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Fish and Shellfish Immunology Reports Dec 2023We report the proteomic profile of Epidermal Mucus (EM) from and identified the differentially abundant proteins (DAPs) against infection through label-free liquid...
We report the proteomic profile of Epidermal Mucus (EM) from and identified the differentially abundant proteins (DAPs) against infection through label-free liquid chromatography-mass spectrometry (LC-MS/MS). Using discovery-based proteomics, a total of 2039 proteins were quantified in nontreated group and 1,328 proteins in the treated group, of which 114 were identified as DAPs in both the groups. Of the 114 DAPs, 68 proteins were upregulated and 46 proteins were downregulated in the treated group compared to nontreated group. Functional annotations of these DAPs shows their association with metabolism, cellular process, molecular process, cytoskeletal, stress, and particularly immune system. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis and Fisher's exact test between the two groups shows that most of the proteins were immune-related, which were significantly associated with the proteasome, phagosome, and infection pathways. Overall, this study shows a basic and primary way for further functional research of the involvement of vitellogenin 2, alpha-2-macroglobulin-like protein, toll-like receptors (TLR-13), calpain, keratin-like proteins, and heat shock proteins against bacterial infection. Nonetheless, this first-ever comprehensive report of a proteomic sketch of EM from after infection provides systematic protein information to broadly understand the biological role of fish EM against bacterial infection.
PubMed: 37771818
DOI: 10.1016/j.fsirep.2023.100115 -
Journal of Cystic Fibrosis : Official... Nov 2023Mucus stasis, a hallmark of muco-obstructive disease, results from impaired mucociliary transport and leads to lung function decline and chronic infection. Although...
BACKGROUND
Mucus stasis, a hallmark of muco-obstructive disease, results from impaired mucociliary transport and leads to lung function decline and chronic infection. Although therapeutics that target mucus stasis in the airway, such as hypertonic saline or rhDNAse, show some therapeutic benefit, they do not address the underlying electrostatic defect apparent in mucins in CF and related conditions. We have previously shown poly (acetyl, arginyl) glucosamine (PAAG, developed as SNSP113), a soluble, cationic polymer, significantly improves mucociliary transport in a rat model of CF by normalizing the charge defects of CF mucin. Here, we report efficacy in the CFTR-sufficient, ENaC hyperactive, Scnn1b-Tg mouse model that develops airway muco-obstruction due to sodium hyperabsorption and airway dehydration.
METHODS
Scnn1b-Tg mice were treated with either 250 µg/mL SNSP113 or vehicle control (1.38% glycerol in PBS) via nebulization once daily for 7 days and then euthanized for analysis. Micro-Optical Coherence Tomography-based evaluation of excised mouse trachea was used to determine the effect on the functional microanatomy. Tissue analysis was performed by routine histopathology.
RESULTS
Nebulized treatment of SNSP113 significantly improved mucociliary transport in the airways of Scnn1b-Tg mice, without altering the airway surface or periciliary liquid layer. In addition, SNSP113 significantly reversed epithelial hypertrophy and goblet cell metaplasia. Finally, SNSP113 significantly ameliorated eosinophilic crystalline pneumonia and lung consolidation in addition to inflammatory macrophage influx in this model.
CONCLUSION
Overall, this study extends the efficacy of SNSP113 as a potential therapeutic to alleviate mucus stasis in muco-obstructive diseases in CF and potentially in related conditions.
Topics: Female; Pregnancy; Mice; Animals; Rats; Mucociliary Clearance; Cystic Fibrosis; Pregnancy-Associated alpha 2-Macroglobulins; Mice, Transgenic; Disease Models, Animal; Airway Obstruction; Mice, Inbred CFTR; Lung; Epithelial Sodium Channels
PubMed: 37714777
DOI: 10.1016/j.jcf.2023.08.011 -
Biomeditsinskaia Khimiia Sep 2023The universal proteinase inhibitor α2-macroglobulin (α₂-MG) exhibiting antiviral and immunomodulatory activities, is considered as an important participant in the...
The universal proteinase inhibitor α2-macroglobulin (α₂-MG) exhibiting antiviral and immunomodulatory activities, is considered as an important participant in the infectious process. The activity of α₂-MG in the new coronavirus infection and post-covid syndrome (long COVID) has not been studied yet. We examined 85 patients diagnosed with community-acquired bilateral polysegmental pneumonia developed under conditions of a new coronavirus infection SARS-CoV-2. For assessment of the post-COVID period, 60 patients were examined 5.0±3.6 months after the coronavirus infection. Among these patients, 40 people had complications, manifested in the form of neurological, cardiological, gastroenterological, dermatological, bronchopulmonary symptoms. The control group included 30 conditionally healthy individuals with a negative PCR result for SARS-CoV-2 RNA and lack of antibodies to the SARS-CoV-2 virus. The α₂-MG activity in serum samples of patients with coronavirus infection dramatically decreased, up to 2.5% of the physiological level. This was accompanied by an increase in the activity of the α₁-proteinase inhibitor, elastase- and trypsin-like proteinases by 2.0-, 4.4- and 2.6-fold respectively as compared with these parameters in conditionally healthy individuals of the control. In the post-COVID period, despite the trend towards normalization of the activity of inhibitors, the activity of elastase-like and especially trypsin-like proteinases in serum remained elevated. In overweight individuals, the increase in the activity of trypsin-like proteinases was most pronounced and correlated with an increase in the antibody titer to the SARS-CoV-2 virus. In the post-COVID period, the α₂-MG activity not only normalized, but also exceeded the control level, especially in patients with dermatological and neurological symptoms. In patients with neurological symptoms or with dermatological symptoms, the α₂-MG activity was 1.3 times and 2.1 times higher than in asymptomatic persons. Low α₂-MG activity in the post-COVID period persisted in overweight individuals. The results obtained can be used to monitor the course of the post-COVID period and identify risk groups for complications.
Topics: Humans; COVID-19; Macroglobulins; Overweight; Pancreatic Elastase; Peptide Hydrolases; Post-Acute COVID-19 Syndrome; RNA, Viral; SARS-CoV-2; Trypsin
PubMed: 37705485
DOI: 10.18097/PBMC20236904240 -
PloS One 2023Extracellular vesicles (EVs) contain a variety of biomolecules and provide information about the cells that produce them. EVs from cancer cells found in urine can be...
Extracellular vesicles (EVs) contain a variety of biomolecules and provide information about the cells that produce them. EVs from cancer cells found in urine can be used as biomarkers to detect cancer, enabling early diagnosis and treatment. The potential of alpha-2-macroglobulin (A2M) and clusterin (CLU) as novel diagnostic urinary EV (uEV) biomarkers for bladder cancer (BC) was demonstrated previously. To validate the diagnostic value of these proteins in uEVs in a large BC cohort, urine handling conditions before uEV isolation should be optimized during sample transportation from medical centers. In this study, we analyzed the uEV protein quantity, EV particle number, and uEV-A2M/CLU after urine storage at 20°C and 4°C for 0-6 days, each. A2M and CLU levels in uEVs were relatively stable when stored at 4°C for a maximum of three days and at 20°C for up to 24 h, with minimal impact on analysis results. Interestingly, pre-processing to remove debris and cells by centrifugation and filtration of urine did not show any beneficial effects on the preservation of protein biomarkers of uEVs during storage. Here, the importance of optimizing shipping conditions to minimize the impact of pre-analytical handling on the uEVs protein biomarkers was emphasized. These findings provide insights for the development of clinical protocols that use uEVs for diagnostic purposes.
Topics: Humans; Pregnancy; Female; Urinary Bladder Neoplasms; Urinary Bladder; Body Fluids; Extracellular Vesicles; Transcription Factors; Pregnancy-Associated alpha 2-Macroglobulins
PubMed: 37676879
DOI: 10.1371/journal.pone.0291198 -
Journal of Biomolecular Structure &... Aug 2023Under simulated physiological conditions, this study investigates the interaction between nutraceutical phycocyanobilin (PCB) and the universal anti-protease protein...
Under simulated physiological conditions, this study investigates the interaction between nutraceutical phycocyanobilin (PCB) and the universal anti-protease protein human alpha-2-macroglobulin (αM). Extensive molecular docking analyses on multiple α2M conformations, spectroscopic techniques, and α2M activity assays were utilized to examine the complex formation. The results revealed that for every protein conformation, two high energy binding sites exist: the first, conformationally independent, at the interface region between two monomer chains and the second, conformationally dependent, in the pocket composed of amino acids from four distinct domains (TED, RBD, CUB, and MG2) of the single protein chain. Spectrofluorimetric measurements indicated a moderate affinity between αM and PCB with a moderately high binding constant of 6.3 10 M at 25 °C. The binding of PCB to αM resulted in minor changes in the secondary structure content of αM. Furthermore, PCB protected αM from oxidation and preserved its anti-protease activity in the oxidative environment. These findings suggest that PCB binding could indirectly impact the body's response to oxidative stress by influencing αM's role in controlling enzyme activity during the inflammatory process.Communicated by Ramaswamy H. Sarma.
PubMed: 37592733
DOI: 10.1080/07391102.2023.2248273 -
Blood Coagulation & Fibrinolysis : An... Oct 2023The fibrinolytic system plays an important role in controlling blood coagulation at each stage, from thrombin generation to fibrin clot cleavage. Currently, long-term...
The fibrinolytic system plays an important role in controlling blood coagulation at each stage, from thrombin generation to fibrin clot cleavage. Currently, long-term multiorgan dysfunction post-coronavirus disease 2019 (COVID-19) may include coagulation disorders. Little information is available about the potential causes of post-COVID-19 coagulopathy, but one of them may be subpopulation IgG produced by the immune system against SARS-CoV-2. This article describes the changes in the main parameters of the fibrinolytic system in donors with various titers of anti-SARS-CoV-2 IgG, which is part of a complex study of the hemostasis system in these donor groups. We determined the most significant parameters of the fibrinolytic system, such as potential activity and amount of plasminogen and tissue plasminogen activator (tPA), amount of plasminogen activator inhibitor-1 (PAI-1), inhibitory potentials of α-2-antiplasmin, α-1-antitrypsin, α-2-macroglobulin in the blood plasma of donor groups. The obtained results represent the maximum and minimum values of measurement parameters among donor groups with titers of anti-SARS-CoV-2 IgG at least 10 ± 3 Index (S/C), and their statistical differences from the reference point [donor group with titer of anti-SARS-CoV-2 IgG 0 Index (S/C)]. We established the changes in fibrinolytic parameters depending on the titers of anti-SARS-CoV-2 IgG. One conclusion can be drawn from this: anti-SARS-CoV-2 IgG population may influence coagulation in the post-COVID-19 period. Further research in-vitro and in-vivo experimental models using selected and purified IgG may confirm our previous findings.
Topics: Humans; Antibodies, Viral; COVID-19; Fibrinolysis; Immunoglobulin G; SARS-CoV-2; Tissue Plasminogen Activator; Blood Coagulation
PubMed: 37577922
DOI: 10.1097/MBC.0000000000001248 -
Clinical & Translational Immunology 2023There is an increasing appreciation for the need to study mucosal antibody responses in humans. Our aim was to determine the utility of different types of samples from...
OBJECTIVES
There is an increasing appreciation for the need to study mucosal antibody responses in humans. Our aim was to determine the utility of different types of samples from the human respiratory tract, specifically nasopharyngeal (NP) swabs obtained for diagnostic purposes and bronchoalveolar lavage (BAL) obtained in outpatient and inpatient settings.
METHODS
We analysed antibody levels in plasma and NP swabs from 67 individuals with acute influenza as well as plasma and BAL from individuals undergoing bronchoscopy, including five control subjects as well as seven moderately and seven severely ill subjects with a respiratory viral infection. Levels of α2-macroglobulin were determined in BAL and plasma to assess plasma exudation.
RESULTS
IgG and IgA were readily detectable in BAL and NP swabs, albeit at different ratios, while IgM levels were low. The total amount of antibody recovered from NP swabs varied greatly between study participants. Accordingly, the levels of influenza HA-specific antibodies varied, and individuals with lower amounts of total Ig in NP swabs had undetectable levels of HA-specific Ig. Similarly, the total amount of antibody recovered from BAL varied between study participants. However, severely ill patients showed evidence of increased plasma exudation, which may confound analysis of their BAL samples for mucosal antibodies.
CONCLUSION
Nasopharyngeal swabs collected for diagnostic purposes may have utility in assessing antibodies from the human nasal mucosa, but variability in sampling should be accounted for. BAL samples can be utilised to study antibodies from the lower respiratory tract, but the possibility of plasma exudation should be excluded.
PubMed: 37564999
DOI: 10.1002/cti2.1460 -
Biochimie Mar 2024Chandipura Virus is an emerging tropical pathogen with a high mortality rate among children. No mode of treatment or antivirals exists against CHPV infection, due to...
Chandipura Virus is an emerging tropical pathogen with a high mortality rate among children. No mode of treatment or antivirals exists against CHPV infection, due to little information regarding its host interaction. Studying viral pathogen interaction with its host can not only provide valuable information regarding its propagation strategy, but also on which host proteins interact with the virus. Identifying these proteins and understanding their role in the infection process can provide more stable anti-viral targets. In this study, we focused on identifying host factors that interact with CHPV and may play a critical role in CHPV infection. We are the first to report the successful identification of Alpha-2-Macroglobulin (A2M), a secretory protein of the host that interacts with CHPV. We also established that LRP1 (Low-density lipoprotein receptor-related protein 1) and GRP78 (Glucose regulated protein 78), receptors of A2M, also interact with CHPV. Furthermore, we could also demonstrate that knocking out A2M has a severe effect on viral infection. We conclusively show the interaction of these host proteins with CHPV. Our findings also indicate that these host proteins could play a role in viral entry into the host cell.
Topics: Child; Humans; Vesiculovirus; Transcription Factors; Macroglobulins; Low Density Lipoprotein Receptor-Related Protein-1
PubMed: 37517577
DOI: 10.1016/j.biochi.2023.07.019 -
Journal of Biomolecular Structure &... Jul 2023In the present study, we investigated the interaction of alpha-2-macroglobulin (α2M) with naringenin using multi-spectroscopic, molecular docking, and molecular...
In the present study, we investigated the interaction of alpha-2-macroglobulin (α2M) with naringenin using multi-spectroscopic, molecular docking, and molecular simulation approaches to identify the functional changes and structural variations in the α2M structure. Our study suggests that naringenin compromised α2M anti-proteinase activity. The results of absorption spectroscopy and fluorescence measurement showed that naringenin-α2M formed a complex with a binding constant of (k)∼10, indicative of moderate binding. The value of ΔG° in the binding indicates the process to be spontaneous and the major force responsible to be hydrophobic interaction. The findings of FRET reveal the binding distance between naringenin and the amino acids of α2M was 2.82 nm. The secondary structural analysis of α2M with naringenin using multi-spectroscopic methods like synchronous fluorescence, red-edge excitation shift (REES), FTIR, and CD spectra further confirmed the significant conformational alterations in the protein. Molecular docking approach reveals the interactions between naringenin and α2M to be hydrogen bonds, van der Waals forces, and pi interactions, which considerably favour and stabilise the binding. Molecular dynamics modelling simulations also supported the steady binding with the least RMSD deviations. Our study suggests that naringenin interacts with α2M to alter its confirmation and compromise its activity.Communicated by Ramaswamy H. Sarma.
PubMed: 37498152
DOI: 10.1080/07391102.2023.2240420 -
Biochemical Pharmacology Sep 2023The brain-derived neurotrophic factor (BDNF) has been recently shown to have activating effects in isolated platelets. However, BDNF circulates in plasma and a mechanism...
The brain-derived neurotrophic factor (BDNF) has been recently shown to have activating effects in isolated platelets. However, BDNF circulates in plasma and a mechanism to preclude constant activation of platelets appears necessary. Hence, we investigated the mechanism regulating BDNF bioavailability in blood. Protein-protein interactions were predicted by molecular docking and validated through immunoprecipitation. Platelet aggregation was assessed using light transmission aggregometry with washed platelets in response to classical agonists or BDNF, in the absence or presence of alpha-2-macroglobulin (αM), and in platelet-rich plasma. BDNF signaling was assessed with phospho-blots. As little as 25% autologous plasma was sufficient to completely abolish platelet aggregation in response to BDNF. Docking predicted two forms of BDNF binding to native or activated αM, in parallel and perpendicular arrangements, and the model suggested that the BDNF-αM complex cannot bind to the high-affinity BDNF receptor, tropomyosin receptor kinase B (TrkB). Experimentally, native and activated αM formed stable complexes with BDNF preventing BDNF-induced TrkB activation and signal transduction. Both native and activated αM inhibited BDNF induced-platelet aggregation in a concentration-dependent manner with comparable half-maximal inhibitory concentrations (IC≈ 125-150 nM). Our study implicates αM as a physiological regulator of BDNF bioavailability, and as an inhibitor of BDNF-induced platelet activation in blood.
Topics: Female; Pregnancy; Humans; Brain-Derived Neurotrophic Factor; Platelet Aggregation; Pregnancy-Associated alpha 2-Macroglobulins; Molecular Docking Simulation; Receptor, trkB; Enzyme Inhibitors
PubMed: 37487878
DOI: 10.1016/j.bcp.2023.115701