Enrichment Of Proteins Research Articles

Integrated proteomics reveals enrichment of oxidative stress and inflammatory proteins in the urine and stone matrix of calcium oxalate stone formers.

Nephrolithiasis is a multifactorial disease associated with urinary and matrix proteins that become a focal point of research for diagnostic and preventative strategies. The functional relevance of these proteins in lithogenesis, along with their origins and impacts, remains a major subject of ongoing lithogenic research. Here, an integrated analysis was done on multiple proteome datasets compiled from various studies of normal urine (NU), urine from calcium oxalate stone formers (SFU), and calcium oxalate stone matrix (SM). Functional annotation and network analysis revealed the profound enrichment of proteins associated with oxidative stress and inflammation only in the stone-related samples (both "SFU but not NU" and "SM but not NU" cohorts). The oxidative stress and inflammation-related proteins were most abundant in the "SM but not NU" cohort and had higher proportions in the "SFU but not NU" cohort than the "NU only" cohort. KEGG pathway analysis corroborated such observation and highlighted the inclusion of proteins in the complement and coagulation pathways, particularly in SM. The findings of this study inform some mechanistic insights into the roles of calcium oxalate stone-related proteins and may help develop effective prevention and treatment strategies for nephrolithiasis.

Targeted detection of endogenous LINE-1 proteins and ORF2p interactions

BackgroundBoth the expression and activities of LINE-1 (L1) retrotransposons are known to occur in numerous cell-types and are implicated in pathobiological contexts such as aging-related inflammation, autoimmunity, and in cancers. L1s encode two proteins that are translated from bicistronic transcripts. The translation product of ORF1 (ORF1p) has been robustly detected by immunoassays and shotgun mass spectrometry (MS). Yet, more sensitive detection methods would enhance the use of ORF1p as a clinical biomarker. In contrast, until now, no direct evidence of endogenous L1 ORF2 translation to protein (ORF2p) has been shown. Instead, assays for ORF2p have been limited to ectopic L1 ORF over-expression contexts and to indirect detection of endogenous ORF2p enzymatic activity, such as by the sequencing of de novo genomic insertions. Immunoassays for endogenous ORF2p have been problematic, producing apparent false positives due to cross-reactivities, and shotgun MS has not yielded reliable evidence of ORF2p peptides in biological samples.ResultsHere we present targeted mass spectrometry assays, selected and parallel reaction monitoring (SRM and PRM, respectively) to detect and quantify L1 ORF1p and ORF2p at their endogenous abundances. We were able to quantify ORF1p and ORF2p present in our samples down to a range in the low attomoles. Confident in our ability to affinity enrich ORF2p, we describe an interactome associated with endogenous ORF2-containing macromolecular assemblies.ConclusionsThis is the first assay to demonstrate sensitive and robust quantitation of endogenous ORF2p. The ability to assay ORF2p directly and quantitatively will improve our understanding of the developmental and diseased cell states where L1 expression and its activity naturally occur. The ability to simultaneously assay endogenous L1 ORF1p and ORF2p is an important step forward for L1 analytical biochemistry. Endogenous ORF2p interactomes can now be presented with confidence that ORF2p is among the enriched proteins.

Novel antibodies detect nucleocytoplasmic O-fucose in protist pathogens, cellular slime molds, and plants.

Cellular adaptations to change often involve post-translational modifications of nuclear and cytoplasmic proteins. An example found in protists and plants is the modification of serine and threonine residues of dozens to hundreds of nucleocytoplasmic proteins with a single fucose (O-fucose). A nucleocytoplasmic O-fucosyltransferase occurs in the pathogen Toxoplasma gondii, the social amoeba Dictyostelium, and higher plants, where it is called Spy because mutants have a spindly appearance. O-fucosylation, which is required for optimal proliferation of Toxoplasma and Dictyostelium, is paralogous to the O-GlcNAcylation of nucleocytoplasmic proteins of plants and animals that are involved in stress and nutritional responses. O-fucose was first discovered in Toxoplasma using Aleuria aurantia lectin, but its broad specificity for terminal fucose residues on N- and O-linked glycans in the secretory pathway limits its use. Here we present affinity-purified rabbit antisera that are selective for the detection and enrichment of proteins bearing fucose-O-Ser or fucose-O-Thr. These antibodies detect numerous nucleocytoplasmic proteins in Toxoplasma, Dictyostelium, and Arabidopsis, as well as O-fucose occurring on secretory proteins of Dictyostelium and mammalian cells except when blocked by further glycosylation. The antibodies label Toxoplasma, Acanthamoeba, and Dictyostelium in a pattern reminiscent of O-GlcNAc in animal cells including nuclear pores. The O-fucome of Dictyostelium is partially conserved with that of Toxoplasma and is highly induced during starvation-induced development. These antisera demonstrate the unique antigenicity of O-fucose, document the conservation of the O-fucome among unrelated protists, and enable the study of the O-fucomes of other organisms possessing O-fucosyltransferase-like genes.IMPORTANCEO-fucose (O-Fuc), a form of mono-glycosylation on serine and threonine residues of nuclear and cytoplasmic proteins of some parasites, other unicellular eukaryotes, and plants, is understudied because it is difficult to detect owing to its neutral charge and lability during mass spectrometry. Yet, the O-fucosyltransferase enzyme (OFT) is required for optimal growth of the agent for toxoplasmosis, Toxoplasma gondii, and an unrelated protist, the social amoeba Dictyostelium discoideum. Furthermore, O-fucosylation is closely related to the analogous process of O-GlcNAcylation of thousands of proteins of animal cells, where it plays a central role in stress and nutritional responses. O-Fuc is currently best detected using Aleuria aurantia lectin (AAL), but in most organisms, AAL also recognizes a multitude of proteins in the secretory pathway that are modified with fucose in different ways. By establishing the potential to induce highly specific rabbit antisera that discriminate O-Fuc from all other forms of protein fucosylation, this study expands knowledge about the protist O-fucome and opens a gateway to explore the potential occurrence and roles of this intriguing posttranslational modification in bacteria and other protist pathogens such as Acanthamoeba castellanii.

Quantitative differences in rumen epithelium proteins and detection of lysine acetylation in lambs fed a low or high metabolizable energy diet.

Thirty-six Merino wethers (10-month-old) were fed ad libitum for 30 days two diets;1) low metabolizable energy diet (LME; 30% lucerne: 70% cereal chaff) and 2) high ME diet (HME; 40% rolled barley grain: 50% lucerne: 10% cereal chaff). Effects of diet on dry matter intake (DMI), metabolizable energy intake (MEI), liveweight (LWT), average daily gain (ADG), carcass lean or fat gain, liver and empty rumen weight and plasma metabolites were analysed. A membrane enriched protein fraction of rumen epithelium isolated enzymatically from whole depth rumen wall was quantified for each sheep using tandem mass tag mass spectrometry (TMT-MS). The presence or absence of acetylation of lysine residues on identified proteins was counted and the position of the lysine acetylation recorded. In lambs fed the HME diet, DMI (P<0.001), MEI (P<0.001), ADG (P<0.001), fat (P<0.001) and lean gain (P<0.001), as well as liver (P<0.001) and empty rumen (P<0.009) weight were greater than those fed the LME diet. Plasma glucose (P<0.001) and β hydroxybutyrate (P<0.001) at 3 and 5h after feeding was greater in HME diet than in the LME fed lambs. Changes in rumen epithelium protein abundance in the LME versus HME fed lambs were associated with metabolism in the peroxisome, protein processing in the endoplasmic reticulum (ER), valine, leucine and isoleucine degradation and carbon metabolism. Acetylation of lysine was detected in enzymes involved in glycolysis, tricarboxylic acid (TCA) cycle and fatty acid (FA) metabolism. Quantitative differences in the abundance of rumen epithelium proteins that carry out intracellular processes of energy expenditure were associated with the concentration of ME (MJ/ kg DM) in the diet of growing lambs. The detection of lysine acetylation sites suggests a difference in the ME of the diet regulates enzymatic activity in central metabolic pathways in the rumen epithelium cells.

Deep, Unbiased, and Quantitative Mass Spectrometry-Based Plasma Proteome Analysis of Individual Responses to mRNA COVID-19 Vaccine.

Global campaign against COVID-19 have vaccinated a significant portion of the world population in recent years. Combating the COVID-19 pandemic with mRNA vaccines played a pivotal role in the global immunization effort. However, individual responses to a vaccine are diverse and lead to varying vaccination efficacy. Despite significant progress, a complete understanding of the molecular mechanisms driving the individual immune response to the COVID-19 vaccine remains elusive. To address this gap, we combined a novel nanoparticle-based proteomic workflow with tandem mass tag (TMT) labeling, to quantitatively assess the proteomic changes in a cohort of 12 volunteers following two doses of the Pfizer-BioNTech mRNA COVID-19 vaccine. This optimized protocol seamlessly integrates comprehensive proteome analysis with enhanced throughput by leveraging the enrichment of low-abundant plasma proteins by engineered nanoparticles. Our data demonstrate the ability of this workflow to quantify over 3,000 proteins, providing the deepest view into COVID-19 vaccine-related plasma proteome study. We identified 69 proteins with boosted responses post-second dose and 74 proteins differentially regulated between individuals who contracted COVID-19 despite vaccination and those who did not. These findings offer valuable insights into individual variability in response to vaccination, demonstrating the potential of personalized medicine approaches in vaccine development.

A Solvatochromic and Photosensitized Lipid Droplet Probe Detects Local Polarity Heterogeneity and Labels Interacting Proteins in Human Liver Disease Tissue.

The intricate morphology, physicochemical properties, and interacting proteins of lipid droplets (LDs) are associated with cell metabolism and related diseases. To uncover these layers of information, a solvatochromic and photosensitized LDs-targeted probe based on the furan-based D-D-π-A scaffold is developed to offer the following integrated functions. First, the turn-on fluorescence of the probe upon selectively binding to LDs allows for direct visualization of their location and morphology. Second, its solvatochromic fluorescence with linear correlation to polarity quantifies micro-environmental heterogeneity among LDs. Third, the unique photosensitized properties enable photocatalytic proximity labeling and enrichment of LDs-interacting proteins, ready for potential downstream proteomic analysis. These functions are exemplified using artificial LDs in buffer, stressed liver cell line, and diseased liver tissues biopsied from patients. While most LD sensors only offer fluorescence imaging functions, the multi-functional LD probe reported herein integrates both singlet fluorescence and triplet photosensitization properties for LDs studies.

The role of SASP in ischemic stroke: a deep dive into cellular mechanisms

BackgroundThe escalating incidence of ischemic stroke (IS) exerts a heavy toll on global health. Aging, a prominent risk factor, implicates the senescence-associated secretory phenotype (SASP) in IS pathogenesis. We postulated that alterations in SASP-related factor expression during IS correlate with remodeling of intercellular interaction networks and disease advancement. The present study endeavored to preliminarily dissect the SASP-IS nexus via combined bulk and single-cell transcriptome analysis.MethodsAggregated expression profiles from human peripheral blood bulk chips and MCAO mouse single-cell sequencing data, followed by SASP gene analysis. Executed protein interaction network and enrichment assays. Investigated immune infiltration in stroke patients, managed quality control and annotation of single-cell data, cherry-picked central cells based on SASP scores, unearthed essential genes via enrichment analysis, conducted pseudo-time and intercellular communication studies, and prognosticated drugs for hub genes. Finally, authenticated core gene expression in serum of MCAO and Sham rats using real-time fluorescent polymerase chain reaction (RT-qPCR).ResultsFourteen hub genes were discerned. Seven cell types were annotated in MCAO mouse peripheral blood single-cell data. Basophils exhibited the highest SASP scores, with Lcp1 upregulated and Ccl3 downregulated in basophils of the MCAO group. Enrichment analysis divulged a significant association of Ccl3 with the cell apoptosis pathway and Lcp1 with immune responses. The Ccl3 gene is pivotal in basophils and basophil-neutrophil crosstalk. Additionally, we forecasted nagrestipen’s regulatory function on Ccl3. RT-qPCR demonstrated a marked elevation in Lcp1 mRNA and a pronounced reduction in Ccl3 in the MCAO group relative to the Sham group.ConclusionThe Ccl3 gene in basophils and its immune cell interaction is a linchpin in the IS immune microenvironment. Ccl3 and Lcp1 might potentially modulate IS progression by influencing SASP, proffering novel prospects for IS clinical diagnosis and treatment.

Secretome and Proteome of Extracellular Vesicles Provide Protein Markers of Lung and Colorectal Cancer

Colorectal cancer (CRC) and lung cancer (LC) are leading causes of cancer-related mortality, highlighting the need for minimally invasive diagnostic, prognostic, and predictive markers for these cancers. Proteins secreted by a tumor into the extracellular space directly, known as the tumor secretome, as well as proteins in the extra-cellular vesicles (EVs), represent an attractive source of biomarkers for CRC and LC. We performed proteomic analyses on secretome and EV samples from LC (A549, NCI-H23, NCI-H460) and CRC (Caco2, HCT116, HT-29) cell lines and targeted mass spectrometry on EVs from plasma samples of 20 patients with CRC and 19 healthy controls. A total of 782 proteins were identified across the CRC and LC secretome and EV samples. Of these, 22 and 44 protein markers were significantly elevated in the CRC and LC samples, respectively. Functional annotation revealed enrichment in proteins linked to metastasis and tumor progression for both cancer types. In EVs isolated from the plasma of patients with CRC, ITGB3, HSPA8, TUBA4A, and TLN1 were reduced, whereas FN1, SERPINA1, and CST3 were elevated, compared to healthy controls. These findings support the development of minimally invasive liquid biopsy methods for the detection, prognosis, and treatment monitoring of LC and CRC.

Electrical Stimulation Generates Induced Tumor-Suppressing Cells, Offering a Potential Option for Combatting Breast Cancer and Bone Metastasis

Treating advanced metastatic cancer, particularly with bone metastasis, remains a significant challenge. In previous studies, induced tumor-suppressing (iTS) cells were successfully generated through genetic, chemical, and mechanical interventions. This study investigates the potential of electrical stimulation to generate iTS cells. Using a custom electrical stimulator with platinum electrodes, mesenchymal stem cells (MSCs) and Jurkat T cells were stimulated under optimized conditions (50 mV/cm, 10–100 Hz, 1 h). Conditioned medium (CM) from electrically stimulated cells demonstrated tumor-suppressing capabilities, inhibiting tumor cell migration, 3D spheroid growth, and cancer tissue fragment viability. Additionally, the CM reduced osteoclast maturation while promoting osteoblast differentiation. Proteomic analysis revealed enrichment of tumor-suppressing proteins, including histone H4, in the CM. Functional studies identified Piezo1 as a key mediator, as its knockdown significantly impaired the tumor-suppressive effects. Mechanistically, the process was distinct from other methods, such as mechanical vibration, with SUN1 inhibition showing no effect on iTS cell generation by electrical stimulation. These findings demonstrate the efficacy of electrical stimulation in enhancing the antitumor capabilities of MSCs and T cells, offering a novel approach to cancer therapy. Further exploration of this strategy could provide valuable insights into developing new treatments for metastatic cancer.

Galangin alleviates gastric mucosal injury in rats with chronic atrophic gastritis by reducing ferroptosis.

Chronic atrophic gastritis (CAG) is a precancerous lesion and is the first stage in a multistep precancerous cascade that can lead to gastric adenocarcinoma. This study aimed to reveal the role and mechanism of galangin in CAG. Rats were intragastrically administered a mixture of 2% sodium salicylate and 30% alcohol, forced to exercise, and fasted irregularly to establish CAG models. To explore the efficacy of galangin on CAG rats, we used Hericium erinaceus (HE) and omeprazole (Ome) as controls. The degree of gastric mucosal injury was assessed by H&E staining and immunohistochemistry. Perls staining and western blot analysis were used to assess iron content and enrichment of ferroptosis-related proteins. Reactive oxygen species and mitochondrial superoxide in the mucosa were visualized by probes. The morphology of cells was examined by transmission electron microscopy. Our data showed that galangin treatment alleviated gastric mucosal damage and reduced ferroptosis in CAG rats, manifested as decreased iron content, iron transporters and storage proteins, decreased ROS and mitochondrial superoxide, and partially restored cellular morphology. Of note, galangin at a high concentration had better treatment efficacy than HE but lower than Ome. This study demonstrated that galangin reduced gastric mucosal injury in CAG rats by inhibiting ferroptosis. These findings provide a theoretical basis for its clinical application and broaden its potential applications.

Proximity labeling and orthogonal nanobody pulldown (ID-oPD) approaches to map the spinophilin interactome uncover a putative role for spinophilin in protein homeostasis.

Spinophilin is a dendritic spine enriched scaffolding and protein phosphatase 1 targeting protein. To detail spinophilin interacting proteins, we created an Ultra-ID and ALFA-tagged spinophilin encoding construct that permits proximity labeling and orthogonal nanobody pulldown (ID-oPD) of spinophilin-associated protein complexes in heterologous cells. We identified 614 specific, and 312 specific and selective, spinophilin interacting proteins in HEK293 cells and validated a subset of these using orthogonal approaches. Many of these proteins are involved in mRNA processing and translation. In the brain, we determined that spinophilin mRNA is highly neuropil localized and that spinophilin may normally function to limit its own expression but promote the expression of other PSD-associated proteins. Overall, our use of an ID-oPD approach uncovers a novel putative role for spinophilin in mRNA translation and synaptic protein expression specifically within dendritic spines.

Extreme Tolerance of Nanoparticle-Protein Corona to Ultra-High Abundance Proteins Enhances the Depth of Serum Proteomics.

The serum nanoparticle-protein corona (NPC) provides specific disease information, thus opening a new avenue for high-throughput in-depth proteomics to facilitate biomarker discovery. Yet, little is known about the interactions between NPs and proteins, and its role in enhanced depth of serum proteomics. Herein, a series of protein spike-in experiments are conducted to systematically investigate protein depletion and enrichment during NPC formation. Proteomic depth is serum concentration-dependent, and NPC exhibits powerful tolerance to ultra-high abundant proteins. In addition, protein-protein interactions (PPI), especially those involving albumin, play a pivotal role in promoting proteomic depth. Furthermore, a triple-protein assay is established to interrogate the relationship between protein binding affinity and concentration. NPC formation is a product of balancing binding affinity, concentration, and PPI. Overall, this study elucidates how NPs achieve protein depletion and enrichment for enhanced serum proteomic depth to gain a better understanding of NPC as an essential tool of proteome profiling.

P0258 APOA1, a promising biomarker for early monitoring recurrence of ulcerative colitis

Abstract Background Individual information transmission is coordinated across tissues by exosomes, and the colonic inflammation response depends on continuous communication with the peripheral blood. Recent report showed exosome derived miRNA could assist in the early diagnosis and monitoring recurrence of ulcerative colitis (UC). However, the role of exosome proteins in colitis remains unknown. Methods We analyzed the clinical information of UC patients in the Department of Gastroenterology of Peking University Third Hospital from January 2022 to October 2023. Then, we assessed plasma exosome mass spectroscope-based proteomics of active UC patients (UCa), UC patients in clinical remission (UCi) and healthy controls (HCs). Finally, we validated the differential proteins expression by western blot in the DSS-induced acute and chronic colitis. Results We retrospectively analyzed the clinical information of 568 UC patients (774 person-time). UC patients whose Mayo endoscopic score was higher had more severe circulating inflammation, and peripheral lower cholesterol, HDL-c and LDL-c were related to a significantly higher degree of colonic inflammation. Using plasma exosome mass spectroscope-based proteomics, we demonstrated that proteome landscape of the plasma exosome was significantly differ among health controls and distinct UC stages, which enriched proteins in UCa group mainly focused on cholesterol metabolism. We further identified a star molecule on the cholesterol metabolism pathway, APOA1, since it was up-regulated both in DSS-induced acute and chronic mice colon, and it might play a significant role in the lipid disturbance of UC. Conclusion We firstly demonstrated that proteome landscape of the plasma exosome among health controls and different UC stages, prompting that cholesterol metabolism might play a significant role in the pathogenesis of UC. In addition, we provided evidence into further investigating of APOA1’s role in the cholesterol metabolism in the context of UC.

P0071 Host fecal proteolytic activity is increased following antibiotic treatment in healthy individuals, but not patients with pouchitis

Abstract Background Antibiotics (Abx) are crucial for treating infections but significantly disrupt the gut microbiome. While fecal metagenomic studies explored microbial taxa affected by Abx, metaproteomic analyses are lacking. This study investigates how Abx treatment influences host and bacterial activities in healthy individuals and in patients with pouch inflammation (pouchitis). Methods We conducted label-free quantitative LC-MS/MS analyses on stool samples from 7 controls and 6 patients with pouchitis, before and after Abx treatment. Proteins were identified using the unified human gut proteome database. Proteolytic activity was measured in fecal extracts, using a fluorescence kit. Statistical analyses were performed using MSstats in R. p-value&amp;lt;0.05 and Log2fold-change&amp;gt;1 were considered significant after adjustment for multiple comparisons. Results Principal component analysis showed that metaproteomes clustered according to disease state and Abx treatment. Before Abx, there was distinct expression of host immune proteins and proteases; as well as of hundreds of microbial proteins, when comparing patients with pouchitis to controls. In healthy individuals, Abx treatment resulted in diminished protein abundance, (e.g., MUC12) and enrichment of several proteases (CELA2A and CTRC). Concurrently, hundreds of microbial proteins were not expressed after Abx. Pouchitis proteins expression was less affected by Abx, possibly due to a pre-existing dysbiosis. Comparison of healthy and pouchitis metaproteomes post-Abx treatment revealed enrichment in immunoglobulins and immune proteins (REG1) among pouchitis patients. Conversely, healthy show enrichment in host proteases (CELA3A, TMPRSS15) and serine protease inhibitors. Microbial proteins were also enriched in healthy compared to patients with pouchitis after Abx, though the effect was less pronounced than in pre-Abx samples. Last, measurements of proteolytic activity in stool extracts, demonstrated that Abx treatment increased proteolytic activity in healthy individuals to levels resembling patients with pouchitis at baseline and post-Abx. Intriguingly, patients with pouchitis showed no further increase in proteolytic activity after Abx treatment. Conclusion Proteolytic activity is increased following Abx treatment in healthy individuals, opposed to patients with pouchitis, who exhibit increased proteolytic activity at baseline and post-Abx. The finding that microbial proteins were enriched in healthy compared to pouchitis, before and after Abx, underscores the importance of the microbiome for the homeostasis of the luminal proteolytic activity. Further research is needed to understand how microbial dysbiosis enriches host protease abundance.

Extracellular Mitochondrial-Derived Vesicles Affect the Progression of Diabetic Foot Ulcer by Regulating Oxidative Stress and Mitochondrial Dysfunction.

Diabetic foot ulcer (DFU) is a common and severe complication of diabetes mellitus, the etiology of which remains insufficiently understood, particularly regarding the involvement of extracellular vesicles (EVs). In this study, nanoflow cytometry to detect EVs in DFU skin tissues is used and found a significant increase in the Translocase of Outer Mitochondrial Membrane 20 (TOM20)+ mitochondrial-derived vesicles (MDVs). The role of MDVs in DFU is yet to be reported. Using single-cell datasets, it is discovered that the increase in MDVs may be regulated by Sorting Nexin 9 (SNX9). In vitro experiments revealed that MDVs secreted by fibroblasts cultured in high glucose medium exhibited similar composition and protein enrichment results to those in DFU tissues, suggesting their potential as an ideal in vitro surrogate. These MDVs promoted apoptosis and intracellular oxidative stress, disrupted mitochondrial structure, and reduced aerobic metabolism in target cells. In vivo experiments also showed that MDV drops hindered wound healing in diabetic mice; however, this effect is rescued by SNX9 inhibitors, restoring mitochondrial dynamics and balance. Under high glucose conditions, MDVs significantly upregulated oxidative stress levels and induced mitochondrial dysfunction. This study proposes targeting MDVs as a potential therapeutic strategy for DFU.

Substrate stiffness dictates unique doxorubicin-induced senescence-associated secretory phenotypes and transcriptomic signatures in human pulmonary fibroblasts.

Cells are subjected to dynamic mechanical environments which impart forces and induce cellular responses. In age-related conditions like pulmonary fibrosis, there is both an increase in tissue stiffness and an accumulation of senescent cells. While senescent cells produce a senescence-associated secretory phenotype (SASP), the impact of physical stimuli on both cellular senescence and the SASP is not well understood. Here, we show that mechanical tension, modeled using cell culture substrate rigidity, influences senescent cell markers like SA-β-gal and secretory phenotypes. Comparing human primary pulmonary fibroblasts (IMR-90) cultured on physiological (2kPa), fibrotic (50kPa), and plastic (approximately 3 GPa) substrates, followed by senescence induction using doxorubicin, we identified unique high-stiffness-driven secretory protein profiles using mass spectrometry and transcriptomic signatures, both showing an enrichment in collagen proteins. Consistently, clusters of p21 + cells are seen in fibrotic regions of bleomycin induced pulmonary fibrosis in mice. Computational meta-analysis of single-cell RNA sequencing datasets from human interstitial lung disease confirmed these stiffness SASP genes are highly expressed in disease fibroblasts and strongly correlate with mechanotransduction and senescence-related pathways. Thus, mechanical forces shape cell senescence and their secretory phenotypes.

Enhancing biosensing of trace tau protein in clinical samples via emergence macroscopic directed aggregation.

Alzheimer's disease (AD) is an irreversible neurodegenerative disorder that poses a significant risk to human health and well-being. The high cost and invasiveness of neuroimaging and cerebrospinal fluid (CSF) analysis underscores the necessity for accessible early screening via blood samples. In this study, we developed an ultrasound-based strategy for emergent macroscopic that enhances the acoustic response enrichment of specific proteins by introducing functionalized microspheres. This customized macroscopic-directed aggregation method combines protein enrichment with specific fluorescent antibody recognition, enabling quantitative detection characterized by high sensitivity and specificity for tau proteins in clinical samples. In comparison with previously reported biosensing platforms, this strategy achieves indirectly driven clustering of tau proteins by integrating an acoustic resonant chamber, realizing facile and low-cost enrichment detection of protein without the need for amplification. Results demonstrate that this method exhibits a linear detection range from 1pg/mL to 10ng/mL, with a detection limit of 0.183pg/mL. AD patients and healthy individuals were successfully distinguished with an accuracy of 90.9%. This ultrasonic biosensing strategy based on protein aggregation exhibits potential as a valuable tool for early screening of AD.

Proteins and pathways involved in inflammation are longitudinally associated with total body bone mineral density among primarily hispanic overweight/obese adolescents and young adults.

Bone mineral density (BMD), an important marker of bone health, is regulated by a complex interaction of proteins. Plasma proteomic analyses can contribute to identification of proteins associated with changes in BMD. This may be especially informative in stages of bone accrual and peak BMD achievement (i.e., adolescence and young adulthood), but existing research has focused on older adults. This analysis in the Study of Latino Adolescents at Risk for Type 2 Diabetes (SOLAR; n = 304; baseline age 8-13, 100% Hispanic) explored associations between baseline proteins (n = 653 proteins) measured with Olink® plasma protein profiling and repeated annual DXA measures of BMD (average of 3.2 visits per participant). Covariate-adjusted linear mixed effect regression models were applied to estimate longitudinal protein-BMD associations using an adjusted p-value cutoff (P < 0.00068). Identified proteins were imported into the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database to determine significantly enriched protein pathways. Forty-four proteins, many of which are involved in inflammatory processes, were associated with longitudinal changes in total body BMD, including several proteins previously linked to bone health such as osteopontin (SPP1) and microfibrillar-associated protein 5 (MFAP5; both P < 0.00068). These 44 proteins were associated with enrichment of pathways including PI3K-Akt Signaling Pathway and Cytokine-Cytokine Receptor Interaction, supporting results from existing proteomics analyses in older adults. To evaluate whether protein associations were consistent into young adulthood, linear mixed effect models were repeated in a young adult cohort (n = 169; baseline age 17-22; 62.1% Hispanic) with 346 available overlapping Olink® protein measures. While there were no significant overlapping longitudinal protein associations between the cohorts, these findings suggest differences in protein regulation at different ages and provide novel insight on longitudinal protein associations with BMD in overweight/obese adolescents and young adults of primarily Hispanic origin, which may inform the development of biomarkers for bone health in youth.

Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation.

Glycosylation plays a critical role in various biological processes, yet identifying specific glycosyltransferase substrates remains a challenge due to the complexity of glycosylation. Here, we employ proximity labeling with biotin ligases BASU and TurboID to map the proximitome of MGAT3, a glycosyltransferase responsible for the biosynthesis of the bisecting GlcNAc structure, in HEK293T cells. This approach enriched 116 and 189 proteins, respectively, identifying 17 common substrates shared with bisecting GlcNAc-bearing proteome obtained via intact glycopeptide enrichment methods. Gene ontology analysis revealed that the enriched proteins were predominantly localized in the exosome, endoplasmic reticulum, and Golgi apparatus, consistent with subcellular localization of MGAT3 substrates. Notably, four novel substrates, GOLM2, CCDC134, ASPH, and ERO1A, were confirmed to bear bisecting GlcNAc modification, validating the utility of the proximity labeling method. Furthermore, we observed that bisecting GlcNAc modification inhibits breast cancer progression by promoting the degradation of α-galactosidase A (GLA). These findings demonstrate the efficacy of proximity labeling in identifying glycosyltransferase substrates and provide insights into the functional impact of bisecting GlcNAc modification.

Possible linking and treatment between Parkinson’s disease and inflammatory bowel disease: a study of Mendelian randomization based on gut–brain axis

BackgroundMounting evidence suggests that Parkinson’s disease (PD) and inflammatory bowel disease (IBD) are closely associated and becoming global health burdens. However, the causal relationships and common pathogeneses between them are uncertain. Furthermore, they are uncurable. Thus, we aimed to identify the causal relationships and novel therapeutic targets shared between them based on their common pathophysiological mechanisms in gut–brain-axis (GBA).MethodsA meta-analysis on bidirectional Mendelian randomization (MR) utilizing various datasets was performed to estimate their causal relationship. Then, pleiotropic analysis under the composite null hypothesis (PLACO) with functional mapping combined with annotation of genetic associations (FUMA) analysis were conducted to identify pleiotropic genes. Next, blood, brain and intestine expression quantitative trait locus (eQTL) were taken to perform drug-target MR finding common causal genes in two diseases. Colocalization analysis ensured the eQTLs of corresponding gene colocalized with disease. Enrichment analysis and protein‒protein interaction (PPI) network were done to explore common pathogenesis pathways. Genes passed all analysis were regarded as drug targets.ResultsOur MR meta-analysis revealed the bidirectional causal relationship between diseases, with combined ORs for PD on IBD, CD, UC (1.050 [95% CI 1.014–1.086], 1.044 [95% CI 0.995–1.095], 1.063 [95% CI 1.016–1.120]); for IBD, CD, UC on PD (1.003 [95% CI 0.973–1.034], 1.035 [95% CI 1.004–1.067], 1.008 [95% CI 0.977–1.040]). Overall, 277, 216 and 201 genes were identified as pleiotropic genes between PD and IBD, CD, UC. Total of 733 genes were classified as tier 3 (found in only one tissue) druggable targets, 57 as tier 2 (found in two tissues, 51 protein-coding genes) and 9 as tier 3 (found in three tissues). Among 60 protein-coding druggable targets over tier 2, 18 overlapped with pleiotropic genes and enriched in mitochondria, antigen presentation, processing and immune cell regulation pathways. Three druggable genes (LRRK2, RAB29 and HLA-DQA2) passed colocalization analysis. LRRK2 and RAB29 were reported to be pleiotropic genes, and RAB29 and HLA-DQA2 were reported for the first time as potential drug targets.ConclusionsThis study established a reliable causal relationship, possible shared drug targets and common pathogenesis pathways of two diseases, which had important implications for intervention and treatment of two diseases simultaneously.