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1.
Pestic Biochem Physiol ; 167: 104592, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32527425

RESUMO

Bacillus thuringiensis (Bt) Cry1Fa and Cry1Ab proteins are important Cry toxins due to their high, selective toxicity against a number of lepidopteran species, including important pests of corn and cotton. Competition binding assays are a classical tool for investigating Cry toxin interactions with target pest insects. We developed a fluorescence-based binding assay and assessed Cry1Fa and Cry1Ab toxin binding to brush border membrane preparations from lepidopteran corn pests including Ostrinia nubilalis (European corn borer, ECB), Diatraea grandiosella (south western corn borer, SWCB), and Helicoverpa zea (corn earworm, CEW). Homologous and heterologous competition binding assays with fluorophore-(Alexa488)-labeled Cry1Fa toxin showed that Cry1Fa shares binding site(s) with Cry1Ab toxin in ECB, and SWCB for which Cry1Ab has higher affinity than Cry1Fa. Apart from the shared binding sites, Cry1Ab and Cry1Fa bind an additional site(s) in ECB and SWCB. In CEW, Cry1Fa and Cry1Ab each, has a high affinity binding site(s), which binds the heterologous toxin with low affinity. The Cry1Ab-Cry1Fa toxin binding models for ECB, SWCB and CEW based on our results are considered in the context of what is known about acquired cross-resistance against Cry1Ab and Cry1Fa toxins.


Assuntos
Bacillus thuringiensis , Mariposas , Animais , Proteínas de Bactérias , Sítios de Ligação , Endotoxinas , Proteínas Hemolisinas , Larva , Microvilosidades , Zea mays
2.
PLoS One ; 15(4): e0231423, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32302323

RESUMO

Recent advances in canine intestinal organoids have expanded the option for building a better in vitro model to investigate translational science of intestinal physiology and pathology between humans and animals. However, the three-dimensional geometry and the enclosed lumen of canine intestinal organoids considerably hinder the access to the apical side of epithelium for investigating the nutrient and drug absorption, host-microbiome crosstalk, and pharmaceutical toxicity testing. Thus, the creation of a polarized epithelial interface accessible from apical or basolateral side is critical. Here, we demonstrated the generation of an intestinal epithelial monolayer using canine biopsy-derived colonic organoids (colonoids). We optimized the culture condition to form an intact monolayer of the canine colonic epithelium on a nanoporous membrane insert using the canine colonoids over 14 days. Transmission and scanning electron microscopy revealed a physiological brush border interface covered by the microvilli with glycocalyx, as well as the presence of mucin granules, tight junctions, and desmosomes. The population of stem cells as well as differentiated lineage-dependent epithelial cells were verified by immunofluorescence staining and RNA in situ hybridization. The polarized expression of P-glycoprotein efflux pump was confirmed at the apical membrane. Also, the epithelial monolayer formed tight- and adherence-junctional barrier within 4 days, where the transepithelial electrical resistance and apparent permeability were inversely correlated. Hence, we verified the stable creation, maintenance, differentiation, and physiological function of a canine intestinal epithelial barrier, which can be useful for pharmaceutical and biomedical researches.


Assuntos
Colo/citologia , Células Epiteliais/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Animais , Diferenciação Celular , Linhagem da Célula , Células Cultivadas , Desmossomos/metabolismo , Cães , Células Epiteliais/citologia , Células Epiteliais/ultraestrutura , Membranas Artificiais , Microvilosidades/fisiologia , Mucinas/metabolismo , Nanoporos , Células-Tronco/citologia , Células-Tronco/metabolismo , Junções Íntimas/metabolismo
3.
Food Chem Toxicol ; 135: 110896, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31654707

RESUMO

Metal oxide nanoparticles (NP) are increasingly used in the food and agriculture industries, making human consumption nearly unavoidable. The goal of this study was to use the Gallus gallus (broiler chicken) intra-amniotic administration of physiologically relevant concentrations of TiO2, SiO2, and ZnO to better understand the effects of NP exposure on gut health and function. Immediately after hatch, blood, cecum, and small intestine were collected for assessment of iron (Fe)-metabolism, zinc (Zn)-metabolism, brush border membrane (BBM) functional, and pro-inflammatory related proteins gene expression; blood Fe and Zn levels; cecum weight; and the relative abundance of intestinal microflora. NP type, dose, and the presence or absence of minerals was shown to result in altered mineral transporter, BBM functional, and pro-inflammatory gene expression. Metal oxide NP also altered the abundance of intestinal bacterial populations. Overall, the data suggest that the in vivo results align with in vitro studies, and that NP have the potential to negatively affect intestinal functionality and health.


Assuntos
Microbioma Gastrointestinal/efeitos dos fármacos , Nanopartículas Metálicas/toxicidade , Microvilosidades/efeitos dos fármacos , Dióxido de Silício/toxicidade , Titânio/toxicidade , Óxido de Zinco/toxicidade , Âmnio , Animais , Ceco/efeitos dos fármacos , Ceco/microbiologia , Galinhas , Injeções , Nanopartículas Metálicas/administração & dosagem , Tamanho do Órgão/efeitos dos fármacos , Dióxido de Silício/administração & dosagem , Titânio/administração & dosagem , Óxido de Zinco/administração & dosagem
4.
Am J Physiol Gastrointest Liver Physiol ; 318(1): G53-G65, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31682159

RESUMO

Detection of nutritional and noxious food components in the gut is a crucial component of gastrointestinal function. Contents in the gut lumen interact with enteroendocrine cells dispersed throughout the gut epithelium. Enteroendocrine cells release many different hormones, neuropeptides, and neurotransmitters that communicate either directly or indirectly with the central nervous system and the enteric nervous system, a network of neurons and glia located within the gut wall. Several populations of enteric neurons extend processes that innervate the gastrointestinal lamina propria; however, how these processes develop and begin to transmit information from the mucosa is not fully understood. In this study, we found that Tuj1-immunoreactive neurites begin to project out of the myenteric plexus at embryonic day (E)13.5 in the mouse small intestine, even before the formation of villi. Using live calcium imaging, we discovered that neurites were capable of transmitting electrical information from stimulated villi to the plexus by E15.5. In unpeeled gut preparations where all layers were left intact, we also mimicked the basolateral release of 5-HT from enteroendocrine cells, which triggered responses in myenteric cell bodies at postnatal day (P)0. Altogether, our results show that enteric neurons extend neurites out of the myenteric plexus early during mouse enteric nervous system development, innervating the gastrointestinal mucosa, even before villus formation in mice of either sex. Neurites are already able to conduct electrical information at E15.5, and responses to 5-HT develop postnatally.NEW & NOTEWORTHY How enteric neurons project into the gut mucosa and begin to communicate with the epithelium during development is not known. Our study shows that enteric neurites project into the lamina propria as early as E13.5 in the mouse, before development of the submucous plexus and before formation of intestinal villi. These neurites are capable of transmitting electrical signals back to their cell bodies by E15.5 and respond to serotonin applied to neurite terminals by birth.


Assuntos
Mucosa Intestinal/inervação , Intestino Delgado/inervação , Microvilosidades/fisiologia , Plexo Mientérico/crescimento & desenvolvimento , Neuritos/fisiologia , Neurogênese , Animais , Células Enteroendócrinas/metabolismo , Células Enteroendócrinas/fisiologia , Potenciais Evocados , Feminino , Idade Gestacional , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Plexo Mientérico/efeitos dos fármacos , Plexo Mientérico/metabolismo , Neuritos/efeitos dos fármacos , Neuritos/metabolismo , Serotonina/farmacologia , Tubulina (Proteína)/metabolismo
5.
J Agric Food Chem ; 68(1): 160-167, 2020 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-31825618

RESUMO

Inflammatory bowel disease (IBD) is a chronic inflammatory disease of intestinal mucosa and submucosa, characterized by the disruption of the intestinal epithelial barrier, increased production of inflammatory mediators, and excessive tissue injury. Intestinal epithelial cells, as well as microvascular endothelial cells, play important roles in IBD. To study the potential effects of kaempferol in IBD progress, we established a novel epithelial-endothelial cells coculture model to investigate the intestinal inflammation and barrier function. Data demonstrated an obvious increased transepithelial electrical resistance (TEER) (1222 ± 60.40 Ω cm2 vs 1371 ± 38.77 Ω cm2), decreased flux of FITC (180.8 ± 20.06 µg/mL vs 136.7 ± 14.78 µg/mL), and up-regulated occludin and claudin-2 expression in Caco-2 that was specifically cocultured with endothelial cells. Meanwhile, 80 µM kaempferol alleviated the drop of TEER, the increase of FITC flux, and the overexpression of interleukin-8 (IL-8) induced by 1 µg/mL lipopolysaccharide (LPS). Additionally, kaempferol also ameliorated the LPS-induced decrease of protein expression of zonula occludens-1 (ZO-1), occludin, and claudin-2, together with the inhibited protein expressions of the phosphorylation level of NF-κB and I-κB induced by LPS. Our results suggest that kaempferol alleviates the IL-8 secretion and barrier dysfunction of the Caco-2 monolayer in the LPS-induced epithelial-endothelial coculture model via inhibiting the NF-κB signaling pathway activation.


Assuntos
Células Endoteliais/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Mucosa Intestinal/citologia , Quempferóis/farmacologia , Lipopolissacarídeos/efeitos adversos , Células CACO-2 , Claudina-2/genética , Claudina-2/metabolismo , Técnicas de Cocultura , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Humanos , Inflamação/tratamento farmacológico , Inflamação/genética , Inflamação/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Lipopolissacarídeos/imunologia , Microvilosidades/efeitos dos fármacos , Microvilosidades/genética , Microvilosidades/metabolismo , Ocludina/genética , Ocludina/metabolismo , Proteína da Zônula de Oclusão-1/genética , Proteína da Zônula de Oclusão-1/metabolismo
6.
F1000Res ; 82019.
Artigo em Inglês | MEDLINE | ID: mdl-31824659

RESUMO

Microvillus inclusion disease (MVID) is a rare congenital severe malabsorptive and secretory diarrheal disease characterized by blunted or absent microvilli with accumulation of secretory granules and inclusion bodies in enterocytes. The typical clinical presentation of the disease is severe chronic diarrhea that rapidly leads to dehydration and metabolic acidosis. Despite significant advances in our understanding of the causative factors, to date, no curative therapy for MVID and associated diarrhea exists. Prognosis mainly relies on life-long total parenteral nutrition (TPN) and eventual small bowel and/or liver transplantation. Both TPN and intestinal transplantation are challenging and present with many side effects. A breakthrough in the understanding of MVID emanated from seminal findings revealing mutations in MYO5B as a cause for MVID. During the last decade, many studies have thus utilized cell lines and animal models with knockdown of MYO5B to closely recapitulate the human disease and investigate potential therapeutic options in disease management. We will review the most recent advances made in the research pertaining to MVID. We will also highlight the tools and models developed that can be utilized for basic and applied research to increase our understanding of MVID and develop novel and effective targeted therapies.


Assuntos
Síndromes de Malabsorção , Microvilosidades/patologia , Mucolipidoses , Animais , Humanos , Cadeias Pesadas de Miosina , Miosina Tipo V
7.
Science ; 366(6463): 326-334, 2019 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-31624206

RESUMO

Collective cell contractions that generate global tissue deformations are a signature feature of animal movement and morphogenesis. However, the origin of collective contractility in animals remains unclear. While surveying the Caribbean island of Curaçao for choanoflagellates, the closest living relatives of animals, we isolated a previously undescribed species (here named Choanoeca flexa sp. nov.) that forms multicellular cup-shaped colonies. The colonies rapidly invert their curvature in response to changing light levels, which they detect through a rhodopsin-cyclic guanosine monophosphate pathway. Inversion requires actomyosin-mediated apical contractility and allows alternation between feeding and swimming behavior. C. flexa thus rapidly converts sensory inputs directly into multicellular contractions. These findings may inform reconstructions of hypothesized animal ancestors that existed before the evolution of specialized sensory and contractile cells.


Assuntos
Coanoflagelados/fisiologia , Luz , Actomiosina/metabolismo , Animais , Evolução Biológica , Coanoflagelados/citologia , GMP Cíclico/metabolismo , Microvilosidades/fisiologia , Movimento , Diester Fosfórico Hidrolases/metabolismo , Proteínas de Protozoários/metabolismo , Rodopsinas Sensoriais/metabolismo
8.
Dev Cell ; 50(5): 529-530, 2019 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-31505173

RESUMO

In this issue of Developmental Cell, Meenderink et al. (2019) describe how treadmilling of the actin core of nascent microvilli on the apical surface of epithelial cells underlies their motility across the cell surface, collision with each other, and ultimately clustering to form the "brush border."


Assuntos
Actinas , Células Epiteliais , Membrana Celular , Análise por Conglomerados , Microvilosidades
9.
J Biomed Sci ; 26(1): 59, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31434568

RESUMO

Increasing evidences have shown strong associations between gut microbiota and many human diseases, and understanding the dynamic crosstalks of host-microbe interaction in the gut has become necessary for the detection, prevention, or therapy of diseases. Many reports have showed that diet, nutrient, pharmacologic factors and many other stimuli play dominant roles in the modulation of gut microbial compositions. However, it is inappropriate to neglect the impact of host factors on shaping the gut microbiota. In this review, we highlighted the current findings of the host factors that could modulate the gut microbiota. Particularly the epithelium-associated factors, including the innate immune sensors, anti-microbial peptides, mucus barrier, secretory IgAs, epithelial microvilli, epithelial tight junctions, epithelium metabolism, oxygen barrier, and even the microRNAs are discussed in the context of the microbiota shaping. With these shaping factors, the gut epithelial cells could select the residing microbes and affect the microbial composition. This knowledge not only could provide the opportunities to better control many diseases, but may also be used for predicting the success of fecal microbiota transplantation clinically.


Assuntos
Peptídeos Catiônicos Antimicrobianos/fisiologia , Epitélio/fisiologia , Microbioma Gastrointestinal/fisiologia , Imunidade Inata , Imunoglobulina A Secretora/fisiologia , Muco/fisiologia , Humanos , MicroRNAs/fisiologia , Microvilosidades/fisiologia , Junções Íntimas/fisiologia
10.
BMC Cancer ; 19(1): 750, 2019 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-31362745

RESUMO

BACKGROUND: Human inflammatory breast cancer (IBC) and canine inflammatory mammary cancer (IMC) are the most lethal mammary cancers. An exacerbated angiogenesis and the existence of vasculogenic mimicry (VM) are hallmarks of these tumors. The information regarding VM and ultrastructural characteristics of mammary cell lines is scant. METHODS: In this study, IBC cell line SUM149 and IMC cell line IPC-366 in adherent (2D) and non-adherent (3D) (mammospheres, cancer stem cells) conditions were analyzed by transmission and scanning electron microscopy (TEM and SEM, respectively). RESULTS: The TEM revealed round to oval shape cells with microvilli on the surface, high numbers of peroxisomes in close apposition to lipid droplets and some extracellular derived vesicles. The TEM and the SEM mammospheres revealed group of cells clumping together with a central lumen (resembling a mammary acini). The cells joint are tight junctions and zonula adherens. By SEM two cell morphologies were observed: spherical and flattened cells. There was evidence endothelial-like cells (ELCs), which is characteristic for this disease, showing several or unique cytoplasmic empty space. ELCs were more frequent in 3D than in 2D culture conditions and contained Weibel-Palade cytoplasmic bodies, which are exclusive structures of endothelial cells. CONCLUSIONS: Both cell lines, IPC-366 and SUM-149, shared ultrastructural characteristics, further supporting canine IMC as a model for the human disease. To the best of our knowledge, this is the first study that demonstrate the morphological differentiation of cultured cancer stem cells from cancer epithelial cell lines into endothelial-like cells, confirming the vasculogenic mimicry phenomenon from an ultrastructural point of view.


Assuntos
Doenças do Cão/patologia , Neoplasias Inflamatórias Mamárias/patologia , Neoplasias Inflamatórias Mamárias/ultraestrutura , Neoplasias Mamárias Animais/patologia , Neoplasias Mamárias Animais/ultraestrutura , Animais , Adesão Celular , Linhagem Celular Tumoral , Desmossomos , Cães , Células Endoteliais/citologia , Feminino , Humanos , Gotículas Lipídicas , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Microvilosidades , Neovascularização Patológica , Peroxissomos , Junções Íntimas
11.
Mol Biol Cell ; 30(19): 2515-2526, 2019 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-31390291

RESUMO

Apical microvilli are critical for the homeostasis of transporting epithelia, yet mechanisms that control the assembly and morphology of these protrusions remain poorly understood. Previous studies in intestinal epithelial cell lines suggested a role for the F-BAR domain protein PACSIN2 in normal microvillar assembly. Here we report the phenotype of PACSIN2 KO mice and provide evidence that through its role in promoting apical endocytosis, this molecule plays a role in controlling microvillar morphology. PACSIN2 KO enterocytes exhibit reduced numbers of microvilli and defects in the microvillar ultrastructure, with membranes lifting away from rootlets of core bundles. Dynamin2, a PACSIN2 binding partner, and other endocytic factors were also lost from their normal localization near microvillar rootlets. To determine whether loss of endocytic machinery could explain defects in microvillar morphology, we examined the impact of PACSIN2 KD and endocytosis inhibition on live intestinal epithelial cells. These assays revealed that when endocytic vesicle scission fails, tubules are pulled into the cytoplasm and this, in turn, leads to a membrane-lifting phenomenon reminiscent of that observed at PACSIN2 KO brush borders. These findings lead to a new model where inward forces generated by endocytic machinery on the plasma membrane control the membrane wrapping of cell surface protrusions.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Mucosa Intestinal/metabolismo , Microvilosidades/metabolismo , Actinas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Proteínas de Transporte/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Endocitose , Enterócitos/metabolismo , Células Epiteliais/metabolismo , Feminino , Humanos , Intestinos , Camundongos , Camundongos Knockout
12.
Macromol Biosci ; 19(9): e1900105, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31386305

RESUMO

Polymeric microcapsules have begun to attract significant interest in biomedical fields. As the interactions between cells and materials are influenced by both cell type and elasticity, silk-based microcapsules are synthesized with desirable mechanical features using layer-by-layer assembly and then the uptake of these microcapsules by BeWo b30 placental cells is investigated. Cellular uptake is enhanced with increasing of elastic modulus of the silk-based microcapsules. More importantly, the distinct microvilli of these cells behaves in a diverse manner when exposed to microcapsules with different mechanical features, including grabbing (rigidity) or random touching (soft) behavior; these factors affect the final uptake. Inspired by oocyte pickup, the grabbing behavior of the microvilli may provide valuable information with which to elucidate the specific characteristics of uptake between cells and man-made particles, particularly in the reproductive system.


Assuntos
Cápsulas/metabolismo , Placenta/citologia , Seda/metabolismo , Cápsulas/síntese química , Linhagem Celular , Feminino , Humanos , Microvilosidades/metabolismo , Gravidez , Seda/síntese química , Solventes , Eletricidade Estática
13.
Pancreatology ; 19(6): 873-879, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31400934

RESUMO

BACKGROUND/OBJECTIVES: Malignant pancreatobiliary strictures are in many cases clinically indistinguishable and present a major problem to endoscopy specialists. Intraductal sampling procedures such as brush cytology are commonly used for diagnosis with a sensitivity that is low for a diagnostic test used in daily clinical practice. MicroRNA (miR) alterations detected in many cancers are disease-specific, which can be utilized in clinical applications. The aim of the present study was to analyze whether determination of miR expression levels in intraductal brush cytology specimens is a feasible approach to improve the diagnosis of pancreatobiliary cancer. METHODS: Brush cytology specimens have been collected during endoscopic retrograde cholangio-pancreatography (ERCP) and analyzed by routine cytology and ancillary miR assays. Total RNA was extracted using the miRNeasy Mini Kit and the expression of miRs frequently dysregulated in pancreatobiliary cancer (miR-16, miR-21, miR-196a, miR-221) were analyzed by quantitative real-time PCR using RNU6B as internal control. RESULTS: Routine cytology resulted in no false positive diagnoses, however, the combined sensitivity remained at 53.8%. Expression (ΔCt values) of miR-16 (p = 0.0039), miR-196a (p = 0.0003) and miR-221 (p = 0.0049) showed a clear statistical significance between malignant and benign pancreatobiliary specimens (n = 35). Malignancy could be detected combining routine cytology and the miR-196a single marker expression levels with a sensitivity of 84.6% (92.9% in biliary strictures) with no false positives. CONCLUSIONS: The results offer the first direct demonstration that microRNAs are readily detectable in brush cytology specimens obtained during ERCP, and have the potential to help the cytological diagnosis of pancreatobiliary malignancy.


Assuntos
Neoplasias dos Ductos Biliares/diagnóstico , MicroRNAs/biossíntese , Microvilosidades/química , Neoplasias Pancreáticas/diagnóstico , Idoso , Neoplasias dos Ductos Biliares/patologia , Colangiopancreatografia Retrógrada Endoscópica , Citodiagnóstico , Reações Falso-Positivas , Feminino , Humanos , Masculino , MicroRNAs/análise , Microvilosidades/patologia , Pessoa de Meia-Idade , Neoplasias Pancreáticas/patologia , Estudos Prospectivos , RNA/análise , RNA/isolamento & purificação , Sensibilidade e Especificidade
14.
Sci Total Environ ; 696: 134035, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31470328

RESUMO

Ammonia (NH3), an inhaled harmful gas, is not only an important volatile in fertilizer production and ranching, but also the main basic component of haze. However, the effect and mechanism of NH3 on the intestines are still unclear. To investigate the intestinal toxicity of NH3 inhalation, morphological changes, transcriptome profiles and oxidative stress indicators of jejunum in broiler chicken exposed to NH3 for 42 days were examined. Results of morphological observation showed that NH3 exposure caused deficiency of jejunal microvilli and neutrophil infiltration. Transcriptomics sequencing identified 677 differential expressed genes (DEGs) including 358 up-regulated genes and 319 down-regulated genes. Enrichment analysis of obtained DEGs by Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) found that biological functions and pathways affected by NH3 included antioxidant function, inflammation, microtubule and nutrition transport. Relative genes validation and chemical detection confirmed that NH3-induced oxidative stress by activating CYPs and inhibiting antioxidant enzymes promoted inflammatory response and decreased microtubule activity, thus destroying the balance of nutritional transporters. Our study perfects the injurious mechanism of NH3 exposure and provides a new insight and method for environmental risk assessment.


Assuntos
Poluentes Atmosféricos/toxicidade , Amônia/toxicidade , Microtúbulos/fisiologia , Microvilosidades/fisiologia , Estresse Oxidativo/fisiologia , Transcriptoma , Animais , Antioxidantes , Galinhas , Regulação para Baixo , Perfilação da Expressão Gênica , Inflamação , Transdução de Sinais , Regulação para Cima
15.
Food Funct ; 10(8): 4834-4843, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31321397

RESUMO

Wheat bran is the by-product of milling wheat flour which is one of the richest sources of dietary fiber, and cellulase that can be used for increasing the soluble dietary fiber. We hypothesize that wheat bran prebiotics would improve mineral-metabolism gene expression, and intestinal functionality, and increase health-promoting bacterial populations in vivo (Gallus gallus). By using the intra-amniotic administration procedure, five groups (non-injected; 18 Ω H2O; 100 mg mL-1 wheat bran prebiotics; 100 mg mL-1 cellulase wheat bran prebiotics; 50 mg mL-1 Arbinose) were utilized. Upon hatch, the cecum, small intestine and liver were collected for assessment. The results indicated that wheat bran prebiotics significantly up regulated the expression of certain brush border membrane functional genes, and mineral metabolism related proteins, in addition to increasing the intestinal beneficial bacterial populations. Prebiotics from wheat bran, cellulase wheat bran and arabinose increased the intestinal villi height, and goblet cell diameters and numbers.


Assuntos
Galinhas/metabolismo , Fibras na Dieta/metabolismo , Microbioma Gastrointestinal , Mucosa Intestinal/metabolismo , Microvilosidades/metabolismo , Prebióticos/análise , Triticum/metabolismo , Animais , Galinhas/crescimento & desenvolvimento , Galinhas/microbiologia , Feminino , Mucosa Intestinal/crescimento & desenvolvimento , Mucosa Intestinal/microbiologia , Masculino , Triticum/química
16.
Arch Insect Biochem Physiol ; 102(1): e21591, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31257641

RESUMO

In Brazil, the use of transgenic plants expressing the insect-toxic Bacillus thuringiensis endotoxin has been successfully used as pest control management since 2013 in transgenic soybean lineages against pest caterpillars such as Helicoverpa armigera. These toxins, endogenously expressed by the plants or sprayed over the crops, are ingested by the insect and bind to receptors in the midgut of these animals, resulting in disruption of digestion and lower insect survival rates. Here, we identified and characterized a membrane-associated alkaline phosphatase (ALP) in the midgut of Anticarsia gemmatalis, the main soybean defoliator pest in Brazil, and data suggested that it binds to Cry1Ac toxin in vitro. Our data showed a peak of ALP activity in homogenate samples of the midgut dissected from the 4th and 5th instars larvae. The brush border membrane vesicles obtained from the midgut of these larvae were used to purify a 60 kDa ALP, as detected by in-gel activity and in vitro biochemical characterization using pharmacological inhibitors and mass spectrometry. When Cry1Ac toxin was supplied to the diet, it was efficient in decreasing larval weight gain and survival. Indeed, in vitro incubation of Cry1Ac toxin with the purified ALP resulted in a 43% decrease in ALP specific activity and enzyme-linked immunosorbent assay showed that ALP interacts with Cry1Ac toxin in vitro, thus suggesting that ALP could function as a Cry toxin ligand. This is a first report characterizing an ALP in A. gemmatalis.


Assuntos
Fosfatase Alcalina/metabolismo , Proteínas de Bactérias/metabolismo , Endotoxinas/metabolismo , Proteínas Hemolisinas/metabolismo , Larva/enzimologia , Mariposas/enzimologia , Fosfatase Alcalina/antagonistas & inibidores , Fosfatase Alcalina/isolamento & purificação , Animais , Proteínas de Bactérias/toxicidade , Endotoxinas/toxicidade , Trato Gastrointestinal/enzimologia , Trato Gastrointestinal/ultraestrutura , Proteínas Hemolisinas/toxicidade , Microvilosidades/enzimologia
17.
J Dairy Sci ; 102(10): 8622-8629, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31351730

RESUMO

This work aimed to study the opioid peptide ß-casomorphin-7 (BCM7) degradation or stability during digestion using human gastrointestinal (GI) juices and porcine jejunal brush border membrane (BBM) peptidases. Synthetic BCM7 was subjected to in vitro digestion by GI fluids obtained from human volunteers for 180 min, and to downstream degradation with porcine BBM vesicles. The BCM7 was sampled at 4 time points over 24 h after BBM addition. The digests were profiled by HPLC-electrospray ionization mass spectrometry (ESI/MS) to monitor BCM7 during GI digestion, and intact BCM7 through BBM digestion was quantified by reverse-phase (RP)-HPLC. We found that BCM7 was partly digested with human GI enzymes, as 3 proteolytic fragments in addition to f(60-66) YPFPGPI were detected: f(62-66) FPGPI, f(60-65) YPFPGP and f(61-66) PFPGPI. The RP-HPLC analysis revealed that 42% of the initial peptide was degraded after only 2 h of BBM digestion, and as much as 79% was degraded after 4-h digestion with supplementation of BBM. In conclusion, this study showed that most of BCM7 was degraded during GI and BBM digestion, although a small amount (5%) was still detected after 24-h digestion. It remains to be studied whether the small amount of intact BCM7 detected after in vitro digestion is transported via active transceptors in the human intestinal epithelial cells and enters blood circulation.


Assuntos
Endorfinas/metabolismo , Trato Gastrointestinal/metabolismo , Jejuno/metabolismo , Microvilosidades/metabolismo , Fragmentos de Peptídeos/metabolismo , Animais , Cromatografia Líquida de Alta Pressão , Digestão , Humanos , Microvilosidades/enzimologia , Peptídeo Hidrolases/metabolismo , Suínos
18.
Dev Biol ; 453(1): 56-67, 2019 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-31158364

RESUMO

Photoreceptor cells (PRCs) across the animal kingdom are characterized by a stacking of apical membranes to accommodate the high abundance of photopigment. In arthropods and many other invertebrate phyla PRC membrane stacks adopt the shape of densely packed microvilli that form a structure called rhabdomere. PRCs and surrounding accessory cells, including pigment cells and lens-forming cells, are grouped in stereotyped units, the ommatidia. In larvae of holometabolan insects, eyes (called stemmata) are reduced in terms of number and composition of ommatidia. The stemma of Drosophila (Bolwig organ) is reduced to a bilateral cluster of subepidermal PRCs, lacking all other cell types. In the present paper we have analyzed the development and fine structure of the Drosophila larval PRCs. Shortly after their appearance in the embryonic head ectoderm, PRC precursors delaminate and lose expression of apical markers of epithelial cells, including Crumbs and several centrosome-associated proteins. In the early first instar larva, PRCs show an expanded, irregularly shaped apical surface that is folded into multiple horizontal microvillar-like processes (MLPs). Apical PRC membranes and MLPs are covered with a layer of extracellular matrix. MLPs are predominantly aligned along an axis that extends ventro-anteriorly to dorso-posteriorly, but vary in length, diameter, and spacing. Individual MLPs present a "beaded" shape, with thick segments (0.2-0.3 µm diameter) alternating with thin segments (>0.1 µm). We show that loss of the glycoprotein Chaoptin, which is absolutely essential for rhabdomere formation in the adult PRCs, does not lead to severe abnormalities in larval PRCs.


Assuntos
Drosophila melanogaster/ultraestrutura , Olho/ultraestrutura , Microscopia Eletrônica , Microvilosidades/ultraestrutura , Células Fotorreceptoras de Invertebrados/ultraestrutura , Animais , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriologia , Desenvolvimento Embrionário , Larva/ultraestrutura , Mutação/genética
19.
World J Gastroenterol ; 25(20): 2402-2415, 2019 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-31171885

RESUMO

The purpose of this review is to provide a definitive account of small intestinal mucosal structure and interpretation. The coeliac lesion has been well known, but not well described to date and this review aims to identify the interpretative difficulties which have arisen over time with the histological assessment of coeliac disease. In early coeliac interpretation, there were significant inaccuracies, particularly surrounding intraepithelial lymphocyte counts and the degree of villous flattening which occurred in the tissue. Many of these interpretive pitfalls are still encountered today, increasing the potential for diagnostic errors. These difficulties are mostly due to the fact that stained 2-dimensional sections can never truly represent the 3-dimensional framework of the intestinal tissue under investigation. Therefore, this review offers a critical account occasioned by these 2-dimensional interpretative errors and which, in our opinion, should in general be jettisoned. As a result, we leave a framework regarding the true 3-dimensional knowledge of mucosal structure accrued over the 70-year period of study, and one which is available for future reference.


Assuntos
Doença Celíaca/diagnóstico , Mucosa Intestinal/patologia , Intestino Delgado/patologia , Microvilosidades/patologia , Doença Celíaca/patologia , Humanos , Mucosa Intestinal/ultraestrutura , Intestino Delgado/ultraestrutura
20.
Dokl Biol Sci ; 485(1): 56-58, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31197596

RESUMO

The ultrastructure of the nephron subcellular organelles was studied in healthy mink kidneys. The data obtained were compared with the results of transmission electron microscopy. The renal cell nanomorphology proved to be similar when electronograms and the atomic force microscopy images were analyzed. The methods used enabled us to visualize the glomerular capillary endotheliocytes with cytolemma pits in the area of fenestrae that provide blood filtration; in the proximal nephron part, on the apical pole of the epithelial cells, brush-border soft microvilli were observed. The microvilli were characterized by a well-organized structure along their entire length and the membrane integrity. The data obtained show morphological parameters of the healthy mink organ and can be helpful in diagnosing of nephropathology.


Assuntos
Vison/anatomia & histologia , Néfrons/ultraestrutura , Animais , Células Endoteliais/ultraestrutura , Microvilosidades/ultraestrutura
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