Disease-specific Protein Research Articles

Exploring immunotherapeutic strategies for neurodegenerative diseases: a focus on Huntington's disease and Prion diseases.

Immunotherapy has emerged as a promising therapeutic approach for the treatment of neurodegenerative disorders, which are characterized by the progressive loss of neurons and impaired cognitive functions. In this review, active and passive immunotherapeutic strategies that help address the underlying pathophysiology of Huntington's disease (HD) and prion diseases by modulating the immune system are discussed. The current landscape of immunotherapeutic strategies, including monoclonal antibodies and vaccine-based approaches, to treat these diseases is highlighted, along with their potential benefits and mechanisms of action. Immunotherapy generally works by targeting disease-specific proteins, which serve as the pathological hallmarks of these diseases. Additionally, the review addresses the challenges and limitations associated with immunotherapy. For HD, immunotherapeutic approaches focus on neutralizing the toxic effects of mutant huntingtin and tau proteins, thereby reducing neurotoxicity. Immunotherapeutic approaches targeting flanking sequences, rather than the polyglutamine tract in the mutant huntingtin protein, have yielded promising outcomes for patients with HD. In prion diseases, therapies attempt to prevent or eliminate misfolded proteins that cause neurodegeneration. The major challenge in prion diseases is immune tolerance. Approaches to overcome the highly tolerogenic nature of the prion protein have been discussed. A common hurdle in delivering antibodies is the blood‒brain barrier, and strategies that can breach this barrier are being investigated. As protein aggregation and neurotoxicity are related, immunotherapeutic strategies being developed for other neurodegenerative diseases could be repurposed to target protein aggregation in HD and prion diseases. While significant advances in this field have been achieved, continued research and development are necessary to overcome the existing limitations, which will help in shaping the future of immunotherapy as a strategy for managing neurological disorders.

VDAC1: A Key Player in the Mitochondrial Landscape of Neurodegeneration.

Voltage-Dependent Anion Channel 1 (VDAC1) is a mitochondrial outer membrane protein that plays a crucial role in regulating cellular energy metabolism and apoptosis by mediating the exchange of ions and metabolites between mitochondria and the cytosol. Mitochondrial dysfunction and oxidative stress are central features of neurodegenerative diseases. The pivotal functions of VDAC1 in controlling mitochondrial membrane permeability, regulating calcium balance, and facilitating programmed cell death pathways, position it as a key determinant in the delicate balance between neuronal viability and degeneration. Accordingly, increasing evidence suggests that VDAC1 is implicated in the pathophysiology of neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), and others. This review summarizes the current findings on the contribution of VDAC1 to neurodegeneration, focusing on its interactions with disease-specific proteins, such as amyloid-β, α-synuclein, and mutant SOD1. By unraveling the complex involvement of VDAC1 in neurodegenerative processes, this review highlights potential avenues for future research and drug development aimed at alleviating mitochondrial-related neurodegeneration.

Comprehensive Mass Spectral Libraries of Human Thyroid Tissues and Cells

Thyroid nodules are a common endocrine condition with an increasing incidence over the decades. Data-independent acquisition has been widely utilized in discovery proteomics to identify disease biomarkers and therapeutic targets. To analyze the thyroid disease-related proteome in a high-throughput, reproducible and reliable manner, we introduce thyroid-specific peptide spectral libraries. Here, we generated four deep-coverage libraries through four mass spectrometers comprising Q Exactive HF, Orbitrap Exploris 480, ZenoTOF 7600, and timsTOF Pro. These libraries encompass over 215,000 precursors, 172,000 peptides, and 12,000 proteins, derived from 245 tissue samples across nine histological types and 50 cell lines across six histological types. Moreover, the spectral libraries are applied to nine types of thyroid samples to facilitate the exploration of disease-specific proteins. Our spectral libraries serve as a valuable resource for analyzing thyroid protein content, facilitating deeper insights into thyroid disorders.

The Protein Corona on Nanoparticles for Tumor Targeting in Prostate Cancer-A Review of the Literature and Experimental Trial Protocol.

The National Cancer Institute (NCI) recognizes the potential of technologies based on the use of nanoparticles (NPs) in revolutionizing clinical approaches to the diagnosis and prognosis of cancer. Recent research suggests that once NPs come into contact with the biological fluid of cancer patients, they are covered by proteins, forming a "protein corona" composed of hundreds of plasma proteins. The concept of a personalized, disease-specific protein corona, demonstrating substantial differences in NP corona profiles between patients with and without cancer, has been introduced. We developed the design of an experimental prospective single-center study with patients allocated in a 1:1:1 ratio of one of three arms: untreated patients with benign prostatic hyperplasia (BPH), untreated patients with non-metastatic prostate cancer (PCa), and metastatic prostate cancer patients starting systemic therapies with new androgen-targeted agents or taxanes. The protocol aims to develop and implement sensitive nanotools with two distinct objectives: First, to design NPs capable of selectively binding and detecting biomarkers in order to build a predictive diagnostic model to effectively discriminate between patient sera affected by BPH and PCa. Secondly, within the population with PCa, in the case of initial advanced metastatic diagnosis, the objective is to find biomarkers capable of predicting the response to systemic treatments to improve the precision and efficiency of monitoring treatment outcomes. For protein and metabolite corona experiments, we developed a cross-reactive sensor array platform with cancer detection capacity made of three liposomal formulations with different surface charges. For proteomic-NP studies, proteins were identified and quantified using nano-high-performance LC (nanoHPLC) coupled with MS/MS (nanoHPLC-MS/MS). Metabolites were instead analyzed using an untargeted metabolomic approach. Compared with previous review articles, the novelty of this review is represented by the analysis of the possible clinical applications of protein corona NPs focused on PCa and the presentation of a new clinical protocol in the metastatic phase of PCa.

Identification of disease-specific extracellular vesicle-associated plasma protein biomarkers for Duchenne Muscular Dystrophy and Facioscapulohumeral Muscular Dystrophy.

Reliable, circulating biomarkers for Duchenne, Becker and facioscapulohumeral muscular dystrophies (DBMD and FSHD) remain unvalidated. Here, we investigated the plasma extracellular vesicle (EV) proteome to identify disease-specific biomarkers that could accelerate therapy approvals. We extracted EVs from the plasma of DBMD and FSHD patients and healthy controls using size-exclusion chromatography, conducted mass spectrometry on the extracted EV proteins, and performed comparative analysis to identify disease-specific biomarkers. We correlated the levels of these biomarkers with clinical outcome measures and confounding factors. The muscle-associated proteins PYGM, MYOM3, FLNC, MYH2 and TTN were exclusively present in DBMD EVs. PYGM, MYOM3, and TTN negatively correlated with age. PYGM and MYOM3 levels were elevated in patients without cardiomyopathy, and PYGM levels were specifically elevated in ambulatory DMD patients. On the other hand, female FSHD patients displayed significantly higher MBL2 and lower GPLD1 levels. However, male FSHD patients exhibited higher C9 and lower C4BPB levels. Additionally, desmosome proteins JUP and DSP were uniquely found in FSHD males. MBL2 positively correlated with age and C4BPB negatively correlated with FSHD severity in male patients. Our findings underscore the sensitivity of analyzing circulating EV content to identify disease-specific protein biomarkers for DBMD and FSHD. Our results also emphasize the potential of EV-based biomarker discovery as a promising approach to monitor disease progression as well as effectiveness of therapies in muscular dystrophy, potentially contributing to their approval. Further research with larger cohorts is needed to validate these biomarkers and explore their clinical implications.

Targeting Protein Misfolding and Aggregation as a Therapeutic Perspective in Neurodegenerative Disorders.

The abnormal deposition and intercellular propagation of disease-specific protein play a central role in the pathogenesis of many neurodegenerative disorders. Recent studies share the common observation that the formation of protein oligomers and subsequent pathological filaments is an essential step for the disease. Synucleinopathies such as Parkinson's disease (PD), dementia with Lewy bodies (DLB) or multiple system atrophy (MSA) are neurodegenerative diseases characterized by the aggregation of the α-synucleinprotein in neurons and/or in oligodendrocytes (glial cytoplasmic inclusions), neuronal loss, and astrogliosis. A similar mechanism of protein Tau-dependent neurodegeneration is a major feature of tauopathies, represented by Alzheimer's disease (AD), corticobasal degeneration (CBD), progressive supranuclear palsy (PSP), and Pick's disease (PD). The specific inhibition of the protein misfolding and their interneuronal spreading represents a promising therapeutic strategy against both disease pathology and progression. The most recent research focuses on finding potential applications targeting the pathological forms of proteins responsible for neurodegeneration. This review highlights the mechanisms relevant to protein-dependent neurodegeneration based on the most common disorders and describes current therapeutic approaches targeting protein misfolding and aggregation.

Identification of Novel Biomarkers for Post-Kasai Portoenterostomy in Biliary Atresia through Shotgun Proteomics Analysis

Introduction: Biliary Atresia (BA) causes neonatal cholestasis jaundice. The primary therapeutic treatment for BA is the Kasai portoenterostomy. Current diagnostic approaches for BA are imprecise and time-consuming, making early diagnosis crucial for successful treatment outcomes. Objective: This study aims to analyze proteins from Peripheral Blood Mononuclear Cells (PBMCs) obtained from children with BA compared with healthy children Methods and Study Design: We employed a large-scale, total shotgun quantitative serum proteomics approach to analyze the protein from PBMC samples from a discovery cohort. This approach allowed for the simultaneous identification and quantification of multiple proteins, enabling the detection of disease-specific protein expression patterns. The study is proteomic-based study. Results: We identified 24 proteins, by Liquid Chromatography-Mass Spectrometry (LC-MS) analysis that exhibited high discriminatory power for five subjects with BA post-Kasai operation compared to ten healthy controls. ATP2A3, LIN28B, SLC25A3, ITGB3, COX5A, and HLA-B identified proteins of upregulation were predicted to associate with BA post-Kasai operation. Discussion: Our findings highlight the utility of proteomic techniques in BA research. The identified proteomic markers offer promise for improving BA diagnostic accuracy and timeliness, leading to enhanced treatment outcomes for affected children. Conclusion: Proteomic analysis revealed a set of potential biomarkers for early and accurate diagnosis of biliary atresia. These biomarkers hold significant clinical value and have the potential to transform the management of biliary atresia by facilitating timely intervention and improving patient outcomes.

Proteomic signatures improve risk prediction for common and rare diseases

For many diseases there are delays in diagnosis due to a lack of objective biomarkers for disease onset. Here, in 41,931 individuals from the United Kingdom Biobank Pharma Proteomics Project, we integrated measurements of ~3,000 plasma proteins with clinical information to derive sparse prediction models for the 10-year incidence of 218 common and rare diseases (81–6,038 cases). We then compared prediction models developed using proteomic data with models developed using either basic clinical information alone or clinical information combined with data from 37 clinical assays. The predictive performance of sparse models including as few as 5 to 20 proteins was superior to the performance of models developed using basic clinical information for 67 pathologically diverse diseases (median delta C-index = 0.07; range = 0.02–0.31). Sparse protein models further outperformed models developed using basic information combined with clinical assay data for 52 diseases, including multiple myeloma, non-Hodgkin lymphoma, motor neuron disease, pulmonary fibrosis and dilated cardiomyopathy. For multiple myeloma, single-cell RNA sequencing from bone marrow in newly diagnosed patients showed that four of the five predictor proteins were expressed specifically in plasma cells, consistent with the strong predictive power of these proteins. External replication of sparse protein models in the EPIC-Norfolk study showed good generalizability for prediction of the six diseases tested. These findings show that sparse plasma protein signatures, including both disease-specific proteins and protein predictors shared across several diseases, offer clinically useful prediction of common and rare diseases.

The role of exosomes derived from stem cells in nerve regeneration: A contribution to neurological repair

Stem cell-derived exosomes have gained attention in regenerative medicine for their role in encouraging nerve regeneration and potential use in treating neurological diseases. These nanosized extracellular vesicles act as carriers of bioactive molecules, facilitating intercellular communication and enhancing the regenerative process in neural tissues. This comprehensive study explores the methods by which exosomes produced from various stem cells contribute to nerve healing, with a particular emphasis on their role in angiogenesis, inflammation, and cellular signaling pathways. By examining cutting-edge developments and exploring the potential of exosomes in delivering disease-specific miRNAs and proteins, we highlight their versatility in tailoring personalized therapeutic strategies. The findings presented here highlight the potential of stem cell-produced exosomes for use in neurological diseases therapy, establishing the door for future research into exosome-based neurotherapies.

Dual-Aptamer Recognition of DNA Logic Gate Sensor-Based Specific Exosomal Proteins for Ovarian Cancer Diagnosis.

Clinical diagnosis of ovarian cancer lacks high accuracy due to the weak selection of specific biomarkers along with the circumstance biomarkers localization. Clustering analysis of proteins transported on exosomes enables a more precise screening of effective biomarkers. Herein, through bioinformatics analysis of ovarian cancer and exosome proteomes, two coexpressed proteins, EpCAM and CD24, specifically enriched, were identified, together with the development of an as-derived dual-aptamer targeted exosome-based strategy for ovarian cancer screening. In brief, a DNA ternary polymer with aptamers targeting EpCAM and CD24 was designed to present a logic gate reaction upon recognizing ovarian cancer exosomes, triggering a rolling circle amplification chemiluminescent signal. A dynamic detection range of 6 orders of magnitude was achieved by quantifying exosomes. Moreover, for clinical samples, this strategy could accurately differentiate exosomes from healthy persons, other cancer patients, and ovarian cancer patients, enabling promising in situ detection. By accurately selecting biomarkers and constructing a dual-targeted exosomal protein detection strategy, the limitation of insufficient specificity of traditional protein markers was circumvented. This work contributed to the development of exosome-based prognosis monitoring in ovarian cancer through the identification of disease-specific exosome protein markers.

Disease and brain region specific immune response profiles in neurodegenerative diseases with pure and mixed protein pathologies

The disease-specific accumulation of pathological proteins has long been the major focus of research in neurodegenerative diseases (ND), including Alzheimer’s disease (AD) and related dementias (RD), but the recent identification of a multitude of genetic risk factors for ND in immune-associated genes highlights the importance of immune processes in disease pathogenesis and progression. Studies in animal models have characterized the local immune response to disease-specific proteins in AD and ADRD, but due to the complexity of disease processes and the co-existence of multiple protein pathologies in human donor brains, the precise role of immune processes in ND is far from understood. To better characterize the interplay between different extracellular and intracellular protein pathologies and the brain’s intrinsic immune system in ND, we set out to comprehensively profile the local immune response in postmortem brain samples of individuals with “pure” beta-Amyloid and tau pathology (AD), “pure” α-Synuclein pathology in Lewy body diseases (LBD), as well as cases with Alzheimer’s disease neuropathological changes (ADNC) and Lewy body pathology (MIX). Combining immunohistochemical profiling of microglia and digital image analysis, along with deep immunophenotyping using gene expression profiling on the NanoString nCounter® platform and digital spatial profiling on the NanoString GeoMx® platform we identified a robust immune activation signature in AD brain samples. This signature is maintained in persons with mixed pathologies, irrespective of co-existence of AD pathology and Lewy body (LB) pathology, while LBD brain samples with “pure” LB pathology exhibit an attenuated and distinct immune signature. Our studies highlight disease- and brain region-specific immune response profiles to intracellular and extracellular protein pathologies and further underscore the complexity of neuroimmune interactions in ND.

Profiling Protein-Protein Interactions in the Human Brain by Refined Cofractionation Mass Spectrometry.

Proteins usually execute their biological functions through interactions with other proteins and by forming macromolecular complexes, but global profiling of protein complexes directly from human tissue samples has been limited. In this study, we utilized cofractionation mass spectrometry (CF-MS) to map protein complexes within the postmortem human brain with experimental replicates. First, we used concatenated anion and cation Ion Exchange Chromatography (IEX) to separate native protein complexes in 192 fractions and then proceeded with Data-Independent Acquisition (DIA) mass spectrometry to analyze the proteins in each fraction, quantifying a total of 4,804 proteins with 3,260 overlapping in both replicates. We improved the DIA's quantitative accuracy by implementing a constant amount of bovine serum albumin (BSA) in each fraction as an internal standard. Next, advanced computational pipelines, which integrate both a database-based complex analysis and an unbiased protein-protein interaction (PPI) search, were applied to identify protein complexes and construct protein-protein interaction networks in the human brain. Our study led to the identification of 486 protein complexes and 10054 binary protein-protein interactions, which represents the first global profiling of human brain PPIs using CF-MS. Overall, this study offers a resource and tool for a wide range of human brain research, including the identification of disease-specific protein complexes in the future.

TransNeT-CGP: A cluster-based comorbid gene prioritization by integrating transcriptomics and network-topological features

The local disruptions caused by the genes of one disease can influence the pathways associated with the other diseases resulting in comorbidity. For gene therapies, it is necessary to prioritize the key genes that regulate common biological mechanisms to tackle the issues caused by overlapping diseases. This work proposes a clustering-based computational approach for prioritising the comorbid genes within the overlapping disease modules by analyzing Protein-Protein Interaction networks. For this, a sub-network with gene interactions of the disease pair was extracted from the interactome. The edge weights are assigned by combining the pairwise gene expression correlation and betweenness centrality scores. Further, a weighted graph clustering algorithm is applied and dominant nodes of high-density clusters are ranked based on clustering coefficients and neighborhood connectivity. Case studies based on neurodegenerative diseases such as Amyotrophic Lateral Sclerosis- Spinal Muscular Atrophy (ALS-SMA) pair and cancers such as Ovarian Carcinoma-Invasive Ductal Breast Carcinoma (OC-IDBC) pair were conducted to examine the efficacy of the proposed method. To identify the mechanistic role of top-ranked genes, we used Functional and Pathway enrichment analysis, connectivity analysis with leave-one-out (LOO) method, analysis of associated disease-related protein complexes, and prioritization tools such as TOPPGENE and Heml2.0. From pathway analysis, it was observed that the top 10 genes obtained using the proposed method were associated with 10 pathways in ALS-SMA comorbidity and 15 in the case of OC-IDBC, while that in similar methods like SAPDSB and S2B were 4, 6 respectively for ALS-SMA and 9, 10 respectively for OC-IDBC. In both case studies, 70 % of the disease-specific benchmark protein complexes were linked to top-ranked genes of the proposed method while that of SAPDSB and S2B were 55 % and 60 % respectively. Additionally, it was found that the removal of the top 10 genes disconnect the network into 14 distinct components in the case of ALS-SMA and 9 in the case of OC-IDBC. The experimental results shows that the proposed method can be effectively used for identifying key genes in comorbidity and can offer insights about the intricate molecular relationship driving comorbid diseases.

Disease-specific plasma protein profiles in patients with fever after traveling to tropical areas.

Fever is common among individuals seeking healthcare after traveling to tropical regions. Despite the association with potentially severe disease, the etiology is often not determined. Plasma protein patterns can be informative to understand the host response to infection and can potentially indicate the pathogen causing the disease. In this study, we measured 49 proteins in the plasma of 124 patients with fever after travel to tropical or subtropical regions. The patients had confirmed diagnoses of either malaria, dengue fever, influenza, bacterial respiratory tract infection, or bacterial gastroenteritis, representing the most common etiologies. We used multivariate and machine learning methods to identify combinations of proteins that contributed to distinguishing infected patients from healthy controls, and each other. Malaria displayed the most unique protein signature, indicating a strong immunoregulatory response with high levels of IL10, sTNFRI and II, and sCD25 but low levels of sCD40L. In contrast, bacterial gastroenteritis had high levels of sCD40L, APRIL, and IFN-γ, while dengue was the only infection with elevated IFN-α2. These results suggest that characterization of the inflammatory profile of individuals with fever can help to identify disease-specific host responses, which in turn can be used to guide future research on diagnostic strategies and therapeutic interventions.

Expression of Toll-like receptors in the cerebellum during pathogenesis of prion disease.

Prion diseases, such as scrapie, entail the accumulation of disease-specific prion protein (PrPSc) within the brain. Toll-like receptors (TLRs) are crucial components of the pattern recognition system. They recognize pathogen-associated molecular patterns (PAMPs) and play a central role in orchestrating host innate immune responses. The expression levels of Toll-like receptors (TLRs) in the central nervous system (CNS) were not well-defined. To establish a model of prion diseases in BALB/C mice, the 22L strain was employed. The features of the 22L strain were analyzed, and the cerebellum exhibited severe pathological changes. TLR1-13 levels in the cerebellum were measured using quantitative polymerase chain reaction (qPCR) at time points of 60, 90, 120, and the final end point (145 days post-infection). During the pathogenesis, the expression levels of Toll-like receptors (TLRs) 1, 2, 7, 8, and 9 increased in a time-dependent manner. This trend mirrored the expression patterns of PrPSc (the pathological isoform of the prion protein) and glial fibrillary acidic protein. Notably, at the end point, TLR1-13 levels were significantly elevated. Protein level of TLR7 and TLR9 showed increasing at the end point of the 22L-infected mice. A deeper understanding of the increased Toll-like receptors (TLRs) in prion diseases could shed light on their role in initiating immune responses at various stages during pathogenesis. This insight is particularly relevant when considering TLRs as potential therapeutic targets for prion diseases.

The Role of Liquid-Liquid Phase Separation in the Accumulation of Pathological Proteins: New Perspectives on the Mechanism of Neurodegenerative Diseases.

It is widely accepted that living organisms form highly dynamic membrane-less organelles (MLOS) with various functions through phase separation, and the indispensable role that phase separation plays in the mechanisms of normal physiological functions and pathogenesis is gradually becoming clearer. Pathological aggregates, regarded as hallmarks of neurodegenerative diseases, have been revealed to be closely related to aberrant phase separation. Specific proteins are assembled into condensates and transform into insoluble inclusions through aberrant phase separation, contributing to the development of diseases. In this review, we present an overview of the progress of phase separation research, involving its biological mechanisms and the status of research in neurodegenerative diseases, focusing on five main disease-specific proteins, tau, TDP-43, FUS, α-Syn and HTT, and how exactly these proteins reside within dynamic liquid-like compartments and thus turn into solid deposits. Further studies will yield new perspectives for understanding the aggregation mechanisms and potential therapeutic strategies, and future research directions are anticipated.

Programmable Atom-Like Nanoparticle Reporters for High-Precision Urinalysis of In Situ Membrane Proteins.

The expression of disease-specific membrane proteins (MPs) is a crucial indicator for evaluating the onset and progression of diseases. Urinalysis of in situ MPs has the potential for point-of-care disease diagnostics, yet remains challenging due to the lack of molecular reporter to transform the expression information of in situ MPs into the measurable urine composition. Herein, a series of tetrahedral DNA frameworks (TDFs) are employed as the cores of programmable atom-like nanoparticles (PANs) to direct the self-assembly of PAN reporters with defined ligand valence and spatial distribution. With the rational spatial organization of ligands, the interaction between PAN reporters and MPs exhibits superior stability on cell-membrane interface under renal tubule-mimic fluid microenvironment, thus enabling high-fidelity conversion of MPs expression level into binding events and reverse assessment of in situ MP levels via measurement of the renal clearance efficiency of PAN reporters. Such PAN reporter-mediated signal transformation mechanism empowers urinalysis of the onset of acute kidney injury at least 6 h earlier than the existing methods with an area under the curve of 100%. This strategy has the potential for urinalysis of a variety of in situ membrane proteins.

Ultrasensitive and High-Resolution Protein Spatially Decoding Framework for Tumor Extracellular Vesicles.

Proteins localized on the surface or within the lumen of tumor-derived extracellular vesicles (EVs) play distinct roles in cancer progression. However, quantifying both populations of proteins within EVs has been hampered due to the limited sensitivity of the existing protein detection methods and inefficient EV isolation techniques. In this study, the eSimoa framework, an innovative approach enabling spatial decoding of EV protein biomarkers with unmatched sensitivity and specificity is presented. Using the luminal eSimoa pipeline, the absolute concentration of luminal RAS or KRASG12D proteins is released and measured, uncovering their prevalence in pancreatic tumor-derived EVs. The pulldown eSimoa pipeline measured absolute protein concentrations from low-abundance EV subpopulations. The eSimoa assays detected EVs in both PBS and plasma samples, confirming their applicability across diverse clinical sample types. Overall, the eSimoa framework offers a valuable tool to (1) detect EVs at concentrations as low as 105 EVmL-1 in plasma, (2) quantify absolute EV protein concentrations as low as fM, and (3) decode the spatial distribution of EV proteins. This study highlights the potential of eSimoa in identifying disease-specific EV protein biomarkers in clinical samples with minimal pre-purification, thereby driving advancements in clinical translation.

Disease-specific corona mediated co-delivery of MTX and siRNA-TNFα by a polypeptide nanoplatform with antigen-scavenging functions in psoriasis

Psoriatic self-antigen LL37 has the ability to stimulate skin-homing T lymphocytes, which is the constitutive factor for the relapse of psoriasis. The local inflammatory modulation in psoriatic skin is not sufficient for the restoration of immune homeostasis. In this study, we found the expression levels of disease specific proteins including LL37, S100A8 and S100A9 were significantly increased in psoriasis. The self-assembled nanoparticles based on methoxy-poly(ethylene glycol)-block-poly(L-histidine) and methoxy-poly(ethylene glycol)-block-poly(L-lactic acid) (HLNPs) were capable of interacting and absorbing these disease specific proteins, leading to the accumulation of HLNPs in peripheral inflammatory immune cells. Therefore, we proposed an “Antigen-scavenging” polypeptide nano-system assembled from the “sandwich” structure composed of HLNPs, siRNA-TNFα, and oligolysine conjugated methotrexate prodrug (HLNP-MNs), with the encapsulation of ROS-responsive methotrexate prodrug and siRNA-TNFα. We evaluated the biological fate of HLNP-MNs in vivo, the ROS-responsive release of methotrexate (MTX) and siRNA-TNFα in inflammatory immune cells, the suppression of immune response triggered by autoantigen LL37 and the inhibition of skin-homing T lymphocytes from the aspects of “nanoparticle-protein” interaction, cellular level and mouse models. The design of “Antigen-scavenging” polypeptide nanoparticles with loading of MTX and siRNA-TNFα, can shed light on restoring immune homeostasis in psoriasis.

Immunofluorescent Labeling in Nasal Mucosa Tissue Sections of Allergic Rhinitis Rats via Multicolor Immunoassay.

Allergic rhinitis (AR) is a chronic, non-infectious inflammatory disease of the nasal mucosa, primarily mediated by specific immunoglobulin E (IgE), affecting approximately 10%-20% of the world's population. While immunofluorescence (IF) staining has long been a standard technique for detecting disease-specific protein expression, conventional IF techniques are limited in their ability to detect the expression levels of three or more proteins in the same sample. Consequently, multicolor IF techniques have been developed in recent years, which allow the simultaneous labeling of multiple targets in cells or tissues. This protocol provides a comprehensive overview of the process for establishing a rat model of AR, obtaining nasal mucosal samples, and the technical procedures for multicolor immunofluorescence. All rats in the AR group exhibited typical symptoms such as sneezing, a runny nose, and an itchy nose, with behavioral observations scoring ≥5 points. Hematoxylin and eosin (H&E) staining revealed increased inflammatory cell counts and disrupted nasal mucosal integrity in the AR group. Multicolor immunofluorescence (mIF) demonstrated increased expression of RORγt and TICAM-1, while Foxp3 expression decreased in the nasal mucosa tissue of AR rats.