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1.
Endocrinology ; 163(4)2022 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-35266539

RESUMO

Enterotoxigenic Escherichia coli (ETEC)-derived purified heat-stable enterotoxin b (STb) is responsible for secretory diarrhea in livestock and humans. STb disrupts intestinal fluid homeostasis, epithelial barrier function, and promotes cell death. Glucagon-like peptide-2 (GLP-2) is a potent intestinotrophic hormone secreted by enteroendocrine L cells. GLP-2 enhances crypt cell proliferation, epithelial barrier function, and inhibits enterocyte apoptosis. Whether STb can affect GLP-2 producing L cells remains to be elucidated. First, secreted-His-labeled STb from transformed E coli was collected and purified. When incubated with L-cell models (GLUTag, NCI-H716, and secretin tumor cell line [STC-1]), fluorescent immunocytochemistry revealed STb was internalized and was differentially localized in the cytoplasm and nucleus. Cell viability experiments with neutral red and resazurin revealed that STb was toxic in all but the GLUTag cells. STb stimulated 2-hour GLP-2 secretion in all cell models. Interestingly, GLUTag cells produced the highest amount of GLP-2 when treated with STb, demonstrating an inverse relationship in GLP-2 secretion and cell toxicity. To demonstrate a protective role for GLP-2, GLUTag-conditioned media (rich in GLP-2) blocked STb toxicity in STC-1 cells. Confirming a protective role of GLP-2, teduglutide was able to improve cell viability in cells treated with H2O2. In conclusion, STb interacts with the L cell, stimulates secretion, and may induce toxicity if GLP-2 is not produced at high levels. GLP-2 or receptor agonists have the ability to improve cell viability in response to toxins. These results suggest that GLP-2 secretion can play a protective role during STb intoxication. This work supports future investigation into the use of GLP-2 therapies in enterotoxigenic-related diseases.


Assuntos
Enterotoxinas , Peptídeo 2 Semelhante ao Glucagon , Animais , Células Enteroendócrinas/metabolismo , Enterotoxinas/metabolismo , Enterotoxinas/toxicidade , Escherichia coli/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Peptídeo 2 Semelhante ao Glucagon/metabolismo , Peptídeo 2 Semelhante ao Glucagon/farmacologia , Temperatura Alta , Humanos , Peróxido de Hidrogênio/metabolismo , Células L , Camundongos
2.
Int J Mol Sci ; 23(3)2022 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-35163196

RESUMO

Inflammatory responses have been shown to induce hyperglycemia, yet the underlying mechanism is still largely unclear. GLP-1 is an important intestinal hormone for regulating glucose homeostasis; however, few studies have investigated the influence of digestive tract Salmonella infection on enteroendocrine L cell secretions. In this study, we established a model of Salmonella-infected piglets by oral gavage in order to analyze the effects of Salmonella infection on enteroendocrine L cell function. Furthermore, in vitro lipopolysaccharide (LPS) was administered to STC-1 cells to clarify its direct effect on GLP-1 secretion. The results showed that significantly increased blood glucose in the group of Salmonella-infected piglets was observed, and Salmonella infection decreased blood GLP-1 content. Then, ileal epithelium damage was observed by histological detection, and this was further verified by TUNEL staining. We identified activation of TLR signaling demonstrating up-regulated expressions of TLR4 and nuclear factor-kappa B (NF-ΚB). Furthermore, it was shown that Salmonella induced pyroptosis of enteroendocrine L cells and enhanced the secretion of IL-1ß through augmenting gene and protein expressions of NOD-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a carboxyl-terminal CARD (ASC), Caspase 1, and gasdermin D (GSDMD). Meanwhile, in vitro LPS treatment induced the pyroptosis of STC-1 cells and reduced the secretion of GLP-1. Altogether, the results demonstrated that Salmonella infection can reduce secretion of GLP-1 by inducing pyroptosis of intestinal L cells, which may eventually result in hyperglycemia. The results provided evidence for the cause of hyperglycemia induced by inflammation and shed new light on glucose homeostasis regulation.


Assuntos
Peptídeo 1 Semelhante ao Glucagon/metabolismo , Hiperglicemia/etiologia , Salmonelose Animal/metabolismo , Animais , Caspase 1/metabolismo , China , Células Enteroendócrinas/citologia , Células Enteroendócrinas/metabolismo , Hiperglicemia/patologia , Inflamassomos/metabolismo , Inflamação , Células L/metabolismo , Camundongos , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Piroptose/efeitos dos fármacos , Piroptose/fisiologia , Salmonella/patogenicidade , Transdução de Sinais , Suínos/microbiologia
3.
Nat Commun ; 13(1): 879, 2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-35169163

RESUMO

Dysregulation of the balance between pro-inflammatory and anti-inflammatory macrophages has a key function in the pathogenesis of Duchenne muscular dystrophy (DMD), a fatal genetic disease. We postulate that an evolutionarily ancient protective mechanism against infection, known as trained immunity, drives pathological inflammation in DMD. Here we show that bone marrow-derived macrophages from a murine model of DMD (mdx) exhibit cardinal features of trained immunity, consisting of transcriptional hyperresponsiveness associated with metabolic and epigenetic remodeling. The hyperresponsive phenotype is transmissible by bone marrow transplantation to previously healthy mice and persists for up to 11 weeks post-transplant. Mechanistically, training is induced by muscle extract in vitro. The functional and epigenetic changes in bone marrow-derived macrophages from dystrophic mice are TLR4-dependent. Adoptive transfer experiments further support the TLR4-dependence of trained macrophages homing to damaged muscles from the bone marrow. Collectively, this suggests that a TLR4-regulated, memory-like capacity of innate immunity induced at the level of the bone marrow promotes dysregulated inflammation in DMD.


Assuntos
Transplante de Medula Óssea , Imunidade Inata/imunologia , Macrófagos/imunologia , Músculo Esquelético/patologia , Distrofia Muscular de Duchenne/patologia , Receptor 4 Toll-Like/imunologia , Animais , Células da Medula Óssea/imunologia , Linhagem Celular , Modelos Animais de Doenças , Inflamação/imunologia , Células L , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Camundongos Knockout , Músculo Esquelético/imunologia , Distrofia Muscular de Duchenne/imunologia , Extratos de Tecidos/farmacologia , Transcrição Genética/genética
4.
J Vet Med Sci ; 84(2): 265-274, 2022 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-34980756

RESUMO

Glucagon-like peptides (GLPs) are secreted from intestinal L cells and stimulate various physiological functions in the gastrointestinal tract. The secretion of GLPs is influenced by macronutrient ingestion. This study aims to clarify the effects of dietary carbohydrate (CHO) on L cells in the chicken ileum. Six-week-old, male White Leghorn chickens were divided into three groups: control, low-CHO and CHO-free, with five chickens in each group. Paraffin sections were made from the proximal and distal ileum of each animal and subjected to immunohistochemistry for GLP-1 and GLP-2 peptides and in situ hybridization for proglucagon (PG) mRNA. A significant reduction of GLP-1- and GLP-2-immunoreactive cells was observed in the two experimental groups compared with that in the control. A reduction of cells expressing PG mRNA was observed in the proximal and distal ileum of the CHO-free group compared with that in the control. The ratio of GLP-1-immunoreactive cells showing Ki-67 immunoreactivity was significantly lower in the distal ileum of the CHO-free group than that in the control group. These data suggest that dietary CHO is an effective stimulator for modifying L cell density in the chicken ileum.


Assuntos
Galinhas , Carboidratos da Dieta , Animais , Carboidratos da Dieta/farmacologia , Células Enteroendócrinas , Íleo , Células L , Masculino , Camundongos
5.
Cell Mol Life Sci ; 79(1): 62, 2022 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-35001155

RESUMO

Availability of iron is a key factor in the survival and multiplication of Mycobacterium tuberculosis (M.tb) within host macrophage phagosomes. Despite host cell iron regulatory machineries attempts to deny supply of this essential micronutrient, intraphagosomal M.tb continues to access extracellular iron. In the current study, we report that intracellular M.tb exploits mammalian secreted Glyceraldehyde 3-phosphate dehydrogenase (sGAPDH) for the delivery of host iron carrier proteins lactoferrin (Lf) and transferrin (Tf). Studying the trafficking of iron carriers in infected cells we observed that sGAPDH along with the iron carrier proteins are preferentially internalized into infected cells and trafficked to M.tb containing phagosomes where they are internalized by resident mycobacteria resulting in iron delivery. Collectively our findings provide a new mechanism of iron acquisition by M.tb involving the hijack of host sGAPDH. This may contribute to its successful pathogenesis and provide an option for targeted therapeutic intervention.


Assuntos
Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Ferro/metabolismo , Lactoferrina/metabolismo , Mycobacterium tuberculosis/metabolismo , Transferrina/metabolismo , Animais , Transporte Biológico/fisiologia , Linhagem Celular Tumoral , Humanos , Células L , Camundongos , Camundongos Endogâmicos C57BL , Fagossomos/metabolismo , Células THP-1 , Tuberculose/patologia
6.
Carbohydr Polym ; 280: 119032, 2022 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-35027134

RESUMO

This study was aimed at preparing O-carboxymethyl chitosan (CM-CTS) fabrics, and examining the wound healing effects on partial-thickness burn. The functional polysaccharides were produced from chitosan needle-punched nonwovens reacted with chloroacetic acid. Then the biocompatibility and biological functions were evaluated through fibroblast L-929 and SD rats. CM-CTS fabrics were obtained with elongation at break more than 42%, tensile strength reaching 0.65 N/mm2, and water vapor transmission rate about 2600 g/m2∙24 h. Moreover, CM-CTS fabrics could effectively promote the mouse L-929 migration in vitro. CM-CTS fabrics yielded satisfactory results in angiogenesis, collagen deposition, interleukin-6 content, transforming growth factor level and healing rate, which were superior to the positive control and model groups after rats suffering with partial-thickness burn. In conclusion, CM-CTS fabrics possessed proper mechanical properties, air permeability, favorable biocompatibility, acceleration on fibroblasts migration and healing capacity for partial-thickness burn injury, and owned good potential as high-quality wound dressing.


Assuntos
Bandagens , Materiais Biocompatíveis , Queimaduras/terapia , Quitosana/análogos & derivados , Cicatrização , Animais , Antígenos CD34/análise , Movimento Celular , Quitosana/química , Quitosana/farmacologia , Quitosana/toxicidade , Feminino , Fibroblastos/citologia , Fibroblastos/fisiologia , Interleucina-6/sangue , Células L , Masculino , Camundongos , Ratos , Ratos Sprague-Dawley , Fator de Crescimento Transformador beta1/sangue
7.
Cell Rep ; 38(1): 110184, 2022 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-34986349

RESUMO

MV130 is an inactivated polybacterial mucosal vaccine that confers protection to patients against recurrent respiratory infections, including those of viral etiology. However, its mechanism of action remains poorly understood. Here, we find that intranasal prophylaxis with MV130 modulates the lung immune landscape and provides long-term heterologous protection against viral respiratory infections in mice. Intranasal administration of MV130 provides protection against systemic candidiasis in wild-type and Rag1-deficient mice lacking functional lymphocytes, indicative of innate immune-mediated protection. Moreover, pharmacological inhibition of trained immunity with metformin abrogates the protection conferred by MV130 against influenza A virus respiratory infection. MV130 induces reprogramming of both mouse bone marrow progenitor cells and in vitro human monocytes, promoting an enhanced cytokine production that relies on a metabolic shift. Our results unveil that the mucosal administration of a fully inactivated bacterial vaccine provides protection against viral infections by a mechanism associated with the induction of trained immunity.


Assuntos
Vacinas Bacterianas/imunologia , Imunidade nas Mucosas/imunologia , Infecções por Orthomyxoviridae/prevenção & controle , Mucosa Respiratória/imunologia , Infecções Respiratórias/prevenção & controle , Administração Intranasal , Animais , Anticorpos Antivirais/imunologia , Bactérias/imunologia , Vacinas Bacterianas/administração & dosagem , Candidíase/prevenção & controle , Linhagem Celular , Chlorocebus aethiops , Citocinas/biossíntese , Humanos , Vírus da Influenza A/imunologia , Células L , Pulmão/imunologia , Metformina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/imunologia , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/virologia , Infecções Respiratórias/microbiologia , Infecções Respiratórias/virologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia
8.
Cell Mol Gastroenterol Hepatol ; 13(4): 1057-1072, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34902628

RESUMO

BACKGROUND & AIMS: Compelling evidence shows that glucagon-like peptide-1 (GLP-1) has a profound effect in restoring normoglycemia in type 2 diabetic patients by increasing pancreatic insulin secretion. Although L-cells are the primary source of circulating GLP-1, the current therapies do not target L-cells to increase GLP-1 levels. Our study aimed to determine the molecular underpinnings of GLP-1 secretion as an impetus to identify new interventions to target endogenous L-cells. METHODS: We used genetic mouse models of intestine-specific overexpression of hypoxia-inducible factor (HIF)-1α and HIF-2α (VhlΔIE), conditional overexpression of intestinal HIF-2α (Hif-2αLSL;Vilin-Cre/ERT2), and intestine-specific HIF-2α knockout mice (Hif-2αΔIE) to show that HIF signaling, especially HIF-2α, regulates GLP-1 secretion. RESULTS: Our data show that intestinal HIF signaling improved glucose homeostasis in a GLP-1-dependent manner. Intestinal HIF potentiated GLP-1 secretion via the lipid sensor G-protein-coupled receptor (GPR)40 enriched in L-cells. We show that HIF-2α regulates GPR40 in L-cells and potentiates fatty acid-induced GLP-1 secretion via extracellular regulated kinase (ERK). Using a genetic model of intestine-specific overexpression of HIF-2α, we show that HIF-2α is sufficient to increase GLP-1 levels and attenuate diet-induced metabolic perturbations such as visceral adiposity, glucose intolerance, and hepatic steatosis. Lastly, we show that intestinal HIF-2α signaling acts as a priming mechanism crucial for postprandial lipid-mediated GLP-1 secretion. Thus, disruption of intestinal HIF-2α decreases GLP-1 secretion. CONCLUSIONS: In summary, we show that intestinal HIF signaling, particularly HIF-2α, regulates the lipid sensor GPR40, which is crucial for the lipid-mediated GLP-1 secretion, and suggest that HIF-2α is a potential target to induce endogenous GLP-1 secretion.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Peptídeo 1 Semelhante ao Glucagon , Intestinos , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Humanos , Células L , Lipídeos , Camundongos
9.
Sci Rep ; 11(1): 23813, 2021 12 10.
Artigo em Inglês | MEDLINE | ID: mdl-34893681

RESUMO

Roux-en-Y gastric bypass (RYGB)-induced glycemic improvement is associated with increases in glucagon-like-peptide-1 (GLP-1) secreted from ileal L-cells. We analyzed changes in ileal bile acids and ileal microbial composition in diet-induced-obesity rats after RYGB or sham surgery to elucidate the early and late effects on L-cells and glucose homeostasis. In early cohorts, there were no significant changes in L-cell density, GLP-1 or glucose tolerance. In late cohorts, RYGB demonstrated less weight regain, improved glucose tolerance, increased L-cell density, and increased villi height. No difference in the expression of GLP-1 genes was observed. There were lower concentrations of ileal bile acids in the late RYGB cohort. Microbial analysis demonstrated decreased alpha diversity in early RYGB cohorts which normalized in the late group. The early RYGB cohorts had higher abundances of Escherichia-Shigella but lower abundances of Lactobacillus, Adlercreutzia, and Proteus while the late cohorts demonstrated higher abundances of Escherichia-Shigella and lower abundances of Lactobacillus. Shifts in Lactobacillus and Escherichia-Shigella correlated with decreases in multiple conjugated bile acids. In conclusion, RYGB caused a late and substantial increase in L-cell quantity with associated changes in bile acids which correlated to shifts in Escherichia-Shigella and Lactobacillus. This proliferation of L-cells contributed to improved glucose homeostasis.


Assuntos
Ácidos e Sais Biliares/metabolismo , Derivação Gástrica/efeitos adversos , Microbioma Gastrointestinal , Glucose/metabolismo , Íleo/metabolismo , Íleo/microbiologia , Células L/metabolismo , Animais , Biomarcadores , Glicemia , Contagem de Células , Suscetibilidade a Doenças , Derivação Gástrica/métodos , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Camundongos , Ratos
10.
World J Microbiol Biotechnol ; 37(11): 193, 2021 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-34642826

RESUMO

The antimicrobial efficacy of rhamnolipid is well established against a wide range of pathogens. However little is known about the enhancement of antimicrobial efficacy of rhamnolipid in the form of nanoparticles. With a curiosity of enhancing antimicrobial activity, a study has been carried out to evaluate the antimicrobial efficacy of rhamnolipid-coated zinc oxide nanoparticles. The zinc oxide nanoparticles were synthesized with rhamnolipid, produced by Pseudomonas aeruginosa JS29. The rhamnolipid-coated zinc oxide nanoparticles were characterized by FTIR, XRD, TGA, TEM, and SAED. The antimicrobial and antibiofilm efficacy of the nanoparticles was evaluated against Staphylococcus aureus MTCC 96. FTIR, XRD, TEM, and SAED analyses confirmed that the nanoparticles contain both rhamnolipid and zinc as constituents and are polycrystalline with sizes ranging from 40 to 50 nm. At a concentration of 250 µg/ml, rhamnolipid-coated zinc oxide nanoparticles exhibited 80% growth inhibition of the pathogen. Again, at the same concentration, the nanoparticle was observed to inhibit 78% of biofilm formation while disrupting 100% of preformed biofilm. The nanoparticles demonstrated an enhanced inhibitory and antibiofilm efficacy against the pathogen compared to the individual effect of both rhamnolipid and zinc oxide nanoparticles. With the established non-toxicity of rhamnolipid-coated zinc oxide nanoparticles in fibroblast cell lines, the nanoparticles could be a promising pharmaceutical alternative.


Assuntos
Antibacterianos/farmacologia , Glicolipídeos/farmacologia , Nanopartículas Metálicas , Staphylococcus aureus/efeitos dos fármacos , Óxido de Zinco/farmacologia , Animais , Antibacterianos/síntese química , Antibacterianos/química , Biofilmes/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Glicolipídeos/química , Células L , Nanopartículas Metálicas/química , Camundongos , Testes de Sensibilidade Microbiana , Viabilidade Microbiana , Staphylococcus aureus/crescimento & desenvolvimento , Óxido de Zinco/química
11.
Front Endocrinol (Lausanne) ; 12: 690387, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34421821

RESUMO

The molecular sensors underlying nutrient-stimulated GLP-1 secretion are currently being investigated. Peripheral administration of melanocortin-4 receptor (MC4R) agonists have been reported to increase GLP-1 plasma concentrations in mice and humans but it is unknown whether this effect results from a direct effect on the GLP-1 secreting L-cells in the intestine, from other effects in the intestine or from extra-intestinal effects. We investigated L-cell expression of MC4R in mouse and human L-cells by reanalyzing publicly available RNA sequencing databases (mouse and human) and by RT-qPCR (mouse), and assessed whether administration of MC4R agonists to a physiologically relevant gut model, isolated perfused mouse and rat small intestine, would stimulate GLP-1 secretion or potentiate glucose-stimulated secretion. L-cell MC4R expression was low in mouse duodenum and hardly detectable in the ileum and MC4R expression was hardly detectable in human L-cells. In isolated perfused mouse and rat intestine, neither intra-luminal nor intra-arterial administration of NDP-alpha-MSH, a potent MC4R agonist, had any effect on GLP-1 secretion (P ≥0.98, n = 5-6) from the upper or lower-half of the small intestine in mice or in the lower half in rats. Furthermore, HS014-an often used MC4R antagonist, which we found to be a partial agonist-did not affect the glucose-induced GLP-1 response in the rat, P = 0.62, n = 6). Studies on transfected COS7-cells confirmed bioactivity of the used compounds and that concentrations employed were well within in the effective range. Our combined data therefore suggest that MC4R-activated GLP-1 secretion in rodents either exclusively occurs in the colon or involves extra-intestinal signaling.


Assuntos
Peptídeo 1 Semelhante ao Glucagon/metabolismo , Intestino Delgado/metabolismo , Células L/metabolismo , Receptor Tipo 4 de Melanocortina/metabolismo , Animais , Células COS , Chlorocebus aethiops , Bases de Dados Factuais , Humanos , Intestino Delgado/efeitos dos fármacos , Células L/efeitos dos fármacos , Masculino , Camundongos , Ratos , Ratos Wistar , Receptor Tipo 4 de Melanocortina/agonistas , Transdução de Sinais/efeitos dos fármacos , alfa-MSH/farmacologia
12.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34206340

RESUMO

Intestinal organoids are used to analyze the differentiation of enteroendocrine cells (EECs) and to manipulate their density for treating type 2 diabetes. EEC differentiation is a continuous process tightly regulated in the gut by a complex regulatory network. However, the effect of chronic hyperglycemia, in the modulation of regulatory networks controlling identity and differentiation of EECs, has not been analyzed. This study aimed to investigate the effect of glucotoxicity on EEC differentiation in small intestinal organoid platforms. Mouse intestinal organoids were cultured in the presence/absence of high glucose concentrations (35 mM) for 48 h to mimic glucotoxicity. Chronic hyperglycemia impaired the expression of markers related to the differentiation of EEC progenitors (Ngn3) and L-cells (NeuroD1), and it also reduced the expression of Gcg and GLP-1 positive cell number. In addition, the expression of intestinal stem cell markers was reduced in organoids exposed to high glucose concentrations. Our data indicate that glucotoxicity impairs L-cell differentiation, which could be associated with decreased intestinal stem cell proliferative capacity. This study provides the identification of new targets involved in new molecular signaling mechanisms impaired by glucotoxicity that could be a useful tool for the treatment of type 2 diabetes.


Assuntos
Diferenciação Celular , Células Enteroendócrinas/metabolismo , Hiperglicemia/complicações , Intestino Delgado/metabolismo , Organoides , Animais , Diabetes Mellitus Tipo 2/complicações , Células Enteroendócrinas/efeitos dos fármacos , Células Enteroendócrinas/fisiologia , Glucose/metabolismo , Glucose/toxicidade , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/fisiopatologia , Células L , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais
13.
Mol Cell Endocrinol ; 535: 111398, 2021 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-34274446

RESUMO

BACKGROUND: The mechanochemical enzyme dynamin mediates endocytosis and regulates neuroendocrine cell exocytosis. Enteroendocrine L cells co-secrete the anorectic gut hormones glucagon-like peptide 1 (GLP-1) and peptide YY (PYY) postprandially and is a potential therapeutic target for metabolic diseases. In the present study, we aimed to determine if dynamin is implicated in human L cell secretion. METHODS: Western blot was performed on the murine L cell line GLUTag. Static incubation of human colonic mucosae with activators and inhibitors of dynamin was carried out. GLP-1 and PYY contents of the secretion supernatants were assayed using ELISA. RESULTS AND CONCLUSION: s: Both dynamin I and II are expressed in GLUTag cells. The dynamin activator Ryngo 1-23 evoked significant GLP-1 and PYY release from human colonic mucosae while the dynamin inhibitor Dynole 3-42 significantly inhibited release triggered by known L cell secretagogues. Thus, the cell signaling regulator dynamin is able to bi-directionally regulate L cell hormone secretion in the human gut and may represent a novel target for gastrointestinal-targeted metabolic drug development.


Assuntos
Dinamina II/metabolismo , Dinamina I/metabolismo , Células Enteroendócrinas/citologia , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Mucosa Intestinal/citologia , Peptídeo YY/metabolismo , Adulto , Idoso , Animais , Células Cultivadas , Meios de Cultura/química , Cianoacrilatos/farmacologia , Células Enteroendócrinas/efeitos dos fármacos , Células Enteroendócrinas/metabolismo , Feminino , Humanos , Indóis/farmacologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Células L , Masculino , Camundongos , Pessoa de Meia-Idade , Tirfostinas/farmacologia
14.
Commun Biol ; 4(1): 753, 2021 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-34140629

RESUMO

Metastatic cancer cells are frequently deficient in WWOX protein or express dysfunctional WWOX (designated WWOXd). Here, we determined that functional WWOX-expressing (WWOXf) cells migrate collectively and expel the individually migrating WWOXd cells. For return, WWOXd cells induces apoptosis of WWOXf cells from a remote distance. Survival of WWOXd from the cell-to-cell encounter is due to activation of the survival IκBα/ERK/WWOX signaling. Mechanistically, cell surface epitope WWOX286-299 (repl) in WWOXf repels the invading WWOXd to undergo retrograde migration. However, when epitope WWOX7-21 (gre) is exposed, WWOXf greets WWOXd to migrate forward for merge. WWOX binds membrane type II TGFß receptor (TßRII), and TßRII IgG-pretreated WWOXf greet WWOXd to migrate forward and merge with each other. In contrast, TßRII IgG-pretreated WWOXd loses recognition by WWOXf, and WWOXf mediates apoptosis of WWOXd. The observatons suggest that normal cells can be activated to attack metastatic cancer cells. WWOXd cells are less efficient in generating Ca2+ influx and undergo non-apoptotic explosion in response to UV irradiation in room temperature. WWOXf cells exhibit bubbling cell death and Ca2+ influx effectively caused by UV or apoptotic stress. Together, membrane WWOX/TßRII complex is needed for cell-to-cell recognition, maintaining the efficacy of Ca2+ influx, and control of cell invasiveness.


Assuntos
Invasividade Neoplásica/fisiopatologia , Metástase Neoplásica/patologia , Neoplasias/patologia , Receptor do Fator de Crescimento Transformador beta Tipo II/metabolismo , Oxidorredutase com Domínios WW/metabolismo , Animais , Apoptose/imunologia , Células COS , Cálcio/metabolismo , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Chlorocebus aethiops , Células HCT116 , Humanos , Células L , Células MCF-7 , Camundongos , Inibidor de NF-kappaB alfa/metabolismo , Neoplasias/genética , Transdução de Sinais/fisiologia , Oxidorredutase com Domínios WW/genética
15.
Front Endocrinol (Lausanne) ; 12: 694284, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34168620

RESUMO

Synthetic glucagon-like peptide-1 (GLP-1) analogues are effective anti-obesity and anti-diabetes drugs. The beneficial actions of GLP-1 go far beyond insulin secretion and appetite, and include cardiovascular benefits and possibly also beneficial effects in neurodegenerative diseases. Considerable reserves of GLP-1 are stored in intestinal endocrine cells that potentially might be mobilized by pharmacological means to improve the body's metabolic state. In recognition of this, the interest in understanding basic L-cell physiology and the mechanisms controlling GLP-1 secretion, has increased considerably. With a view to home in on what an L-cell is, we here present an overview of available data on L-cell development, L-cell peptide expression profiles, peptide production and secretory patterns of L-cells from different parts of the gut. We conclude that L-cells differ markedly depending on their anatomical location, and that the traditional definition of L-cells as a homogeneous population of cells that only produce GLP-1, GLP-2, glicentin and oxyntomodulin is no longer tenable. We suggest to sub-classify L-cells based on their differential peptide contents as well as their differential expression of nutrient sensors, which ultimately determine the secretory responses to different stimuli. A second purpose of this review is to describe and discuss the most frequently used experimental models for functional L-cell studies, highlighting their benefits and limitations. We conclude that no experimental model is perfect and that a comprehensive understanding must be built on results from a combination of models.


Assuntos
Células L/fisiologia , Via Secretória/fisiologia , Animais , Endocrinologia/métodos , Humanos , Células L/metabolismo , Camundongos , Projetos de Pesquisa
16.
Endocr Pathol ; 32(4): 433-441, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34041698

RESUMO

This morphological and immunohistochemical study demonstrates that tumors currently known as "middle ear adenomas" are truly well-differentiated epithelial neuroendocrine tumors (NETs) composed of cells comparable to normal intestinal L cells, and therefore, these tumors resemble hindgut NETs. These tumors show consistent expression of glucagon, pancreatic polypeptide, PYY, and the transcription factor SATB2, as well as generic neuroendocrine markers and keratins. The same L cell markers are expressed by cells within the normal middle ear epithelium. These markers define a valuable immunohistochemical profile that can be used for differential diagnosis of middle ear neoplasms, particularly in distinguishing epithelial NETs from paragangliomas. The discovery of neuroendocrine cells expressing the same markers in non-neoplastic middle ear mucosa opens new areas of investigation into the physiology of the normal middle ear and the pathophysiology of middle ear disorders.


Assuntos
Adenoma/diagnóstico , Neoplasias da Orelha/diagnóstico , Orelha Média/patologia , Células L/fisiologia , Tumores Neuroendócrinos/diagnóstico , Adenoma/classificação , Adenoma/metabolismo , Adenoma/patologia , Adulto , Idoso , Animais , Diferenciação Celular , Diagnóstico Diferencial , Neoplasias da Orelha/classificação , Neoplasias da Orelha/metabolismo , Neoplasias da Orelha/patologia , Orelha Média/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Células L/metabolismo , Células L/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Tumores Neuroendócrinos/classificação , Tumores Neuroendócrinos/metabolismo , Tumores Neuroendócrinos/patologia , Estudos Retrospectivos , Terminologia como Assunto
17.
J Virol Methods ; 293: 114164, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33864853

RESUMO

Lumpy skin disease virus (LSDV), sheeppox virus (SPPV) and goatpox (GTPV) virus have been usually grown on primary cells for diagnosis, production and titration purposes. The use of primary cells present several inconvenient, heavy preparation, heterogeneous cell population, non-reproducible viral titration and presence of potential endogenous contaminants. Therefore investigating sensitivity of candidate continuous cell lines is needed. In this study, we compared the above Capripox viruses (CaPVs) sensitivity of primary cells of four origin (heart, skin, testis and kidney), with three cell lines (Vero, OA3.Ts and ESH-L). We tested sensitivity for virus isolation, replication cycle and titration, revealed by cytopathic effect (CPE), immunoenzymatic staining and immunofluorescence. Our results show that ESH-L cells and primary fetal heart cells present the highest sensitivity for CaPVs growth and detection. Vero cells can replicate those viruses but without showing any CPE while the titer obtained on OA3.Ts is lower than primary and ESH-L cells. ESH-L cells are an effective alternative to primary cells use for growing Capripoxviruses and their diagnosis.


Assuntos
Capripoxvirus , Doenças das Cabras , Doença Nodular Cutânea , Doenças dos Ovinos , Animais , Bovinos , Chlorocebus aethiops , Cabras , Células L , Masculino , Camundongos , Filogenia , Ovinos , Células Vero
18.
Genes Cells ; 26(3): 136-151, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33501714

RESUMO

The molecular mechanisms underlying neurodevelopmental disorders (NDDs) remain unclear. We previously identified Down syndrome cell adhesion molecule like 1 (Dscaml1) as a responsible gene for Ihara epileptic rat (IER), a rat model for human NDDs with epilepsy. However, the relationship between NDDs and DSCAML1 in humans is still elusive. In this study, we screened databases of autism spectrum disorders (ASD), intellectual disability (ID)/developmental disorders (DD) and schizophrenia for genomic mutations in human DSCAML1. We then performed in silico analyses to estimate the potential damage to the mutated DSCAML1 proteins and chose three representative mutations (DSCAML1C729R , DSCAML1R1685* and DSCAML1K2108Nfs*37 ), which lacked a cysteine residue in the seventh Ig domain, the intracellular region and the C-terminal PDZ-binding motif, respectively. In overexpression experiments in a cell line, DSCAML1C729R lost its mature N-glycosylation, whereas DSCAML1K2108Nfs*37 was abnormally degraded via proteasome-dependent protein degradation. Furthermore, in primary hippocampal neurons, the ability of the wild-type DSCAML1 to regulate the number of synapses was lost with all mutant proteins. These results provide insight into understanding the roles of the domains in the DSCAML1 protein and further suggest that these mutations cause functional changes, albeit through different mechanisms, that likely affect the pathophysiology of NDDs.


Assuntos
Moléculas de Adesão Celular/genética , Mutação/genética , Transtornos do Neurodesenvolvimento/genética , Animais , Transtorno do Espectro Autista/genética , Adesão Celular , Membrana Celular/metabolismo , Espinhas Dendríticas/metabolismo , Feminino , Glicosilação , Hipocampo/patologia , Humanos , Células L , Masculino , Camundongos , Anotação de Sequência Molecular , Proteínas Mutantes/metabolismo , Proteólise , Ratos Wistar , Esquizofrenia/genética , Sinapses/metabolismo
19.
Cell Death Dis ; 12(1): 113, 2021 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-33479193

RESUMO

In the status of obesity, the glucagon-like peptide-1 (GLP-1) level usually declines and results in metabolic syndrome. This study aimed to investigate the intracellular mechanism of GLP-1 synthesis in L cells from the perspective of microRNA (miRNA). In the present study, we found that GLP-1 level was down-regulated in the plasma and ileum tissues of obese mice, while the ileac miR-194 expression was up-regulated. In vitro experiments indicated that miR-194 overexpression down-regulated GLP-1 level, mRNA levels of proglucagon gene (gcg) and prohormone convertase 1/3 gene (pcsk1), and the nuclear protein level of beta-catenin (ß-catenin). Further investigation confirmed that ß-catenin could promote gcg transcription through binding to transcription factor 7-like 2 (TCF7L2). miR-194 suppressed gcg mRNA level via negatively regulating TCF7L2 expression. What's more, forkhead box a1 (Foxa1) could bind to the promoter of pcsk1 and enhanced its transcription. miR-194 suppressed pcsk1 transcription through targeting Foxa1. Besides, the interference of miR-194 reduced palmitate (PA)-induced cell apoptosis and the anti-apoptosis effect of miR-194 inhibitor was abolished by TCF7L2 knockdown. Finally, in HFD-induced obese mice, the silence of miR-194 significantly elevated GLP-1 level and improved the metabolic symptoms caused by GLP-1 deficiency. To sum up, our study found that miR-194 suppressed GLP-1 synthesis in L cells via inhibiting TCF7L2-mediated gcg transcription and Foxa1-mediated pcsk1 transcription. Meanwhile, miR-194 took part in the PA-induced apoptosis of L cells.


Assuntos
Células Enteroendócrinas/metabolismo , Peptídeo 1 Semelhante ao Glucagon/biossíntese , MicroRNAs/metabolismo , Obesidade/metabolismo , Animais , Peptídeo 1 Semelhante ao Glucagon/genética , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Células L , Masculino , Camundongos , MicroRNAs/genética , Obesidade/genética , Transfecção
20.
Res Microbiol ; 172(1): 103787, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33049327

RESUMO

Staphylococcus aureus and Staphylococcus epidermidis are among the most important bacterial species responsible for biofilm formation on indwelling medical devices, including orthopaedic implants. The increasing resistance to antimicrobials, partly attributed to the ability to form biofilms, is a challenge for the development of new antimicrobial agents. In this study, the cell-free supernatant obtained from sponge-associated Enterobacter strain 84.3 culture inhibited biofilm formation (>65%) and dissociated mature biofilm (>85%) formed by S. aureus and S. epidermidis strains. The culture supernatant was subjected to solvent partitioning and the aqueous extract presented a concentration-dependent antibiofilm activity for each strain with a minimum biofilm eradication concentration (MBEC) ranging from 16 to 256 µg/mL. The effect of the aqueous extract on mature S. aureus biofilm was analyzed by confocal scanning laser microscopy, showing a significant reduction of the biofilm layer as well as diminished interactions among the cells. This extract is not toxic for mammalian cells (L929 cell line). Studies targeting substances with antibiofilm activity gained significant attention in recent years due to difficult-to-treat biofilm infections. Here, sponge-associated Enterobacter 84.3 proved to be a source of substances capable of eradicating staphylococcal biofilm, with potential medical use in the future.


Assuntos
Antibacterianos/farmacologia , Biofilmes/crescimento & desenvolvimento , Extratos Celulares/farmacologia , Enterobacter/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus epidermidis/efeitos dos fármacos , Animais , Infecções Relacionadas a Cateter/tratamento farmacológico , Infecções Relacionadas a Cateter/microbiologia , Cateteres de Demora/microbiologia , Linhagem Celular , Infecção Hospitalar/tratamento farmacológico , Infecção Hospitalar/microbiologia , Células L , Camundongos , Testes de Sensibilidade Microbiana , Poríferos/microbiologia , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/prevenção & controle
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