept evaluation results from deploying iPRACTISE to supplement in-person and online classroom teaching in real-life settings. Strengths, practical difficulties, and potentials for future applications of iPRACTISE to crowdsource and sustain personalized SMDS education are discussed.

Abstract Senescence is a cellular response having physiological and reparative functions to preserve tissue homeostasis and suppress tumor growth. However, the accumulation of senescent cells would cause deleterious effects that lead to age‐related dysfunctions and cancer progression. Hence, selective detection and elimination of senescent cells are crucial yet remain a challenge. A β‐galactosidase (β‐gal)‐activated boron dipyrromethene (BODIPY)‐based photosensitizer (compound 1) is reported here that can selectively detect and eradicate senescent cells. It contains a galactose moiety connected to a pyridinium BODIPY via a self‐immolative nitrophenylene linker, of which the photoactivity is effectively quenched. Upon interactions with the senescence‐associated β‐gal, it undergoes enzymatic hydrolysis followed by self‐immolation, leading to the release of an activated BODIPY moiety by which the fluorescence emission and singlet oxygen generation are restored. The ability of 1 to detect and eliminate senescent cells is demonstrated in vitro and in vivo, using SK‐Mel‐103 tumor‐bearing mice treated with senescence‐inducing therapy. The results demonstrate that 1 can be selectively activated in senescent cells to trigger a robust senolytic effect upon irradiation. This study breaks new ground in the design and application of new senolytic agents based on photodynamic therapy.

Abstract For almost 200 years, the taxonomy of cutthroat trout (Oncorhynchus clarkii), a salmonid native to Western North America, has been in flux as ichthyologists and fisheries biologists have tried to describe the diversity within these fishes. Starting in the 1950s, Robert Behnke reexamined the cutthroat trout and identified 14 subspecies based on morphological traits, Pleistocene events, and modern geographic ranges. His designations became instrumental in recognizing and preserving the remaining diversity of cutthroat trout. Over time, molecular techniques (i.e. karyotypes, allozymes, mitochondrial DNA, SNPs, and microsatellite arrays) have largely reinforced Behnke's phylogenies, but have also revealed that some relationships are consistently weakly supported. To further resolve these relationships, we generated de novo transcriptomes for nine cutthroat subspecies, as well as a Bear River Bonneville form and two Colorado River lineages (blue and green). We present phylogenies of these subspecies generated from multiple sets of orthologous genes extracted from our transcriptomes. We confirm many of the relationships identified in previous morphological and molecular studies, as well as discuss the importance of significant differences apparent in our phylogenies from these studies within a geological perspective. Specific findings include three distinct clades: (1) Bear River Bonneville form and Yellowstone cutthroat trout; (2) Bonneville cutthroat trout (n = 2); and (3) Greenback and Rio Grande cutthroat trout. We also identify potential gene transfer between Bonneville cutthroat trout and a population of Colorado River green lineage cutthroat trout. Using these findings, it appears that additional groups warrant species‐level consideration if other recent species elevations are retained.

Singlet oxygen (1O2) is a potent biocide potentially deployable for integrated control of tropical diseases and their insect vectors. This very short-lived free radical is highly destructive of cellular molecules when generated intracellularly. Most organisms, including parasites and vectors, are defenseless against 1O2 except for plants, which produce it abundantly during photosynthesis, hence, the acquisition of specific mechanisms for its detoxification. In the presence of O2 under physiological conditions, certain dyes or photosensitizers (PS), e.g., porphyrins and phthalocyanines (PC), are excitable by light to produce biocidal 1O2. Its half-life is in the order of microseconds, necessitating its intracellular generation in order to harness its biocidal activity most effectively. This is achievable by loading cells with PS for excitation with light to produce 1O2in situ. One example to achieve this is the genetic engineering of Leishmania to complement its inherent defects in porphyrin biosynthesis, resulting in cytosolic accumulation of abundant PS in the form of uroporphyrin 1 (URO). Another example is the chemical engineering of PC for hydrophilicity, thereby facilitating the endocytosis of such PS by cells. Leishmania loaded with cytosolic URO and endosomal PC are inactivated by the 1O2 produced via light-activation of these PS in the two different cell compartments. The inactivated Leishmania are nonviable, but have their natural vaccines and adjuvants well-preserved for prophylactic vaccination against experimental leishmaniasis. 1O2-inactivated Leishmania is potentially useful to serve as a platform for the safe and effective delivery of transgenically add-on vaccines against malignant and viral diseases in experimental models. Hydrophilic and cationic PC were also shown experimentally to act as a new type of dim light-activable insecticides, i.e., their mosquito larvicidal activities with

Species delimitation can be based on the consideration of several different criteria, including the differentiation of ecological or functional traits. Two species of Pacific rockfish, the dark rockfish (Sebastes ciliatus) and the dusky rockfish (Sebastes variabilis), appear to represent recently divergent evolutionary lineages. We evaluate evidence for the differentiation of these two species in somatic shape using geometric morphometrics at two locations in the northeast Pacific where they occur in sympatry. The somatic shape was significantly different between species, but the species’ shape did not vary between the two locations. Sebastes ciliatus had an upturned and relatively smaller head, eye, and jaw, and an elongated midbody, whereas S. variabilis had a downturned and larger head, eye, and jaw, and a shorter midbody. These results suggest that S. ciliatus and S. variabilis are morphometrically differentiated in a similar way in both locations. The somatic shape differentiation between these two sympatric species is similar to genus-wide patterns of somatic shape differentiation.

Summary: Heterochrony—alteration to the rate or timing of development—is an important mechanism of trait differentiation associated with speciation. Heterochrony may explain the morphological divergence between two polyploid species, June sucker (Chasmistes liorus) and Utah sucker (Catostomus ardens). The larvae of both species have terminal mouths; however, as adults, June sucker and Utah sucker develop subterminal and ventral mouths, respectively. We document a difference in the timing of shape development and a corresponding change in the timing of gene expression, suggesting the distinctive mouth morphology in June suckers may result from paedomorphosis. Specifically, adult June suckers exhibit an intermediate mouth morphology between the larval (terminal) and ancestral (ventral) states. Endemic and sympatric Chasmistes/Catostomus pairs in two other lakes also are morphologically divergent, but genetically similar. These species pairs could have resulted from the differential expression of genes and corresponding divergence in trait development. Paedomorphosis may lead to adaptive diversification in Catostomids.

Abstract As an effective and non‐invasive treatment modality for cancer, photodynamic therapy (PDT) has attracted considerable interest. With the recent advances in the photosensitizing agents, the fiber‐optic systems, and other aspects, its application is extended to a wide range of superficial and localized cancers. However, for the few clinically used photosensitizers, most of them suffer from the drawback of causing prolonged photosensitivity after the treatment. As a result, post‐PDT management is also a crucial issue. Herein, a facile bioorthogonal approach is reported that can effectively suppress this common side effect of PDT in nude mice. It involves the use of an antidote that contains a black‐hole quencher BHQ‐3 conjugated with a bicyclo[6.1.0]non‐4‐yne (BCN) moiety and a tetrazine‐substituted boron dipyrromethene‐based photosensitizer. By using tumor‐bearing nude mice as an animal model, it is demonstrated that after PDT with this photosensitizer, the administration of the antidote can effectively quench the photodynamic activity of the residual photosensitizer by bringing the BHQ‐3 quencher close to the photosensitizing unit through a rapid click reaction. It results in substantial reduction in skin damage upon light irradiation. The overall results demonstrate that this simple and facile strategy can provide an effective means for minimizing the photosensitivity after PDT.

Abstract Background Acute exacerbations of chronic inflammatory lung diseases, such as chronic obstructive pulmonary disease (COPD), are frequently associated with rhinovirus (RV) infections. Despite these associations, the pathogenesis of virus-induced exacerbations is incompletely understood. We aimed to investigate effects of cigarette smoke (CS), a primary risk factor for COPD, on RV infection in airway epithelium and identify novel mechanisms related to these effects. Methods Primary bronchial epithelial cells (PBEC) from COPD patients and controls were differentiated by culture at the air–liquid interface (ALI) and exposed to CS and RV-A16. Bulk RNA sequencing was performed using samples collected at 6 and 24 h post infection (hpi), and viral load, mediator and l-lactate levels were measured at 6, 24 and 48hpi. To further delineate the effect of CS on RV-A16 infection, we performed growth differentiation factor 15 (GDF15) knockdown, l-lactate and interferon pre-treatment in ALI-PBEC. We performed deconvolution analysis to predict changes in the cell composition of ALI-PBEC after the various exposures. Finally, we compared transcriptional responses of ALI-PBEC to those in nasal epithelium after human RV-A16 challenge. Results CS exposure impaired antiviral responses at 6hpi and increased viral replication at 24 and 48hpi in ALI-PBEC. At 24hpi, CS exposure enhanced expression of RV-A16-induced epithelial interferons, inflammation-related genes and CXCL8. CS exposure increased expression of oxidative stress-related genes, of GDF15, and decreased mitochondrial membrane potential. GDF15 knockdown experiments suggested involvement of this pathway in the CS-induced increase in viral replication. Expression of glycolysis-related genes and l-lactate production were increased by CS exposure, and was demonstrated to contribute to higher viral replication. No major differences were demonstrated between COPD and non-COPD-derived cultures. However, cellular deconvolution analysis predicted higher secretory cells in COPD-derived cultures at baseline. Conclusion Altogether, our findings demonstrate that CS exposure leads to higher viral infection in human bronchial epithelium by altering not only interferon responses, but likely also through a switch to glycolysis, and via GDF15-related pathways.

Abstract Background The 95-95-95 UNAIDS global strategy was adapted to end the AIDS epidemic by 2030. The target is based on the premise that early detection of HIV-infected persons and linking them to treatment regardless of their CD4 counts will lead to sustained viral suppression. HIV testing strategies to increase uptake of testing in Western and Central Africa remain inadequate. Hence, a high proportion of people living with HIV in this region do not know their status. This report describes the implementation of a community based multi-disease health screening (also known as “Know Your Status” -KYS), as part of basic science research, in a way that contributed to achieving public health goals. Methods A community based multi-disease health screening was conducted in 7 communities within the Eastern region of Ghana between November 2017 and April 2018, to recruit and match HIV seronegative persons to HIV seropositive persons in a case-control HIV gut microbiota study. Health assessments included blood pressure, body mass index, blood sugar, Hepatitis B virus, syphilis, and HIV testing for those who consented. HIV seronegative participants who consented were consecutively enrolled in an ongoing HIV gut microbiota case-control study. Descriptive statistics (percentages) were used to analyze data. Results Out of 738 people screened during the exercise, 700 consented to HIV testing and 23 (3%) were HIV positive. Hepatitis B virus infection was detected in 4% (33/738) and Syphilis in 2% (17/738). Co-infection of HIV and HBV was detected in 4 persons. The HIV prevalence of 3% found in these communities is higher than both the national prevalence of 1.7% and the Eastern Regional prevalence of 2.7 in 2018. Conclusion Community based multi-disease health screening, such as the one undertaken in our study could be critical for identifying HIV infected persons from the community and linking them to care. In the case of HIV, it will greatly contribute to achieving the first two 95s and working towards ending AIDS by 2030.

Abstract Objectives Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is a member of a class of highly pathogenic coronaviruses. The large family of coronaviruses, however, also includes members that cause only mild symptoms, like human coronavirus‐229E (HCoV‐229E) or OC43 (HCoV‐OC43). Unravelling how molecular (and cellular) pathophysiology differs between highly and low pathogenic coronaviruses is important for the development of therapeutic strategies. Methods Here, we analysed the transcriptome of primary human bronchial epithelial cells (PBEC), differentiated at the air–liquid interface (ALI) after infection with SARS‐CoV‐2, SARS‐CoV, Middle East Respiratory Syndrome (MERS)‐CoV and HCoV‐229E using bulk RNA sequencing. Results ALI‐PBEC were efficiently infected by all viruses, and SARS‐CoV, MERS‐CoV and HCoV‐229E infection resulted in a largely similar transcriptional response. The response to SARS‐CoV‐2 infection differed markedly as it uniquely lacked the increase in expression of immediate early genes, including FOS, FOSB and NR4A1 that was observed with all other coronaviruses. This finding was further confirmed in publicly available experimental and clinical datasets. Interfering with NR4A1 signalling in Calu‐3 lung epithelial cells resulted in a 100‐fold reduction in extracellular RNA copies of SARS‐CoV‐2 and MERS‐CoV, suggesting an involvement in virus replication. Furthermore, a lack in induction of interferon‐related gene expression characterised the main difference between the highly pathogenic coronaviruses and low pathogenic viruses HCoV‐229E and HCoV‐OC43. Conclusion Our results demonstrate a previously unknown suppression of a host response gene set by SARS‐CoV‐2 and confirm a difference in interferon‐related gene expression between highly pathogenic and low pathogenic coronaviruses.

Abstract In randomized controlled trials of influenza vaccination, 550 children received trivalent-inactivated influenza vaccine, permitting us to explore relationship between vaccine response and host single nucleotide polymorphisms (SNPs) in 23 candidate genes with adjustment of multiple testing. For host SNPs in TLR7–1817G/T (rs5741880), genotype GT was associated with lower odds (OR: 0.22, 95% CI: 0.09, 0.53) of have post-vaccination hemagglutination-inhibiting (HAI) titers ≥40, compared with genotype GG and TT combined under the over-dominant model. For host SNPs in TLR8–129G/C (rs3764879), genotype GT was associated with lower odds (OR: 0.47; 95% CI: 0.28, 0.80) of have post vaccination HAI titers ≥40 compared with genotype GG and AA combined under the over-dominant model. Our results could contribute to the development of better vaccines that may offer improved protection to all recipients.

Lung epithelial cells represent the first line of host defence against foreign inhaled components, including respiratory pathogens. Their responses to these exposures may direct subsequent immune activation to these pathogens. The epithelial response to mycobacterial infections is not well characterized and may provide clues to why some mycobacterial infections are cleared, while others are persistent and pathogenic. We have utilized an air-liquid interface model of human primary bronchial epithelial cells (ALI-PBEC) to investigate the epithelial response to infection with a variety of mycobacteria: Mycobacterium tuberculosis (Mtb), M. bovis (BCG), M. avium, and M. smegmatis. Airway epithelial cells were found to be infected by all four species, albeit at low frequencies. The proportion of infected epithelial cells was lowest for Mtb and highest for M. avium. Differential gene expression analysis revealed a common epithelial host response to mycobacteria, including upregulation of BIRC3, S100A8 and DEFB4, and downregulation of BPIFB1 at 48 h post infection. Apical secretions contained predominantly pro-inflammatory cytokines, while basal secretions contained tissue growth factors and chemokines. Finally, we show that neutrophils were attracted to both apical and basal secretions of infected ALI-PBEC. Neutrophils were attracted in high numbers to apical secretions from PBEC infected with all mycobacteria, with the exception of secretions from M. avium-infected ALI-PBEC. Taken together, our results show that airway epithelial cells are differentially infected by mycobacteria, and react rapidly by upregulation of antimicrobials, and increased secretion of inflammatory cytokines and chemokines which directly attract neutrophils. Thus, the airway epithelium may be an important immunological component in controlling and regulating mycobacterial infections.

Human lung function is intricately linked to blood flow and breathing cycles, but it remains unknown how these dynamic cues shape human airway epithelial biology. Here we report a state-of-the-art protocol for studying the effects of dynamic medium and airflow as well as stretch on human primary airway epithelial cell differentiation and maturation, including mucociliary clearance, using an organ-on-chip device. Perfused epithelial cell cultures displayed accelerated maturation and polarization of mucociliary clearance, and changes in specific cell-types when compared to traditional (static) culture methods. Additional application of airflow and stretch to the airway chip resulted in an increase in polarization of mucociliary clearance towards the applied flow, reduced baseline secretion of interleukin-8 and other inflammatory proteins, and reduced gene expression of matrix metalloproteinase (MMP) 9, fibronectin, and other extracellular matrix factors. These results indicate that breathing-like mechanical stimuli are important modulators of airway epithelial cell differentiation and maturation and that their fine-tuned application could generate models of specific epithelial pathologies, including mucociliary (dys)function.

Abstract Background Despite the well-known detrimental effects of cigarette smoke (CS), little is known about the complex gene expression dynamics in the early stages after exposure. This study aims to investigate early transcriptomic responses following CS exposure of airway epithelial cells in culture and compare these to those found in human CS exposure studies. Methods Primary bronchial epithelial cells (PBEC) were differentiated at the air–liquid interface (ALI) and exposed to whole CS. Bulk RNA-sequencing was performed at 1 h, 4 h, and 24 h hereafter, followed by differential gene expression analysis. Results were additionally compared to data retrieved from human CS studies. Results ALI-PBEC gene expression in response to CS was most significantly changed at 4 h after exposure. Early transcriptomic changes (1 h, 4 h post CS exposure) were related to oxidative stress, xenobiotic metabolism, higher expression of immediate early genes and pro-inflammatory pathways (i.e., Nrf2, AP-1, AhR). At 24 h, ferroptosis-associated genes were significantly increased, whereas PRKN, involved in removing dysfunctional mitochondria, was downregulated. Importantly, the transcriptome dynamics of the current study mirrored in-vivo human studies of acute CS exposure, chronic smokers, and inversely mirrored smoking cessation. Conclusion These findings show that early after CS exposure xenobiotic metabolism and pro-inflammatory pathways were activated, followed by activation of the ferroptosis-related cell death pathway. Moreover, significant overlap between these transcriptomic responses in the in-vitro model and human in-vivo studies was found, with an early response of ciliated cells. These results provide validation for the use of ALI-PBEC cultures to study the human lung epithelial response to inhaled toxicants.

Abstract Background Pathogenic variants in AXIN2 have been associated with tooth agenesis, colon polyps, and colon cancer. Given the rare nature of this phenotype, we set out to collect additional genotypic and phenotypic information. Methods Data were collected via a structured questionnaire. Sequencing was performed in these patients mostly due to diagnostic purpose. A little more than half of the AXIN2 variant carriers were identified by NGS; other six were family members. Results Here, we report 13 individuals with a heterozygous AXIN2 pathogenic/likely pathogenic variant who have a variable expression of oligodontia‐colorectal cancer syndrome (OMIM 608615) or oligodontia‐cancer predisposition syndrome (ORPHA 300576). Three individuals from one family also had cleft palate, which might represent a new clinical feature of AXIN2 phenotype, also given the fact that AXIN2 polymorphisms have been found in association with oral clefting in population studies. AXIN2 has already been added to multigene cancer panel tests; further research should be conducted to determine whether it should be added to cleft lip/palate multigene panels. Conclusion More clarity about oligodontia‐colorectal cancer syndrome, about the variable expression, and associated cancer risks is needed to improve clinical management and to establish guidelines for surveillance. We collected information about the surveillance that was advised, which might support clinical management of these patients.

Background: The objective of this study is to find novel antineoplastic agents that display greater toxicity to malignant cells than to neoplasms. In addition, the mechanisms of action of representative compounds are sought. This report describes the cytotoxicity of a number of dimers of 3,5-bis(benzylidene)-4-piperidones against human malignant cells (promyelocytic leukemia HL-60 and squamous cell carcinoma HSC-2, HSC-3, and HSC-4). Methods: Tumor specificity was evaluated by the selectivity index (SI), that is the ratio of the mean CC50 for human non-malignant oral cells (gingival fibroblasts, pulp cells, periodontal ligament fibroblasts) to that for malignant cells. Results: The compounds were highly toxic to human malignant cells. On the other hand, these molecules were less toxic to human non-malignant cells. In particular, a potent lead molecule, 3b, was identified. A QSAR study revealed that the placement of electron-releasing and hydrophilic substituents into the aryl rings led to increases in cytotoxic potencies. The modes of action of a lead compound discovered in this study designated 3b were the activation of caspases-3 and -7, as well as causing PARP1 cleavage and G2 arrest, followed by sub-G1 accumulation in the cell cycle. This compound also depolarized the mitochondrial membrane and generated reactive oxygen species in human colon carcinoma HCT116 cells. In conclusion, this study has revealed that, in general, the compounds described in this report are tumor-selective cytotoxins.

Undiagnosed and untreated tyrosinemia type 1 (TT1) individuals carry a significant risk for developing liver fibrosis, cirrhosis and hepatocellular carcinoma (HCC). Elevated succinylacetone (SA) is pathognomonic for TT1 and therefore often used as marker for TT1 newborn screening (NBS). While SA was long considered to be elevated in every TT1 patient, here we present a recent false-negative SA TT1 screen. A nine-year-old boy presented with HCC in a cirrhotic liver. Additional tests for the underlying cause unexpectedly revealed TT1. Nine years prior, the patient was screened for TT1 via SA NBS with a negative result: SA 1.08 µmol/L, NBS cut-off 1.20 µmol/L. To our knowledge, this report is the first to describe a false-negative result from the TT1 NBS using SA. False-negative TT1 NBS results may be caused by milder TT1 variants with lower SA excretion. Such patients are more likely to be missed in NBS programs and can be asymptomatic for years. Based on our case, we advise TT1 to be considered in patients with otherwise unexplained liver pathology, including fibrosis, cirrhosis and HCC, despite a previous negative TT1 NBS status. Moreover, because the NBS SA concentration of this patient fell below the Dutch cut-off value (1.20 µmol/L at that time), as well as below the range of cut-off values used in other countries (1.29–10 µmol/L), it is likely that false-negative screening results for TT1 may also be occurring internationally. This underscores the need to re-evaluate TT1 SA NBS programs.

The three most important commercial bacterial insecticides are all derived from subspecies of Bacillus thuringiensis (Bt). Specifically, Bt subsp. kurstaki (Btk) and Bt subsp. aizawai (Bta) are used to control larval lepidopteran pests. The third, Bt subsp. israelensis (Bti), is primarily used to control mosquito and blackfly larvae. All three subspecies produce a parasporal body (PB) during sporulation. The PB is composed of insecticidal proteins that damage the midgut epithelium, initiating a complex process that results in the death of the insect. Among these three subspecies of Bt, Bti is unique as it produces the most complex PB consisting of three compartments. Each compartment is bound by a multilaminar fibrous matrix (MFM). Two compartments contain one protein each, Cry11Aa1 and Cyt1Aa1, while the third contains two, Cry4Aa1/Cry4Ba1. Each compartment is packaged independently before coalescing into the mature spherical PB held together by additional layers of the MFM. This distinctive packaging process is unparalleled among known bacterial organelles, although the underlying molecular biology is yet to be determined. Here, we present structural and molecular evidence that the MFM has a hexagonal pattern to which Bti proteins Bt152 and Bt075 bind. Bt152 binds to a defined spot on the MFM during the development of each compartment, yet its function remains unknown. Bt075 appears to be derived from a bacteriophage major capsid protein (MCP), and though its sequence has markedly diverged, it shares striking 3-D structural similarity to the Escherichia coli phage HK97 Head 1 capsid protein. Both proteins are encoded on Bti’s pBtoxis plasmid. Additionally, we have also identified a six-amino acid motif that appears to be part of a novel molecular process responsible for targeting the Cry and Cyt proteins to their cytoplasmic compartments. This paper describes several previously unknown features of the Bti organelle, representing a first step to understanding the biology of a unique process of sorting and packaging of proteins into PBs. The insights from this research suggest a potential for future applications in nanotechnology.

Abstract Ocular aerial surveys allow efficient coverage of large areas and can be used to monitor abundance and distribution of wild populations. However, uncertainty around resulting population estimates can be large due to difficulty in visually identifying and counting animals from aircraft, as well as logistical challenges in estimating detection probabilities. Photographic aerial surveys can mitigate these challenges and can allow flight at higher altitudes to minimize disturbance of birds and improve safety for surveyors. We evaluated a photographic aerial survey that incorporated a systematic sampling design with automated photo capture and processing for fall‐staging geese at Izembek Lagoon, Alaska, in 2017–2019. Ocular aerial surveys have been completed at Izembek Lagoon for >40 years. For the new photo survey, we used a commercial system to automatically trigger cameras at preset points. We then applied a machine‐learning algorithm trained to automatically identify and count geese in our photos, manually corrected those counts, and quantified the algorithm's accuracy. We translated corrected counts into density and extrapolated mean density across the entire lagoon to estimate total population size for Pacific brant (Branta bernicla) and cackling geese (B. hutchinsii). The automated algorithm undercounted geese, but successfully identified the small subset of photos containing geese. Manual correction was therefore needed only for photos automatically identified as containing geese, allowing substantial reduction of workload. Manually‐corrected, photo‐based estimates of Pacific brant and cackling goose population sizes were larger and more precise than ocular estimates in all 3 years. To reduce costs with little penalty for variance around population estimates, the photographic survey design could be optimized by reducing the number of transects to ~67% of the current number while still manually correcting all photos in which the automated algorithm detected geese. Further years of both ocular and photo surveys would be needed to calibrate the photo estimates against the >40‐year timeseries of the ocular survey, after which the photo series could successfully guide management of Pacific brant. As technologies continue to advance, we expect photographic surveys with automated counting to be easily implemented and advantageous to many monitoring programs.

The consequences of infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can range from asymptomatic to fatal disease. Variations in epithelial susceptibility to SARS-CoV-2 infection depend on the anatomical location from the proximal to distal respiratory tract. However, the cellular biology underlying these variations is not completely understood. Thus, air-liquid interface (ALI) cultures of well-differentiated primary human tracheal and bronchial epithelial cells were employed to study the impact of epithelial cellular composition and differentiation on SARS-CoV-2 infection by transcriptional (RNA sequencing) and immunofluorescent analyses. Changes of cellular composition were investigated by varying time of differentiation or by using specific compounds. We found that SARS-CoV-2 primarily infected ciliated cells but also goblet cells and transient secretory cells. Viral replication was impacted by differences in cellular composition, which depended on culturing time and anatomical origin. A higher percentage of ciliated cells correlated with a higher viral load. However, DAPT-treatment, which increased number of ciliated cells and reduced goblet cells, decreased viral load, indicating the contribution of goblet cells to infection. Cell-entry factors, especially cathepsin L and transmembrane protease serine 2, were also affected by differentiation time. In conclusion, our study demonstrates that viral replication is affected by changes in cellular composition, especially in cells related to the mucociliary system. This could explain in part the variable susceptibility to SARS-CoV-2 infection between individuals and between anatomical locations in the respiratory tract.

We report the complete mitochondrial genomes of two rockfish: Sebastes maliger and Sebastes norvegicus. The mitogenomes consist of 13 protein-coding regions, 22 tRNAs, two rRNAs, and one control region. Sebastes mitogenome control regions are highly variable due to the presence of repeat sequences. The mitogenomes for S. maliger and S. norvegicus are 16,403 and 16,401 bp, respectively. Using these two mitogenomes and 25 additional Sebastes mitogenomes from GenBank, we examine the phylogenetic relationships in Sebastes. Sebastes maliger is sister to a clade including S. rubrivinctus, S. nigrocinctus, S. umbrosus, and S. oculatus, while S. norvegicus is sister to S. fasciatus.

Serological classification of influenza infection has classically been based on a four-fold or higher increase in antibody levels, but this approach may not be optimal. Here, the authors develop a Bayesian model to improve identification of infections in serological samples by accounting for individual antibody dynamics.

This work studies the emulsifying and antioxidant properties of potato protein hydrolysates (PPHs) fractions obtained through enzymatic hydrolysis of potato protein using trypsin followed by ultrafiltration. Unfractionated (PPH1) and fractionated (PPH2 as >10 kDa, PPH3 as 10–5 kDa, PPH4 as 5–0.8 kDa, and PPH5 as 10 kDa showed the highest ability to decrease oil–water interfacial tension. All PPH fractions predominantly provided elastic, weak, and easily stretchable interfaces. PPH2 provided a more rigid interfacial layer than the other hydrolysates. Radical scavenging and metal chelating activities of PPHs were also tested and the highest activities were provided by the unfractionated hydrolysate and the fractions with peptides >5 kDa. Furthermore, the ability of PPHs to form physically and oxidatively stable 5% fish oil-in-water emulsions (pH 7) was investigated during 8-day storage at 20 °C. Our results generally show that the fractions with peptides >5 kDa provided the highest physicochemical stability, followed by the fraction with peptides between 5 and 0.8 kDa. Lastly, promising sensory results with mostly mild attributes were obtained even at protein concentration levels that are higher than needed to obtain functional properties. The more prominent attributes (e.g., bitterness and astringency) were within an acceptable range for PPH3 and PPH4.

Various approaches have been discussed in the literature for the clustering of data, such as partitioning, hierarchical, and machine learning methods. Most of the approaches require some prior knowledge about the clusters, such as their total number. Furthermore, some previous algorithms are not robust enough to process higher-dimensional data or require a large amount of memory for computations. We propose, herein, a data clustering algorithm, Piecemeal Clustering, that successfully clusters data without prior knowledge of the number of clusters. The proposed clustering algorithm uses the similarity and density of the data to identify the number of clusters in the data set and works with both low- and high-dimensional data. We demonstrate the power of the proposed Piecemeal Clustering algorithm with two real-world data sets. It is found that the proposed algorithm outperforms seven other state-of-the-art algorithms on both of these data sets.

Abstract Conservation translocations have become an important management tool, particularly for large wildlife species such as the lion (Panthera leo). When planning translocations, the genetic background of populations needs to be taken into account; failure to do so risks disrupting existing patterns of genetic variation, ultimately leading to genetic homogenization, and thereby reducing resilience and adaptability of the species. We urge wildlife managers to include knowledge of the genetic background of source/target populations, as well as species‐wide patterns, in any management intervention. We present a hierarchical decision‐making tool in which we list 132 lion populations/lion conservation units and provide information on genetic assignment, uncertainty and suitability for translocation for each source/target combination. By including four levels of suitability, from ‘first choice’ to ‘no option’, we provide managers with a range of options. To illustrate the extent of international trade of lions, and the potential disruption of natural patterns of intraspecific diversity, we mined the CITES Trade Database for estimated trade quantities of live individuals imported into lion range states during the past 4 decades. We identified 1056 recorded individuals with a potential risk of interbreeding with wild lions, 772 being captive‐sourced. Scoring each of the records with our decision‐making tool illustrates that only 7% of the translocated individuals were ‘first choice’ and 73% were ‘no option’. We acknowledge that other, nongenetic factors are important in the decision‐making process, and hence a pragmatic approach is needed. A framework in which source/target populations are scored based on suitability is not only relevant to lion, but also to other species of wildlife that are frequently translocated. We hope that the presented overview supports managers to include genetics in future management decisions and contributes towards conservation of the lion in its full diversity.

In this study, integrated geostatistical techniques were applied to appraise the groundwater quality for various purposes in the Lower Anayari Catchment (LAC), Ghana. Physico-chemical and trace metals were analysed for 41 groundwater samples. Groundwater in LAC was generally found to be moderately hard to fresh and the information entropy-based groundwater quality index (IEGQI) ranked approximately 80.49% of the groundwater as excellent. Partial Least square regression analysis suggests that PO43−, As, Cd and Fe are important parameters for the prediction of IEGQI values. The irrigation water quality index (IWQI) suggest that 2.44% of groundwater in LAC is suitable for moderate permeability soils, 65.85% for moderate to high permeability soils and 31.71% for moderate to high salt tolerance plants. The LSI and RSI indices rate groundwater in LAC to be aggressive water with a high probability of being corrosive to metals, depending on the levels of dissolved oxygen present.

BackgroundLittle is known about patient preferences and the value of pharmacy-collaborative disease management with primary care using technology-driven interprofessional communication under real-world conditions. Discrete Choice Experiments (DCEs) are useful for quantifying preferences for non-market services.Objectives1) To explore variation in patient preferences and estimate willingness-to-accept annual cost to the National Health Service (NHS) for attributes of a collaborative intervention trial between pharmacies and primary care using a trial exit DCE interview; 2) to incorporate a DCE into an economic evaluation using cost-benefit analysis (CBA).MethodsWe performed a DCE telephone interview with a sample of hypertension and hyperlipidemia trial patients 12 months after trial onset. We used five attributes (levels): waiting time to get urgent/not urgent medical appointment (7 days/45 days; 48 hrs./30 days; same day/15 days), model of pharmacy intervention (5-min. counter basic check; 15-min. office every 3 months for BP and medication review of selected medicines; 30-min. office every 6 months for comprehensive measurements and medication review of all medicines), integration with primary care (weak; partial; full), chance of having a stroke in 5 years (same; slightly lower; much lower), and annual cost to the NHS (0€; 30€; 51€; 76€). We used an experimental orthogonal fractional factorial design. Data were analyzed using conditional logit. We subtracted the estimated annual incremental trial costs from the mean WTA (Net Benefit) for CBA.ResultsA total of 122 patients completed the survey. Waiting time to get medical appointment-on the same day (urgent) and within 15 days (non-urgent)-was the most important attribute, followed by 30-minute pharmacy intervention in private office every 6 months for point-of-care measurements and medication review of all medicines, and full integration with primary care. The cost attribute was not significant. Intervention patients were willing to accept the NHS annual cost of €877 for their preferred scenario. The annual net benefit per patient is €788.20 and represents the monetary value of patients' welfare surplus for this model.ConclusionsThis study is the first conducted in Portugal alongside a pharmacy collaborative trial, incorporating DCE into CBA. The findings can be used to guide the design of pharmacy collaborative interventions with primary care with the potential for reimbursement for uncontrolled or at-risk chronic disease patients informed by patient preferences. Future DCE studies conducted in community pharmacy may provide additional contributions.Trial registrationCurrent Controlled Trials (ISRCTN): ISRCTN13410498, retrospectively registered on 12 December 2018.

BackgroundInfluenza causes considerable disease burden each year, particularly in children. Monitoring school absenteeism has long been proposed as a surveillance tool of influenza activity in the community, but the practice of school absenteeism could be varying, and the potential of such usage remains unclear. ObjectiveThe aim of this paper is to determine the potential of monitoring school absenteeism as a surveillance tool of influenza. MethodsWe conducted a systematic review of the published literature on the relationship between school absenteeism and influenza activity in the community. We categorized the types of school absenteeism and influenza activity in the community to determine the correlation between these data streams. We also extracted this correlation with different lags in community surveillance to determine the potential of using school absenteeism as a leading indicator of influenza activity. ResultsAmong the 35 identified studies, 22 (63%), 12 (34%), and 8 (23%) studies monitored all-cause, illness-specific, and influenza-like illness (ILI)–specific absents, respectively, and 16 (46%) used quantitative approaches and provided 33 estimates on the temporal correlation between school absenteeism and influenza activity in the community. The pooled estimate of correlation between school absenteeism and community surveillance without lag, with 1-week lag, and with 2-week lag were 0.44 (95% CI 0.34, 0.53), 0.29 (95% CI 0.15, 0.42), and 0.21 (95% CI 0.11, 0.31), respectively. The correlation between influenza activity in the community and ILI-specific absenteeism was higher than that between influenza activity in community all-cause absenteeism. Among the 19 studies that used qualitative approaches, 15 (79%) concluded that school absenteeism was in concordance with, coincided with, or was associated with community surveillance. Of the 35 identified studies, only 6 (17%) attempted to predict influenza activity in the community from school absenteeism surveillance. ConclusionsThere was a moderate correlation between school absenteeism and influenza activity in the community. The smaller correlation between school absenteeism and community surveillance with lag, compared to without lag, suggested that careful application was required to use school absenteeism as a leading indicator of influenza epidemics. ILI-specific absenteeism could monitor influenza activity more closely, but the required resource or school participation willingness may require careful consideration to weight against the associated costs. Further development is required to use and optimize the use of school absenteeism to predict influenza activity. In particular, the potential of using more advanced statistical models and validation of the predictions should be explored.

Abstract Background Drug checking uses chemical analytical technologies to analyze drugs from the unregulated market to reduce substance use-related risks. We aim to examine the frequency of third party use of a community drug checking service to explore the potential for harm reduction to extend beyond the individual into the community, increase service accessibility, and to contribute to upstream interventions in the supply. Methods Over 31 months, data were collected from a point-of-care drug checking service operated in Victoria, Canada. Through the implementation of survey questions at the intake of the service, data were collected about whether the drug check was for the individual, to sell, and/or for others. Results Just over half (52%) of service users were checking for reasons that extended beyond individual use. When checking for others, friends were the most common response, representing 52% of responses, and outreach/support workers checking for others was the second most at 32%. Twelve percent of service users reported checking to sell or for a supplier. Conclusions Third party checking is a frequent, and important aspect of drug checking services, which through facilitating community engagement and increasing accessibility, has expanded the reach of interventions beyond individuals to reduce risks within the unregulated market. Therefore, drug checking as an overdose response should be responsive and accessible for those using the service on the behalf of others.

This article describes a suite of student-created Shiny apps for teaching statistics and a field test of their short-term effectiveness. To date, more than 50 Shiny apps and a growing collection of associated lesson plans, designed to enrich the teaching of both introductory and upper division statistics courses, have been developed. The apps are available for free use and their open source code can be adapted as desired. We report on the experimental testing of four of these Shiny apps to examine short-term learning outcomes in an introductory statistical concepts course.

Abstract Ascoviruses are large dsDNA viruses characterized by the extraordinary changes they induce in cellular pathogenesis and architecture whereby after nuclear lysis and extensive hypertrophy, each cell is cleaved into numerous vesicles for virion reproduction. However, the level of viral replication and transcription in vesicles compared to other host tissues remains uncertain. Therefore, we applied RNA-Sequencing to compare the temporal transcriptome of Spodoptera frugiperda ascovirus (SfAV) and Trichoplusia ni ascovirus (TnAV) at 7, 14, and 21 days post-infection (dpi). We found most transcription occurred in viral vesicles, not in initial tissues infected, a remarkably novel reproduction mechanism compared to all other viruses and most other intracellular pathogens. Specifically, the highest level of viral gene expression occurred in hemolymph, for TnAV at 7 dpi, and SfAV at 14 dpi. Moreover, we found that host immune genes were partially down-regulated in hemolymph, where most viral replication occurred in highly dense accumulations of vesicles.

Background: There is little experience in the economic evaluation of pharmacy/primary care collaborative health interventions using interprofessional technology-driven communication under real-world conditions. This study aimed to conduct cost-effectiveness and cost-utility analyses of a collaborative care intervention in hypertension and hyperlipidemia management between pharmacies and primary care versus usual (fragmented) care alongside a trial.Methods: An economic evaluation was conducted alongside a 6-month pragmatic quasi-experimental controlled trial. Data sources included primary care clinical software; pharmacy dispensing software; patient telephone surveys; and published literature. The target population was adult patients on hypertension and/or lipid-lowering medication. The perspective was societal. We collected patient-level data on resource use to estimate trial costs. Effect outcomes included blood pressure (BP) and quality-adjusted life years (QALYs). Bootstrapping was used to estimate uncertainty around the incremental cost-effectiveness and cost-utility ratios. Cost-effectiveness planes and acceptability curves were estimated.Results: The intervention was not shown to have reasonable levels of cost-effectiveness or cost-utility when compared to usual care as denoted by the levels of uncertainty expressed in wide confidence intervals. The probability of the intervention being cost-effective is 28% at the threshold of €20,000 per QALY gained and 57% at the threshold of €500 per mmHg systolic BP decrease.Conclusion: Considering the limitations of the trial which affected effectiveness and economic outcomes, our results are not generalizable for community pharmacy and primary care in Portugal. This research offers, however, valuable lessons on methods and strategies that can be used in future economic evaluations of collaborative public health interventions with the potential for reimbursement.Clinical trial registration:https://www.isrctn.com/ISRCTN13410498, identifier ISRCTN13410498

Summary: The extracellular RNA communication consortium (ERCC) is an NIH-funded program aiming to promote the development of new technologies, resources, and knowledge about exRNAs and their carriers. After Phase 1 (2013–2018), Phase 2 of the program (ERCC2, 2019–2023) aims to fill critical gaps in knowledge and technology to enable rigorous and reproducible methods for separation and characterization of both bulk populations of exRNA carriers and single EVs. ERCC2 investigators are also developing new bioinformatic pipelines to promote data integration through the exRNA atlas database. ERCC2 has established several Working Groups (Resource Sharing, Reagent Development, Data Analysis and Coordination, Technology Development, nomenclature, and Scientific Outreach) to promote collaboration between ERCC2 members and the broader scientific community. We expect that ERCC2’s current and future achievements will significantly improve our understanding of exRNA biology and the development of accurate and efficient exRNA-based diagnostic, prognostic, and theranostic biomarker assays.

The relationship between June sucker (Chasmistes liorus, Jordan, 1878) and Utah sucker (Catostomus ardens, Jordan & Gilbert, 1881) has been a matter of controversy since the mid 1900s. Chasmistes liorus is endemic to Utah Lake, UT and has a subterminal mouth adapted for pelagic feeding. Catostomus ardens is widely distributed throughout the Bonneville Basin and Upper Snake River Basin and has a ventral mouth adapted for benthic feeding. Chasmistes has been recognized as a separate ancient genus. Despite being morphologically distinct, no study has successfully identified residual genetic markers that separate these species. Of these studies, several have used a subset of mitochondrial genes, but no study has analyzed the complete mitochondrial genomes (mitogenomes) of these suckers (Pisces: Catostomidae). To further explore the genetic relationships between these species, we report the complete mitogenomes of Chasmistes liorus and Catostomus ardens. DNA was sequenced using an Illumina HiSeq 2500 system and mitogenomes were assembled and annotated using Geneious v. 2021.2 and MitoAnnotator, respectively. The mitogenomes of Chasmistes liorus and Catostomus ardens are both 16,623 bp and are ∼0.072% divergent. We examine the phylogenetic relationship between Chasmistes liorus and Catostomus ardens using 33 mitogenomes, representing 16 species, from Catostomidae. Our data suggest that Chasmistes liorus is sister to Catostomus ardens. Additional samples from multiple localities and/or cohorts of these species will allow us to better resolve the complicated phylogenetic relationships between these species.

A digital twin is a simulator of a physical system, which is built upon a series of models and computer programs with real-time data (from sensors or devices). Digital twins are used in various industries, such as manufacturing, healthcare, and transportation, to understand complex physical systems and make informed decisions. However, predictions and optimizations with digital twins can be time-consuming due to the high computational requirements and complexity of the underlying computer programs. This poses significant challenges in making well-informed and timely decisions using digital twins. This paper proposes a novel methodology, called the “digital triplet”, to facilitate real-time prediction and decision-making. A digital triplet is an efficient representation of a digital twin, constructed using statistical models and effective experimental designs. It offers two noteworthy advantages. Firstly, by leveraging modern statistical models, a digital triplet can effectively capture and represent the complexities of a digital twin, resulting in accurate predictions and reliable decision-making. Secondly, a digital triplet adopts a sequential design and modeling approach, allowing real-time updates in conjunction with its corresponding digital twin. We conduct comprehensive simulation studies to explore the application of various statistical models and designs in constructing a digital triplet. It is shown that Gaussian process regression coupled with sequential MaxPro designs exhibits superior performance compared to other modeling and design techniques in accurately constructing the digital triplet.

Objective: The aim of this study was to explore the expression and effects of tumor necrosis factor alpha (TNFα) in maternal endothelial cells in preeclampsia (PE). Methods: Expression levels in primary microvascular endothelial cells (MVEC) isolated from patients with severe preeclampsia (PE) and normal pregnancies were determined by RT-qPCR with or without treatment of TNFα and inhibitors for downstream signaling. Results: PE MVEC exhibited increased basal TNFα expression. TNFα treatment increased TNFα, VCAM, and endocan expression in MVEC. Conclusion: TNFα expression is increased in PE MVEC and the treatment of these cells with exogenous TNFα modifies their gene expression.

Objective Four-factor prothrombin complex concentrate (4F-PCC) was approved by the US Food and Drug Administration in 2013 for management of severely bleeding patients on warfarin therapy. We describe use of 4F-PCC at a large, suburban academic center. Methods We retrospectively reviewed all patients receiving 4F-PCC from its introduction through 2016 at a large level 1 trauma center. Clinical and demographic data were obtained, including indications for anticoagulation and antiplatelet agents, comorbidities, concomitant medications, etiology and site of bleeding, as well as disposition, length of stay, mortality, and thrombotic events. Results One hundred eighty-four patients received 4F-PCC. Mean age was 72 years; 40.8% were female. Indications for 4F-PCC administration included: active bleeding (74%), reversal prior to a procedure (14%), and elevated international normalized ratio (12%). Warfarin was the most common concomitant medication (71.1%). Most patients were receiving anticoagulation for atrial fibrillation (63%). Concomitant treatments for bleeding included vitamin K (58.2%), packed red blood cells (50%), fresh frozen plasma (38%), and platelets (26.1%), amongst others. Median length of hospital stay was 8.4 days. Nine patients (4.9%) developed thrombosis within 90 days of 4F-PCC. Mortality was 24.5%, with notably higher rates amongst those who received 4F-PCC for off-label indications (19.1% on-label mortality vs. 37.7% off-label mortality on chi-square analysis, P=0.01). Conclusion This study demonstrates that 4F-PCC is being utilized for indications other than the reversal of warfarin-induced coagulopathy. Further investigation is warranted to determine the efficacy and safety of 4F-PCC for these potential indications.

Abstract We have recently reported the discovery of a new distinct nanoparticle by ultracentrifugation of the supernatant of exomeres at 376,000 x g for 16 hours. We termed the resulting pellet, supermeres (supernatant of exomeres). The Witwer team raised the legitimate question that these two membraneless nanoparticles may arise from a continuous population of extracellular particles. We have reviewed our data here and present additional analyses that reinforce supermeres as distinct nanoparticles.

Immunoglobulin A provides a major line of defence against pathogens and plays a key role in the maintenance of the commensal microbiota in the intestinal tract. Having been shown to be more effective at tumour cell killing than IgG and strongly active against pathogens present in the mucosae, IgA antibodies have been attracting significant attention in recent years for use as therapeutic antibodies. To improve their therapeutic potential, bioengineered IgA forms with increased serum half-life and neutralizing abilities have been developed but the IgA hinge, which impacts susceptibility to bacterial proteases and ability to bridge between target and effector cells, has not yet been explored. The European rabbit has 15 IgA subclasses with exclusive hinge region motifs and varying lengths, constituting a unique model to evaluate the functional capabilities offered by incorporation of longer IgA hinges into immunoglobulins. Hinge regions from rabbit IgAs, featuring different lengths and sequences, were inserted into human IgA1 heavy chain to substitute the IgA1 hinge. These hinges did not appear to affect antigen binding nor the ability of the engineered chimeric IgA1 to bind and trigger FcαRI, as detected by IgA-mediated cell agglutination and release of superoxide by neutrophils. All rabbit hinge-human IgA1 hybrids were resistant to Clostridrum ramosum IgA protease enzyme digestion, as predicted by the lack of the cleavage site in the rabbit hinges. Some IgA1s featuring long rabbit hinges were cleaved by Neisseria meningitidis IgA1 protease cleavage type 1 or 2 enzymes, despite the lack of the predicted cleavage sites. More interestingly, the hybrid featuring the rabbit IgA15 hinge was not affected by any of the IgA proteases. The IgA15 hinge is longer than that found in human IgA1 and is composed by a unique motif with a stretch of nine consecutive Ser residues. These characteristics allow the preservation of a long hinge, with associated ability to bridge distantly spaced antigens and provide higher avidity binding, while remaining resistant to IgA protease degradation. The data suggest that the rabbit Cα15 hinge represents an interesting alternative hinge sequence for therapeutic human IgA antibodies that remains resistant to proteolytic cleavage.

Abstract The vaccine efficacy of standard-dose seasonal inactivated influenza vaccines (S-IIV) can be improved by the use of vaccines with higher antigen content or adjuvants. We conducted a randomized controlled trial in older adults to compare cellular and antibody responses of S-IIV versus enhanced vaccines (eIIV): MF59-adjuvanted (A-eIIV), high-dose (H-eIIV), and recombinant-hemagglutinin (HA) (R-eIIV). All vaccines induced comparable H3-HA-specific IgG and elevated antibody-dependent cellular cytotoxicity (ADCC) activity at day 30 post vaccination. H3-HA-specific ADCC responses were greatest following H-eIIV. Only A-eIIV increased H3-HA-IgG avidity, HA-stalk IgG and ADCC activity. eIIVs also increased polyfunctional CD4+ and CD8+ T cell responses, while cellular immune responses were skewed toward single-cytokine-producing T cells among S-IIV subjects. Our study provides further immunological evidence for the preferential use of eIIVs in older adults as each vaccine platform had an advantage over the standard-dose vaccine in terms of NK cell activation, HA-stalk antibodies, and T cell responses.

The SARS-CoV-2 pandemic highlighted the need for broad-spectrum antivirals to increase our preparedness. Patients often require treatment by the time that blocking virus replication is less effective. Therefore, therapy should not only aim to inhibit the virus, but also to suppress pathogenic host responses, e.g., leading to microvascular changes and pulmonary damage. Clinical studies have previously linked SARS-CoV-2 infection to pathogenic intussusceptive angiogenesis in the lungs, involving the upregulation of angiogenic factors such as ANGPTL4. The β-blocker propranolol is used to suppress aberrant ANGPTL4 expression in the treatment of hemangiomas. Therefore, we investigated the effect of propranolol on SARS-CoV-2 infection and the expression of ANGPTL4. SARS-CoV-2 upregulated ANGPTL4 in endothelial and other cells, which could be suppressed with R-propranolol. The compound also inhibited the replication of SARS-CoV-2 in Vero-E6 cells and reduced the viral load by up to ~2 logs in various cell lines and primary human airway epithelial cultures. R-propranolol was as effective as S-propranolol but lacks the latter’s undesired β-blocker activity. R-propranolol also inhibited SARS-CoV and MERS-CoV. It inhibited a post-entry step of the replication cycle, likely via host factors. The broad-spectrum antiviral effect and suppression of factors involved in pathogenic angiogenesis make R-propranolol an interesting molecule to further explore for the treatment of coronavirus infections.