Integrated proteogenomic analysis for inherited bone marrow failure syndrome.

Recent advances in in-depth data-independent acquisition proteomic analysis have enabled comprehensive quantitative analysis of >10,000 proteins. Herein, an integrated proteogenomic analysis for inherited bone marrow failure syndrome (IBMFS) was performed to reveal their biological features and to develop a proteomic-based diagnostic assay in the discovery cohort; dyskeratosis congenita (n = 12), Fanconi anemia (n = 11), Diamond-Blackfan anemia (DBA, n = 9), Shwachman-Diamond syndrome (SDS, n = 6), ADH5/ALDH2 deficiency (n = 4), and other IBMFS (n = 18). Unsupervised proteomic clustering identified eight independent clusters (C1-C8), with the ribosomal pathway specifically downregulated in C1 and C2, enriched for DBA and SDS, respectively. Six patients with SDS had significantly decreased SBDS protein expression, with two of these not diagnosed by DNA sequencing alone. Four patients with ADH5/ALDH2 deficiency showed significantly reduced ADH5 protein expression. To perform a large-scale rapid IBMFS screening, targeted proteomic analysis was performed on 417 samples from patients with IBMFS-related hematological disorders (n = 390) and healthy controls (n = 27). SBDS and ADH5 protein expressions were significantly reduced in SDS and ADH5/ALDH2 deficiency, respectively. The clinical application of this first integrated proteogenomic analysis would be useful for the diagnosis and screening of IBMFS, where appropriate clinical screening tests are lacking.

Effects of Anti-Fibrotic Drugs on Transcriptome of Peripheral Blood Mononuclear Cells in Idiopathic Pulmonary Fibrosis.

Two anti-fibrotic drugs, pirfenidone (PFD) and nintedanib (NTD), are currently used to treat idiopathic pulmonary fibrosis (IPF). Peripheral blood mononuclear cells (PBMCs) are immunocompetent cells that could orchestrate cell-cell interactions associated with IPF pathogenesis. We employed RNA sequencing to examine the transcriptome signature in the bulk PBMCs of patients with IPF and the effects of anti-fibrotic drugs on these signatures. Differentially expressed genes (DEGs) between "patients with IPF and healthy controls" and "before and after anti-fibrotic treatment" were analyzed. Enrichment analysis suggested that fatty acid elongation interferes with TGF-ß/Smad signaling and the production of oxidative stress since treatment with NTD upregulates the fatty acid elongation enzymes ELOVL6. Treatment with PFD downregulates COL1A1, which produces wound-healing collagens because activated monocyte-derived macrophages participate in the production of collagen, type I, and alpha 1 during tissue damage. Plasminogen activator inhibitor-1 (PAI-1) regulates wound healing by inhibiting plasmin-mediated matrix metalloproteinase activation, and the inhibition of PAI-1 activity attenuates lung fibrosis. DEG analysis suggested that both the PFD and NTD upregulate SERPINE1, which regulates PAI-1 activity. This study embraces a novel approach by using RNA sequencing to examine PBMCs in IPF, potentially revealing systemic biomarkers or pathways that could be targeted for therapy.

Strong evolutionary constraints against amino acid changes in the P2 subdomain of sapovirus GI.1 capsid protein VP1.

Sapovirus (SaV) is a nonenveloped RNA virus that causes acute gastroenteritis in humans. Although SaV is a clinically important pathogen in children, an effective vaccine is currently unavailable. The capsid protein VP1 of SaVs forms the outer shell of the virion and is highly diverse, as often seen in the virion-surface proteins of RNA viruses, creating an obstacle for vaccine development. We here report a unique phenomenon pertaining to the variation of SaV VP1. Phylogenetic and information entropy analyses using full-length VP1 sequences from a public database consistently showed that the amino acid sequences of the VP1 protein have been highly conserved over more than 40 years in the major epidemic genotype GI.1 but not in GI.2. Structural modeling showed that even the VP1 P2 subdomain, which is arranged on the outermost shell of the virion and presumably exposed to anti-SaV antibodies, remained highly homogeneous in GI.1 but not in GI.2. These results suggest strong evolutionary constraints against amino acid changes in the P2 subdomain of the SaV GI.1 capsid and illustrate a hitherto unappreciated mechanism, i.e., preservation of the VP1 P2 subdomain, involved in SaV survival. Our findings could have important implications for the development of an anti-SaV vaccine.

Blood pressure outcomes at 18 months in primary care patients prescribed remote physiological monitoring for hypertension: a prospective cohort study.

This pragmatic matched cohort study using EHR data extended the follow up to 18 months for BP outcomes comparing individuals prescribed remote patient monitoring (n = 288) and temporally-matched controls (n = 1152) from six primary care practices. After 18 months, the RPM-prescribed cohort had greater BP control < 140/90 mm Hg (RPM cohort: 71.5%, control cohort: 51.9%, p < 0.001) and lower systolic BP (131.6 versus 136.0 mm Hg, p = 0.004) using office and home measurements. BP control at 18 months assessed by office measurements only was also higher in the RPM group (62.2% versus 51.9%, p = 0.004).

Genetic regions affecting the replication and pathogenicity of dengue virus type 2.

Dengue is a mosquito-borne disease that has spread to over 100 countries. Its symptoms vary from the relatively mild acute febrile illness called dengue fever to the much more severe dengue shock syndrome. Dengue is caused by dengue virus (DENV), which belongs to the Flavivirus genus of the family Flaviviridae. There are four serotypes of DENV, i.e., DENV1 to DENV4, and each serotype is divided into distinct genotypes. Thailand is an endemic area where all four serotypes of DENV co-circulate. Genome sequencing of the DENV2 that was isolated in Thailand in 2016 and 2017 revealed the emergence of the Cosmopolitan genotype and its co-circulation with the Asian-I genotype. However, it was unclear whether different genotypes have different levels of viral replication and pathogenicity. Focus-forming assay (FFA) results showed that clinical isolates of these genotypes differed in focus size and proliferative capacity. Using circular polymerase extension reaction, we generated parental and chimeric viruses with swapped genes between these two DENV2 genotypes, and compared their focus sizes and infectivity titers using FFA. The results showed that the focus size was larger when the structural proteins and/or non-structural NS1-NS2B proteins were derived from the Cosmopolitan virus. The infectious titers were consistent with the focus sizes. Single-round infectious particle assay results confirmed that chimeric viruses with Cosmopolitan type structural proteins, particularly prM/E, had significantly increased luciferase activity. Replicon assay results showed that Cosmopolitan NS1-NS2B proteins had increased reporter gene expression levels. Furthermore, in interferon-receptor knock-out mice, viruses with Cosmopolitan structural and NS1-NS2B proteins had higher titers in the blood, and caused critical disease courses. These results suggested that differences in the sequences within the structural and NS1-NS2B proteins may be responsible for the differences in replication, pathogenicity, and infectivity between the Asian-I and Cosmopolitan viruses.

Development of Small-Molecule Anti-HIV-1 Agents Targeting HIV-1 Capsid Proteins.

The capsid of human immunodeficiency virus type 1 (HIV-1) forms a conical structure by assembling oligomers of capsid (CA) proteins and is a virion shell that encapsulates viral RNA. The inhibition of the CA function could be an appropriate target for suppression of HIV-1 replication because the CA proteins are highly conserved among many strains of HIV-1, and the drug targeting CA, lenacapavir, has been clinically developed by Gilead Sciences, Inc. Interface hydrophobic interactions between two CA molecules via the Trp184 and Met185 residues in the CA sequence are indispensable for conformational stabilization of the CA multimer. Our continuous studies found two types of small molecules with different scaffolds, MKN-1 and MKN-3, designed by in silico screening as a dipeptide mimic of Trp184 and Met185 have significant anti-HIV-1 activity. In the present study, MKN-1 derivatives have been designed and synthesized. Their structure-activity relationship studies found some compounds having potent anti-HIV activity. The present results should be useful in the design of novel CA-targeting molecules with anti-HIV activity.

Case Report: A case of TAFRO syndrome with severe and prolonged thrombocytopenia: diagnostic pitfalls.

Thrombocytopenia, anasarca, myelofibrosis, renal dysfunction, and organomegaly (TAFRO) syndrome is a rare condition with diverse clinical and pathological characteristics related to multi-organ damage. We report a case of TAFRO syndrome complicated by immune thrombocytopenia with prolonged fever and thrombocytopenia for several weeks. A 61-year-old man was transferred with sepsis caused by Enterococcus faecalis, and developed disseminated intravascular coagulation. Antibiotics treatment was initiated: however, low-grade fever and thrombocytopenia persisted despite the adequate antimicrobial treatment. Systemic edema, pleural effusion, and ascites had developed before hospitalization, and renal and liver function had deteriorated, resulting in progressive multi-organ damage. Prednisolone 40 mg/day was initiated based on the assumption of a condition in which excessive production of inflammatory cytokines would lead to systemic deterioration and fatal organ damage. Subsequently, the fever resolved, and renal function began to normalize. However, thrombocytopenia did not show much recovery trend after Helicobacter pylori eradication therapy and initiation of thrombopoietin receptor agonists. Bone marrow biopsy results showed normal bone marrow with no malignant findings. Alternatively, significant clinical signs met the diagnostic criteria for TAFRO syndrome, and a renal biopsy revealed thrombotic microangiopathy, which is also reasonable for renal involvement in TAFRO syndrome. The use of cyclosporine remarkably corrected the thrombocytopenia. We considered this a case of TAFRO syndrome that developed after sepsis with disseminated intravascular coagulation and performed the differential diagnosis of prolonged thrombocytopenia and excluded it. Although TAFRO syndrome is a unique disease concept, diagnostic criteria may consist of nonspecific elements such as generalized edema, thrombocytopenia, persistent fever, and elevated inflammatory response, and there are many differential conditions to exclude, requiring caution in diagnosing TAFRO syndrome.

Digital hypertension management: clinical and cost outcomes of a pilot implementation of the OMRON hypertension management platform.

Importance: Home monitoring of blood pressure (BP) in hypertensive patients can improve outcomes, but challenges to both patient compliance and the effective transmission of home BP readings to physicians can limit the extent to which physicians can use this information to improve care. The OMRON Hypertension Management Platform (OMRON HMP) pairs a home BP cuff with a digital product that tracks data, provides reminders to improve patient compliance, and provides a streamlined source of information to physicians. Objective: The primary objective of the quality improvement (QI) project was to test the hypothesis that use of the OMRON HMP could reduce the number and cost of hypertension related claims, relative to a retrospectively matched cohort of insured members. A secondary objective was to demonstrate improvement in control of BP among patients. Design: Eligible members were recruited to the QI project between December 1, 2018 and December 30, 2020 and data collected for six months following recruitment. All members received the OMRON HMP intervention. Setting: Enrollment and data collection were coordinated on-site at selected PCP partner providers in Western Pennsylvania. Eligible members were identified from insurance claims data as those receiving care for primary hypertension from participating primary care physicians and/or cardiologists. Participants: Eligible members were between the ages of 35 and 85, with a diagnosis of primary hypertension. The retrospective cohort was selected from electronic medical records of Highmark-insured patients with hypertension who received care at Allegheny Health Network (AHN), a subsidiary of Highmark Health. Members were matched on baseline BP and lipid measures, age, smoking status, diabetes status, race and sex. Intervention: Daily home BP readings were recorded by the OMRON HMP app. Patient data was reviewed by clinical staff on a weekly basis and treatment plans could be adjusted in response to this data. Results: OMRON HMP users showed a significant increase in the number and cost of hypertension-related claims, contrary to the hypothesis, but did display improvements in control of BP. Conclusions and Relevance: The use of a digital platform to facilitate at-home BP monitoring appeared to improve BP control but led to increased hypertension-related costs in the short-term.

Blood pressure outcomes at 12 months in primary care patients prescribed remote physiological monitoring for hypertension: a prospective cohort study.

Remote patient monitoring (RPM) for hypertension enables automatic transmission of blood pressure (BP) and pulse into the electronic health record (EHR), but its effectiveness in primary care is unknown. This pragmatic matched cohort study using EHR data compared BP outcomes between individuals prescribed RPM and temporally-matched controls from six primary care practices. We retrospectively created a cohort of 288 Medicare-enrolled patients prescribed BP RPM (cases) and 1152 propensity score-matched controls (1:4). Matching was based on age, sex, systolic blood pressure (SBP), marital status, and other characteristics. Outcomes at 3, 6, 9 and 12 months were: controlling high BP (most recent BP < 140/90 mm Hg), antihypertensive medication intensification, and most recent SBP assessed using: all measurements, and office measurements only. At baseline, RPM-prescribed patients and controls had similar ages and systolic BP. BP control diverged at 3 months (RPM: 72.2%, control: 51%, p < 0.001). This difference persisted but decreased over follow-up. After 12 months, the RPM-prescribed cohort had greater BP control (RPM: 71.5%, control: 58.1%, p < 0.001) and lower SBP (132.3 versus 136.5 mm Hg, p = 0.003) using all measurements, but they did not differ using only office measurements (12 month BP control: 60.8% versus 58.1%, p = 0.44; SBP: 135.9 versus 136.5 mm Hg, p = 0.91). At 12 months, the most recent BP measurements were more current for RPM-prescribed patients (median [IQR] 8 [0-109] versus 134 [56-239] days). Net increases in antihypertensive medications by 12 months were similar. Implementation of RPM in primary care could inform hypertension management strategies and increase hypertension control. Registration: ClinicalTrials.gov identifier: NCT05562921.

In-depth proteomic analysis of juvenile dermatomyositis serum reveals protein expression associated with muscle-specific autoantibodies.

OBJECTIVES: The clinical symptoms and complications of JDM differ depending on the type of muscle-specific autoantibodies (MSAs) present. We aimed to identify protein expression profiles specific for MSAs that characterize various clinical features by comprehensively analyzing the proteins present in the serum of patients with JDM. METHODS: We analysed sera from patients with JDM that were positive for anti-melanoma differentiation-associated protein 5 (MDA5) antibodies (n = 5), anti-nuclear matrix protein 2 (NXP2) antibodies (n = 5) and anti-transcriptional intermediary factor 1 alpha or gamma subunit (TIF1-γ) antibodies (n = 5), and evaluated healthy controls (n = 5) via single-shot liquid chromatography-tandem mass spectrometry (MS) in data-independent acquisition mode, which is superior for comparative quantitative analysis. We identified different protein groups based on MSAs and performed pathway analysis to understand their characteristics. RESULTS: We detected 2413 proteins from serum MS analysis; 508 proteins were commonly altered in MSAs, including many myogenic enzymes and IFN-regulated proteins. Pathway analysis using the top 50 proteins that were upregulated in each MSA group revealed that the type I IFN and proteasome pathways were significantly upregulated in the anti-MDA5 antibody group alone. CONCLUSION: Although JDM serum contains many proteins commonly altered in MSAs, the pathways associated with clinical features of MSAs differ based on protein accumulation. In-depth serum protein profiles associated with MSAs may be useful for developing therapeutic target molecules and biomarkers.

Prospective Cohort Study of Remote Patient Monitoring with and without Care Coordination for Hypertension in Primary Care.

BACKGROUND: Out-of-office blood pressure (BP) measurements contribute valuable information for guiding clinical management of hypertension. Measurements from home devices can be directly transmitted to patients' electronic health record for use in remote monitoring programs. OBJECTIVE: This study aimed to compare in primary care practice care coordinator-assisted implementation of remote patient monitoring (RPM) for hypertension to RPM implementation alone and to usual care. METHODS: This was a pragmatic observational cohort study. Patients aged 65 to 85 years with Medicare insurance from two populations were included: those with uncontrolled hypertension and a general hypertension group seeing primary care physicians (PCPs) within one health system. Exposures were clinic-level availability of RPM plus care coordination, RPM alone, or usual care. At two clinics (13 PCPs), nurse care coordinators with PCP approval offered RPM to patients with uncontrolled office BP and assisted with initiation. At two clinics (39 PCPs), RPM was at PCPs' discretion. Twenty clinics continued usual care. Main measures were controlling high BP (<140/90 mm Hg), last office systolic blood pressure (SBP), and proportion with antihypertensive medication intensification. RESULTS: Among the Medicare cohorts with uncontrolled hypertension, 16.7% (39/234) of patients from the care coordination clinics were prescribed RPM versus <1% (4/600) at noncare coordination sites. RPM-enrolled care coordination group patients had higher baseline SBP than the noncare coordination group (148.8 vs. 140.0 mm Hg). After 6 months, in the uncontrolled hypertension cohorts the prevalences of controlling high BP were 32.5% (RPM with care coordination), 30.7 % (RPM alone), and 27.1% (usual care); multivariable adjusted odds ratios (95% confidence interval) were 1.63 (1.12-2.39; p = 0.011) and 1.29 (0.98-1.69; p = 0.068) compared with usual care, respectively. CONCLUSION: Care coordination facilitated RPM enrollment among poorly controlled hypertension patients and may improve hypertension control in primary care among Medicare patients.

Remote physiologic monitoring for hypertension in primary care: a prospective pragmatic pilot study in electronic health records using propensity score matching.

Objectives: Since 2019, the Centers for Medicare and Medicaid Services covers remote physiologic monitoring (RPM) for blood pressure (BP) per hypertension diagnosis and treatment guidelines. Here, we integrated Omron VitalSight RPM into the health system's electronic health record to transmit BP and pulse without manual entry, assessed feasibility, and used pragmatic prospective matched cohort studies to assess initial effects in (1) uncontrolled (last two office BP ≥140/90 mmHg) and (2) general (diagnosed hypertension or last office BP ≥140/90 mmHg) hypertension patient populations. Materials and Methods: Seventeen clinicians at two internal medicine practices were oriented. Eligible patients were aged 65-85 years had Medicare insurance with ≥1 office visit in the previous year. We prospectively identified matched controls (age, sex, BP, and number of office visits in previous year) from other primary care practices within the health system and estimated the association between RPM availability (clinic-level) and patient BP outcomes after 6 months. ClinicalTrials.gov: NCT04604925. Results: Feasibility. Uptake was low at pilot clinics: 10 physicians prescribed RPM to 118 patients during the 6-month pilot. This included 7% (14/207) of the prespecified uncontrolled hypertension cohort and 3.3% (78/2356) of the general hypertension cohort. Surveyed clinicians (n = 4) reported changing their patients' medical treatment in response to RPM BPs, although they recommended having a dedicated RN or LPN to review BP readings. Effectiveness. At 6 months, BP control was greater at pilot practices than among matched controls (uncontrolled: 31.4% vs 22.8%; P = .007; general: 64.0% vs 59.7%; P < .001). Systolic BP at last office visit did not differ (mean [SD] 146.0 [15.7] vs 147.1 [15.6]; P = .48) in the uncontrolled population, and was lower in the general population (131.8 [15.7] vs 132.8 [15.9]; P = .04).The frequency of antihypertensive medication changes was similar in both groups (uncontrolled P = .986; general P = .218). Discussion and Conclusions: Uptake notwithstanding, RPM may have improved BP control. A potential mechanism is increased physician awareness of and attention to uncontrolled hypertension. Barriers to RPM use among physicians require further study.

Low-molecular-weight anti-HIV-1 agents targeting HIV-1 capsid proteins.

The HIV-1 capsid is a shell that encapsulates viral RNA, and forms a conical structure by assembling oligomers of capsid (CA) proteins. Since the CA proteins are highly conserved among many strains of HIV-1, the inhibition of the CA function could be an appropriate goal for suppression of HIV-1 replication, but to date, no drug targeting CA has been developed. Hydrophobic interactions between two CA molecules through Trp184 and Met185 in the protein are known to be indispensable for conformational stabilization of the CA multimer. In our previous study, a small molecule designed by in silico screening as a dipeptide mimic of Trp184 and Met185 in the interaction site was synthesized and found to have significant anti-HIV-1 activity. In the present study, molecules with different scaffolds based on a dipeptide mimic of Trp184 and Met185 have been designed and synthesized. Their significant anti-HIV activity and their advantages compared to the previous compounds were examined. The present results should be useful in the design of novel CA-targeting anti-HIV agents.

Different Transcriptome Features of Peripheral Blood Mononuclear Cells in Non-Emphysematous Chronic Obstructive Pulmonary Disease.

Non-emphysematous chronic obstructive pulmonary disease (COPD), which is defined based on chest computed tomography findings, presented different transcriptome features of peripheral blood mononuclear cells (PBMCs) compared with emphysematous COPD. Enrichment analysis of transcriptomic data in COPD demonstrated that the "Hematopoietic cell lineage" pathway in Kyoto Encyclopedia of Genes and Genomes pathway analysis was highly upregulated, suggesting that cellular dynamic dysregulation in COPD lungs is affected by pathologically modified PBMCs. The differentially expressed genes (DEGs) upregulated in PBMCs reflected the disease state of non-emphysematous COPD. Upregulated DEGs such as XCL1, PRKCZ, TMEM102, CD200R1, and AQP1 activate T lymphocytes and eosinophils. Upregulating keratan sulfate biosynthesis and metabolic processes is associated with protection against the destruction of the distal airways. ITGA3 upregulation augments interactions with extracellular matrix proteins, and COL6A1 augments the profibrotic mast cell phenotype during alveolar collagen VI deposition. Upregulating HSPG2, PDGFRB, and PAK4 contributes to the thickening of the airway wall, and upregulating SERPINF1 expression explains the better-preserved vascular bed. Therefore, gene expression and pathway analysis in PBMCs in patients with non-emphysematous COPD represented type 2 immune responses and airway remodeling features. Therefore, these patients have asthmatic potential despite no clinical signs of asthma, in contrast to those with emphysematous COPD.

Optimization of Ultrafast Proteomics Using an LC-Quadrupole-Orbitrap Mass Spectrometer with Data-Independent Acquisition.

Proteomics has become an increasingly important tool in medical and medicinal applications. It is necessary to improve the analytical throughput for these applications, particularly in large-scale drug screening to enable measurement of a large number of samples. In this study, we aimed to establish an ultrafast proteomic method based on 5-min gradient LC and quadrupole-Orbitrap mass spectrometer (Q-Orbitrap MS). We precisely optimized data-independent acquisition (DIA) parameters for 5-min gradient LC and reached a depth of >5000 and 4200 proteins from 1000 and 31.25 ng of HEK293T cell digest in a single-shot run, respectively. The throughput of our method enabled the measurement of approximately 80 samples/day, including sample loading, column equilibration, and wash running time. We demonstrated that our method is applicable for the screening of chemical responsivity via a cell stimulation assay. These data show that our method enables the capture of biological alterations in proteomic profiles with high sensitivity, suggesting the possibility of large-scale screening of chemical responsivity.

Single-Shot 10K Proteome Approach: Over 10,000 Protein Identifications by Data-Independent Acquisition-Based Single-Shot Proteomics with Ion Mobility Spectrometry.

The evolution of mass spectrometry (MS) and analytical techniques has led to the demand for proteome analysis with high proteome coverage in single-shot measurements. Focus has been placed on data-independent acquisition (DIA)-MS and ion mobility spectrometry as techniques for deep proteome analysis. We aimed to expand the proteome coverage by single-shot measurements using optimizing high-field asymmetric waveform ion mobility spectrometry parameters in DIA-MS. With our established proteome analysis system, more than 10,000 protein groups were identified from HEK293 cell digests within 120 min of MS measurement time. Additionally, we applied our approach to the analysis of host proteins in mouse feces and detected as many as 892 host protein groups (771 upregulated/121 downregulated proteins) in a mouse model of repeated social defeat stress (R-SDS) used in studying depression. Interestingly, 285 proteins elevated by R-SDS were related to mental disorders. The fecal host protein profiling by deep proteome analysis may help us understand mental illness pathologies noninvasively. Thus, our approach will be helpful for an in-depth comparison of protein expression levels for biological and medical research because it enables the analysis of highly proteome coverage in a single-shot measurement.

Unique Mode of Antiviral Action of a Marine Alkaloid against Ebola Virus and SARS-CoV-2.

Lamellarin α 20-sulfate is a cell-impenetrable marine alkaloid that can suppress infection that is mediated by the envelope glycoprotein of human immunodeficiency virus type 1. We explored the antiviral action and mechanisms of this alkaloid against emerging enveloped RNA viruses that use endocytosis for infection. The alkaloid inhibited the infection of retroviral vectors that had been pseudotyped with the envelope glycoprotein of Ebola virus and SARS-CoV-2. The antiviral effects of lamellarin were independent of the retrovirus Gag-Pol proteins. Interestingly, although heparin and dextran sulfate suppressed the cell attachment of vector particles, lamellarin did not. In silico structural analyses of the trimeric glycoprotein of the Ebola virus disclosed that the principal lamellarin-binding site is confined to a previously unappreciated cavity near the NPC1-binding site and fusion loop, whereas those for heparin and dextran sulfate were dispersed across the attachment and fusion subunits of the glycoproteins. Notably, lamellarin binding to this cavity was augmented under conditions where the pH was 5.0. These results suggest that the final action of the alkaloid against Ebola virus is specific to events following endocytosis, possibly during conformational glycoprotein changes in the acidic environment of endosomes. Our findings highlight the unique biological and physicochemical features of lamellarin α 20-sulfate and should lead to the further use of broadly reactive antivirals to explore the structural mechanisms of virus replication.

Evaluation of the Suitability of Dried Saliva Spots for In-Depth Proteome Analyses for Clinical Applications.

Previously, we performed nontargeted proteome analysis using dried blood spots (DBSs) that are widely used in newborn screening for the clinical diagnosis of congenital genetic diseases and immunodeficiency. We have developed an efficient and simple pretreatment method for DBSs that can detect more than 1000 proteins. To complement proteins that are difficult to detect via DBS analysis with less invasive alternative body fluids, we conducted this study to investigate the proteins detected from dried saliva spots (DSSs) using single-shot LC-MS/MS, which is practical in clinical settings. We also clarified whether DSSs have the same advantages as DBSs, and we investigated the influence of saliva collection conditions and the storage environment on their protein profile. As a result, we detected approximately 5000 proteins in DSSs and whole saliva, and we concluded that they were sufficient to complement the proteins lacking in the blood analysis. DSSs could be used as an alternative tool to DBSs for detecting the presence of causative proteins.

In-Depth Serum Proteomics by DIA-MS with In Silico Spectral Libraries Reveals Dynamics during the Active Phase of Systemic Juvenile Idiopathic Arthritis.

In serum proteomics using mass spectrometry, the number of detectable proteins is reduced due to high-abundance proteins, such as albumin. However, recently developed data-independent acquisition mass spectrometry (DIA-MS) proteomics technology has made it possible to remarkably improve the number of proteins in a serum analysis by removing high-abundance proteins. Using this technology, we analyzed sera from patients with systemic juvenile idiopathic arthritis (sJIA), a rare pediatric disease. As a result, we identified 2727 proteins with a wide dynamic range derived from various tissue leakages. We also selected 591 proteins that differed significantly in their active phases. These proteins were involved in many inflammatory processes, and we also identified immunoproteasomes, which were not previously found in serum, suggesting that they may be involved in the pathogenesis of sJIA. A detailed high-depth DIA-MS proteomic analysis of serum may be useful for understanding the pathogenesis of sJIA and may provide clues for the development of new biomarkers.