Effect of chronic stress on gel-forming mucins in the small intestine of BALB/c mice.

Intestinal homeostasis involves the collaboration of gut barrier components, such as goblet cells and IgA-microbiota complexes, that are under the control of stress that promotes inflammatory responses addressed primarily in the colon. The aim of this study was to evaluate the effect of stress on mucins, goblet cells, and proinflammatory parameters in the proximal and distal regions of the small intestine. A group (n = 6) of female 8-week-old BALB/c mice underwent board immobilization stress (2 h per day for 4 days) and were sacrificed with isoflurane. Samples from proximal and distal small segments were collected to analyze the following: 1) goblet cells stained with periodic acid-Schiff (PAS) and with alcian blue (AB) to visualize histologically neutral and acidic mucins, respectively; 2) IgA-microbiota complexes identified by flow cytometry in intestinal lavages; and 3) MUC2, MUC5AC, and IL-18 mRNA levels in whole mucosal scrapings by reverse transcription-qPCR. Regarding the unstressed group, in the proximal region of small intestine both PAS+ and AB+ goblet cells were unchanged; however, MUC5AC and IL-18 mRNA levels were increased, and the percentage of IgA-microbiota complexes was reduced. In the distal segment, the number of PAS+ goblet cells was increased, whereas the number of AB+ goblet cells was reduced and did not affect the remaining parameters. The data suggest that stress induces inflammation in the proximal small intestine; these findings may provide an experimental reference for human diseases that may affect the proximal small intestine, such as Crohn's disease, in which stress contributes to the progression of intestinal inflammation or relapse.

Cigarette Smoke Extract Induces MUC5AC Expression Through the ROS/ IP3R/Ca2+ Pathway in Calu-3 Cells.

Background: Chronic obstructive pulmonary disease (COPD) is caused by exposure to noxious external particles, air pollution, and the inhalation of cigarette smoke. Airway mucus hypersecretion particularly mucin5AC (MUC5AC), is a crucial pathological feature of COPD and is associated with its initiation and progression. In this study, we aimed to investigate the effects of cigarette smoke extract (CSE) on MUC5AC expression, particularly the mechanisms by which reactive oxygen species (ROS) induce MUC5AC expression. Methods: The effects of CSE on the expression of MUC5AC and mucin5B (MUC5B) were investigated in vitro in Calu-3 cells. MUC5AC and MUC5B expression levels were measured using quantitative reverse transcription-polymerase chain reaction (qRT-PCR), immunofluorescence staining, and enzyme-linked immunosorbent assay (ELISA). Total cellular levels of ROS and Ca2+ were determined using DCFH-DA and Fluo-4 AM. Subsequently, the expression levels of IP3R, IRE1α, p-IRE1α and XBP1s were measured by Western blotting. Gene silencing was achieved by using small-interfering RNAs. Results: Our findings revealed that exposure to CSE increased MUC5AC levels and upregulated ROS, IP3R/Ca2+ and unfolded protein response (UPR)-associated factors. In addition, knockdown of IP3R using siRNA decreased CSE-induced Ca2+ production, UPR-associated factors, and MUC5AC expression. Furthermore, 10 mM N-acetyl-l-cysteine (NAC) treatment suppressed the effects of CSE, including ROS generation, IP3R/ Ca2+, UPR activation, and MUC5AC overexpression. Conclusion: Our results suggest that ROS regulates CSE-induced UPR and MUC5AC overexpression through IP3R/ Ca2+ signaling. Additionally, we identified NAC as a promising therapeutic agent for mitigating CSE-induced MUC5AC overexpression.

Teleost Muc2 and Muc5ac: Key guardians of mucosal immunity in flounder (Paralichthys olivaceus).

Mucins secreted by mucous cells constitute a core part of the defense line against the invasion of pathogens. However, mucins' structure and immunological functions remain largely unknown in teleost fish. In this study, two typical mucins, Muc2 and Muc5ac of flounder (Paralichthys olivaceus), were cloned and their physicochemical properties, structure and conservation were analyzed. Notably, specific antibodies against flounder Muc2 and Muc5ac were developed. It was verified at the gene and protein level that Muc2 was expressed in the hindgut and gills but not in the skin, while Muc5ac was expressed in the skin and gills but not in the hindgut. After flounders were immunized by immersion with inactivated Edwardsiella tarda, Muc2 and Muc5ac were significantly upregulated at both the gene expression and protein levels, and Muc2+/Muc5ac+ mucous cells proliferated and increased secretion of Muc2 and Muc5ac. Moreover, Muc2 and Muc5ac exerted retention and clearance effects on E. tarda in a short period (within 1 dpi). These results revealed the characterization of fish mucins Muc2 and Muc5ac at the protein level and clarified the role of mucins as key guardians to maintain the mucus barrier, which advanced our understanding of teleost mucosal barrier.

Pathological features of gastric­type endocervical adenocarcinoma: A report of two cases.

Gastric-type endocervical adenocarcinoma (GEA) is an uncommon form of uterine cervical adenocarcinoma with an unfavorable prognosis. The tumor consists of glands exhibiting a morphological resemblance to gastric cells and occasionally manifests features akin to pancreaticobiliary mucinous adenocarcinoma. GEA differs from the typical cervical cancer, particularly in its lack of association with the human papillomavirus. Immunophenotypic analysis suggests intestinal differentiation. The present study reports two cases of GEA occurring in postmenopausal individuals who were diagnosed in Lishui Central Hospital (Lishui, China) between January 2015 and January 2023. Microscopic examination revealed cysts lined with mucinous cells within the tumors. Immunohistochemical assays confirmed the positivity of the tumors for cytokeratin 7, mucin (MUC)5AC, and mutant tumor protein p53, while the results were negative for tumor suppressor p16, and in one case for paired box protein 8, consistent with characteristics of mucinous adenocarcinoma originating from the gastrointestinal tract. Programmed death-ligand 1 expression was also negative. The proto-oncogene K-ras was identified using amplification refractory mutation system polymerase chain reaction. Both cases were negative for mutations in codons 12 and 13 of exon 2, codon 61 of exon 3 and codon 146 of exon 4, but were positive for wild-type K-ras. Clinical follow-up revealed a potential association between histopathological features and resistance to chemotherapeutic drugs. The infrequency of this tumor type may contribute to diagnostic challenges.

Effects of Bimin Formula （鼻敏方） on the Nasal Mucosa TMEM16A/NF-κB/MUC5AC Signaling Pathway in a Rat Model of Allergic Rhinitis with Lung-Spleen Qi Deficiency / 中医杂志

ObjectiveTo explore the possible mechanism of Bimin Formula （鼻敏方） in treating lung-spleen qi deficiency syndrome of allergic rhinitis （AR） with high mucin secretion. MethodsThirty-four SD rats were randomly divided into a blank group （8 rats）， a model group （8 rats）， a low-dose Bimin Formula group （8 rats）， and a high-dose Bimin Formula group （10 rats）. Except for the blank group， the other groups were subjected to AR lung-spleen qi deficiency rat models induced by smoking， gavage of Ginkgo biloba leaf extract， and ovalbumin. After modeling， rats in the low- and high-dose Bimin Formula groups were given Bimin Formula concentrate （concentration of 2.16 g/ml） by gavage at doses of 1.08 g/100 g and 2.16 g/100 g， respectively， while rats in the model group were given 0.5 ml/100 g of normal saline by gavage， once daily for 28 days； the blank group was not intervened. Behavioral assessments were performed after intervention. ELISA was used to detect the levels of peripheral blood total immunoglobulin E （IgE）. HE staining was used to observe the pathological changes of nasal mucosa epithelium in rats， while immunohistochemistry was used to detect the expression of transmembrane protein 16A （TMEM16A） and mucin 5AC （MUC5AC） protein in nasal mucosa. Western Blot was used to detect the expression of nuclear factor kappa B （NF-κB） protein， and RT-PCR was used to detect the expression of TMEM16A， MUC5AC， and NF-κB mRNA in nasal mucosa. ResultsHE staining showed that the nasal mucosa epithelial cell structure in the blank group was intact without shedding， swelling， or necrosis； the nasal mucosa epithelial tissue of rats in the model group was thickened and partially shed， with infiltration of eosinophils and lymphocytes visible； the pathological changes in nasal mucosa tissue of rats in the high- and low-dose Bimin Formulagroups were improved， and more improvement was showen in the high-dose group. Compared with those in the blank group， the behavioral scores and peripheral blood total IgE levels of rats in the model group significantly increased， as well as the expression of TMEM16A， MUC5AC， and NF-κB proteins and mRNA in nasal mucosa （P＜0.05 or P＜0.01）. Compared with those in the model group， the behavioral scores and peripheral blood total IgE levels of rats in the high-dose Bimin Formula group decreased， and the expression of TMEM16A， MUC5AC， and NF-κB proteins and mRNA in nasal mucosaalso decreased （P＜0.05 or P＜0.01）； the behavioral scores and peripheral blood total IgE levels of rats in the low-dose Bimin Formula group were reduced， and the expression of TMEM16A and MUC5AC proteins and mRNA in nasal mucosa， as well as the expression of NF-κB protein decreased （P＜0.05 or P＜0.01）， but the difference in NF-κB mRNA expression was not statistically significant （P＞0.05）. Compared with the low-dose Bimin Formula group， the expression of NF-κB protein in the high-dose group decreased （P＜0.01）. ConclusionBimin Formula may improve the symptoms and high mucus secretion of AR lung-spleen qi deficiency by regulating the TMEM16A/NF-κB/MUC5AC signaling pathway in nasalmucosa.

Application value of MEX3A,CDX2,MUC2 and MUC5AC in judging cancerous gastric mucosal intestinal metaplasia / 中国现代医生

@#Objective To investigate the correlation between MEX3A and differentiation characteristics of gastric cancer and intestinal metaplasia,and its combination with caudal-related homeobox transcription factor 2(CDX2)and mucin 2(MUC2)and mucin 5AC(MUC5AC)to determine the role of carcinogenic intestinal metaplasia.Methods From January 2010 to December 2014,a total of 410 cases of gastric cancer and paracarcinoma paraffin-embedded tissue samples were selected from the Central Hospital Affiliated to Shenyang Medical College and the Second Hospital Affiliated to Shenyang Medical College.According to pathological diagnosis,they were divided into control group(mild superficial gastritis,79 cases),intestinal metaplasia group(149 cases)and gastric cancer group(182 cases).The expressions of MEX3A,CDX2,MUC2 and MUC5AC were detected by immunohistochemistry.Results MEX3A was highly expressed in gastric cancer group and intestinal metaplasia group,especially diffuse gastric cancer,poorly differentiated gastric cancer and type Ⅲ intestinal metaplasia(P<0.05).CDX2 and MUC2 were highly expressed in gastric cancer group and intestinal metaplasia group,especially intestinal type gastric cancer,highly and moderately differentiated gastric cancer,type Ⅰ and type Ⅱ intestinal metaplasia(P<0.05).The expression of MUC5AC was high in control group and low in gastric cancer group and intestinal metaplasia group,especially in intestinal type gastric cancer,type Ⅰ and type Ⅲ intestinal metaplasia(P<0.05).Gastric cancer and intestinal metaplasia differentiation were negatively correlated with MEX3A and MUC5AC expression,but positively correlated with CDX2 and MUC2 expression(P<0.05).MEX3A was negatively correlated with the expression of CDX2 and MUC2,and positively correlated with the expression of MUC5AC in gastric cancer(P<0.05).MEX3A was negatively correlated with the expression of CDX2 and MUC2 in intestinal metaplasia(P<0.05),while CDX2 was positively correlated with the expression of MUC2(P<0.05).Conclusion MEX3A is negatively correlated with gastric cancer and intestinal metaplasia differentiation.Gastric cancer is characterized by high MEX3A expression and low CDX2 and MUC2 expression.

Fine particulate matter promotes airway inflammation and mucin production by activating endoplasmic reticulum stress and the IRE1α/NOD1/NF­κB pathway.

Fine particulate matter (PM2.5) is a type of small particle that is <2.5 µm in diameter that may cause airway inflammation. Thus, the present study aimed to explore the effects of PM2.5 on endoplasmic reticulum (ER) stress and airway inflammation in human airway epithelial cells. For this purpose, HBE135­E6E7 airway epithelial cells were cultured and exposed to specific concentrations of PM2.5 for various periods of time, and cell viability was determined using a Cell Counting Kit­8 assay. The results of the present study demonstrated that exposure to PM2.5 increased the mRNA and protein expression levels of interleukin (IL)­6, tumor necrosis factor (TNF)­α and mucin 5AC (MUC5AC). Moreover, the expression levels of ER stress­related proteins, such as glucose­regulated protein 78, CCAAT­enhancer binding protein homologous protein, activating transcription factor 6, protein kinase R­like ER kinase (PERK), phosphorylated (p­)PERK, inositol­requiring enzyme 1α (IRE1α) and p­IRE1α, and nucleotide­binding oligomerization domain 1 (NOD1) expression levels were increased following exposure to PM2.5. Transfection with IRE1α small interfering RNA (siRNA) led to the increased production of IL­6, TNF­α and MUC5AC. Moreover, the expression of NOD1 and the translocation of NF­κB p65 were inhibited following transfection with IRE1α siRNA. In addition, the results of the present study demonstrated that transfection with NOD1 siRNA decreased the production of IL­6, TNF­α and MUC5AC, and decreased the translocation of NF­κB p65. The expression levels of IL­6, TNF­α and MUC5AC were increased in the HBE135­E6E7 cells following treatment with C12­iE­DAP, a NOD1 agonist. Moreover, treatment with C12­iE­DAP led to the activation of NF­κB p65. Collectively, the results of the present study suggest that PM2.5 promotes airway inflammation and mucin production by activating ER stress in HBE135­E6E7 airway epithelial cells, and that the IRE1α/NOD1/NF­κB pathway may be involved in this process.

Proteins: Neglected active ingredients in edible bird's nest.

Edible bird's nest (EBN) is a kind of natural invigorant with a long history of consumption in Asia, especially in China. EBN is formed by mixing the saliva of swiftlets (Aerodramus) with feathers and other components during the breeding season. Proteins are the most important nutrient in EBN. By studying proteins in EBN, we can not only elucidate their components at the molecular level, but also study their bioactivities. Therefore, it is of great significance to study the proteins in EBN. Previous research on the proteins in EBN was preliminary and cursory, and no one has summarized and analyzed the proteins in EBN and correlated the bioactivities of these proteins with the biological functions of EBN. This article focused on the proteins in EBN, listed the proteins identified in different proteomic studies, and introduced the sources, structures and bioactivities of the most frequently identified proteins, including acidic mammalian chitinase, lysyl oxidase homolog 3, mucin-5AC, ovoinhibitor, nucleobindin-2, calcium-binding protein (MW: 4.5 × 104) and glucose-regulated protein (MW: 7.8 × 104). The properties of these proteins are closely related to the bioactivities of EBN. Therefore, this article can provide inspiration for further research on the efficacy of EBN.

Inhibition of microRNA-328 Increases Ocular Mucin Expression and Conjunctival Goblet Cells.

We previously reported anti-miR-328 therapy for dry eye disease (DED). Since decreased mucin secretion is a risk factor for DED, we aimed to explore whether anti-miR-328 affects mucin expression and goblet cells. MiR-328 was increased in goblet cells when they were under desiccating stress or treated with benzalkonium chloride (BAC), both of which are risk factors for DED. Based on bioinformatics tool results, miR-328 was predicted to directly target the transcription factor CREB1 that has been known to promote the expression of mucin5AC. The inhibitory effect of miR-328 on CREB1 was confirmed by the transfection assay. A miR-328 binding site on the CREB1 gene was confirmed by the luciferase assay. Furthermore, anti-miR-328 increased CREB1 and mucin5AC in cultured goblet cells according to qPCR, Western blot, and IF staining experiments. Anti-miR-328 increased mucin5AC secretion from the cultured goblet cells based on an ELISA assay for the cultured medium. Finally, impression cytology data revealed anti-miR-328 increased conjunctival goblet cells in the DED rabbits induced by BAC. In conclusion, anti-miR-328 increases CREB1 expression leading to an increase in mucin5AC production and secretion. Furthermore, anti-miR-328 also increases conjunctival goblet cells. These results warrant the further development of anti-miR-328 therapy for DED.

[Dickkopf-1 inhibits the secretion of MUC5AC induced by Mycoplasma pneumoniae P1-C in mouse lung epithelial cells].

Mycoplasma pneumoniae is the most common pathogen of respiratory tract infection in children and adults. Clinical observation shows that M. pneumoniae infection can cause massive mucus secretion in the respiratory tract, which makes the breathing of patients difficult. Studies have shown that M. pneumoniae infection can cause massive secretion of mucin 5AC (MUC5AC). Adhesin P1 plays an important role in the pathogenesis of M. pneumoniae infection by mediating the adhesion of pathogens to host cells, and the C-terminal residues of P1 (P1-C) are immunogenic. This study investigated the molecular mechanism of Wnt/ß-catenin signaling pathway inhibitor Dickkopf-1 (DKK1) in the secretion of MUC5AC in mouse airway epithelial cells (MAECs) induced by P1-C. Scanning electron microscope and hematoxylin-eosin staining were used to observe the effect of P1-C on mucus secretion of MAECs. Protein chip was used to detect the secretion of cytokines and analyse the enrichment of related signaling pathways induced by P1-C in MAECs. Periodic acid schiff stain (PAS) staining, Tunel staining and Masson staining were used to detect the damage of the lungs of mouse exposed to P1-C. Immunohistochemistry was used to detect the secretion of MUC5AC expression, and Western blotting was used to reveal the molecular mechanism of DKK1-regulated secretion of MUC5AC induced by P1-C protein in MACES. The results showed that P1-C induced the massive secretion of mucus and inflammatory factors in MAECs. During P1-C infection, DKK1 down-regulated janus kinase 2 (JAK2), phosphorylation signaling and transcription activator 1 (p-STAT1) and phosphorylation signaling and activator of transcription 3 (p-STAT3) expression. Overexpression of DKK1 significantly up-regulated the expression of MUC5AC repressor transcription factor fork-head box protein A2 (FOXA2). At the same time, the expression of MUC5AC induced by P1-C was inhibited significantly. It is speculated that DKK1 can effectively reduce the secretion of MUC5AC in MAECs induced by P1-C by inhibiting the JAK/STAT1-STAT3 signaling pathway and up-regulating the expression of FOXA2.

Effects of Z-VaD-Ala-Asp-Fluoromethyl Ketone (Z-VAD-FMK) and Acetyl-Asp-Glu-Val-Asp-Aldehyde(Ac-DEVD-CHO) on Inflammation and Mucus Secretion in Mice Exposed to Cigarette Smoke.

Background and Objectives: Smoking can lead to airway inflammation and mucus secretion through the nucleotide-binding domain-like receptor protein 3/caspase-1 pathway. In this study, z-VaD-Ala-Asp-fluoromethyl ketone(Z-VAD), a pan-caspase inhibitor, and acetyl-Asp-Glu-Val-Asp-aldehyde(Ac-DEVD), a caspase-3 inhibitor, were used to investigate the effect of caspase inhibitors on the expression of interleukin(IL)-1ß and IL-8, airway inflammation, and mucus secretion in mice exposed to cigarette smoke(CS). Methods: Thirty-two C57BL/6J male mice were divided into a control group, Smoke group, Z-VAD group, and Ac-DEVD group. Except for the control group, the animals were all exposed to CS for three months. After the experiment, lung function was measured and hematoxylin and eosin staining and periodic acid-Schiff staining were performed. The levels of IL-1ß, IL-8, and mucin 5ac(Muc5ac) in serum and bronchoalveolar lavage fluid(BALF) were determined by enzyme-linked immunosorbent assay. Results: Compared with the control group, the lung function of mice exposed to smoke was poorer, with a large number of inflammatory cells infiltrating around the airway, collapse of alveoli, expansion and fusion of distal alveoli, and formation of emphysema. The Z-VAD group was relieved compared with the smoke group. Airway inflammation was also reduced in the Ac-DEVD group compared with the Smoke group, but the degree of emphysema was not significantly improved. Although Z-VAD relieved airway inflammation and emphysema, Ac-DEVD only relieved inflammation. Z-VAD and Ac-DEVD decreased serum IL-1ß and IL-8 levels. In BALF, IL-1ß was decreased in Z-VAD group and IL-8 was highest in Smoke +Ac-DEVD group compared with control group and Ac-DEVD group. There was no significant difference in the expression of Muc5ac in serum. However, in BALF, levels of Muc5ac were higher in the smoking group and the lowest in the Ac-DEVD group. Conclusion: Mice exposed to smoke had decreased lung function and significant cilia lodging, epithelial cell shedding, and inflammatory cell infiltration, with significant emphysema formation. The pan-caspase inhibitor, Z-VAD, improved airway inflammation and emphysema lesions in the mice exposed to smoke and reduced IL-1ß and IL-8 levels in serum. The caspase-3 inhibitor, Ac-DEVD, reduced airway inflammation, serum IL-1ß and IL-8 levels, and Muc5ac levels in BALF, but it did not improve emphysema.

Understanding the Clinical Significance of MUC5AC in Biliary Tract Cancers.

Biliary tract cancers (BTC) arise from biliary epithelium and include cholangiocarcinomas or CCA (including intrahepatic (ICC) and extrahepatic (ECC)) and gallbladder cancers (GBC). They often have poor outcomes owing to limited treatment options, advanced presentations, frequent recurrence, and poor response to available systemic therapy. Mucin 5AC (MUC5AC) is rarely expressed in normal biliary epithelium, but can be upregulated in tissues of benign biliary disease, premalignant conditions (e.g., biliary intraepithelial neoplasia), and BTCs. This mucin's numerous glycoforms can be divided into less-glycosylated immature and heavily-glycosylated mature forms. Reported MUC5AC tissue expression in BTC varies widely, with some associations based on cancer location (e.g., perihilar vs. peripheral ICC). Study methods were variable regarding cancer subtypes, expression positivity thresholds, and MUC5AC glycoforms. MUC5AC can be detected in serum of BTC patients at high concentrations. The hesitancy in developing MUC5AC into a clinically useful biomarker in BTC management is due to variable evidence on the diagnostic and prognostic value. Concrete conclusions on tissue MUC5AC are difficult, but serum detection might be relevant for diagnosis and is associated with poor prognosis. Future studies are needed to further the understanding of the potential clinical value of MUC5AC in BTC, especially regarding predictive and therapeutic value.

Dickkopf-1 inhibits the secretion of MUC5AC induced by Mycoplasma pneumoniae P1-C in mouse lung epithelial cells / 生物工程学报

Mycoplasma pneumoniae is the most common pathogen of respiratory tract infection in children and adults. Clinical observation shows that M. pneumoniae infection can cause massive mucus secretion in the respiratory tract, which makes the breathing of patients difficult. Studies have shown that M. pneumoniae infection can cause massive secretion of mucin 5AC (MUC5AC). Adhesin P1 plays an important role in the pathogenesis of M. pneumoniae infection by mediating the adhesion of pathogens to host cells, and the C-terminal residues of P1 (P1-C) are immunogenic. This study investigated the molecular mechanism of Wnt/β-catenin signaling pathway inhibitor Dickkopf-1 (DKK1) in the secretion of MUC5AC in mouse airway epithelial cells (MAECs) induced by P1-C. Scanning electron microscope and hematoxylin-eosin staining were used to observe the effect of P1-C on mucus secretion of MAECs. Protein chip was used to detect the secretion of cytokines and analyse the enrichment of related signaling pathways induced by P1-C in MAECs. Periodic acid schiff stain (PAS) staining, Tunel staining and Masson staining were used to detect the damage of the lungs of mouse exposed to P1-C. Immunohistochemistry was used to detect the secretion of MUC5AC expression, and Western blotting was used to reveal the molecular mechanism of DKK1-regulated secretion of MUC5AC induced by P1-C protein in MACES. The results showed that P1-C induced the massive secretion of mucus and inflammatory factors in MAECs. During P1-C infection, DKK1 down-regulated janus kinase 2 (JAK2), phosphorylation signaling and transcription activator 1 (p-STAT1) and phosphorylation signaling and activator of transcription 3 (p-STAT3) expression. Overexpression of DKK1 significantly up-regulated the expression of MUC5AC repressor transcription factor fork-head box protein A2 (FOXA2). At the same time, the expression of MUC5AC induced by P1-C was inhibited significantly. It is speculated that DKK1 can effectively reduce the secretion of MUC5AC in MAECs induced by P1-C by inhibiting the JAK/STAT1-STAT3 signaling pathway and up-regulating the expression of FOXA2.

Effect of sitagliptin on expression of airway mucin 5AC in mice with endotoxin-induced lung injury / 中华麻醉学杂志

Objective:To evaluate the effect of sitagliptin on the expression of airway mucin 5AC (MUC5AC) in mice with endotoxin-induced lung injury.Methods:Thirty-six healthy male SPF C57BL/6 mice, aged 6-8 weeks, weighing 20-25 g, were divided into 3 groups ( n=12 each) using a random number table method: control group (group C), endotoxin-induced lung injury group (group L), and endotoxin-induced lung injury+ sitagliptin group (group S). Lipopolysaccharide (LPS) 3 mg/kg was intratracheally infused to prepare endotoxin-induced lung injury model in L and S groups, while the equal volume of normal saline was given instead in group C. Sitagliptin 100 mg/kg was intraperitoneally injected at 1 h before LPS infusion in group S, and normal saline was intraperitoneally injected at 1 h before endotracheal infusion in C and L groups. The arterial blood samples from femoral artery were taken at 24 h of LPS or normal saline infusion for measurement of PaO 2 and glucose levels.The mice were then sacrificed, and broncho-alveolar lavage fluid (BALF) and lung tissues were collected for determination of the concentrations of interleukin-6 (IL-6), interleukin-1beta (IL-1β), and tumor necrosis factor-alpha (TNF-α)in serum and BALF (by enzyme-linked immunosorbent assay), wet/dry weight ratio (W/D ratio), expression of MUC5AC (by immunohistochemistry and immunohistochemical comprehensive score), and expression of MUC5AC mRNA in lung tissues (by quantitative real-time polymerase chain reaction) and for examination of the pathological changes of lung tissues (using haematoxylin and eosin staining) which were scored. Results:Compared with group C, PaO 2 was significantly decreased, the glucose levels, W/D ratio and lung injury score were increased, the concentrations of IL-6, IL-1β and TNF-α in serum and BALF were increased, and the expression of MUC5AC mRNA in lung tissues was up-regulated in L and S groups( P<0.05). Compared with group L, PaO 2 was significantly increased, the glucose levels, W/D ratio and lung injury score were decreased, the concentrations of IL-6, IL-1β and TNF-α in serum and BALF were decreased, and the expression of MUC5AC mRNA in lung tissues was down-regulated in group S( P<0.05). Conclusions:The mechanism by which sitagliptin alleviates endotoxin-induced lung injury is related to down-regulation of MUC5AC expression in mice.

Proteins: Neglected active ingredients in edible bird's nest / 中草药·英文版

Edible bird's nest (EBN) is a kind of natural invigorant with a long history of consumption in Asia, especially in China. EBN is formed by mixing the saliva of swiftlets (Aerodramus) with feathers and other components during the breeding season. Proteins are the most important nutrient in EBN. By studying proteins in EBN, we can not only elucidate their components at the molecular level, but also study their bioactivities. Therefore, it is of great significance to study the proteins in EBN. Previous research on the proteins in EBN was preliminary and cursory, and no one has summarized and analyzed the proteins in EBN and correlated the bioactivities of these proteins with the biological functions of EBN. This article focused on the proteins in EBN, listed the proteins identified in different proteomic studies, and introduced the sources, structures and bioactivities of the most frequently identified proteins, including acidic mammalian chitinase, lysyl oxidase homolog 3, mucin-5AC, ovoinhibitor, nucleobindin-2, calcium-binding protein (MW: 4.5 × 104) and glucose-regulated protein (MW: 7.8 × 104). The properties of these proteins are closely related to the bioactivities of EBN. Therefore, this article can provide inspiration for further research on the efficacy of EBN.

Adeno-associated virus-based caveolin-1 delivery via different routes for the prevention of cholesterol gallstone formation.

BACKGROUND: Hepatic caveolin-1 (CAV1) is reduced in cholesterol gallstone disease (CGD). Mice with CAV1 deficiency were prone to develop CGD. However, it remains unknown whether restored hepatic CAV1 expression prevents the development of CGD. METHODS: C57BL/6 mice were injected with adeno-associated virus 2/8 (AAV2/8) vectors carrying the CAV1 gene (AAV2/8CAV1) via intravenous (i.v.) or intraperitoneal (i.p.) route and then subjected to a lithogenic diet (LD) for 8 weeks. Uninjected mice were used as controls. The functional consequences of rescuing CAV1 expression by either i.v. or i.p. AAV2/8CAV1 treatment for CGD prevention and its subsequent molecular mechanisms were examined. RESULTS: CAV1 expression was reduced in the liver and gallbladder of LD-fed CGD mice. We discovered that AAV2/8CAV1 i.p. delivery results in higher transduction efficiency in the gallbladder than tail vein administration. Although either i.v. or i.p. injection of AAV2/8CAV1 improved liver lipid metabolic abnormalities in CGD mice but did not affect LD feeding-induced bile cholesterol supersaturation. In comparison with i.v. administration route, i.p. administration of AAV2/8CAV1 obviously increased CAV1 protein levels in the gallbladder of LD-fed mice, and i.p. delivery of AAV2/8CAV1 partially improved gallbladder cholecystokinin receptor (CCKAR) responsiveness and impeded bile cholesterol nucleation via the activation of adenosine monophosphate-activated protein kinase (AMPK) signaling, which induced a reduction in gallbladder mucin-1 (MUC1) and MUC5ac expression and gallbladder cholesterol accumulation. CONCLUSION: CGD prevention by i.p. AAV2/8CAV1 injection in LD-fed mice was associated with the improvement of gallbladder stasis, which again supported the notion that supersaturated bile is required but not sufficient for the formation of cholesterol gallstones. Additionally, AAV treatment via the local i.p. injection offers particular advantages over the systemic i.v. route for much more effective gallbladder gene delivery, which will be an excellent tool for conducting preclinical functional studies on the maintenance of normal gallbladder function to prevent CGD.

Serum MUC5AC protein levels are correlated with the development and severity of connective tissue disease-associated pulmonary interstitial lesions.

Background: Mucin 5AC (MUC5AC) and mucin 5B (MUC5B) are the major components of airway mucins. The expression levels of MUC5AC and MUC5B are related to connective tissue disease-associated interstitial lung disease (CTD-ILD) in the promoter region of MUC5AC and MUC5B and the relevant bronchoalveolar lavage fluid. However, the serum protein levels of MUC5AC and MUC5B have not been tested in CTD-ILD patients. In this study, we tested the serum levels of MUC5AC and MUC5B proteins in CTD-ILD patients and assessed their relationship with the occurrence and development of ILD. Methods: Serum samples were obtained from 168 CTD and 80 healthy participants from the First Affiliated Hospital of Xiamen University. The serum levels of MUC5AC and MUC5B proteins were measured by enzyme-linked immunosorbent assay. Results: Of the 168 individuals with CTD, 70 had primary Sjögren's syndrome (pSS), 64 had systemic sclerosis (SSc), and 34 had polymyositis/dermatomyositis (PM/DM). There were 116 cases with concurrent ILD; ILD scores were 1 (n=23), 2 (n=41), and 3 (n=52). Serum MUC5AC and MUC5B protein levels were considerably higher in CTD-ILD than CTD-only individuals or healthy controls (both p<0.005). Among the CTD subgroups, MUC5AC was higher in individuals with concurrent ILD than in those without ILD (all p<0.05). MUC5AC was positively correlated with ILD severity in all three CTD subgroups (all R>0.47 and all p<0.05). The MUC5B levels varied substantially between SSc and SSc patients with concurrent ILD (p=0.032) and were related to ILD severity only in PM/DM patients (R=0.346 and p=0.045). Conclusion: MUC5AC is correlated with the occurrence and development of ILD, while MUC5B is associated with ILD diagnosis and severity in CTD subgroups. Serum MUC5AC levels present a definite diagnostic utility for CTD-ILD and as proxies for its severity.

Basiliximab Does Not Impair Airway Mucociliary Clearance of Rats.

Previous studies have shown that immunosuppressive drugs impair the airway mucociliary clearance of rats. However, considering the high specificity of basiliximab (BSX) and the absence of studies reporting its side effects, our aim was to investigate whether BSX, associated or not with triple therapy, impairs the mucociliary system. Forty rats were divided into 4 groups: Control, BSX, Triple, and BSX + Triple. After 15 days of treatment, animals were euthanized and the ciliary beating frequency (CBF), mucociliary transport velocity (MCTV), neutral and acid mucin production, Muc5ac and Muc5b gene expression, inflammatory cell number, and interleukin (IL)-6 concentration were analyzed. CBF and MCTV were lower in Triple and BSX + Triple groups (p < 0.05). Neutral mucin percentage was higher in Triple group (p < 0.05), and acid mucin percentage was higher in Triple and BSX + Triple groups (p < 0.05). The Muc5ac and Muc5b gene expression was higher in Triple and BSX + Triple groups (p < 0.05). Animals from Triple and BSX + Triple groups presented fewer mononuclear cells (p < 0.05). The number of polymorphonuclear cells was higher in the Triple group (p < 0.05). In the analysis of inflammatory cells in the blood, there was a decrease in lymphocytes and an increase in neutrophils in the Triple and BSX + Triple groups (p < 0.05). The concentration of IL-6 significantly increased in the animals of the Triple and BSX + Triple groups (p < 0.05). BSX did not change the mucociliary apparatus of rats.

Mucin 5AC expression is common but unrelated to tumor progression in pancreatic adenocarcinoma.

Introduction: Mucin 5AC (MUC5AC) belongs to the family of secreted gel-forming mucins. It is physiologically expressed in some normal mucin producing epithelial cells but also in pancreatic, ovarian, and colon cancer cells. The role of MUC5AC expression in cancer is not fully understood. This study was designed to explore the role of MUC5AC for pancreatic cancer progression, its association to microsatellite instability, and its diagnostic utility. Methods: Mucin 5AC expression was studied immunohistochemically in a tissue microarray (TMA) from 532 pancreatic cancers, 61 cancers of the ampulla Vateri, six acinar cell carcinomas and 12 large sections of pancreatitis. Results: Mucin 5AC staining was interpretable in 476 of 599 (79%) arrayed cancers. Staining was completely absent in normal pancreas and pancreatitis, but frequent in pancreatic cancer. Membranous and cytoplasmic MUC5AC expression was most common in pancreatic adenocarcinomas (71% of 423), followed by carcinomas of the ampulla Vateri (43% of 47), and absent in six acinar cell carcinomas. Mucin 5AC expression was unrelated to tumor phenotype (tumor stage, tumor grade, lymph node, and distant metastasis), and microsatellite instability in ductal adenocarcinomas and carcinomas of the ampulla Vateri. Conclusion: Our study indicates that MUC5AC is an excellent biomarker for pancreatic cancer diagnosis, especially to support the sometimes-difficult diagnosis on small biopsies. Mucin 5AC expression is unrelated to pancreatic cancer aggressiveness.

Expression of Mucins in Different Entities of Salivary Gland Cancer: Highest Expression of Mucin-1 in Salivary Duct Carcinoma : Mucin-1 - highest expression in Salivary Duct Carcinoma.

Therapeutic options for advanced salivary gland cancer (SGC) are rare. Therefore, it was the aim of this study to investigate the extent and intensity of Mucin-1 (MUC1), Mucin-16 (MUC16), and Mucin-5AC (MUC5AC) as potential molecular targets using immunohistochemistry. The medical records of all patients who underwent primary surgery for salivary gland cancer with curative intent in a tertiary referral center between 1990 and 2018 were reviewed. Immunohistochemical staining for MUC1, MUC16, and MUC5AC was performed for all patients with sufficient formalin-fixed paraffin-embedded material, and a semi-quantitative combined score derived from the H-score for the cytoplasmatic, the membranous and the apical membrane was built for the most common entities of SGC. 107 patients with malignancies of the parotid (89.7%) and the submandibular gland (10.3%) were included. The most common entities were mucoepidermoid carcinoma (MuEp; n = 23), adenoid cystic carcinoma (AdCy; n = 22), and salivary duct carcinoma (SaDu; n = 21). The highest mean MUC1 combined score was found in SaDu with 223.6 (±91.7). The highest mean MUC16 combined score was found in MuEp with 177.0 (±110.0). The mean MUC5AC score was low across all entities. A higher MUC1 combined score was significantly associated with male gender (p = 0.03), lymph node metastasis (p < 0.01), lymphovascular invasion (p = 0.045), and extracapsular extension (p = 0.03). SaDu patients with MUC16 expression showed a significantly worse 5-year progression-free survival than those without MUC16 expression (p = 0.02). This is the first study to give a comprehensive overview of the expression of MUC1, MUC16, and MUC5AC in SGC. Since advanced SGCs lack therapeutic options in many cases, these results warrant in vitro research on therapeutic targets against MUC1 in SaDu cell lines and xenograft models.