PROTEOMICS – Clinical Applications最新文献

Purpose: Anti-CRISPR (Acr) proteins can evade CRISPR-Cas immunity, yet their molecular determinants remain poorly understood. This study aimed to uncover key features driving Acr activity, thereby advancing both fundamental knowledge and the rational design of robust CRISPR-based tools.

Experimental design: We compiled a binary-encoded matrix of 761 InterPro-annotated domains and binding-site features for known Acr proteins. Seven feature ranking algorithms were applied to prioritize determinant features, and an incremental feature selection strategy, coupled with four distinct classifiers, was used to identify optimal subsets. Consensus key features were defined by intersecting the top subsets across all methods.

Results: Key identified features include the DUF2829 domain, the Lambda repressor-like domain and Sulfolobus islandicus virus proteins, the Cro/C1-type helix-turn-helix domain, phage protein, and replication initiator A. These findings illuminate novel structural modules and regulatory motifs that underpin Acr inhibition.

Conclusions: This study provides critical theoretical support for deciphering Acr mechanisms and offers actionable insights for engineering next-generation CRISPR-Cas applications in clinical and biotechnological settings.

Summary: The CRISPR system is a part of the antiviral immune defense initially discovered in bacteria and archaea. At present, the CRISPR system has become the cornerstone of genome editing technologies such as CRISPR-Cas9, widely used in clinical, agricultural, and biological research. Anti-CRISPR proteins are a group of proteins that inhibit the normal activity of CRISPR-Cas system in certain bacteria or archaea and avoid having the phages' genomes destroyed by the prokaryotic cells. The anti-CRISPR protein family has various components, but with similar functions to help exogenous DNA escape from the immune system. This study tried to uncover molecular mechanisms for anti-CRISPR proteins.

Background: Capillary blood collection has a number of advantages over venous collection, especially in the context of increasing decentralized clinical trials and field-based testing. No studies are available comparing venous versus capillary blood collection for proteomics measurement. The aim of this study was to compare venous versus capillary blood collection methods for proteomic measurement in peripheral blood.

Methods: The expression of 368 different proteins from the Olink Explore 384 Inflammation panel was measured in both venous and capillary blood samples collected from 22 individuals at a single time point. Protein levels from venous and capillary blood samples were compared with descriptive statistics and correlation calculations. Correlations were examined for a subset of proteins identified in recent reports as associated with morbidity and mortality.

Results: Strong positive correlation (r ≥ 0.7) between protein concentrations measured in venous and capillary blood samples was observed for two in three proteins tested (215 of 327, 66%). A further 47 (14%) showed a moderate positive correlation (0.4 ≤ r < 0.7), with weak or very weak correlation (r < 0.4) observed for the remaining 65 (20%). Protein expression was consistently higher in capillary blood samples for proteins with lower correlation (r < 0.6) between sampling methods. Further work is required to understand the underlying reasons why proteins were consistently under-expressed in venous samples/over-expressed in capillary samples in a minority of proteins tested.

Conclusions: Proteomic measurement utilising capillary blood collection provides very similar results to using venous blood collection. This is a promising sign for the validity and reliability of studies using capillary blood collection, including decentralised and remote studies.

Biomolecules preserved in dental pulp are increasingly being used to identify individuals in the context of forensics and archaeology. Despite the vast amount of research into host and pathogen DNA, the potential use of physiologically informative proteins preserved in dental pulp has rarely been studied. Here, we hypothesized that pregnancy-specific proteins circulating in the blood could be identified from the dental pulp of postpartum individuals and this was investigated using eight human third molars extracted from four postpartum and three control individuals during clinical treatment. A total of 885 proteins were identified from these eight dental pulp samples using liquid chromatography coupled tandem mass spectrometry, whose gene ontology compositions were similar to previous studies. However, despite our hypothesis, pregnancy-specific proteins were not identified from the dental pulp of postpartum individuals (n = 5, 4-12 months postpartum). Although the dental pulp proteomes obtained from three individuals postpartum ≤6 months were distinct from those of other individuals by principal component analysis (PCA), their driving proteins were less evident. Although our hypothesis was not supported, sample collection, protein extraction, and mass spectrometry analysis could be improved to explore the forensic application of detecting pregnancy-specific proteins in dental pulp.

Introduction: The human retina relies on a complex network of proteins, many of which exhibit intrinsic disorder and liquid-liquid phase separation (LLPS), enabling dynamic interactions for retinal function. Disruptions in these properties, along with missense mutations, have been linked to retinal diseases. This study aims to characterize and compare retinal proteins categorized by their expression specificity and tissue distribution using bioinformatics tools to explore relationships between intrinsic protein disorder, phase separation potential, and mutation pathogenicity.

Methods: We analyzed retinal proteins classified by the Human Protein Atlas (HPA) into two major groups based on gene expression specificity (degree of unique retinal expression) and gene expression distribution (extent of expression across tissues). We analyzed nine retinal proteomes categorized by gene expression specificity and distribution. Intrinsic protein disorder was assessed using per-residue and global disorder predictors from the Rapid Intrinsic Disorder Analysis Online (RIDAO) platform, LLPS potential was evaluated with ParSe v2, and missense mutation pathogenicity was predicted using AlphaMissense.

Results: Significant differences in per-residue intrinsic protein disorder were found within the specificity and distribution subgroups (p < 0.0001). In addition, global disorder predictions from the RIDAO platform demonstrated non-random distributions of protein species across the proteomes analyzed in both subgroups (p < 0.0001). Furthermore, proteins specifically elevated in the retina exhibited higher intrinsic disorder and greater phase separation propensity (ParSe v2, AUC up to 0.650), compared to those more broadly expressed. Lastly, AlphaMissense analysis showed significant variations in the average pathogenicity scores of missense mutations within subgroups (p < 0.0001).

Conclusion: Our results show that intrinsic disorder, LLPS, and mutational tendencies are not evenly distributed among retinal proteomes. Our study demonstrates a link between intrinsic disorder, LLPS potential, and pathogenic vulnerability among retinal proteins, underscoring the unique structural and functional landscape of retinal proteomes. Proteins with higher specificity to the retina exhibit greater disorder and phase separation potential, highlighting their potential role in dynamic cellular processes that support retinal function. Conversely, proteins widely distributed across multiple tissues tend to be more ordered, suggesting a need for structural stability in their broader functional roles.

Purpose: Anxiety is the most common underlying cause of behavioral problems in dogs, which remain a top reason for relinquishment and euthanasia. Despite its high prevalence, anxiety is often underdiagnosed, partly due to a limited understanding of biological processes and absence of diagnostic biomarkers. Our study aims to address this knowledge gap.

Experimental design: Plasma from 10 anxious and 10 matched control dogs were analyzed following a label-free quantitation proteomics workflow based on data-dependent acquisition using a Thermo Q Exactive Plus coupled to an EASY-nLC 1200, Vanquish UHPLC, or Evosep One. Data were processed with Proteome Discoverer 2.4 (Thermo), Perseus (Max Planck Institute), Cytoscape and other bioinformatic tools.

Results: Between 279 and 350 proteins were identified, and proteins such as fibrinogen, apolipoproteins, and complement system and coagulation cascade proteins were significantly different between groups. Additionally, we identified two putative subgroups of anxious dogs, suggesting potentially different underlying pathophysiological mechanisms for a single anxiety phenotype.

Conclusions and clinical relevance: To our knowledge, this is the first comprehensive clinical in-depth proteomic profiling of plasma from anxious dogs. Our findings lay the foundation for elucidating the pathophysiology of canine anxiety and for the future validation and establishment of novel candidate biomarkers for disease diagnosis. Novel biomarkers would allow for a more effective and objective diagnosis of anxiety, even when not phenotypically apparent.

Summary: Previous mass spectrometry (MS) studies have found proteomic profile differences in other diseases and other animal species. This is to our knowledge, the first unbiased and comprehensive clinical in-depth proteomic profiling of plasma from dogs suffering from anxiety disorders. These findings have an impact on animal health as they set the foundation to elucidate the pathophysiology of canine anxiety so that in the future novel candidate biomarkers can be established and validated, furthering the potential development of new drugs and guiding patient-specific therapeutic interventions based on biomarker profiles. In the clinic, novel biomarkers could allow for a more effective and objective diagnosis of anxiety disorders, even when not phenotypically apparent. Detection and measurement of early stages of anxiety disorders as well as treatment monitoring in pet dogs would allow patients to be treated quicker, before the potential onset of aggression, and a faster recovery, thus improving the welfare of companion animals.

Purpose: Chronic kidney disease (CKD) causes detrimental systemic effects, including inflammation or apoptosis, which lead to substantial morbidity and mortality. However, the causal effect of reduced kidney function on systemic proteomic signatures is incompletely understood.

Methods: We performed an integrated Mendelian randomization (MR) and observational analyses to identify the causal association between kidney function and plasma protein levels, based on 1815 plasma protein profiles in 50,407 UK Biobank participants and the CKDGen Phase 4 genome-wide association study (GWAS) meta-analysis for the genetic instruments of eGFR.

Results: The MR analysis revealed 383 plasma proteins causally associated with eGFR. Reduced kidney function was found to be causally associated with an increase in the plasma levels of 381 proteins, among which TNF and IGFBP4 were increased, while the level of two proteins, NPHS1 and SPOCK1, decreased. Apoptosis-related pathway was significantly enriched in the gene-set enrichment analysis. In network analysis, TNF was identified as a hub protein with multiple linkages to molecules included in the TNF-signaling pathways, involved in inflammation, fibrosis, and apoptosis.

Conclusions: In this proteo-genomic analysis, we identified 383 plasma proteins causally associated with eGFR, highlighting TNF-associated pathways as pathologically relevant processes in kidney disease progression, systemic inflammation, and organ fibrosis, warranting further investigation.

Purpose: Diabetes mellitus (DM) is a major cause of end-stage kidney disease (ESKD), with kidney transplantation being the preferred treatment. However, post-transplant diabetes mellitus (PTDM) increases mortality and graft loss. While PTDM and Type 2 diabetes mellitus (T2DM) share risk factors, their mechanisms differ, particularly in diabetic nephropathy (DN). This study aimed to investigate the molecular differences in PTDM by mapping the proteomes of proximal tubuli and serum in normoglycemic (NG), pre-transplant T2DM, and PTDM patients one year post-transplantation. Experimental Design Proteomic analysis was performed on microdissected proximal tubular cells and serum samples from kidney transplant recipients categorized as NG, pre-transplant T2DM, or PTDM at one year post-transplantation. Mass spectrometry was used to identify differentially expressed proteins. Data analyses were performed using gene ontology databases and pathway analysis.

Results: Proteomic analysis revealed key differences, including significant dysregulation of mitochondrial proteins and lipid metabolism pathways in PTDM patients compared to T2DM and NG groups. Additionally, we observed distinct serum patterns of cholesterol metabolism dysregulation in PTDM, highlighting a complex interplay between fatty acid metabolism, mitochondrial dysfunction and systemic lipid dysregulation that may drive renal injury in PTDM-related DN.

Conclusions and clinical relevance: This pilot study is the first to perform proteomic analysis on both microdissected tubular cells and serum from post-transplant PTDM, pre-transplant T2DM and NG transplant recipients. The proteomic differences between PTDM and T2DM could help to develop targeted therapies and early diagnostic markers, ultimately improving transplant outcomes and patient management. Further research is needed to validate these findings and explore their therapeutic potential.

Introduction: Percutaneous transluminal coronary angioplasty (PTCA) is an effective procedure to decrease the severity of stenotic coronary atherosclerotic lesions. However, its long-term success is limited by the formation of restenosis or neointima by increased proliferation of smooth muscle cells (SMCs) and synthesis of extracellular matrix. Polypeptide growth factors are potent SMC mitogens and are involved in SMC proliferation and extracellular matrix (ECM) synthesis. In this line, inhibition of de novo protein synthesis might be beneficial.

Methods: We examined the effects of different concentrations of translational inhibitor puromycin on SMC proliferation and apoptosis, in vitro. Further, we examined the effects of local administration of puromycin in a rabbit balloon injury model of the iliac artery.

Results: Injection of puromycin or its vehicle was performed with an infusion-balloon catheter directly into the vessel wall during angioplasty. PTA in the vehicle group resulted in neointima formation 3 weeks after the vascular intervention. In contrast, puromycin treatment resulted in a significant reduction of intima-media ratio. We observed decreased elastin and collagen III synthesis in puromycin-treated animals. With proteomics, we could demonstrate reduced protein expression of lamin, vimentin, alpha-1 antitrypsin, alpha-actin allowing puromycin treatment. In in vitro experiments, puromycin decreased SMCs proliferation (i.e., BrdU incorporation) following FCS stimulation.

Perspective: Based on the data from our animal experiments, aministration of puromycin directly into the vessel wall during angioplasty may be effective in preventing or reducing restenosis in humans.

Purpose: Parkinson's disease (PD) is neuropathologically characterized by the progressive degeneration of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc), affecting 10 million people worldwide. Rosmarinic acid (RA), a polyphenol found in plants like rosemary (Rosmarinus officinalis), is known for its intriguing biological properties and potential antioxidant and neuroprotective effects. In a previous study we showed that RA treatment prevented hyperlocomotion in mice with MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine)-induced parkinsonism and improved the monoaminergic system in healthy animals. However, the molecular mechanisms underlying RA's action in PD remain unclear.

Experimental design: In this study, we treated MPTP-induced PD animals (C57BL/6 male mice) with RA orally at a dose of 100 mg/kg for 15 days and examined the proteome of substantia nigra (SN) to identify possible regulatory targets of RA treatment to shed some lights into its neuroprotective effects.

Results: Quantitative proteomics revealed that RA treatment regulated proteins associated with oxidative phosphorylation (OXPHOS), glutamatergic synapse, and vesicular cycle signaling pathway. We identified 371 proteins significantly regulated in response to RA administration (255 upregulated and 116 downregulated). Notably, some cellular targets of RA treatment reported here, including mGluR2/mGluR3/EAAT-proteins from the glutamatergic system-and proteins from the Complex I of the electron transport chain are promising targets for therapeutic intervention.

Conclusions and clinical relevance: These findings highlight the molecular differences between MPTP-induced PD mice and those treated with RA, providing insights on the molecular basis behind the neuroprotective effects of RA and revealing potential PD signatures that warrant further investigation.

Maternal and neonatal health (women during pregnancy, childbirth, and the postnatal period) presents a spectrum of healthcare challenges, including preterm birth, preeclampsia, intrauterine growth restriction, polycystic ovarian syndrome, and gestational diabetes mellitus. While genomic investigations have shed light on many of these topics, protein biomarker discovery, a pivotal aspect of such research, holds promise in offering insights into disease diagnosis, progression, and prognosis. This review paper aims to explore the landscape of proteomics research pertaining to the aforementioned disorders. In the search for viable biomarkers, existing ones are either outdated or lack specificity and new ones being investigated do not commonly make it to the validation stage. In this review, the reasons for the gap between the biomarker discovery stage and the clinical validation stage are evaluated, in addition to what steps are being taken to mitigate the unexpectedly slow scientific and clinical progress. Notably, this paper also delves into the ethnic disparities found in maternal and neonatal health research, as well as how AI is currently being used to alleviate socioeconomic and ethnic disparities, as well as its advantages for the analysis of large "omics" datasets. We anticipate this investigation will provide critical, invaluable information for researchers, medical professionals, and policy decision-makers in this field to improve overall maternal and neonatal health outcomes.