Elife. 2023 Apr 27;12:e84757. doi: 10.7554/eLife.84757.

ABSTRACT

Morphogen gradients can instruct cells about their position in a patterned tissue. Non-linear morphogen decay has been suggested to increase gradient precision by reducing the sensitivity to variability in the morphogen source. Here, we use cell-based simulations to quantitatively compare the positional error of gradients for linear and non-linear morphogen decay. While we confirm that non-linear decay reduces the positional error close to the source, the reduction is very small for physiological noise levels. Far from the source, the positional error is much larger for non-linear decay in tissues that pose a flux barrier to the morphogen at the boundary. In light of this new data, a physiological role of morphogen decay dynamics in patterning precision appears unlikely.

PMID:37102505 | DOI:10.7554/eLife.84757

iScience. 2023 Mar 31;26(4):106541. doi: 10.1016/j.isci.2023.106541. eCollection 2023 Apr 21.

ABSTRACT

Skeletal muscle dysfunction or reprogramming due to the effects of the cancer secretome is observed in multiple malignancies. Although mouse models are routinely used to study skeletal muscle defects in cancer, because of species specificity of certain cytokines/chemokines in the secretome, a human model system is required. Here, we establish simplified multiple skeletal muscle stem cell lines (hMuSCs), which can be differentiated into myotubes. Using single nuclei ATAC-seq (snATAC-seq) and RNA-seq (snRNA-seq), we document chromatin accessibility and transcriptomic changes associated with the transition of hMuSCs to myotubes. Cancer secretome accelerated stem to myotube differentiation, altered the alternative splicing machinery and increased inflammatory, glucocorticoid receptor, and wound healing pathways in hMuSCs. Additionally, cancer secretome reduced metabolic and survival pathway associated miR-486, AKT, and p53 signaling in hMuSCs. hMuSCs underwent myotube differentiation when engrafted into NSG mice and thus providing a humanized in vivo skeletal muscle model system to study cancer cachexia.

PMID:37102148 | PMC:PMC10123345 | DOI:10.1016/j.isci.2023.106541

Biomed Res Int. 2023 Apr 17;2023:3728131. doi: 10.1155/2023/3728131. eCollection 2023.

ABSTRACT

PURPOSE: As a scientific field, bioinformatics has drawn remarkable attention from various fields, such as information technology, mathematics, and modern biological sciences, in recent years. The topic models originating from the field of natural language processing have become the focus of attention with the rapid accumulation of biological datasets. Thus, this research is aimed at modeling the topic content of the bioinformatics literature presented by Iranian researchers in the Scopus Citation Database. Methodology. This research was a descriptive-exploratory study, and the studied population included 3899 papers indexed in the Scopus database, which had been indexed in this database until March 9, 2022. The topic modeling was then performed on the abstracts and titles of the papers. A combination of LDA and TF-IDF was utilized for topic modeling. Findings. The data analysis with topic modeling resulted in identifying seven main topics "Molecular Modeling," "Gene Expression," "Biomarker," "Coronavirus," "Immunoinformatics," "Cancer Bioinformatics," and "Systems Biology." Moreover, "Systems Biology" and "Coronavirus" had the largest and smallest clusters, respectively.

CONCLUSION: The present investigation demonstrated an acceptable performance for the LDA algorithm in classifying the topics included in this field. The extracted topic clusters indicated excellent consistency and topic connection with each other.

PMID:37101687 | PMC:PMC10125747 | DOI:10.1155/2023/3728131

ACS Bio Med Chem Au. 2022 Feb 16;2(3):196-204. doi: 10.1021/acsbiomedchemau.1c00061. eCollection 2022 Jun 15.

ABSTRACT

The anaerobic biosyntheses of heme, heme d 1, and bacteriochlorophyll all require the action of radical SAM enzymes. During heme biosynthesis in some bacteria, coproporphyrinogen III dehydrogenase (CgdH) catalyzes the decarboxylation of two propionate side chains of coproporphyrinogen III to the corresponding vinyl groups of protoporphyrinogen IX. Its solved crystal structure was the first published structure for a radical SAM enzyme. In bacteria, heme is inserted into enzymes by the cytoplasmic heme chaperone HemW, a radical SAM enzyme structurally highly related to CgdH. In an alternative heme biosynthesis route found in archaea and sulfate-reducing bacteria, the two radical SAM enzymes AhbC and AhbD catalyze the removal of two acetate groups (AhbC) or the decarboxylation of two propionate side chains (AhbD). NirJ, a close homologue of AhbC, is required for propionate side chain removal during the formation of heme d 1 in some denitrifying bacteria. Biosynthesis of the fifth ring (ring E) of all chlorophylls is based on an unusual six-electron oxidative cyclization step. The sophisticated conversion of Mg-protoporphyrin IX monomethylester to protochlorophyllide is facilitated by an oxygen-independent cyclase termed BchE, which is a cobalamin-dependent radical SAM enzyme. Most of the radical SAM enzymes involved in tetrapyrrole biosynthesis were recognized as such by Sofia et al. in 2001 (Nucleic Acids Res.2001, 29, 1097-1106) and were biochemically characterized thereafter. Although much has been achieved, the challenging tetrapyrrole substrates represent a limiting factor for enzyme/substrate cocrystallization and the ultimate elucidation of the corresponding enzyme mechanisms.

PMID:37101575 | PMC:PMC10114771 | DOI:10.1021/acsbiomedchemau.1c00061

Aging Cell. 2023 Apr 26:e13852. doi: 10.1111/acel.13852. Online ahead of print.

ABSTRACT

Perturbed metabolism of ammonia, an endogenous cytotoxin, causes mitochondrial dysfunction, reduced NAD+ /NADH (redox) ratio, and postmitotic senescence. Sirtuins are NAD+ -dependent deacetylases that delay senescence. In multiomics analyses, NAD metabolism and sirtuin pathways are enriched during hyperammonemia. Consistently, NAD+ -dependent Sirtuin3 (Sirt3) expression and deacetylase activity were decreased, and protein acetylation was increased in human and murine skeletal muscle/myotubes. Global acetylomics and subcellular fractions from myotubes showed hyperammonemia-induced hyperacetylation of cellular signaling and mitochondrial proteins. We dissected the mechanisms and consequences of hyperammonemia-induced NAD metabolism by complementary genetic and chemical approaches. Hyperammonemia inhibited electron transport chain components, specifically complex I that oxidizes NADH to NAD+ , that resulted in lower redox ratio. Ammonia also caused mitochondrial oxidative dysfunction, lower mitochondrial NAD+ -sensor Sirt3, protein hyperacetylation, and postmitotic senescence. Mitochondrial-targeted Lactobacillus brevis NADH oxidase (MitoLbNOX), but not NAD+ precursor nicotinamide riboside, reversed ammonia-induced oxidative dysfunction, electron transport chain supercomplex disassembly, lower ATP and NAD+ content, protein hyperacetylation, Sirt3 dysfunction and postmitotic senescence in myotubes. Even though Sirt3 overexpression reversed ammonia-induced hyperacetylation, lower redox status or mitochondrial oxidative dysfunction were not reversed. These data show that acetylation is a consequence of, but is not the mechanism of, lower redox status or oxidative dysfunction during hyperammonemia. Targeting NADH oxidation is a potential approach to reverse and potentially prevent ammonia-induced postmitotic senescence in skeletal muscle. Since dysregulated ammonia metabolism occurs with aging, and NAD+ biosynthesis is reduced in sarcopenia, our studies provide a biochemical basis for cellular senescence and have relevance in multiple tissues.

PMID:37101412 | DOI:10.1111/acel.13852

Mol Cancer. 2023 Apr 26;22(1):76. doi: 10.1186/s12943-023-01777-z.

NO ABSTRACT

PMID:37101282 | DOI:10.1186/s12943-023-01777-z

Orphanet J Rare Dis. 2023 Apr 26;18(1):95. doi: 10.1186/s13023-023-02683-9.

ABSTRACT

BACKGROUND: Inherited Metabolic Disorders (IMDs) are rare diseases where one impaired protein leads to a cascade of changes in the adjacent chemical conversions. IMDs often present with non-specific symptoms, a lack of a clear genotype-phenotype correlation, and de novo mutations, complicating diagnosis. Furthermore, products of one metabolic conversion can be the substrate of another pathway obscuring biomarker identification and causing overlapping biomarkers for different disorders. Visualization of the connections between metabolic biomarkers and the enzymes involved might aid in the diagnostic process. The goal of this study was to provide a proof-of-concept framework for integrating knowledge of metabolic interactions with real-life patient data before scaling up this approach. This framework was tested on two groups of well-studied and related metabolic pathways (the urea cycle and pyrimidine de-novo synthesis). The lessons learned from our approach will help to scale up the framework and support the diagnosis of other less-understood IMDs.

METHODS: Our framework integrates literature and expert knowledge into machine-readable pathway models, including relevant urine biomarkers and their interactions. The clinical data of 16 previously diagnosed patients with various pyrimidine and urea cycle disorders were visualized on the top 3 relevant pathways. Two expert laboratory scientists evaluated the resulting visualizations to derive a diagnosis.

RESULTS: The proof-of-concept platform resulted in varying numbers of relevant biomarkers (five to 48), pathways, and pathway interactions for each patient. The two experts reached the same conclusions for all samples with our proposed framework as with the current metabolic diagnostic pipeline. For nine patient samples, the diagnosis was made without knowledge about clinical symptoms or sex. For the remaining seven cases, four interpretations pointed in the direction of a subset of disorders, while three cases were found to be undiagnosable with the available data. Diagnosing these patients would require additional testing besides biochemical analysis.

CONCLUSION: The presented framework shows how metabolic interaction knowledge can be integrated with clinical data in one visualization, which can be relevant for future analysis of difficult patient cases and untargeted metabolomics data. Several challenges were identified during the development of this framework, which should be resolved before this approach can be scaled up and implemented to support the diagnosis of other (less understood) IMDs. The framework could be extended with other OMICS data (e.g. genomics, transcriptomics), and phenotypic data, as well as linked to other knowledge captured as Linked Open Data.

PMID:37101200 | DOI:10.1186/s13023-023-02683-9

Cell Death Differ. 2023 Apr 26. doi: 10.1038/s41418-023-01153-w. Online ahead of print.

ABSTRACT

Apoptosis is a form of regulated cell death (RCD) that involves proteases of the caspase family. Pharmacological and genetic strategies that experimentally inhibit or delay apoptosis in mammalian systems have elucidated the key contribution of this process not only to (post-)embryonic development and adult tissue homeostasis, but also to the etiology of multiple human disorders. Consistent with this notion, while defects in the molecular machinery for apoptotic cell death impair organismal development and promote oncogenesis, the unwarranted activation of apoptosis promotes cell loss and tissue damage in the context of various neurological, cardiovascular, renal, hepatic, infectious, neoplastic and inflammatory conditions. Here, the Nomenclature Committee on Cell Death (NCCD) gathered to critically summarize an abundant pre-clinical literature mechanistically linking the core apoptotic apparatus to organismal homeostasis in the context of disease.

PMID:37100955 | DOI:10.1038/s41418-023-01153-w

Nature. 2023 Apr 26. doi: 10.1038/s41586-023-05990-0. Online ahead of print.

ABSTRACT

The approximately 120 MDa mammalian nuclear pore complex (NPC) acts as a gatekeeper for the transport between the nucleus and cytosol1. The central channel of the NPC is filled with hundreds of intrinsically disordered proteins (IDPs) called FG-nucleoporins (FG-NUPs)2,3. Although the structure of the NPC scaffold has been resolved in remarkable detail, the actual transport machinery built up by FG-NUPs-about 50 MDa-is depicted as an approximately 60-nm hole in even highly resolved tomograms and/or structures computed with artificial intelligence4-11. Here we directly probed conformations of the vital FG-NUP98 inside NPCs in live cells and in permeabilized cells with an intact transport machinery by using a synthetic biology-enabled site-specific small-molecule labelling approach paired with highly time-resolved fluorescence microscopy. Single permeabilized cell measurements of the distance distribution of FG-NUP98 segments combined with coarse-grained molecular simulations of the NPC allowed us to map the uncharted molecular environment inside the nanosized transport channel. We determined that the channel provides-in the terminology of the Flory polymer theory12-a 'good solvent' environment. This enables the FG domain to adopt expanded conformations and thus control transport between the nucleus and cytoplasm. With more than 30% of the proteome being formed from IDPs, our study opens a window into resolving disorder-function relationships of IDPs in situ, which are important in various processes, such as cellular signalling, phase separation, ageing and viral entry.

PMID:37100914 | DOI:10.1038/s41586-023-05990-0

Commun Chem. 2023 Apr 26;6(1):80. doi: 10.1038/s42004-023-00879-5.

ABSTRACT

By facilitating a water/water phase separation (w/wPS), crowded biopolymers in cells form droplets that contribute to the spatial localization of biological components and their biochemical reactions. However, their influence on mechanical processes driven by protein motors has not been well studied. Here, we show that the w/wPS droplet spontaneously entraps kinesins as well as microtubules (MTs) and generates a micrometre-scale vortex flow inside the droplet. Active droplets with a size of 10-100 µm are generated through w/wPS of dextran and polyethylene glycol mixed with MTs, molecular-engineered chimeric four-headed kinesins and ATP after mechanical mixing. MTs and kinesin rapidly created contractile network accumulated at the interface of the droplet and gradually generated vortical flow, which can drive translational motion of a droplet. Our work reveals that the interface of w/wPS contributes not only to chemical processes but also produces mechanical motion by assembling species of protein motors in a functioning manner.

PMID:37100870 | DOI:10.1038/s42004-023-00879-5

Sci Rep. 2023 Apr 26;13(1):6835. doi: 10.1038/s41598-023-33862-0.

ABSTRACT

In gene therapy, potential integration of therapeutic transgene into host cell genomes is a serious risk that can lead to insertional mutagenesis and tumorigenesis. Viral vectors are often used as the gene delivery vehicle, but they are prone to undergoing integration events. More recently, non-viral delivery of linear DNAs having modified geometry such as closed-end linear duplex DNA (CELiD) have shown promise as an alternative, due to prolonged transgene expression and less cytotoxicity. However, whether modified-end linear DNAs can also provide a safe, non-integrating gene transfer remains unanswered. Herein, we compare the genomic integration frequency upon transfection of cells with expression vectors in the forms of circular plasmid, unmodified linear DNA, CELiDs with thioester loops, and Streptavidin-conjugated blocked-end linear DNA. All of the forms of linear DNA resulted in a high fraction of the cells being stably transfected-between 10 and 20% of the initially transfected cells. These results indicate that blocking the ends of linear DNA is insufficient to prevent integration.

PMID:37100816 | DOI:10.1038/s41598-023-33862-0

Curr Top Dev Biol. 2023;154:99-129. doi: 10.1016/bs.ctdb.2023.02.005. Epub 2023 Mar 14.

ABSTRACT

Actin networks are central to shaping and moving cells during animal development. Various spatial cues activate conserved signal transduction pathways to polarize actin network assembly at sub-cellular locations and to elicit specific physical changes. Actomyosin networks contract and Arp2/3 networks expand, and to affect whole cells and tissues they do so within higher-order systems. At the scale of tissues, actomyosin networks of epithelial cells can be coupled via adherens junctions to form supracellular networks. Arp2/3 networks typically integrate with distinct actin assemblies, forming expansive composites which act in conjunction with contractile actomyosin networks for whole-cell effects. This review explores these concepts using examples from Drosophila development. First, we discuss the polarized assembly of supracellular actomyosin cables which constrict and reshape epithelial tissues during embryonic wound healing, germ band extension, and mesoderm invagination, but which also form physical borders between tissue compartments at parasegment boundaries and during dorsal closure. Second, we review how locally induced Arp2/3 networks act in opposition to actomyosin structures during myoblast cell-cell fusion and cortical compartmentalization of the syncytial embryo, and how Arp2/3 and actomyosin networks also cooperate for the single cell migration of hemocytes and the collective migration of border cells. Overall, these examples show how the polarized deployment and higher-order interactions of actin networks organize developmental cell biology.

PMID:37100525 | DOI:10.1016/bs.ctdb.2023.02.005

Life Sci Alliance. 2023 Apr 26;6(7):e202201876. doi: 10.26508/lsa.202201876. Print 2023 Jul.

ABSTRACT

Although ubiquitin is found only in eukaryotes, several pathogenic bacteria and viruses possess proteins that hinder the host ubiquitin system. Legionella, a gram-negative intracellular bacterium, possesses an ovarian tumor (OTU) family of deubiquitinases (Lot DUBs). Herein, we describe the molecular characteristics of Lot DUBs. We elucidated the structure of the LotA OTU1 domain and revealed that entire Lot DUBs possess a characteristic extended helical lobe that is not found in other OTU-DUBs. The structural topology of an extended helical lobe is the same throughout the Lot family, and it provides an S1' ubiquitin-binding site. Moreover, the catalytic triads of Lot DUBs resemble those of the A20-type OTU-DUBs. Furthermore, we revealed a unique mechanism by which LotA OTU domains cooperate together to distinguish the length of the chain and preferentially cleave longer K48-linked polyubiquitin chains. The LotA OTU1 domain itself cleaves K6-linked ubiquitin chains, whereas it is also essential for assisting the cleavage of longer K48-linked polyubiquitin chains by the OTU2 domain. Thus, this study provides novel insights into the structure and mechanism of action of Lot DUBs.

PMID:37100438 | DOI:10.26508/lsa.202201876

Brain Behav Immun. 2023 Apr 24:S0889-1591(23)00106-X. doi: 10.1016/j.bbi.2023.04.008. Online ahead of print.

ABSTRACT

One-sentence summary. We find that IFN-γ induces transcription of MHC class I antigen processing and presentation machinery in all major parenchymal cell types in the ventral midbrain in vivo; however, neuronal responses are low amplitude and limited to a small set of genes, MHC class II expression and cellular proliferation are restricted to microglia, and dopamine neuronal responses require cell autonomous expression of IFNGR1. Dysregulated inflammation within the central nervous system (CNS) contributes to neuropathology in infectious, autoimmune, and neurodegenerative disease. With the exception of microglia, however, major histocompatibility complex (MHC) proteins are virtually undetectable in the mature, healthy central nervous system (CNS). Neurons in particular have generally been considered incapable of antigen presentation, and although interferon gamma (IFN-γ) can elicit neuronal MHC class I (MHC-I) expression and antigen presentation in vitro, it has been unclear whether similar responses occur in vivo or if such responses might be cell-autonomous or dependent on peripheral inflammation. Here we directly inject IFN-γ into the ventral midbrain of mature mice and analyze gene expression profiles of specific CNS cells. We found that IFN-γ upregulated MHC-I and associated mRNAs in ventral midbrain microglia, astrocytes, oligodendrocytes, and GABAergic, glutamatergic, and dopaminergic neurons. The core set of IFN-γ-induced genes and their response kinetics were present in both neurons and glia, with a lower amplitude of expression in neurons. A diverse repertoire of genes was upregulated in glia, particularly microglia, which were the only cells to undergo cellular proliferation and express MHC-II and associated genes. To determine if neurons respond to in a cell-autonomous manner, we produced mutant mice with a deletion of the IFN-γ-binding domain of IFNGR1 only in dopaminergic neurons, which resulted in a complete loss of dopaminergic neuronal response to IFN-γ. Our results demonstrate that neurons can respond to IFN-γ in a cell-autonomous manner by upregulating MHC-I and its related genes in vivo, although the expression level is low compared to oligodendrocytes, astrocytes, and microglia.

PMID:37100211 | DOI:10.1016/j.bbi.2023.04.008

ACS Macro Lett. 2023 Apr 26:632-638. doi: 10.1021/acsmacrolett.3c00101. Online ahead of print.

ABSTRACT

Amphiphilic molecules that can crystallize often form molecularly thin nanosheets in aqueous solutions. The possibility of atomic-scale corrugations in these structures has not yet been recognized. We have studied the self-assembly of amphiphilic polypeptoids, a family of bio-inspired polymers that can self-assemble into various crystalline nanostructures. Atomic-scale structure of the crystals in these systems has been inferred using both X-ray diffraction and electron microscopy. Here we use cryogenic electron microscopy to determine the in-plane and out-of-plane structures of a crystalline nanosheet. Data were collected as a function of tilt angle and analyzed using a hybrid single-particle crystallographic approach. The analysis reveals that adjacent rows of peptoid chains, which are separated by 4.5 Å in the plane of the nanosheet, are offset by 6 Å in the direction perpendicular to the plane of the nanosheet. These atomic-scale corrugations lead to a doubling of the unit cell dimension from 4.5 to 9 Å. Our work provides an alternative interpretation for the observed Å X-ray diffraction peak often reported in polypeptoid crystals.

PMID:37099693 | DOI:10.1021/acsmacrolett.3c00101

Plant Physiol. 2023 Apr 26:kiad252. doi: 10.1093/plphys/kiad252. Online ahead of print.

NO ABSTRACT

PMID:37099487 | DOI:10.1093/plphys/kiad252

J Proteome Res. 2023 Apr 26. doi: 10.1021/acs.jproteome.3c00069. Online ahead of print.

ABSTRACT

Viruses can elicit varying types and severities of symptoms during plant host infection. We investigated changes in the proteome and transcriptome of Nicotiana benthamiana plants infected by grapevine fanleaf virus (GFLV) with an emphasis on vein clearing symptom development. Comparative, time-course liquid chromatography tandem mass spectrometry and 3' ribonucleic acid sequencing analyses of plants infected by two wildtype GFLV strains, one symptomatic and one asymptomatic, and their asymptomatic mutant strains carrying a single amino acid change in the RNA-dependent RNA polymerase (RdRP) were conducted to identify host biochemical pathways involved in viral symptom development. During peak vein clearing symptom display at 7 days post-inoculation (dpi), protein and gene ontologies related to immune response, gene regulation, and secondary metabolite production were overrepresented when contrasting wildtype GFLV strain GHu and mutant GHu-1EK802GPol. Prior to the onset of symptom development at 4 dpi and when symptoms faded away at 12 dpi, protein and gene ontologies related to chitinase activity, hypersensitive response, and transcriptional regulation were identified. This systems biology approach highlighted how a single amino acid of a plant viral RdRP mediates changes to the host proteome (∼1%) and transcriptome (∼8.5%) related to transient vein clearing symptoms and the network of pathways involved in the virus-host arms race.

PMID:37099450 | DOI:10.1021/acs.jproteome.3c00069

Nucleic Acids Res. 2023 Apr 26:gkad296. doi: 10.1093/nar/gkad296. Online ahead of print.

ABSTRACT

e-RNA is a collection of web-servers for the prediction and visualisation of RNA secondary structures and their functional features, including in particular RNA-RNA interactions. In this updated version, we have added novel tools for RNA secondary structure prediction and have significantly updated the visualisation functionality. The new method CoBold can identify transient RNA structure features and their potential functional effects on a known RNA structure during co-transcriptional structure formation. New tool ShapeSorter can predict evolutionarily conserved RNA secondary structure features while simultaneously taking experimental SHAPE probing evidence into account. The web-server R-Chie which visualises RNA secondary structure information in terms of arc diagrams, can now be used to also visualise and intuitively compare RNA-RNA, RNA-DNA and DNA-DNA interactions alongside multiple sequence alignments and quantitative information. The prediction generated by any method in e-RNA can be readily visualised on the web-server. For completed tasks, users can download their results and readily visualise them later on with R-Chie without having to re-run the predictions. e-RNA can be found at http://www.e-rna.org.

PMID:37099369 | DOI:10.1093/nar/gkad296

JAMA Netw Open. 2023 Apr 3;6(4):e2310068. doi: 10.1001/jamanetworkopen.2023.10068.

ABSTRACT

IMPORTANCE: Calciphylaxis is a rare disease with high mortality mainly involving patients with chronic kidney disease (CKD). Sodium thiosulphate (STS) has been used as an off-label therapeutic in calciphylaxis, but there is a lack of clinical trials and studies that demonstrate its effect compared with those without STS treatment.

OBJECTIVE: To perform a meta-analysis of the cohort studies that provided data comparing outcomes among patients with calciphylaxis treated with and without intravenous STS.

DATA SOURCES: PubMed, Embase, Cochrane Library, Web of Science, and ClinicalTrials.gov were searched using relevant terms and synonyms including sodium thiosulphate and calci* without language restriction.

STUDY SELECTION: The initial search was for cohort studies published before August 31, 2021, that included adult patients diagnosed with CKD experiencing calciphylaxis and could provide a comparison between patients treated with and without intravenous STS. Studies were excluded if they reported outcomes only from nonintravenous administration of STS or if the outcomes for CKD patients were not provided.

DATA EXTRACTION AND SYNTHESIS: Random-effects models were performed. The Egger test was used to measure publication bias. Heterogeneity was assessed using the I2 test.

MAIN OUTCOMES AND MEASURES: Skin lesion improvement and survival, synthesized as ratio data by a random-effects empirical Bayes model.

RESULTS: Among the 5601 publications retrieved from the targeted databases, 19 retrospective cohort studies including 422 patients (mean age, 57 years; 37.3% male) met the eligibility criteria. No difference was observed in skin lesion improvement (12 studies with 110 patients; risk ratio, 1.23; 95% CI, 0.85-1.78) between the STS and the comparator groups. No difference was noted for the risk of death (15 studies with 158 patients; risk ratio, 0.88; 95% CI, 0.70-1.10) and overall survival using time-to-event data (3 studies with 269 participants; hazard ratio, 0.82; 95% CI, 0.57-1.18). In meta-regression, lesion improvement associated with STS negatively correlated with publication year, implying that recent studies are more likely to report a null association compared with past studies (coefficient = -0.14; P = .008).

CONCLUSIONS AND RELEVANCE: Intravenous STS was not associated with skin lesion improvement or survival benefit in patients with CKD experiencing calciphylaxis. Future investigations are warranted to examine the efficacy and safety of therapies for patients with calciphylaxis.

PMID:37099293 | DOI:10.1001/jamanetworkopen.2023.10068

Microbiol Spectr. 2023 Apr 26:e0020623. doi: 10.1128/spectrum.00206-23. Online ahead of print.

ABSTRACT

Carryover contamination during amplicon sequencing workflow (AMP-Seq) put the accuracy of the high-throughput detection for pathogens at risk. The purpose of this study is to develop a carryover contaminations-controlled AMP-Seq (ccAMP-Seq) workflow to enable accurate qualitative and quantitative detection for pathogens. By using the AMP-Seq workflow to detect SARS-CoV-2, Aerosols, reagents and pipettes were identified as potential sources of contaminations and ccAMP-Seq was then developed. ccAMP-Seq used filter tips and physically isolation of experimental steps to avoid cross contamination, synthetic DNA spike-ins to compete with contaminations and quantify SARS-CoV-2, dUTP/uracil DNA glycosylase system to digest the carryover contaminations, and a new data analysis procedure to remove the sequencing reads from contaminations. Compared to AMP-Seq, the contamination level of ccAMP-Seq was at least 22-folds lower and the detection limit was also about an order of magnitude lower-as low as one copy/reaction. By testing the dilution series of SARS-CoV-2 nucleic acid standard, ccAMP-Seq showed 100% sensitivity and specificity. The high sensitivity of ccAMP-Seq was further confirmed by the detection of SARS-CoV-2 from 62 clinical samples. The consistency between qPCR and ccAMP-Seq was 100% for all the 53 qPCR-positive clinical samples. Seven qPCR-negative clinical samples were found to be positive by ccAMP-Seq, which was confirmed by extra qPCR tests on subsequent samples from the same patients. This study presents a carryover contamination-controlled, accurate qualitative and quantitative amplicon sequencing workflow that addresses the critical problem of pathogen detection for infectious diseases. IMPORTANCE Accuracy, a key indicator of pathogen detection technology, is compromised by carryover contamination in the amplicon sequencing workflow. Taking the detection of SARS-CoV-2 as case, this study presents a new carryover contamination-controlled amplicon sequencing workflow. The new workflow significantly reduces the degree of contamination in the workflow, thereby significantly improving the accuracy and sensitivity of the SARS-CoV-2 detection and empowering the ability of quantitative detection. More importantly, the use of the new workflow is simple and economical. Therefore, the results of this study can be easily applied to other microorganism, which has great significance for improving the detection level of microorganism.

PMID:37098913 | DOI:10.1128/spectrum.00206-23