Changes In Protein Profiles Research Articles

Preventing microbe colonization on avocado (Persea nubigena var. guatemalensis) through metabiotic treatment, a promising postharvest safety improvement.

Lactic acid bacteria (LAB) produce various metabolites (i.e. metabiotics) with inhibitory capacity towards harmful foodborne pathogens. This study aimed to design several antimicrobial formulations based on metabiotics obtained from different native LAB species (Lactobacillus pentosus UTNGt5, Lactococcus lactis UTNGt28, and Weissella cibaria UTNGt21O) and to detect the possible mode of action towards two multidrug resistant Staphylococcus spp. strains isolated from avocado (Persea nubigena var. guatemalensis) fruits. Additionally, the formulation with the highest inhibitory activity was tested ex vitro on avocados at the immature (firm) ripeness stage to evaluate their effect on microorganisms' growth and fruit quality attributes post-harvest. Out of the top five formulations showing the highest bactericidal effect in vitro at their minimum inhibitory concentration (1 x MIC) on both Staphylococcus spp. targets one candidate annotated P11 (consisting of UTNGt21O and UTNGt28; 1:3, v/v) was selected. Co-cultivation of Staphylococcus strains with P11 formulation results in cell viability reduction by 98%, by impairing the integrity of the cell membrane inducing cytoplasm molecule content leakage, protein profile changes, and finally bacterial death. Even though the total coliforms, Staphylococcus spp., Enterobacter spp., molds, and yeasts counts were not fully eliminated by day 13 of storage, a statistically significant reduction (p < 0.05) in viable cell counts were observed by day 8 upon the P11 treatment compared with non-treated control (C) and treated with a commercial disinfectant (T1) samples, suggesting that P11 formulation inhibited microbial colonization during storage. Likewise, no visible dark spots were observed on the mesocarp (pulp) upon the treatment with P11, whereas T1 and C fruits showed greater dark spots on the pulp as indicative of damage. The quality attributes, such as pH, total soluble solids, total titratable acidity, antioxidant capacity, and total polyphenol content, were not affected by the treatment. Principal Component Analysis (PCA) conducted on these five variables showed a clear separation of samples according to the maturity stage regardless of the treatment. These results suggest that the active metabolites from LAB strains might create a barrier between the exocarp and mesocarp, inhibiting the microorganisms colonization, reducing fruit damage, and lengthening the fruit quality and safety after harvest.

Comprehensive analysis of systemic, metabolic, and molecular changes following prospective change to low-carbohydrate diet in adults with type 2 diabetes mellitus in India.

South Asians, especially Indians, face higher diabetes-related risks despite lower body mass index (BMI) compared with the White population. Limited research connects low-carbohydrate high-fat (LCHF)/ketogenic diets to metabolic changes in this group. Systematic studies are needed to assess the long-term effects of the diet, such as ocular health. In this prospective, observational study, 465 candidates aged 25-75 years with type 2 diabetes included with institutional ethics approval. A total of 119 subjects were included in the final study assessment based on the availability of pathophysiological reports, tears, and blood samples collected at baseline, 3rd, and 6th months. Serum and tear samples were analyzed by an enzyme-linked lectinsorbent assay, to examine secreted soluble protein biomarkers, such as IL-1β (interleukin 1 Beta), IL-6 (interleukin 6), IL-10 (interleukin 10), IL-17A (interleukin 17A), MMP-9 (matrix metalloproteinase 9), ICAM-1 (intercellular adhesion molecule 1), VEGF-A (vascular endothelial growth factor A), and TNF-α (tumor necrosis factor-alpha). A Wilcoxon test was performed for paired samples. Spearman's correlation was applied to test the strength and direction of the association between tear biomarkers and HbA1c. p-value of < 0.05 was considered significant. After a 3- and 6-month LCHF intervention, fasting blood sugar decreased by 10% (Δ: -14 mg/dL; p < 0.0001) and 7% (Δ: -8 mg/dL; p < 0.0001), respectively. Glycated hemoglobin A1c levels decreased by 13% (Δ: -1%; p < 0.0001) and 9% (Δ: -0.6%; p < 0.0001). Triglycerides reduced by 22% (Δ: -27 mg/dL; p < 0.0001) and 14% (Δ: -19 mg/dL; p < 0.0001). Total cholesterol reduced by 5.4% (Δ: -10.5 mg/dL; p < 0.003) and 4% (Δ: -7 mg/dL; p < 0.03), while low-density lipoprotein decreased by 10% (Δ: -11.5 mg/dL; p < 0.003) and 9% (Δ: -11 mg/dL; p < 0.002). High-density lipoprotein increased by 11% (Δ: 5 mg/dL; p < 0.0001) and 17% (Δ: 8 mg/dL; p < 0.0001). At the first follow-up, tear proteins such as ICAM-1, IL-17A, and TNF-α decreased by 30% (Δ: -2,739 pg/mL; p < 0.01), 22% (Δ: -4.5 pg/mL; p < 0.02), and 34% (Δ: -0.9 pg/mL; p < 0.002), respectively. At the second follow-up, IL-1β and TNF-α reduced by 41% (Δ: -2.4 pg/mL; p < 0.05) and 34% (Δ: -0.67 pg/mL; p < 0.02). Spearman's correlation between HbA1c and tear analytes was not statistically significant. The LCHF diet reduces the risk of hyperglycemia and dyslipidemia. Changes in tear fluid protein profiles were observed, but identifying promising candidate biomarkers requires validation in a larger cohort.

Growth, ultrastructural and physiological characteristics of Abelmoschus cytotypes under elevated ozone stress: a study on ploidy-specific responses.

Tropospheric ozone (O3 ) is a significant abiotic stressor whose rising concentration negatively influences plant growth. Studies related to the differential response of Abelmoschus cytotypes to elevated O3 treatment are scarce and need further exploration to recognise the role of polyploidisation in stress tolerance. In this study, we analysed the changes in growth pattern, ultrastructure, physiology and foliar protein profile occurring under O3 stress in Abelmoschus moschatus (monoploid), Abelmoschus esculentus (diploid) and Abelmoschus caillei (triploid). Our findings showed that higher stomatal conductance in A. moschatus triggered higher O3 intake, causing damage to stomatal cells and photosynthetic pigments. Additionally, it caused a reduction in photosynthetic rates, leading to reduced plant growth, total biomass and economic yield. This O3 -induced toxicity was less in diploid and triploid cytotypes of Abelmoschus . Protein profiling by sodium dodecyl sulpate-polyacrylamide gel electrophoresis showed a significant decrease in the commonly found RuBisCO larger and smaller subunits. The decrease was more prominent in monoploid compared to diploid and triploid. This study provides crucial data for research that aim to enhance plant ability to withstand O3 induced oxidative stress. Our findings may help in developing a tolerant variety through plant breeding techniques, which will be economically more advantageous in reaching the objective of sustainable production at the high O3 levels projected under a climate change scenario.

Urine metabolomics phenotyping and urinary biomarker exploratory in mild cognitive impairment and Alzheimer's disease.

Alzheimer's disease is a prevalent disease with a heavy global burden and is suggested to be a metabolic disease in the brain in recent years. The metabolome is considered to be the most promising phenotype which reflects changes in genetic, transcript, and protein profiles as well as environmental effects. Aiming to obtain a comprehensive understanding and convenient diagnosis of MCI and AD from another perspective, researchers are working on AD metabolomics. Urine is more convenient which could reflect the change of disease at an earlier stage. Thus, we conducted a cross-sectional study to investigate novel diagnostic panels. We first enrolled participants from China-Japan Friendship Hospital from April 2022 to November 2022, collected urine samples and conducted an LC-MS/MS analysis. In parallel, clinical data were collected and clinical examinations were performed. After statistical and bioinformatics analyzes, significant risk factors and differential urinary metabolites were determined. We attempt to investigate diagnostic panels based on machine learning including LASSO and SVM. Fifty-seven AD patients, 43 MCI patients and 62 CN subjects were enrolled. A total of 2,140 metabolites were identified among which 125 significantly differed between the AD and CN groups, including 46 upregulated ones and 79 downregulated ones. In parallel, there were 93 significant differential metabolites between the MCI and CN groups, including 23 upregulated ones and 70 downregulated ones. AD diagnostic panel (30 metabolites+ age + APOE) achieved an AUC of 0.9575 in the test set while MCI diagnostic panel (45 metabolites+ age + APOE) achieved an AUC of 0.7333 in the test set. Atropine, S-Methyl-L-cysteine-S-oxide, D-Mannose 6-phosphate (M6P), Spiculisporic Acid, N-Acetyl-L-methionine, 13,14-dihydro-15-keto-tetranor Prostaglandin D2, Pyridoxal 5'-Phosphate (PLP) and 17(S)-HpDHA were considered valuable for both AD and MCI diagnosis and defined as hub metabolites. Besides, diagnostic metabolites were weakly correlated with cognitive functions. In conclusion, the procedure is convenient, non-invasive, and useful for diagnosis, which could assist physicians in differentiating AD and MCI from CN. Atropine, M6P and PLP were evidence-based hub metabolites in AD.

A systemic approach to identify non-abundant immunogenic proteins in Lyme disease pathogens.

Borrelia burgdorferi, the pathogen of Lyme disease, differentially produces many outer surface proteins (Osp), some of which represent the most abundant membrane proteins, such as OspA, OspB, and OspC. In cultured bacteria, these proteins can account for a substantial fraction of the total cellular or membrane proteins, posing challenges to the identification and analysis of non-abundant proteins, which could serve as novel pathogen detection markers or as vaccine candidates. Herein, we introduced serial mutations to remove these abundant Osps and generated a B. burgdorferi mutant deficient in OspA, OspB, and OspC in an infectious 297-isolate background, designated as OspABC- mutant. Compared to parental isolate, the mutant did not reflect growth defects in the cultured medium but showed differential mRNA expression of representative tested genes, in addition to gross changes in cellular and membrane protein profiles. The analysis of differentially detectable protein contents of the OspABC- mutant, as compared to the wild type, by two-dimensional gel electrophoresis followed by liquid chromatography-mass spectrometry, identified several spirochete proteins that are dominated by proteins of unknown functions, as well as membrane transporters, chaperons, and metabolic enzymes. We produced recombinant forms of two of these represented proteins, BBA34 and BB0238, and showed that these proteins are detectable during spirochete infection in the tick-borne murine model of Lyme borreliosis and thus serve as potential antigenic markers of the infection.IMPORTANCEThe present manuscript employed a systemic approach to identify non-abundant proteins in cultured Borrelia burgdorferi that are otherwise masked or hidden due to the overwhelming presence of abundant Osps like OspA, OspB, and OspC. As these Osps are either absent or transiently expressed in mammals, we performed a proof-of-concept study in which their removal allowed the analysis of otherwise less abundant antigens in OspABC-deficient mutants and identified several immunogenic proteins, including BBA34 and BB0238. These antigens could serve as novel vaccine candidates and/or genetic markers of Lyme borreliosis, promoting new research in the clinical diagnosis and prevention of Lyme disease.

Comparative proteomic analysis of the protein profile in the cuttlefish Sepia pharaonis associated with skin ulceration syndrome

Skin ulceration syndrome (SUS) is becoming a severe problem in the breeding and culturing process of the cuttlefish Sepia pharaonis. However, limited knowledge is available about the occurrence of this devastating disease. In this study, proteomic analysis was used to identify the differentially expressed proteins (DEPs) and the biological pathways enriched in SUS-diseased S. pharaonis. Both the healthy group and diseased group were analyzed in triplicate, with 4 cuttlefish in each replicate. The results showed that 85 DEPs were identified between the two groups, including 36 upregulated proteins and 49 downregulated proteins in the diseased group compared to the healthy group. GO enrichment analysis revealed that the DEPs were mainly enriched in cellular component organization or biogenesis, nucleus and ion binding processes. The results of the KEGG pathway analysis indicated that extracellular matrix (ECM)-receptor interaction was the most enriched upregulated pathway. Real-time reverse transcriptase PCR was used to identify the expression of two differentially expressed matrix metalloproteinases (MMPs), and the results showed that the mRNA expression of MMP14 and MMP19 was significantly upregulated in the skin tissue of the diseased group. Furthermore, the protease activity of the diseased group was higher than that of the healthy group. Our results offer basic knowledge on the changes in protein profiles during the occurrence of SUS in the cuttlefish S. pharaonis.

Effects of processing on component interactions and peptide profiles of preterm infant formula based on peptidomics by Q Exactive plus analysis

The study aimed to investigate the effects of processing on the component interactions, the peptide profile as well as the bioactive peptides of preterm infant formula after digestion through EASY-nLC1200 Q Exactive Plus system. Results indicated that whey proteins and caseins transferred from serum and pellet fractions to fat fraction to recombine the milk fat globule membrane destroyed after homogenization. Protein interactions caused the changes of nitrogen distribution, protein profiles, and morphology after pasteurization and spray drying. Peptidomics analysis demonstrated that the component interactions of preterm infant formula affected the peptide profiles after digestion shown by the different peptide intensities of certain amino acid compositions and the totally different abundance of the peptides in heatmap. The hydrolysates of five groups exhibited 334, 274, 421, 214, and 265 unique peptides belonging to 144, 116, 185, 101, and 231 master accession proteins respectively. The top 8 abundant proteins identified were β-casein, Lipocln, αS1-casein, αS2-casein, κ-casein, bovine Glycosylation-dependent cell adhesion molecule, α-lactalbumin, and Albumin. The number of bioactive peptides increased after processing but were different from each other. In total, component interactions during processing of preterm infant formula could change the peptide profile and bioactive peptides after in vitro digestion.

Fibrinogen in mice cerebral microvessels induces blood–brain barrier dysregulation with aging via a dynamin-related protein 1–dependent pathway

We previously reported evidence that oxidative stress during aging leads to adverse protein profile changes of brain cortical microvessels (MVs: end arterioles, capillaries, and venules) that affect mRNA/protein stability, basement membrane integrity, and ATP synthesis capacity in mice. As an extension of our previous study, we also found that proteins which comprise the blood–brain barrier (BBB) and regulate mitochondrial quality control were also significantly decreased in the mice’s cortical MVs with aging. Interestingly, the neuroinflammatory protein fibrinogen (Fgn) was increased in mice brain MVs, which corresponds with clinical reports indicating that the plasma Fgn concentration increased progressively with aging. In this study, protein–protein interaction network analysis indicated that high expression of Fgn is linked with downregulated expression of both BBB- and mitochondrial fission/fusion–related proteins in mice cortical MVs with aging. To investigate the mechanism of Fgn action, we observed that 2 mg/mL or higher concentration of human plasma Fgn changed cell morphology, induced cytotoxicity, and increased BBB permeability in primary human brain microvascular endothelial cells (HBMECs). The BBB tight junction proteins were significantly decreased with increasing concentration of human plasma Fgn in primary HBMECs. Similarly, the expression of phosphorylated dynamin-related protein 1 (pDRP1) and other mitochondrial fission/fusion–related proteins were also significantly reduced in Fgn-treated HBMECs. Interestingly, DRP1 knockdown by shRNA(h) resulted in the reduction of both BBB- and mitochondrial fission/fusion–related proteins in HBMECs. Our results suggest that elevated Fgn downregulates DRP1, leading to mitochondrial-dependent endothelial and BBB dysfunction in the brain microvasculature.

Deciphering neuronal deficit and protein profile changes in human brain organoids from patients with creatine transporter deficiency.

Creatine transporter deficiency (CTD) is an X-linked disease caused by mutations in the SLC6A8 gene. The impaired creatine uptake in the brain results in intellectual disability, behavioral disorders, language delay, and seizures. In this work, we generated human brain organoids from induced pluripotent stem cells of healthy subjects and CTD patients. Brain organoids from CTD donors had reduced creatine uptake compared with those from healthy donors. The expression of neural progenitor cell markers SOX2 and PAX6 was reduced in CTD-derived organoids, while GSK3β, a key regulator of neurogenesis, was up-regulated. Shotgun proteomics combined with integrative bioinformatic and statistical analysis identified changes in the abundance of proteins associated with intellectual disability, epilepsy, and autism. Re-establishment of the expression of a functional SLC6A8 in CTD-derived organoids restored creatine uptake and normalized the expression of SOX2, GSK3β, and other key proteins associated with clinical features of CTD patients. Our brain organoid model opens new avenues for further characterizing the CTD pathophysiology and supports the concept that reinstating creatine levels in patients with CTD could result in therapeutic efficacy.

Deciphering neuronal deficit and protein profile changes in human brain organoids from patients with creatine transporter deficiency

Creatine transporter deficiency (CTD) is an X-linked disease caused by mutations in the SLC6A8 gene. The impaired creatine uptake in the brain results in intellectual disability, behavioral disorders, language delay, and seizures. In this work, we generated human brain organoids from induced pluripotent stem cells of healthy subjects and CTD patients. Brain organoids from CTD donors had reduced creatine uptake compared with those from healthy donors. The expression of neural progenitor cell markers SOX2 and PAX6 was reduced in CTD-derived organoids, while GSK3β, a key regulator of neurogenesis, was up-regulated. Shotgun proteomics combined with integrative bioinformatic and statistical analysis identified changes in the abundance of proteins associated with intellectual disability, epilepsy, and autism. Re-establishment of the expression of a functional SLC6A8 in CTD-derived organoids restored creatine uptake and normalized the expression of SOX2, GSK3β, and other key proteins associated with clinical features of CTD patients. Our brain organoid model opens new avenues for further characterizing the CTD pathophysiology and supports the concept that reinstating creatine levels in patients with CTD could result in therapeutic efficacy.

Thiamine-induced resistance in carnation against Fusarium oxysporum f. sp dianthi and mode of action studies based on the proteomics analysis of root tissue

Colombia is the main producer of carnations worldwide. However, its production is affected by vascular wilting caused by Fusarium oxysporum f. sp dianthi (Fod). Previous studies have proposed thiamine as a potential inducer of resistance against this pathogen, but the biochemical mechanisms involved in its action have not been elucidated. The objective of the present work was to investigate the underlying mechanism of thiamine–induced resistance by analyzing changes in the root protein profile. A greenhouse experiment was performed with two cultivars, evaluating the following treatments: control, Fod–inoculated plants, foliar application of thiamine, and application of thiamine and subsequent Fod inoculation. Incidence and severity were determined for each treatment. Applicating thiamine reduced the incidence (22%) and severity (62%) of vascular wilting in a susceptible cultivar confirming the effect as Resistance Inducer. In addition, a shotgun (nLC–MS/MS) proteomic analysis of root tissues was carried out at 96 h post–inoculation. The application of thiamine and subsequent inoculation with Fod, increased the abundance of 44 proteins in the susceptible cultivar and 70 in the resistant one. These proteins belonged to different functional groups, including protein synthesis and degradation, redox homeostasis, transport, cell organization, stress, and defense. In conclusion, the results obtained suggest that the treatment with thiamine previously to Fod inoculation allows the early activation of defense mechanisms that protect the plant against the pathogen. These results increase the knowledge of thiamine use as a biostimulant agent in plants and its protein–level mode of action against pathogens.

Complement System Proteins in the Human Aqueous Humor and Their Association with Primary Open-Angle Glaucoma.

This study discovers the complement protein profile in the aqueous humor (AH) of human subjects and investigates its association with primary open-angle glaucoma (POAG) pathogenesis. Among the 32 complement proteins identified, 22 were highly abundant and detected in more than 50% of AH samples. The most predominant active complement proteins in the AH are C3, C4B, C4A, CFB, CFD, and C9. Additionally, the most prevalent complement regulators and receptors include CLU, SERPING1, F2, CFH, CFI, and VTN. Significant alterations in complement proteins were observed in individuals with POAG compared to those with cataracts. Specifically, complement protein F2 was upregulated, while C8G, C6, and CFH were downregulated in POAG samples. Stratification of the samples by race and sex revealed distinct alterations of complement proteins in patients with POAG. In the African American cohort, five complement proteins (C4A, C4B, F2, C7, and C3) were upregulated in POAG compared to cataract patients. In the Caucasian cohort, eight complement proteins (C3, SERPING1, CFI, CLU, CFHR1, C8G, C6, and CFH) were downregulated in the POAG samples compared to the cataract samples. Within the male cohort, three complement proteins (CLU, C6, and CFH) were downregulated in POAG patients compared to those with cataracts. Whereas, within the female cohort, two complement proteins (C4B and F2) were upregulated and one (C8G) downregulated in the POAG samples when compared to cataracts. Discerning these changes in the AH complement protein profile will assist in the development of tailored therapies to modulate the complement system for managing ocular disorders. These insights may also lead to novel biomarkers for diagnosing and monitoring disease progression.

Abstract PO-071: Pseudonormal morphology of adenoid cystic carcinoma cells subverts the antitumor reactivity of immune cells

Abstract Objectives: Adenoid cystic carcinoma (ACC) is a rare salivary gland cancer most frequently arising in the head and neck region. The ACC microenvironment exhibits low immunogenicity (low tumor infiltrating lymphocytes and dendritic cells). This scenario favours immune evasion and partially explains poor prognosis because of the activation of immune inhibitory proteins. Gipie is a hook-related protein, that is involved in protein trafficking. Gipie (HkRP3, encoded by CCDC88B) is associated with T cell maturation and inflammatory functions and target cell killing in natural killer (NK) cells. Gipie is present in ACC patient samples and cells. Therefore, we investigated proliferation, apoptosis, and morphologic changes caused by Gipie in oral and salivary gland cancer cells in presence of immune cells. Methods: We assayed UM-HACC-2A (adenoid cystic carcinoma), A-253 (mucoepidermoid carcinoma, MEC), SCC4, SCC9, SCC25, CAL27 (oral squamous cell carcinoma, OSCC), OKF6 (normal oral) cells cocultured with, NK-92 (natural killer), and Jurkat, Clone E6-1 (acute T cell leukemia) cells, to construct multiple salivary gland and oral cancer-immune cell co-culture models. Gipie silenced cancer cells (silencing maintained for 48 hours) in the cancer immune coculture models were compared with their unaltered subsets. Cancer-immune cells interactions were maintained for 16 hours. Overall three hundred thirty-six (336) co-culture models were used to determine apoptosis, immune activation, morphology, transmigration, and protein expression. Additionally, we performed targeted mass spectrometry analysis to identify changes in protein profiles. Results: ACC cells exhibited Pseudonormal morphology after interacting with immune cells in the 3D coculture model. Gipie-silenced ACC cells in coculture model transformed to “lymphoblast-like” morphology. Gipie-silenced ACC cells in co-culture model (n = 24) showed significant increase of apoptotic cells compared to unaltered ACC cells, lowering of T regulatory cells (FoxP3+/IL-2Rα+/CD25+) (n = 24), and increase of activated NK cells (NKp30+/IFN-γ+) (n = 24) with significantly higher release of granzyme (n = 12) and perforin (n = 12). Increased activation of immune cells (transmigration and fluorescence intensity) via 3D-z-stack images were observed in Gipie-silenced subsets (n = 24). MEC cells responses were significantly less compared to ACC in co-culture models followed by silencing of Gipie, while OSCC and OKF6 cell responses were negligible, under identical conditions. Conclusions: The presence of Gipie confers pseudonormal morphology to ACC cells thereby reducing immune attack. Thus, trafficking proteins may impact ACC cancer-immune cell interaction during conventional chemotherapy or monoclonal immunotherapy. Citation Format: Rajdeep Chakraborty, Charbel Darido, Giuseppe Palmisano, Arthur Chien, Aidan Tay, Thiri Zaw, Shoba Ranganathan. Pseudonormal morphology of adenoid cystic carcinoma cells subverts the antitumor reactivity of immune cells [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Innovating through Basic, Clinical, and Translational Research; 2023 Jul 7-8; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2023;29(18_Suppl):Abstract nr PO-071.

Peculiarities of the morphological composition, protein profile, and activity of aminotransferases in the blood of turkeys in the presence of keel “namins”

A timely blood test is essential to detect possible abnormalities in the body. Blood somewhat objectively reflects the state of the internal environment in which all metabolic processes take place. The blood test is one of the most informative laboratory diagnostic tools, which provides essential information about the functional state of various organs and systems of the body. The work aimed to investigate the changes in morphological parameters, protein profile, and activity of aminotransaminases in the blood of turkeys in the presence of keel “namins”. The research was carried out at “Indykat” Ltd. in the village Kadubivtsi of the Lviv region on turkeys of the Big-6 breed on two groups of turkeys, 10 birds each. The turkeys of the control group were clinically healthy, and those of the experimental group had visible clinical signs of “namin” on the keel. The results of the research showed that the number of erythrocytes in the blood of clinically healthy birds was 2.5 ± 0.19 T/l, leukocytes – 21.8 ± 1.36 G/l, hemoglobin content – 90.5 ± 2.54 g/l, and the hematocrit was equal to 29.4 ± 1.44 %. In the presence of “namins” in the blood of turkeys, the number of leukocytes and hematocrit, compared to the control, was higher by 15.2 and 3.8 %, respectively, and the number of erythrocytes and hemoglobin content was lower by 16.1 and 11.3 %. Analyzing the leukogram of the blood of the turkeys of the research group, the appearance of basophilic leukocytes and young neutrophils was established, the number of which was 1.1 ± 0.08 and 0.2 ± 0.01 %, respectively. Compared to the control, the number of rod-shaped and segmented neutrophils was higher by 1.3 and 6.3 %, monocytes by 16.3 %, and eosinophils and lymphocytes were lower by 1.3 and 16.8 % and amounted to 6, 3 ± 0.27 and 41.1 ± 2.41 %. The level of total protein in the blood of birds in the pectoral muscles that developed and formed “namins” was 26.4 ± 1.35 g/l and was 6.7 % lower than that of the experimental group. In the presence of “namins” on the keels of turkeys, a 4.4 % higher level of globulins, which perform a protective function, and a 17.1 % lower amount of albumins were found. The pathological process that developed in the pectoral muscles of turkeys led to an increase in the activities of alanine aminotransferase and aspartate aminotransferase in the blood of turkeys to 32.4 ± 1.23 and 314.2 ± 14.2 units/l, which was more significant than indicators of the control group by 8.6 and 8.3 %, respectively.

Non-thermal plasma radiation-induced changes in antibiotic susceptibility and protein profile of Staphylococcus aureus.

Plasma radiation is a widely used technique for sterilization or decontamination in various industries, as well as in some healthcare settings such as dentistry. The primary aim of this study was to assess the potential of plasma radiation to create a new population of Staphylococcus aureus cells with distinct characteristics that could lead to novel healthcare challenges. A homemade non-thermal plasma apparatus was applied and the effects of plasma treatment on S. aureus ATCC25923 was assessed. Plasma radiation was applied under controlled conditions to ensure that some bacterial cells remained viable. The treatment was repeated 10 times, with each round followed by a recovery phase to collect any surviving bacterial cells. To assess the potential changes in the bacterial population, we examined the antibiotic susceptibility pattern, micro-structural characteristics using scanning electron microscopy (SEM), and total protein profile using the matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) technique. The experimental results revealed slight variations in the antibiotic susceptibility patterns of certain cell wall agents (imipenem, cephalothin, and cefepime), as well as in the MALDI-TOF spectra. However, no changes were observed in the SEM images. The insufficient application of non-thermal plasma in bacterial decontamination may lead to physiological changes that could enrich or select certain subpopulations of S. aureus.

Assessment the toxicological perturbations of zinc oxide nanoparticles on histopathology, biochemistry and protein profile in marine mussels Brachidontes pharaonis

Zinc oxide nanoparticles (ZnO NPs) pose a major threat to aquatic life and their presence in the aquatic environment requires attention. This study intends to examine the impacts of ZnO NPs on Brachidontes pharaonis. Brachidontes pharaonis were divided into four groups, the control group and experimental groups exposed to ZnO NPs at concentrations of 1 mg/L, 5 mg/L, and 8 mg/L. The results revealed a decrease in protein and glucose, along with an increase in cholesterol, HDL, LDL, and triglycerides. The SDS-PAGE showed changes in the concentration of protein. In histological observations, there was necrosis in the epithelial cells, infiltration in the connective tissue between ovarian follicles, hemorrhage of blood vessels in gill filaments, and fibrosis in muscle myofibrils. Histochemical studies showed a rise in mucous cells in gill filaments and an increase in collagen fibers in the muscle. As concluded, ZnO NPs lead to alteration in biochemical parameters, protein profile, and histopathological changes in B. pharaonis.

Complementary analysis of proteome-wide proteomics reveals changes in RNA binding protein-profiles during prostate cancer progression.

Accumulating evidence indicates importance of RNA regulation in cancer. This includes events such as splicing, translation, and regulation of noncoding RNAs, functions which are governed by RNA binding proteins (RBPs). To find which RBPs could be relevant for prostate cancer, we performed systematic screening of RBP expression in clinical prostate cancer. We interrogated four proteome-wide proteomics datasets including tumor samples of primary, castration resistant, and metastatic prostate cancer. We found that, while the majority of RBPs are expressed but not significantly altered during prostate cancer development and progression, expression of several RBPs increases in advanced disease. Interestingly, most of the differentially expressed RBPs are not targets of differential posttranscriptional phosphorylation during disease progression. The RBPs undergoing expression changes have functions in, especially, poly(A)-RNA binding, nucleocytoplasmic transport, and cellular stress responses, suggesting that these may play a role in formation of castration resistance. Pathway analyzes indicate that increased ribosome production and chromatin-related functions of RBPs are also linked to castration resistant and metastatic prostate cancers. We selected a group of differentially expressed RBPs and studied their role in cultured prostate cancer cells. With siRNA screens, several of these were indicated in survival (DDX6, EIF4A3, PABPN1), growth (e.g., EIF5A, HNRNPH2, LRRC47, and NVL), and migration (e.g., NOL3 and SLTM) of prostate cancer cells. Our analyzes further show that RRP9, a U3 small nucleolar protein essential for ribosome formation, undergoes changes at protein level during metastasis in prostate cancer. In this work, we recognized significant molecular alterations in RBP profiles during development and evolution of prostate cancer. Our study further indicates several functionally significant RBPs warranting further investigation for their functions and possible targetability in prostate cancer.

Miniaturized microarray-format digital ELISA enabled by lithographic protein patterning

The search for reliable protein biomarker candidates is critical for early disease detection and treatment. However, current immunoassay technologies are failing to meet increasing demands for sensitivity and multiplexing. Here, the authors have created a highly sensitive protein microarray using the principle of single-molecule counting for signal amplification, capable of simultaneously detecting a panel of cancer biomarkers at sub-pg/mL levels. To enable this amplification strategy, the authors introduce a novel method of protein patterning using photolithography to subdivide addressable arrays of capture antibody spots into hundreds of thousands of individual microwells. This allows for the total sensor area to be miniaturized, increasing the total possible multiplex capacity. With the immunoassay realized on a standard 75x25 mm form factor glass substrate, sample volume consumption is minimized to <10 μL, making the technology highly efficient and cost-effective. Additionally, the authors demonstrate the power of their technology by measuring six secretory factors related to glioma tumor progression in a cohort of mice. This highly sensitive, sample-sparing multiplex immunoassay paves the way for researchers to track changes in protein profiles over time, leading to earlier disease detection and discovery of more effective treatment using animal models.

Role of blaTEM and OmpC in the piperacillin-tazobactam resistance evolution by E. coli in patients with complicated intra-abdominal infection

Piperacillin-tazobactam resistance (P/T-R) is increasingly reported among Escherichia coli isolates. Although in vitro experiments have suggested that blaTEM gene plays a key role in the P/T-R acquisition, no clinical in vivo study has yet confirmed the role of blaTEM or other genes. Therefore, we aimed to identify the mechanisms underlying P/T-R by following up patients with E. coli complicated intra-abdominal infections (cIAI) who experienced P/T treatment failure. Four pairs of strains, clonally related from four patients, were isolated both before and after treatment with P/T dosed at 4g/0.5g intravenously. The P/T MIC was tested using broth microdilution, and β-lactamase activity was determined in these isolates. Whole-genome sequencing (WGS) was performed to decipher the role of blaTEM and other genes associated with P/T-R. Changes in the outer membrane protein (OMP) profile were analyzed using SDS-PAGE, and blaTEM and ompC transcription levels were measured by RT-qPCR. In addition, in vitro competition fitness was performed between each pairs of strains (P/T-susceptible vs. P/T-resistant). We found a higher copy number of blaTEM gene in P/T-R isolates, generated by three different genetic events: (1) IS26-mediated duplication of the blaTEM gene, (2) generation of a small multicopy plasmid (ColE-like) carrying blaTEM, and (3) adaptive evolution via reduction of plasmid size, leading to a higher plasmid copy number. Moreover, two P/T-R strains showed reduced expression of OmpC. This study describes the mechanisms involved in the acquisition of P/T-R by E. coli in patients with cIAI. The understanding of P/T-R evolution is crucial for effectively treating infected patients and preventing the spread of resistant microorganisms.

Investigation of Taxonomic Relationship and Effect of Seasonal Temperature Changes Based on Protein Profiles of Fishes From Beyşehir, Suğla Lakes and Dam Apa

Besides traditional methods based on morphological characters, electrophoretic methods such as SDS-PAGE are preferred by taxonomists to make the right decision in the species identification process. In addition, the effect of environmental factors, such as pH, salinity, heat, and temperature on protein profiles are essential in various studies. In this study, we aimed to determine the degree of relationship in some fish species, such as Squalius lepidus, Cyprinus carpio, Carassius gibelio, Pseudophoxinus anatolicus, Tinca tinca, Alburnus orontis, Scardinius erythrophthalmus, Capoeta capoeta, Vimba vimba, Sander lucioperca living in Beyşehir, Suğla lakes and Apa Dam by SDS-PAGE method, and to examine seasonal differences by evaluating the effect of hot/cold water on protein profiles in fish. Although there were common major protein bands in all fish species studied, the presence of species-specific minor protein bands led to the separation of the species. The same fish species distributed in different lakes and dams were different both in minor bands, and changes in protein profiles were observed consequently on the same fish species synthesizing different proteins in different seasons. The data obtained from this study can contribute to systematic classification studies of fish.