RESUMEN
This study introduces the nanobromhexine lipid particle (NBL) platform designed for effective pulmonary drug delivery. Inspired by respiratory virus transport mechanisms, NBL address challenges associated with mucus permeation and inflammation in pulmonary diseases. Composed of low molecular weight polyethylene glycol-coated lipid nanoparticles with bromhexine hydrochloride, NBL exhibit a size of 118 ± 24 nm, a neutral zeta potential, osmolarity of 358 ± 28 mOsmol/kg, and a pH of 6.5. Nebulizing without leakage and showing no toxicity to epithelial cells, NBL display mucoadhesive properties with a 60% mucin-binding efficiency. They effectively traverse the dense mucus layer of Calu-3 cultures in an air-liquid interface, as supported by a 55% decrease in MUC5AC density and a 29% increase in nanoparticles internalization compared to non-exposed cells. In assessing immunomodulatory effects, NBL treatment in SARS-CoV-2-infected lung cells leads to a 40-fold increase in anti-inflammatory MUC1 gene expression, a proportional reduction in pro-inflammatory IL-6 expression, and elevated anti-inflammatory IL-10 expression. These findings suggest a potential mechanism to regulate the excessive IL-6 expression triggered by virus infection. Therefore, the NBL platform demonstrates promising potential for efficient pulmonary drug delivery and immunomodulation, offering a novel approach to addressing mucus permeation and inflammation in pulmonary diseases.
Asunto(s)
Pulmón , Nanopartículas , SARS-CoV-2 , Nanopartículas/administración & dosificación , Humanos , Pulmón/metabolismo , SARS-CoV-2/efectos de los fármacos , Sistemas de Liberación de Medicamentos , Inmunomodulación , Línea Celular , Mucina-1/metabolismo , COVID-19 , Lípidos/química , Lípidos/administración & dosificación , Moco/metabolismo , Polietilenglicoles/química , Células Epiteliales/metabolismo , Células Epiteliales/efectos de los fármacos , Tratamiento Farmacológico de COVID-19 , Mucina 5AC/metabolismo , LiposomasRESUMEN
Numerous factors can increase the risk of severe influenza; however, a majority of severe cases occur in previously healthy children. Identification of high-risk children is important for targeted preventive interventions and prompt treatment. The aim of this study was to evaluate MUC5AC as a biomarker for influenza disease severity in children. For this, a prospective cohort study was conducted in 2019. Children hospitalized with acute respiratory infection (ARI) with confirmed positive influenza infection were enrolled. Influenza cases were identified by reverse transcriptase-polymerase chain reaction. Life-threatening disease (LTD) was defined by the need for intensive care and ventilatory support. MUC5AC, epidemiologic, and clinical risk factors were assessed. Three hundred and forty-two patients were hospitalized with ARI, of which 49 (14%) had confirmed influenza infection and 6 (12%) of them developed LTD. MUC5AC levels were higher in those patients with mild disease compared to cases with poorer outcomes. Our results show that the severity of influenza infection in children is significantly associated with low levels of MUC5AC. These findings suggest its potential as a suitable biomarker for predicting disease severity.
Asunto(s)
Biomarcadores , Gripe Humana , Mucina 5AC , Índice de Severidad de la Enfermedad , Humanos , Gripe Humana/diagnóstico , Gripe Humana/virología , Masculino , Femenino , Biomarcadores/sangre , Mucina 5AC/metabolismo , Estudios Prospectivos , Preescolar , Lactante , Niño , Factores de Riesgo , Hospitalización , Adolescente , Infecciones del Sistema Respiratorio/virología , Infecciones del Sistema Respiratorio/diagnósticoRESUMEN
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.
Asunto(s)
Basiliximab , Inmunosupresores , Mucina 5AC , Depuración Mucociliar , Animales , Ratas , Basiliximab/farmacología , Inmunosupresores/farmacología , Mucina 5AC/genéticaRESUMEN
Pre-harvest burning of sugarcane fields produces large amounts of air pollutants which are known to cause health problems, including ocular surface abnormalities. In this study, we evaluated the effect of biomass burning on mucus quality and mucin gene expression (MUC1, MUC5AC, MUC16) in the conjunctiva of sugarcane workers (SWs) and residents of an adjacent town (RTs). Impression cytology samples of the inferior tarsal and bulbar conjunctiva of 78 SWs and 32 RTs were collected before (T1) and immediately after (T2) a 6-month harvest period. The neutral, acid and total mucus content of goblet cells was determined by PAS and AB staining. The levels of MUC5AC, MUC1 and MUC16 mRNA in the conjunctiva were measured by real-time PCR. Compared to RTs, SWs had higher levels of bulbar acid mucus and MUC16 mRNA and tarsal MUC5AC mRNA at T2 and lower levels of neutral mucus at T1 and T2. In the SW group, MUC1 mRNA levels were higher at T2 than at T1, but the levels of neutral and acid mucus were similar. In the RT group, acid mucus decreased and neutral mucus increased in the bulbar and tarsal conjunctiva at T2. In conclusion, our findings show that sugarcane harvesting is associated with abnormalities in mucus quality and content and changes in mucin mRNA levels on the ocular surface. This may help explain the ocular inflammatory signs and symptoms observed in subjects exposed to air pollutants and high temperatures from sugarcane biomass burning.
Asunto(s)
Contaminantes Ocupacionales del Aire/toxicidad , Antígeno Ca-125/genética , Conjuntiva/efectos de los fármacos , Proteínas de la Membrana/genética , Mucina 5AC/genética , Mucina-1/genética , Exposición Profesional/efectos adversos , Saccharum , Adulto , Agricultura , Biomasa , Brasil , Conjuntiva/metabolismo , Conjuntivitis/inducido químicamente , Conjuntivitis/diagnóstico , Conjuntivitis/metabolismo , Regulación de la Expresión Génica/fisiología , Células Caliciformes/efectos de los fármacos , Células Caliciformes/patología , Humanos , Masculino , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Población Rural , Adulto JovenRESUMEN
Clara cells are the main airway secretory cells able to regenerate epithelium in the distal airways through transdifferentiating into goblet cells, a process under negative regulation of the Notch pathway. Pneumocystis is a highly prevalent fungus in humans occurring between 2 and 5 months of age, a period when airways are still developing and respiratory morbidity typically increases. Pneumocystis induces mucus hyperproduction in immunocompetent host airways and whether it can stimulate Clara cells is unknown. Markers of Clara cell secretion and Notch1 activation were investigated in lungs of immunocompetent rats at 40, 60, and 80 days of age during Pneumocystis primary infection with and without Valproic acid (VPA), a Notch inducer. The proportion of rats expressing mucin increased in Pneumocystis-infected rats respect to controls at 60 and 80 days of age. Frequency of distal airways Clara cells was maintained while mRNA levels for the mucin-encoding genes Muc5B and Muc5ac in lung homogenates increased 1.9 and 3.9 times at 60 days of infection (P. = 0.1609 and P. = 0.0001, respectively) and protein levels of the Clara cell marker CC10 decreased in the Pneumocystis-infected rats at 60 and 80 days of age (P. = 0.0118 & P. = 0.0388). CC10 and Muc5b co-localized in distal airway epithelium of Pneumocystis-infected rats at day 60. Co-localization of Muc5b and Ki67 as marker of mitosis in distal airways was not observed suggesting that Muc5b production by Clara cells was independent of mitosis. Notch levels remained similar and no transnucleation of activated Notch associated to Pneumocystis infection was detected. Unexpectedly, mucus was greatly increased at day 80 in Pneumocystis-infected rats receiving VPA suggesting that a Notch-independent mechanism was triggered. Overall, data suggests a Clara to goblet cell transdifferentiation mechanism induced by Pneumocystis and independent of Notch.
Asunto(s)
Pulmón/metabolismo , Pulmón/microbiología , Mucina 5AC/biosíntesis , Mucina 5B/biosíntesis , Infecciones por Pneumocystis/metabolismo , Infecciones por Pneumocystis/microbiología , Pneumocystis/patogenicidad , Receptores Notch/metabolismo , Animales , Transdiferenciación Celular/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Femenino , Antígeno Ki-67/metabolismo , Mitosis/efectos de los fármacos , Mucina 5AC/genética , Mucina 5AC/metabolismo , Mucina 5B/genética , Mucina 5B/metabolismo , Pneumocystis/efectos de los fármacos , Infecciones por Pneumocystis/patología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas Sprague-Dawley , Transducción de Señal , Uteroglobina/metabolismo , Ácido Valproico/farmacologíaRESUMEN
To assess MUC5AC as a biomarker for respiratory syncytial virus (RSV) disease severity, we tested nasal aspirates from RSV+ children with mild, moderate, and severe disease. Levels were significantly higher in those in the severe and moderate groups compared to mild group, indicating MUC5AC may be a useful biomarker for RSV disease severity.
Asunto(s)
Mucina 5AC/análisis , Infecciones por Virus Sincitial Respiratorio/diagnóstico , Virus Sincitial Respiratorio Humano/aislamiento & purificación , Índice de Severidad de la Enfermedad , Argentina , Biomarcadores/análisis , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Nariz/virología , Curva ROC , Infecciones por Virus Sincitial Respiratorio/complicaciones , Virus Sincitial Respiratorio Humano/genéticaRESUMEN
Secreted mucins constitute a crucial part of the gel that protects respiratory and digestive epithelia, being MUC2/Muc2 the predominant gel-forming mucin of the intestine while MUC5AC/Muc5ac is one of the gel-forming mucins most expressed at the airways. In this study, we have analyzed Muc2 and Muc5ac during rat development by using immunohistochemistry, Western blotting and RT-PCR. We demonstrated that rat Muc2 was expressed in fetal intestinal goblet cells of surface epithelium of villi and developing Lieberkühn crypts. In neonates and adults, Muc2 was expressed at luminal goblet cells of small and large intestine and at gastric mucous and glandular cells. Muc5ac protein was observed in embryonic gastric and lung samples; expression increased during development and postnatal and adult life. After birth, a low reaction was detected at the tracheal surface epithelium and glands, which increased in adults.
Asunto(s)
Tracto Gastrointestinal/metabolismo , Expresión Génica , Mucina 5AC/genética , Mucina 2/genética , Ratas/genética , Sistema Respiratorio/metabolismo , Animales , Embrión de Mamíferos/metabolismo , Desarrollo Embrionario , Tracto Gastrointestinal/crecimiento & desarrollo , Mucina 5AC/metabolismo , Mucina 2/metabolismo , Ratas/crecimiento & desarrollo , Ratas/metabolismo , Sistema Respiratorio/crecimiento & desarrolloRESUMEN
Chronic obstructive pulmonary disease (COPD) is an inflammatory process characterized by airway mucus hypersecretion. Lipopolysaccharides (LPS) are known to stimulate the production of mucin 5AC (MUC5AC) via epidermal growth factor receptor (EGFR) in human airway cells. Noteworthy, we have previously demonstrated that EGFR/Rac1/reactive oxygen species (ROS)/matrix metalloproteinase 9 (MMP-9) is a key signaling cascade regulating MUC5AC production in airway cells challenged with LPS. Various reports have shown an inverse association between the intake of polyunsaturated fatty acids (PUFA) of the n-3 (omega-3) family or fish consumption and COPD. In the present study, we investigated the influence of docosahexaenoic acid (DHA), one of the most important omega-3 PUFA contained in fish oil, on the production of MUC5AC in LPS-challenged human airway cells NCI--H292. Our results indicate that DHA is capable of counteracting MUC5AC overproduction in LPS-stimulated cells by abrogating both EGFR phosphorylation and its downstream signaling pathway. This signaling pathway not only includes Rac1, ROS and MMP-9, but also NF-κB, since we have found that ROS require NF-κB activity to induce MMP-9 secretion and activation.
Asunto(s)
Ácidos Docosahexaenoicos/farmacología , Receptores ErbB/metabolismo , Lipopolisacáridos/antagonistas & inhibidores , Mucina 5AC/biosíntesis , Transducción de Señal/efectos de los fármacos , Línea Celular , Humanos , Lipopolisacáridos/farmacología , Metaloproteinasa 9 de la Matriz/metabolismo , FN-kappa B/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
BACKGROUND: Diesel exhaust particles (DEPs) are deposited into the respiratory tract and are thought to be a risk factor for the development of diseases of the respiratory system. In healthy individuals, the timing and mechanisms of respiratory tract injuries caused by chronic exposure to air pollution remain to be clarified. METHODS: We evaluated the effects of chronic exposure to DEP at doses below those found in a typical bus corridor in Sao Paulo (150 µg/m3). Male BALB/c mice were divided into mice receiving a nasal instillation: saline (saline; n = 30) and 30 µg/10 µL of DEP (DEP; n = 30). Nasal instillations were performed five days a week, over a period of 90 days. Bronchoalveolar lavage (BAL) was performed, and the concentrations of interleukin (IL)-4, IL-10, IL-13 and interferon-gamma (INF-γ) were determined by ELISA-immunoassay. Assessment of respiratory mechanics was performed. The gene expression of Muc5ac in lung was evaluated by RT-PCR. The presence of IL-13, MAC2+ macrophages, CD3+, CD4+, CD8+ T cells and CD20+ B cells in tissues was analysed by immunohistochemistry. Bronchial thickness and the collagen/elastic fibers density were evaluated by morphometry. We measured the mean linear intercept (Lm), a measure of alveolar distension, and the mean airspace diameter (D0) and statistical distribution (D2). RESULTS: DEP decreased IFN-γ levels in BAL (p = 0.03), but did not significantly alter IL-4, IL-10 and IL-13 levels. MAC2+ macrophage, CD4+ T cell and CD20+ B cell numbers were not altered; however, numbers of CD3+ T cells (p ≤ 0.001) and CD8+ T cells (p ≤ 0.001) increased in the parenchyma. Although IL-13 (p = 0.008) expression decreased in the bronchiolar epithelium, Muc5ac gene expression was not altered in the lung of DEP-exposed animals. Although respiratory mechanics, elastic and collagen density were not modified, the mean linear intercept (Lm) was increased in the DEP-exposed animals (p ≤ 0.001), and the index D2 was statistically different (p = 0.038) from the control animals. CONCLUSION: Our data suggest that nasal instillation of low doses of DEP over a period of 90 days results in alveolar enlargement in the pulmonary parenchyma of healthy mice.
Asunto(s)
Contaminantes Atmosféricos/toxicidad , Neumonía/inducido químicamente , Alveolos Pulmonares/efectos de los fármacos , Emisiones de Vehículos/toxicidad , Animales , Brasil , Líquido del Lavado Bronquioalveolar/inmunología , Colágeno/metabolismo , Citocinas/inmunología , Citocinas/metabolismo , Tejido Elástico/metabolismo , Mediadores de Inflamación/metabolismo , Subgrupos Linfocitarios/efectos de los fármacos , Subgrupos Linfocitarios/inmunología , Subgrupos Linfocitarios/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Ratones Endogámicos BALB C , Mucina 5AC/genética , Mucina 5AC/metabolismo , Neumonía/inmunología , Neumonía/metabolismo , Neumonía/patología , Neumonía/fisiopatología , Alveolos Pulmonares/inmunología , Alveolos Pulmonares/metabolismo , Alveolos Pulmonares/patología , Alveolos Pulmonares/fisiopatología , ARN Mensajero/metabolismo , Mecánica Respiratoria/efectos de los fármacos , Factores de TiempoRESUMEN
BACKGROUND: MUC2 and MUC5AC overproduction is considered to be associated with hepatolithiasis and related to inflammation. However, mechanisms underlying MUC upregulation under inflammatory stimulation in human intrahepatic biliary epithelial cells (HIBECs) are not completely understood. MATERIAL AND METHODS: Expression of MUC2 and MUC5AC mRNA in HIBECs was detected by real-time PCR. Expression of COX-2, EP4, and phosphorylated ERK, JNK and p38MAPK protein was detected by Western blot. Concentrations of PGE2, IL-1ß and TNF-α in cell culture supernatants were measured using the Quantikine Elisa kit. RESULTS: COX-2 expression as well as PGE2 production in HIBECs was upregulated significantly by LPS, which was completely blocked by either TLR4 antagonist or NFκB inhibitor. Selective COX-2 inhibitor suppressed LPS-induced MUC2 and MUC5AC mRNA expression remarkably. Exogenous PGE2 increased MUC2 and MUC5AC mRNA expression in a dosage-dependent manner independent of IL-1ß and TNF-α. PGE2 receptor EP4 agonist elevated MUC2 and MUC5AC expression, whereas EP4 antagonist had the opposite effect. Expression of phosphorylated p38MAPK was upregulated by exogenous PGE2, and p38MAPK inhibitor reduced MUC2 and MUC5AC expression in HIBECs. In addition, it was found that levels of PGE2, MUC2 and MUC5AC in bile samples from the hepatic ducts affected by intrahepatic stones were significantly higher than those from the unaffected hepatic ducts of patients with hepatolithiasis. CONCLUSIONS: Our findings indicate that PGE2 induces MUC2 and MUC5AC expression in HIBECs via EP4-p38MAPK signaling.
Asunto(s)
Conductos Biliares/efectos de los fármacos , Dinoprostona/farmacología , Células Epiteliales/efectos de los fármacos , Mucina 5AC/metabolismo , Mucina 2/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/agonistas , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Bilis/metabolismo , Conductos Biliares/enzimología , Conductos Biliares/inmunología , Western Blotting , Células Cultivadas , Colelitiasis/metabolismo , Colelitiasis/patología , Ciclooxigenasa 2/metabolismo , Inhibidores de la Ciclooxigenasa 2/farmacología , Dinoprostona/metabolismo , Relación Dosis-Respuesta a Droga , Activación Enzimática , Ensayo de Inmunoadsorción Enzimática , Células Epiteliales/enzimología , Células Epiteliales/inmunología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Mediadores de Inflamación/metabolismo , Interleucina-1beta/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Lipopolisacáridos/farmacología , Mucina 5AC/genética , Mucina 2/genética , FN-kappa B/metabolismo , Fosforilación , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 4/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Regulación hacia ArribaRESUMEN
BACKGROUND: Mucociliary transport (MCT) is a defense mechanism of the airway. To study the underlying mechanisms of MCT, we have both developed an experimental model of cultures, from human adenoid tissue of ciliated and secretory cells, and characterized the response to local chemical signals that control ciliary activity and the secretion of respiratory mucins in vitro. MATERIALS AND METHODS: In ciliated cell cultures, ciliary beat frequency (CBF) and intracellular Ca(2+) levels were measured in response to ATP, UTP, and adenosine. In secretory cultures, mucin synthesis and secretion were identified by using immunodetection. Mucin content was taken from conditioned medium and analyzed in the presence or absence of UTP. RESULTS: Enriched ciliated cell monolayers and secretory cells were obtained. Ciliated cells showed a basal CBF of 10.7 Hz that increased significantly after exposure to ATP, UTP, or adenosine. Mature secretory cells showed active secretion of granules containing different glycoproteins, including MUC5AC. CONCLUSION: Culture of ciliated and secretory cells grown from adenoid epithelium is a reproducible and feasible experimental model, in which it is possible to observe ciliary and secretory activities, with a potential use as a model to understand mucociliary transport control mechanisms.
Asunto(s)
Tonsila Faríngea/citología , Tonsila Faríngea/metabolismo , Modelos Biológicos , Mucina 5AC/metabolismo , Adenosina/metabolismo , Adenosina Trifosfato/metabolismo , Calcio/metabolismo , Células Cultivadas , Niño , Preescolar , Cilios/metabolismo , Femenino , Humanos , Masculino , Transporte de Proteínas/fisiología , Uridina Trifosfato/metabolismoRESUMEN
PURPOSE: To report the outcomes of transplantation of autologous conjunctival epithelial cells cultivated ex vivo (EVCAU) in patients with total limbal stem cell deficiency (LSCD). METHODS: EVCAU were cultivated on denuded human amniotic membrane and transplanted in 12 eyes of 10 patients with total LSCD. We evaluated the improvement in the defined clinical parameters of LSCD (loss of corneal epithelial transparency, superficial corneal neovascularization and epithelial irregularity/recurrent epithelial breakdown), vision acuity, impression cytology, immunocytochemical analysis (CK3/CK19), and the appearance of a regular hexagonal basal layer of cells on corneal confocal microscopy. Histologic and immunohistochemical features were studied in 3 corneal buttons of patients submitted to penetrating keratoplasty after EVCAU. RESULTS: Cultivated conjunctival epithelium formed 4 to 5 layers with the formation of basement membrane-like structures. Immunocytochemical analysis showed positivity for CK3, CK19, MUC5AC, Ki-67, P63, and ABCG2. The improvement of the clinical parameters for this treatment in our cohort was 10 of 12 (83.3%) in a mean follow-up time of 18.5 months (range, 15-26 months), and these eyes showed an improvement in impression cytology, immunocytochemistry, and in vivo confocal analysis. Corneal buttons showed a well-formed epithelium with 5 to 6 layers, with rare cells periodic acid-Schiff+, and positivity for CK3, CK19, P63, connexin 43, and MUC5AC. CONCLUSION: We demonstrated the preliminary results of transplantation of EVCAU for corneal surface reconstruction in cases with total LSCD. Future studies are needed to further assess the long-term efficacy of this procedure.
Asunto(s)
Conjuntiva/citología , Enfermedades de la Córnea/cirugía , Células Epiteliales/trasplante , Células Madre/patología , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP/metabolismo , Adulto , Anciano , Biomarcadores/metabolismo , Trasplante de Células , Células Cultivadas , Epitelio Corneal/metabolismo , Epitelio Corneal/patología , Femenino , Humanos , Inmunohistoquímica , Queratina-19/metabolismo , Queratina-3/metabolismo , Queratoplastia Penetrante , Antígeno Ki-67/metabolismo , Limbo de la Córnea/patología , Masculino , Microscopía Confocal , Persona de Mediana Edad , Mucina 5AC/metabolismo , Proteínas de Neoplasias/metabolismo , Estudios Prospectivos , Células Madre/metabolismo , Trasplante Autólogo , Agudeza Visual/fisiología , Adulto JovenRESUMEN
BACKGROUND: Pneumocystis without obvious accompanying pathology is occasionally reported in autopsied infant lungs. Its prevalence and significance are unknown. Interestingly, this mild infection induces a strong activation of mucus secretion-related genes in young immunocompetent rodents that has not been explored in infants. Excess mucus is induced by multiple airway offenders through nonspecific pathways and would explain a cofactor role of Pneumocystis in respiratory disease. We undertook characterization of the prevalence of Pneumocystis and associated mucus in infant lungs. METHODS: Samples from 128 infants (mean age, 101 days) who died suddenly and unexpectedly in Santiago during 1999-2004 were examined for Pneumocystis using nested polymerase chain reaction (nPCR) amplification of the P. jirovecii mtLSU ribosomal RNA gene and immunofluorescence microscopy (IF). Pneumocystis-negative infants 28 days and older and their age-closest positives were studied for MUC5AC expression and Pneumocystis burden by Western blot and quantitative PCR, respectively. RESULTS: Pneumocystis DNA was detected by nPCR in 105 of the 128 infants (82.0%) and Pneumocystis organisms were visualized by IF in 99 (94.3%) of the DNA-positive infants. The infection was commonest at 3-4 months with 40 of 41 (97.6%) infants of that age testing positive. MUC5AC was significantly increased in Pneumocystis-positive tissue specimens (P = .013). Death was unexplained in 113 (88.3%) infants; Pneumocystis was detected in 95 (84.0%) of them vs 10 of 15 (66.7%) with explained death (P = .28). CONCLUSIONS: A highly focal Pneumocystis infection associated to increased mucus expression is almost universally present in the lungs of infants dying unexpectedly in the community regardless of autopsy diagnosis.
Asunto(s)
Pulmón/metabolismo , Moco/metabolismo , Pneumocystis/aislamiento & purificación , Neumonía por Pneumocystis/epidemiología , Muerte Súbita del Lactante/epidemiología , Autopsia , Recuento de Colonia Microbiana , ADN de Hongos/genética , Femenino , Humanos , Lactante , Recién Nacido , Pulmón/microbiología , Pulmón/patología , Masculino , Microscopía , Mucina 5AC/metabolismo , Técnicas de Amplificación de Ácido Nucleico , Pneumocystis/genética , Neumonía por Pneumocystis/diagnóstico , Neumonía por Pneumocystis/microbiología , Prevalencia , Sensibilidad y Especificidad , Muerte Súbita del Lactante/diagnósticoRESUMEN
OBJECTIVE: Recently, hepatocyte antigen (Hep) was introduced as a sensitive and reliable marker of intestinal metaplasia (IM). However, the distribution of Hep expression in subtypes of IM was not described. METHODS: We examined the expression of Hep in 58 cases of chronic gastritis associated with IM by immunohistochemical staining. Cases were classified as: 19 of IM Type I (complete) cases, 16 cases of IM Type II (incomplete) and 23 cases of IM Type III (incomplete). The distribution of Hep expression was classified into four groups according to the intensity of Hep expressing metaplastic cells: negative, low, moderate and high. We also compared expression of Hep with that of MUC-1, MUC-2 and MUC-5AC. RESULTS: Hep expression showed granular cytoplasmic staining and was specifically identified in columnar cells, but not in goblet cells. There was no significant difference between Hep expression and subtypes of IM (p > 0.005). However the difference between the distribution of Hep expression among three subtypes of IM was significant (p < 0.001). No relationship was observed among the expression of Hep, MUC-1, MUC-2 and MUC-5AC. CONCLUSION: Results of the present study revealed that the distribution of Hep expression is high in the majority of the complete type (Type I) IM cases, moderate in the majority of the incomplete Type II IM cases and low in all of the incomplete Type III IM cases and suggest that besides its role as a sensitive marker in IM, the evaluation of the distribution of Hep expression might be useful in the classification of IM.
Asunto(s)
Adenocarcinoma/metabolismo , Gastritis/metabolismo , Hepatocitos/inmunología , Lesiones Precancerosas/metabolismo , Neoplasias Gástricas/metabolismo , Adenocarcinoma/clasificación , Adenocarcinoma/patología , Adulto , Anciano , Biomarcadores de Tumor/metabolismo , Femenino , Mucosa Gástrica/inmunología , Mucosa Gástrica/patología , Gastritis/patología , Humanos , Inmunohistoquímica , Masculino , Metaplasia/inmunología , Persona de Mediana Edad , Mucina 5AC/metabolismo , Mucina-1/metabolismo , Mucina 2/metabolismo , Lesiones Precancerosas/inmunología , Lesiones Precancerosas/patología , Neoplasias Gástricas/inmunología , Neoplasias Gástricas/patologíaRESUMEN
Pyloric gland adenoma (PGA), also called adenoma with gastric differentiation, is a rare neoplasm of the gastric mucosa that can appear as gastric heterotopia in several organs. A 49-year-old woman presented with gastric reflux and chronic elevation of liver enzymes. She had a history of type 2 diabetes mellitus, hypothyroidism and an unspecified allergy treated with deflazacor, and a family history of autoimmune diseases. A liver biopsy showed macro- and microvesicular steatohepatitis. Hepatitis B and virus serum tests were negative. Autoimmune hepatitis was suspected and investigated. As an evaluation for dyspeptic symptoms an upper gastrointestinal endoscopy was performed, showing diffuse gastroduodenitis. A few polyps were found and resected from the gastric fundus; histopathology revealed a pyloric gland adenoma. There is very few clinical data on this tumor type because it is frequently underdiagnosed and reported as dysplasia. Further research is needed on the pathophysiology of this disease.
Asunto(s)
Adenoma/patología , Mucosa Gástrica/patología , Neoplasias Gástricas/patología , Femenino , Reflujo Gastroesofágico , Humanos , Inmunohistoquímica , Persona de Mediana Edad , Mucina 5AC/metabolismo , Mucina 6/metabolismoRESUMEN
Naegleria fowleri is an amoeboflagellate responsible for the fatal central nervous system (CNS) disease primary amoebic meningoencephalitis (PAM). This amoeba gains access to the CNS by invading the olfactory mucosa and crossing the cribriform plate. Studies using a mouse model of infection have shown that the host secretes mucus during the very early stages of infection, and this event is followed by an infiltration of neutrophils into the nasal cavity. In this study, we investigated the role of N. fowleri trophozoites in inducing the expression and secretion of airway mucin and pro-inflammatory mediators. Using the human mucoepidermal cell line NCI-H292, we demonstrated that N. fowleri induced the expression of the MUC5AC gene and protein and the pro-inflammatory mediators interleukin-8 (IL-8) and interleukin-1 beta (IL-1 beta), but not tumour necrosis factor-alpha or chemokine c-c motif ligand 11 (eotaxin). Since the production of reactive oxygen species (ROS) is a common phenomenon involved in the signalling pathways of these molecules, we analysed if trophozoites were capable of causing ROS production in NCI-H292 cells by detecting oxidation of the fluorescent probe 2,7-dichlorofluorescein diacetate. NCI-H292 cells generated ROS after 15-30 min of trophozoite stimulation. Furthermore, the expression of MUC5AC, IL-8 and IL-1 beta was inhibited in the presence of the ROS scavenger DMSO. In addition, the use of an epidermal growth factor receptor inhibitor decreased the expression of MUC5AC and IL-8, but not IL-1 beta. We conclude that N. fowleri induces the expression of some host innate defence mechanisms, such as mucin secretion (MUC5AC) and local inflammation (IL-8 and IL-1 beta) in respiratory epithelial cells via ROS production and suggest that these innate immune mechanisms probably prevent most PAM infections.
Asunto(s)
Infecciones Protozoarias del Sistema Nervioso Central/inmunología , Receptores ErbB/metabolismo , Interleucina-1beta/inmunología , Interleucina-8/inmunología , Mucina 5AC/inmunología , Naegleria fowleri/inmunología , Especies Reactivas de Oxígeno/metabolismo , Animales , Línea Celular , Dimetilsulfóxido/farmacología , Células Epiteliales/inmunología , Células Epiteliales/metabolismo , Humanos , Inmunidad Innata , Masculino , Ratones , Ratones Endogámicos BALB C , Neutrófilos/inmunología , Quinazolinas , Trofozoítos/inmunología , Tirfostinos/farmacologíaRESUMEN
Chronic obstructive pulmonary disease (COPD) is an inflammatory process characterized by airway mucus hypersecretion. Previous studies have reported that lipopolysaccharides (LPS) stimulate mucin 5AC (MUC5AC) production via epidermal growth factor receptor (EGFR) in human airway cells. Moreover, this production was shown to depend on the expression and activity of matrix metalloproteinase 9 (MMP-9), which is increased in COPD patients' serum. In the present study we investigated the signaling pathway mediating LPS-stimulated secretion and activation of MMP-9, and the regulatory effects of this pathway on the production of MUC5AC in the human airway cells NCI-H292. Using specific inhibitors, we found that LPS-stimulated cells secreted and activated MMP-9 via EGFR. Our results also indicate that signaling events downstream of EGFR involved PI3K-dependent activation of Rac1, which mediated the NADPH-generated reactive oxygen species responsible for MMP-9 secretion and activation. Finally, we observed that EGFR/PI3K/Rac1/NADPH/ROS/MMP-9 regulate MUC5AC production in LPS-challenged NCI-H292 cells.
Asunto(s)
Lipopolisacáridos/inmunología , Metaloproteinasa 9 de la Matriz/metabolismo , Mucina 5AC/metabolismo , Mucosa Respiratoria/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Línea Celular Tumoral , Receptores ErbB/metabolismo , Humanos , Moco/metabolismo , NADP/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Enfermedad Pulmonar Obstructiva Crónica/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Mucosa Respiratoria/inmunología , Transducción de SeñalRESUMEN
BACKGROUND: Mucin immunoexpression in adenocarcinoma arising in Barrett's esophagus (BE) may indicate the carcinogenesis pathway. The aim of this study was to evaluate resected specimens of adenocarcinoma in BE for the pattern of mucins and to correlate to the histologic classification. METHODS: Specimens were retrospectively collected from thirteen patients who underwent esophageal resection due to adenocarcinoma in BE. Sections were scored for the grade of intestinal metaplasia. The tissues were examined by immunohistochemistry for MUC2 and MUC5AC antibodies. RESULTS: Eleven patients were men. The mean age was 61 years old (varied from 40 to 75 years old). The tumor size had a mean of 4.7 +/- 2.3 cm, and the extension of BE had a mean of 7.7 +/- 1.5 cm. Specialized epithelium with intestinal metaplasia was present in all adjacent mucosas. Immunohistochemistry for MUC2 showed immunoreactivity in goblet cells, while MUC5AC was extensively expressed in the columnar gastric cells, localizing to the surface epithelium and extending to a variable degree into the glandular structures in BE. Tumors were classified according to the mucins in gastric type in 7/13 (MUC5AC positive) and intestinal type in 4/13 (MUC2 positive). Two tumors did not express MUC2 or MUC5AC proteins. The pattern of mucin predominantly expressed in the adjacent epithelium was associated to the mucin expression profile in the tumors, p = 0.047. CONCLUSION: Barrett's esophagus adenocarcinoma shows either gastric or intestinal type pattern of mucin expression. The two types of tumors developed in Barrett's esophagus may reflect the original cell type involved in the malignant transformation.
Asunto(s)
Adenocarcinoma/etiología , Esófago de Barrett/complicaciones , Neoplasias Esofágicas/etiología , Mucina 5AC/análisis , Mucina 2/análisis , Adenocarcinoma/química , Adenocarcinoma/patología , Adulto , Anciano , Neoplasias Esofágicas/química , Neoplasias Esofágicas/patología , Femenino , Mucosa Gástrica/patología , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Estudios RetrospectivosRESUMEN
UNLABELLED: Mucins are related to infectious and non-infectious diseases in Veterinary and Human Medicine. MUC1 mucin is a transmembrane glycoprotein expressed on the apical surface of human epithelia while MUC5AC is the predominant secreted mucin expressed in human gastric epithelium and goblet cells of lung and eyes. MUC5AC C-terminus cysteine rich regions and the cytoplasmic tail of MUC1 domains are conserved among several mammalian species. OBJECTIVE: to compare the expression of MUC1 and MUC5AC mucins in mammalian epithelia. CT33 anti-MUC1 cytoplasmic tail (MUC1CT) polyclonal antibody and 45M1 anti-MUC5AC monoclonal antibody were employed. By immunohistochemistry, MUC1CT was expressed in most tissues while MUC5AC was restricted to gastric surface epithelium and goblet cells from trachea and lung. By western blot, MUC1CT showed a band at approximately 35 kDa in most tissues; MUC5AC revealed bands at >180 kDa in stomach and lung secretions from rat, cat, pig and cow. When rat MUC5AC was immunoprecipitated, a band at about 180 kDa was obtained.
Asunto(s)
Mamíferos/inmunología , Mucina 5AC/biosíntesis , Mucina-1/biosíntesis , Animales , Anticuerpos Monoclonales/química , Western Blotting/veterinaria , Epitelio/inmunología , Humanos , Inmunohistoquímica/veterinaria , Estructura Terciaria de ProteínaRESUMEN
Naegleria fowleri is the aetiological agent of primary amoebic meningoencephalitis (PAM). This parasite invades its host by penetrating the olfactory mucosa. During the initial stages of infection, the host response is initiated by the secretion of mucus that traps the trophozoites. Despite this response, some trophozoites are able to reach, adhere to and penetrate the epithelium. In the present work, we evaluated the effect of mucins on amoebic adherence and cytotoxicity to Madin-Darby canine kidney (MDCK) cells and the MUC5AC-inducing cell line NCI-H292. We showed that mucins inhibited the adhesion of amoebae to both cell lines; however, this inhibition was overcome in a time-dependent manner. N. fowleri re-established the capacity to adhere faster than N. gruberi. Moreover, mucins reduced the cytotoxicity to target cells and the progression of the illness in mice. In addition, we demonstrated mucinolytic activity in both Naegleria strains and identified a 37 kDa protein with mucinolytic activity. The activity of this protein was inhibited by cysteine protease inhibitors. Based on these results, we suggest that mucus, including its major mucin component, may act as an effective protective barrier that prevents most cases of PAM; however, when the number of amoebae is sufficient to overwhelm the innate immune response, the parasites may evade the mucus by degrading mucins via a proteolytic mechanism.