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Cytokine Jun 2024Laryngopharyngeal reflux (LPR) is one of the most common disorders in otorhinolaryngology, affecting up to 10% of outpatients visiting otolaryngology departments. In...
BACKGROUND
Laryngopharyngeal reflux (LPR) is one of the most common disorders in otorhinolaryngology, affecting up to 10% of outpatients visiting otolaryngology departments. In addition, 50% of hoarseness cases are related to LPR. Pepsin reflux-induced aseptic inflammation is a major trigger of LPR; however, the underlying mechanisms are unclear. The nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome has become an important bridge between stimulation and sterile inflammation and is activated by intracellular reactive oxygen species (ROS) in response to danger signals, leading to an inflammatory cascade. In this study, we aimed to determine whether pepsin causes LPR-associated inflammatory injury via mediating inflammasome activation and explore the potential mechanism.
METHODS
We evaluated NLRP3 inflammasome expression and ROS in the laryngeal mucosa using immunofluorescence and immunohistochemistry. Laryngeal epithelial cells were exposed to pepsin and analyzed using flow cytometry, western blotting, and real-time quantitative PCR to determine ROS, NLRP3, and pro-inflammatorycytokine levels.
RESULTS
Pepsin expression was positively correlated with ROS as well as caspase-1 and IL-1β levels in laryngeal tissues. Intracellular ROS levels were elevated by increased pepsin concentrations, which were attenuated by apocynin (APO)-a ROS inhibitor-in vitro. Furthermore, pepsin significantly induced the mRNA and protein expression of thioredoxin-interacting protein, NLRP3, caspase-1, and IL-1β in a dose-dependent manner. APO and the NLRP3 inhibitor, MCC950, inhibited NLRP3 inflammasome formation and suppressed laryngeal epithelial cell damage.
CONCLUSION
Our findings verified that pepsin could regulate the NLRP3/IL-1β signaling pathway through ROS activation and further induce inflammatory injury in LPR. Targeting the ROS/NLRP3 inflammasome signaling pathway may help treat patients with LPR disease.
Topics: Humans; NLR Family, Pyrin Domain-Containing 3 Protein; Inflammasomes; Reactive Oxygen Species; Pepsin A; Laryngopharyngeal Reflux; Signal Transduction; Inflammation; Caspase 1; Interleukin-1beta
PubMed: 38471420
DOI: 10.1016/j.cyto.2024.156568 -
The Analyst Apr 2024Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is a versatile bioanalytical technique for protein analysis. Since the reliability of HDX-MS analysis considerably...
Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is a versatile bioanalytical technique for protein analysis. Since the reliability of HDX-MS analysis considerably depends on the retention of deuterium labels in the post-labeling workflow, deuterium/hydrogen (D/H) back exchange prevention strategies, including decreasing the pH, temperature, and exposure time to protic sources of the deuterated samples, are widely adopted in the conventional HDX-MS protocol. Herein, an alternative and effective back exchange prevention strategy based on the encapsulation of a millimeter droplet of a labeled peptide solution in a water-immiscible organic solvent (cyclohexane) is proposed. Cyclohexane was used to prevent the undesirable uptake of water by the droplet from the atmospheric vapor through the air-water interface. Using the pepsin digest of deuterated myoglobin, our results show that back exchange kinetics of deuterated peptides is retarded in a millimeter droplet as compared to that in the bulk solution. Performing pepsin digestion directly in a water-in-oil droplet at room temperature (18-21 °C) was found to preserve more deuterium labels than that in the bulk digestion with an ice-water bath. Based on the present findings, it is proposed that keeping deuterated peptides in the form of water-in-oil droplets during the post-labelling workflow will facilitate the preservation of deuterium labels on the peptide backbone and thereby enhance the reliability of the H/D exchange data.
Topics: Deuterium; Pepsin A; Water; Reproducibility of Results; Mass Spectrometry; Deuterium Exchange Measurement; Peptides; Hydrogen; Myoglobin; Cyclohexanes
PubMed: 38462973
DOI: 10.1039/d4an00179f -
Journal of Agricultural and Food... Mar 2024Coffee is one of the most popular beverages around the world and its consumption contributes to the daily intake of dietary melanoidins. Despite the emerging...
Coffee is one of the most popular beverages around the world and its consumption contributes to the daily intake of dietary melanoidins. Despite the emerging physiological role of food melanoidins, their effect on digestive processes has not been studied so far. In this study, the activity of the gastrointestinal enzymes pepsin and trypsin was investigated in the presence of water-soluble coffee melanoidins. The gastric enzyme pepsin is only slightly affected, whereas the intestinal enzyme trypsin is severely inhibited by coffee melanoidins. The intestinal digestibility of casein was significantly inhibited by coffee melanoidins at a concentration achievable by regular coffee consumption. The inhibition of proteolytic enzymes by coffee melanoidins might decrease the nutritional value of dietary proteins.
Topics: Coffee; Pepsin A; Peptide Hydrolases; Trypsin; Dietary Proteins; Polymers
PubMed: 38456211
DOI: 10.1021/acs.jafc.3c09654 -
Journal of Nuclear Medicine Technology Mar 2024In 2009, the Society of Nuclear Medicine and Molecular Imaging published a standardized protocol guideline for gastric emptying scintigraphy that contains specific...
In 2009, the Society of Nuclear Medicine and Molecular Imaging published a standardized protocol guideline for gastric emptying scintigraphy that contains specific instructions on the exact meal and preparation procedure. Previous research has shown that the standardized meal and proper preparation of the meal for gastric emptying scintigraphy are not being adopted by some facilities. This research explores the differences of radiolabeling in the method of preparation of Tc-sulfur colloid (SC)-radiolabeled eggs. Liquid egg whites were mixed with Tc-SC before cooking in conjunction with the standardized protocol. A second sample set was prepared by adding the Tc-SC to eggs after they were cooked. Each sample set was placed in a solution of HCl and pepsin to simulate gestation. Radiolabeling efficacy was tested on each sample set at 2 and 4 h after gestating in HCl and pepsin. Tc-SC added to the liquid egg whites before microwave cooking yielded radiolabeling efficacy of 70% Tc-SC after 2 and 4 h of simulated gastric fluid gestation. In contrast, radiolabeling after cooking the egg whites yielded 50% radiolabeling after simulated gestation. The results from this experiment showed that the method of mixing the Tc-SC with liquid egg whites before microwave cooking has higher binding efficacy than when adding Tc-SC onto already cooked egg whites. These results highlight the importance of following the standardized protocol for the meal preparation of a gastric emptying study.
Topics: Egg White; Pepsin A; Albumins; Colloids; Sulfur
PubMed: 38443106
DOI: 10.2967/jnmt.123.266794 -
International Journal of Biological... Apr 2024Pepsin is one of the major enzymes with significant importance in the food industry, biomedicines, and pharmaceutical formulations. In this work, the main objective was...
Pepsin is one of the major enzymes with significant importance in the food industry, biomedicines, and pharmaceutical formulations. In this work, the main objective was to biochemically characterize a pepsin-like enzymatic extract obtained from Pygocentrus nattereri, a predatory freshwater fish, focusing on their potential industrial application. The obtained extract exhibited optimal activity at 45 °C and pH 1.0-2.0. These proteases remained stable after 2 h of incubation at temperatures ranging from 0° to 45 °C and within pH range of 1.0 to 7.0. Their activity was significantly affected in presence of pepstatin A and SDS, 10 μM and 3.46 mM respectively, while EDTA and PMSF showed partial inhibitory effects. Divalent cations (Ca2+ and Mg2+) did not inhibit the proteolytic activity of the extract; in fact, it improved at a 5 mM CaCl2 concentration. As the NaCl concentration increased, the enzyme activity decreased. However, after desalination, 90 % of the activity was recovered within the tested exposure time. Besides, this extract demonstrated exceptional versatility across diverse industrial applications, including collagen extraction augmentation, IgG hydrolysis facilitation, and silver and polyester recovery from X-ray films. Our results suggest that the obtained enzymatic extract has a wide range of potential applications.
Topics: Animals; Peptide Hydrolases; Pepsin A; Characidae; Stomach; Perciformes; Hydrogen-Ion Concentration
PubMed: 38431015
DOI: 10.1016/j.ijbiomac.2024.130548 -
Meat Science Jun 2024The aim of this study was to assess whether ultrasound treatment (sonification time: 5, 15, and 30 min; constants: ∼40 kHz, ∼2.5 W cm2) can be applied prior to...
The aim of this study was to assess whether ultrasound treatment (sonification time: 5, 15, and 30 min; constants: ∼40 kHz, ∼2.5 W cm2) can be applied prior to hydrolysis to enhance the anti-radical and angiotensin converting enzyme inhibiting (anti-ACE) effect of the hydrolysates from fermented pork loins. Enzymatic hydrolysis was performed using pepsin, followed by pancreatin. The influence of meat matrix on the course of hydrolysis, shaped using a lactic acid bacteria (LAB)-based starter culture, was also analyzed. It was found that proteases caused a systematic increase in the content of peptides, while pancreatin limited the peptide content in the protein hydrolysate from the loins subjected to spontaneous fermentation. Moreover, for these tests, sonication time had a negligible effect on the peptides content of the hydrolysates. On the other hand, for the sample of LAB-fermented products, both sonication time and stage of hydrolysis promoted the biological activity of the hydrolysates. Samples from the LAB-fermented meat had more peptides at the stage of digestion with pepsin and pancreatin, exhibiting much faster antiradical and anti-ACE activity compared to the control sample. The obtained results suggest that the use of LAB promotes the release of antiradical peptides during the two-step enzymatic hydrolysis, the duration of which can be shortened to achieve satisfactory biofunctionalities. Additional application of sonication pretreatment allows controlling the course of the hydrolysis, as the pro-health, biological effect of some protein-derived sequences is associated with the content of peptides.
Topics: Animals; Swine; Peptidyl-Dipeptidase A; Protein Hydrolysates; Pepsin A; Pork Meat; Red Meat; Pancreatin; Sonication; Peptides; Hydrolysis; Lactobacillales
PubMed: 38422590
DOI: 10.1016/j.meatsci.2024.109472 -
Marine Drugs Feb 2024The effects of ultrasonic power (0, 150, 300, 450, and 600 W) on the extraction yield and the structure and rheological properties of pepsin-soluble collagen (PSC) from...
The effects of ultrasonic power (0, 150, 300, 450, and 600 W) on the extraction yield and the structure and rheological properties of pepsin-soluble collagen (PSC) from albacore skin were investigated. Compared with the conventional pepsin extraction method, ultrasonic treatment (UPSC) significantly increased the extraction yield of collagen from albacore skin, with a maximum increase of 8.56%. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed that peptides of low molecular weight were produced when the ultrasonic power exceeded 300 W. Meanwhile, secondary structure, tertiary structure, and X-ray diffraction analyses showed that the original triple helix structure of collagen was intact after the ultrasonic treatment. The collagen solutions extracted under different ultrasonic powers had significant effects on the dynamic frequency sweep, but a steady shear test suggested that the collagen extracted at 150 W had the best viscosity. These results indicate that an ultrasonic power between 150 and 300 W can improve not only the extraction yield of natural collagen, but also the rheological properties of the collagen solution without compromising the triple helix structure.
Topics: Animals; Ultrasonics; Pepsin A; Fish Proteins; Collagen; Perciformes; Skin
PubMed: 38393055
DOI: 10.3390/md22020084 -
Zhonghua Er Bi Yan Hou Tou Jing Wai Ke... Feb 2024To explore the clinical characteristics of children with adenoid hypertrophy (AH) and laryngopharyngeal reflux (LPR) by detecting the expression of pepsin in adenoids...
To explore the clinical characteristics of children with adenoid hypertrophy (AH) and laryngopharyngeal reflux (LPR) by detecting the expression of pepsin in adenoids as a standard for AH with LPR. A total of 190 children who were admitted for surgical treatment due to AH were included in the study. The main clinical symptoms of the patients were recorded, and the degree of adenoid hypertrophy was evaluated. Before the surgery, Reflux Symptom Index (RSI) and Reflux Finding Score (RFS) were used to evaluate the reflux symptoms. After the surgery, pepsin immunohistochemical staining was performed on the adenoid tissue, and according to the staining results, the patients were divided into study group (pepsin staining positive) and control group (pepsin staining negative). SPSS 19.0 software was used for statistical analysis. Quantitative data conforming to normal distribution between the two groups were tested by two-independent sample test, and quantitative data with skewed distribution were tested by test. The positive rate of pepsin staining in the 190 AH patients was 78.4% (149/190). The study group had higher levels of preoperative symptoms such as erythema and/or congestion of the pharynx(2.1±0.7 . 1.8±0.6,=2.23), vocal cord edema[1.0(0, 1.0) . 1.0(0, 1.0), =2.00], diffuse laryngeal edema[0(0, 1.0) . 0(0, 0), =2.48], posterior commissure hypertrophy[(1.4±0.6 . 1.1±0.5), =2.63], and a higher total score on the RFS scale than the control group(6.2±2.7 . 5.0±2.6, =2.47), with statistical differences (<0.05). The sensitivity and specificity of RFS score in diagnosing AH with LPR were 24.8% and 80.5%, respectively. When RFS>5 was used as the positive threshold, the sensitivity and specificity of RFS score in diagnosing AH with LPR were 61.1% and 58.5%, respectively. There was a statistical difference in the number of positive cases of RFS score between the study group and the control group(91 17,χ=5.04,=0.032). LPR is common in AH children. Children with AH and LPR have specific performance in electronic laryngoscopy, such as erythema with edema in the pharynx, posterior commissure hypertrophy, and vocal cord edema.
Topics: Child; Humans; Adenoids; Pepsin A; Laryngopharyngeal Reflux; Laryngeal Edema; Edema; Hypertrophy; Erythema
PubMed: 38369792
DOI: 10.3760/cma.j.cn115330-20231221-00318 -
EFSA Journal. European Food Safety... Feb 2024The food enzyme containing chymosin (EC 3.4.23.4) and pepsin (EC 3.4.23.1) is prepared from the abomasum of suckling calves, goats, lambs and buffaloes by Caglificio...
The food enzyme containing chymosin (EC 3.4.23.4) and pepsin (EC 3.4.23.1) is prepared from the abomasum of suckling calves, goats, lambs and buffaloes by Caglificio Clerici S.p.A. It is intended to be used in the production of cheese. As no concerns arise from the source of the food enzyme, from its manufacture and based on the history of safe use and consumption, the Panel considered that toxicological data were not required and no exposure assessment was necessary. The similarity of the amino acid sequences of the two proteins (chymosin and pepsin A) to those of known allergens was searched and two matches were found with respiratory allergens. The Panel considered that the risk of allergic reactions by dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.
PubMed: 38361797
DOI: 10.2903/j.efsa.2024.8607 -
Journal of Hazardous Materials Apr 2024Cadmium (Cd) and arsenic (As) co-contamination is widespread and threatens human health, therefore it is important to investigate the bioavailability of Cd and As...
Cadmium (Cd) and arsenic (As) co-contamination is widespread and threatens human health, therefore it is important to investigate the bioavailability of Cd and As co-exposure. Currently, the interactions of Cd and As by in vitro assays are unknown. In this work, we studied the concurrent Cd-As release behaviors and interactions with in vitro simulated gastric bio-fluid assays. The studies demonstrated that As bioaccessibility (2.04 to 0.18 ± 0.03%) decreased with Cd addition compared to the As(V) single system, while Cd bioaccessibility (11.02 to 39.08 ± 1.91%) increased with As addition compared to the Cd single system. Release of Cd and As is coupled to proton-promoted and reductive dissolution of ferrihydrite. The As(V) is released and reduced to As(Ⅲ) by pepsin. Pepsin formed soluble complexes with Cd and As. X-ray photoelectron spectroscopy showed that Cd and As formed Fe-As-Cd ternary complexes on ferrihydrite surfaces. The coordination intensity of As-O-Cd is lower than that of As-O-Fe, resulting in more Cd release from Fe-As-Cd ternary complexes. Our study deepens the understanding of health risks from Cd and As interactions during environmental co-exposure of multiple metal(loid)s.
Topics: Humans; Cadmium; Arsenic; Pepsin A; Digestion; Ferric Compounds
PubMed: 38335617
DOI: 10.1016/j.jhazmat.2024.133633