Specific binding of aspartic protease and enterocytes promotes Trichinella spiralis invasion of murine intestinal epithelium cells

@#Trichinella spiralis is an important foodborne zoonotic parasite and it is necessary to develop vaccine to prevent T. spiralis infection in food animals. T. spiralis aspartic protease-2 (TsASP2) has been demonstrated to play a crucial role in larval invasion of intestinal epithelium cells (IECs). The purpose of this study was to assess the interaction between TsASP2 and IECs and to investigate the immune protection elicited by vaccination with rTsASP2. The results showed that the enzymatic activity of native aspartic protease was detected in crude proteins of all T. spiralis development stages other than NBL stage, the highest activity was observed in the IIL stage. The results of Western blot showed that TsASP2 protein was expressed at ML, IIL and AW but not NBL, and the TsASP2 expression level at IIL stage was significantly higher than those of other three worm stages (P < 0.05). The specific binding between rTsASP2 and IECs was observed by immunofluorescence test (IFT) and confocal microscopy, and the binding site was localized at the IEC membrane and this binding ability was inhibited by aspartic protease specific inhibitor pepstain A. The results of ELISA showed that the binding ability was protein dose-dependent. Vaccination with rTsASP2 triggered a mixed Th1/Th2 humoral and mucosal immune responses, as demonstrated by the elevation levels of Th1/Th2 cytokines (IFN-γ and IL-4) secreted by the spleen and mesenteric lymph nodes (MLNs) of immunized mice. The mice vaccinated with rTsASP2 exhibited a 54.17% reduction in enteral adult worms and a 54.58% reduction in muscle larvae after T. spiralis challenge. The results demonstrated that TsASP2 might be a potential molecular target for anti-Trichinella vaccines.

Epithelial-microbial diplomacy: escalating border tensions drive inflammation in inflammatory bowel disease

Inflammatory bowel diseases (IBD) are chronic conditions of the gastrointestinal tract-the main site of host-microbial interaction in the body. Development of IBD is not due to a single event but rather is a multifactorial process where a patient's genetic background, behavioral habits, and environmental exposures contribute to disease pathogenesis. IBD patients exhibit alterations to gut bacterial populations “dysbiosis” due to the inflammatory microenvironment, however whether this alteration of the gut microbiota precedes inflammation has not been confirmed. Emerging evidence has highlighted the important role of gut microbes in developing measured immune responses and modulating other host responses such as metabolism. Much of the work on the gut microbiota has been correlative and there is an increasing need to understand the intimate relationship between host and microbe. In this review, we highlight how commensal and pathogenic bacteria interact with host intestinal epithelial cells and explore how altered microenvironments impact these connections.

Implication of Porphyromonas gingivalis in colitis and homeostasis of intestinal epithelium / 한국실험동물학회지

Emerging evidences have reported that periodontitis can be a risk factor for the pathogenesis of various systemic diseases. Porphyromonas gingivalis (Pg), one of the crucial pathogens in chronic periodontitis, has been spotlighted as a potential cause for the promotion and acceleration of periodontitis-associated systemic disorders. To investigate the association between Pg and intestinal disease or homeostasis, we treated Pg-derived lipopolysaccharide (LPS) in murine colitis model or intestinal organoid, respectively. Pg-derived LPS (Pg LPS) was administrated into chemically induced murine colitis model and disease symptoms were monitored compared with the infusion of LPS derived from E. coli (Ec LPS). Organoids isolated and cultured from mouse small intestine were treated with Pg or Ec LPS and further analyzed for the generation and composition of organoids. In vivo observations demonstrated that both Pg and Ec LPS exerted slight protective effects against murine colitis. Pg LPS did not affect the generation and growth of intestinal epithelial organoids. Among subtypes of epithelial cells, markers for stem cells, goblet cells or Paneth cells were changed in response to Pg LPS. Taken together, these results indicate that Pg LPS leads to partial improvement in colitis and that its treatment does not significantly affect the self-organization of intestinal organoids but may regulate the epithelial composition.

Physiological understanding of host-microbial pathogen interactions in the gut

The gut epithelial barrier, which is composed of the mucosal layer and the intestinal epithelium, has multiple defense mechanisms and interconnected regulatory mechanisms against enteric microbial pathogens. However, many bacterial pathogens have highly evolved infectious stratagems that manipulate mucin production, epithelial cell-cell junctions, cell death, and cell turnover to promote their replication and pathogenicity in the gut epithelial barrier. In this review, we focus on current knowledge about how bacterial pathogens regulate mucin levels to circumvent the epithelial mucus barrier and target cell-cell junctions to invade deeper tissues and increase their colonization. We also describe how bacterial pathogens manipulate various modes of epithelial cell death to facilitate bacterial dissemination and virulence effects. Finally, we discuss recent investigating how bacterial pathogens regulate epithelial cell turnover and intestinal stem cell populations to modulate intestinal epithelium homeostasis.

Emprovement of Rhodiola rosea on intestinal immune function of Drosophila melanogaster / 中草药

Objective: To study the effect of Rhodiola rosea extracts on intestinal immune function of Drosophila melanogaster. Methods: D. melanogaster was treated by SDS and fungal biological solution with or without R. rosea extracts, and the effect of R. rosea extracts on the survival rate of D. melanogaster, intestinal epithelium cell death, relative contents of reactive oxygen species (ROS) in intestinal epithelium cells, and intestinal morphology changes were analyzed. Results: R. rosea extracts could significantly improve the survival rate of SDS and fungus (Beauveria bassiana)-infected D. melanogaster (P < 0.05), reduce the intestinal epithelial cell death and the level of intestinal ROS levels, and protect and maintain the intestinal morphology. Conclusion: R. rosea extracts could significantly improve the intestinal immune function of D. melanogaster with the amelioration and protection of intestinal immunologic function injury induced by fungi and SDS.

Análisis estructural y funcional de proteínas solubles que unen lípidos de intestino e hígado / Structure-function analysis of soluble lipid binding proteins from intestine and liver / Análise estrutural e funcional de proteínas solúveis que ligam lipídeos de intestino e fígado

Luego de la ingesta, el epitelio del intestino delgado está encargado de asimilar grandes cantidades de nutrientes, como aminoácidos, glúcidos y ácidos grasos. Las proteínas solubles que unen lípidos cumplirían un rol determinante en este proceso, sobre todo protegiendo la integridad del tejido contra el efecto simil-detergente de los ácidos grasos provenientes de la dieta. En enterocitos se expresan dos proteínas que unen ácidos grasos de cadena larga, IFABP y LFABP, para las cuales no se conocen bien aún sus funciones específicas, o el porqué de la necesidad de dos proteínas aparentemente equivalentes. Este laboratorio se ha enfocado en el estudio comparativo de estas dos proteínas empleando distintas variantes estructurales y métodos bioquímicos, biofísicos, y de biología molecular y celular. Así, se han podido definir los determinantes moleculares de cada proteína responsables de la interacción con membranas, los mecanismos de transferencia de ligandos y los factores que modulan estas propiedades. Más recientemente, se han extendido estos ensayos a cultivos celulares donde se ha correlacionado la expresión de estas proteínas con la secreción de citoquinas, la proliferación y la diferenciación celular. El estudio de estas proteínas es de gran importancia por su potencial como blancos terapéuticos y su utilidad en el diagnóstico de injurias tisulares.

After ingestion, the epithelium of the small intestine is responsible for assimilating large amounts of nutrients such as amino acids, sugars and fatty acids. Soluble lipid binding proteins fulfill a determining role in this process, especially protecting the tissue integrity against the detergent-like effect of fatty acids from the diet. Two proteins that bind long-chain fatty acids are expressed in enterocytes, IFABP and LFABP, whose specific functions are still poorly understood, or the reason for the need of two apparently equivalent proteins. Our laboratory has focused on the comparative study of these two proteins using structural variants and biochemical, biophysical, and molecular and cellular biology approaches. Thus, the molecular determinants responsible for the interaction with membranes were defined for each protein, their ligand transfer mechanism and the factors that modulate these properties. More recently, these assays have been extended to cell culture studies which correlate the expression of these proteins with cytokine secretion, cell proliferation and differentiation. The study of these proteins is of great importance due to their potential as therapeutic targets and their usefulness in the diagnosis of tissue injury.

Após a ingestão, o epitélio do intestino delgado é responsável pela assimilação de uma grande quantidade de nutrientes, tais como aminoácidos, glicídios e ácidos graxos. As proteínas solúveis que ligam lipídeos desempenhariam um papel determinante neste processo, principalmente protegendo a integridade do tecido contra o efeito detergente dos ácidos graxos da dieta. Nos enterócitos se expressam duas proteínas que ligam ácidos graxos de cadeia longa, IFABP e LFABP; cujas funções específicas ainda não são muito conhecidas, ou não se conhece o motivo pelo qual são necessárias duas proteínas aparentemente equivalentes. Nosso laboratório tem se focado no estudo comparativo destas duas proteínas utilizando variantes estruturais e métodos bioquímicos, biofísicos, e de biologia molecular e celular. Assim, foi possível definir os determinantes moleculares de cada proteína responsáveis pela interação com membranas, os mecanismos da transferência de ligantes e os fatores que modulam essas propriedades. Mais recentemente, estendemos estes ensaios para culturas celulares, correlacionando a expressão destas proteínas com a secreção de citocinas, a proliferação e a diferenciação celular. O estudo destas proteínas é de grande importância por seu potencial como alvos terapêuticos e sua utilidade no diagnóstico de lesões teciduais.

Renewal versus Retention: Isotopic Composition of Intestinal Epithelium and Eye Lens.

Aims: To compare isotopic signatures of contrasting (due to the structure and metabolism) organs in mice of two contrasting ages. Study Design: Cross-sectional study. Place and Duration of Study: Biology Department of Moscow State University; Institute of Geochemistry and Analytical Chemistry, Russian Academy of Sciences; 2007–2011. Methodology: Mass spectrometric measurements of carbon and nitrogen isotopic ratios of jejunal epithelium and lens in 1- and 22-mo mice fed a monotonous diet. Results: The lenses are enriched in carbon and nitrogen as compared with intestinal epithelium (by 5.5% and 4.5% in 1-mo mice and 8.3% and 6% in 22-mo mice, respectively). The 15N content is also higher in lenses than in intestinal epithelium (8.97‰ vs. 7.62‰ in 1-mo mice, and 7.40‰ vs. 6.58‰ in 22-mo mice). The 13C content of lenses exceeds that of intestinal epithelium in 1-mo mice (-20.27‰ vs. -21.69‰), although 13C content is equal in 22-mo mice (-22.56‰ vs. -22.67‰). 15N content is depleted in the intestinal epithelium of 22-mo mice (-1.04‰), whereas 13C depletion (-0.98‰) is non-significant. 13C and 15N content in lenses is also significantly decreased in 22-mo mice (-2.29‰ and -1.57‰). Conclusion: The intestinal epithelium represents a structure with short-term isotopic memory lasting a few days, whereas the events of the organism’s entire lifetime are retained in lens isotopic memory. The difference of the parameters measured is evidently determined by structural contrast, metabolic rate, and rejuvenation modes of the tissues. The 15N depletion in both the intestinal epithelium and lenses, as well as 13C depletion in lenses of 22-mo mice might be considered as a sign of ageing. In contrast, the depletion of 15N in lenses of 22-mo mice should be considered primarily as a result of dilution of breastmilk isotopic signature that probably obscures age-related alterations of the organ. Comparison of isotopic compositions of these contrasting organs may be useful for physiological and ecological determinations.

Efeitos da infecção crônica por Toxoplasma gondii sobre a parede intestinal de gatos domésticos / The effects of the infection caused by Toxoplasma gondii on the cat duodenal wall

O objetivo deste trabalho foi analisar os efeitos da infecção causada pelo Toxoplasma gondii sobre a parede do duodeno de gatos. Foram utilizados seis gatos (Felis catus), com cerca de três meses de vida, distribuídos aleatoriamente em Grupo controle (G1; n = 3) e Grupo infectado (G2; n = 3). Os animais do G2 receberam, por via oral, 200 cistos teciduais da cepa ME49 (tipo II) do T. gondii. Após 40 dias, os animais foram submetidos à eutanásia, laparotomia e retirada do duodeno, que foi fixado em solução de Bouin e submetido à rotina histológica para obtenção de cortes transversais de 3 µm. Os cortes foram corados com Hematoxilina-Eosina (HE), Azan, Ácido Periódico de Schiff (PAS), Alcian-Blue e Tricrômio de Mallory. Realizou-se uma avaliação qualitativa da parede intestinal e medidas comparativas entre os dois grupos, com relação: à espessura da túnica mucosa, túnica muscular, parede total, altura dos vilos, profundidade das criptas e altura dos enterócitos e seus núcleos. As células caliciformes, os linfócitos intraepiteliais e as células de Paneth foram quantificados. Os resultados mostraram que a infecção levou à atrofia da túnica mucosa, túnica muscular e parede intestinal do duodeno de gatos do G2 (p < 0,05). A altura dos enterócitos apresentou um aumento significativo (p < 0,05) nos animais do G2. Na avaliação qualitativa, as fibras colágenas ocupavam visivelmente uma maior área dos estratos da parede intestinal, o que sugere que estejam aumentadas. Observou-se a redução da secreção de sulfomucinas e o aumento das células de Paneth nesses mesmos animais (p < 0,05).

This paper analyzes the effects of the infection caused by Toxoplasma gondii on the cat duodenal wall. Six cats (Felis catus) with 3-month-old were randomly divided into Control Group (G1; n = 3) and Infected Group (G2; n = 3). The animals from G2 received orarilly 200 T. gondii tissye cysts of ME49-strain (type II). After 40 days, the animals were submitted to euthanasia, laparotomy and had their duodenum removed, fixed in Bouin solution and submitted to histological routine obtaining 3 µm transverse cuts. The cuts were stained with Hematoxylin-Eosin (HE), Azan, Periodic acid - Schiff (PAS), Alcian-Blue, and Mallory trichrome. Qualitative assessment of the intestine wall as well as comparative measurements with respect to the thickness of mucosa, muscle tunic, total wall, the height of the villous, the depth of the crypts, and the height of the enterocytes and their nuclei were carried out. Calciform cells, the intraepithelial lymphocytes, and the Paneth cells were quantified. The results showed that the infection led to the atrophy of the mucosa, muscle tunic, and the intestinal wall of the duodenum of G2 cats (p < 0.05). The enterocytes height presented significant (p < 0.05) increase for G2 animals. According to the qualitative analysis, the collagen fibers were visibly taken a broader area on the intestinal wall layers, what suggests they have increased in size. Decrease in the sulphomucins secretion and the increase of Paneth cells were observed for these animals (p < 0.05).

The effect of methylmercury on the epithelial apoptosis of fetal mice intestine in vitro / 重庆医科大学学报

Objective:To study the effect of methylmercury chloride(MMC) on the apoptosis and the expression of apoptosis-associated proteins in the intestinal epithelium of fetal mice.Methods:The duodenum and colon were taken from E13.5d and E14.5d fetal mice respectively,and cultured with different doses of MMC(0 ?g/L,200?g/L,400?g/L,800?g/L) for 24h.The samples were sliced into the paraffin sections.Some sections were stained with HE to count the numbers of apoptotic bodies(NAB) in the fetal intestinal epithelium with stereological method.The others were stained with the immunohistochemical method to observe the expression of apoptosis-associated proteins Bcl-2,Bax,P53 and c-Fos.Results:①In experimental groups,all the NAB in the intestinal epithelium were higher than that in control groups(P

Molecule mechanisms of the pro-inflammatory cytokine's effect on intestinal epithelium tight junctions / 肠外与肠内营养

The pro-inflammation cytokine is secreted by cells,and the function is to promote and regulate inflammatory reactions.They include IL-1,IL-6,TNF-? and IFN-?.Researchs have revealed that the levels of serum pro-inflammation cytokinemight increase notably in inflammatory bowel disease patients and they could cause the change of intestinal epithelium tight junctions.

Effects and related mechanism of low-dosed methyl-mercury chloride in promoting apoptosis of intestine epithelial cells of fetal mice / 第三军医大学学报

Objective To study the effects and related mechanism of low-dosed methyl-mercury chloride (MMC) on the epithelial apoptosis of fetal mice intestine. Methods Pregnant mice on E12.5 d and E13.5 d were injected with different doses of MMC (0, 1, 2, 4 mg/kg) intraperitoneally. After 48 h, their duodenum and colon were dissected out and sliced into the paraffin sections. Some sections were stained with HE to count the numbers of apoptotic bodies (NAB) with stereological method. The others were stained with the immunohistochemical method to observe the expression of apoptosis-associated proteins Bcl-2, Bax and immediate early gene c-fos. Results ①In all experimental groups, the NAB in the epithelium were higher than that of control groups (P

The effect of hydrocortisone on epithelial apoptosis in fetal mice intestine / 重庆医科大学学报

Objective:To study the effect of hydrocortisone (HC) on apoptosis and expression of apoptosis- associated proteins in the intestinal epithelium of fetal mice.Methods:The duodena and colons were taken from E13.5d and E14.5d fetal mice respectively,and cultured with different doses of HC (0 ?g/ml,0.5?g/ml,1.0?g/ml,2.0?g/ml).After 24h,the samples were sliced up into the paraffin sections.Some sections were stained with HE to count the numbers of apoptotic bodies (NAB)in the fetal intestinal epithelium with stereological method.Others were stained with the immunohistochemical method to observe the expression of Bcl-2,Bax,P53 and c-Fos.Results:①In each experimental group,the NAB were higher than that of control group( P