Institut für Ernährungsmedizin

Permanent URI for this collection

https://hohpublica.uni-hohenheim.de/handle/123456789/80

Browse

Now showing 1 - 20 of 27

Vitamin A- and D-deficient diets disrupt intestinal antimicrobial peptide defense involving Wnt and STAT5 signaling pathways in mice
(2023) Filipe Rosa, Louisa; Petersen, Patricia P.; Görtz, Lisa F.; Stolzer, Iris; Kaden-Volynets, Valentina; Günther, Claudia; Bischoff, Stephan C.
Vitamin A and D deficiencies are associated with immune modulatory effects and intestinal barrier impairment. However, the underlying mechanisms remain unclear. C57BL/6J mice were fed either a diet lacking in vitamin A (VAd), vitamin D (VDd) or a control diet (CD) for 12 weeks. Gut barrier function, antimicrobial peptide (AMP) defense and regulatory pathways were assessed. VAd mice compared to CD mice showed a reduced villus length in the ileum (p < 0.01) and decreased crypt depth in the colon (p < 0.05). In both VAd- and VDd-fed mice, ileal α-defensin 5 (p < 0.05/p < 0.0001 for VAd/VDd) and lysozyme protein levels (p < 0.001/p < 0.0001) were decreased. Moreover, mRNA expression of lysozyme (p < 0.05/p < 0.05) and total cryptdins (p < 0.001/p < 0.01) were reduced compared to controls. Furthermore, matrix metalloproteinase-7 (Mmp7) mRNA (p < 0.0001/p < 0.001) as well as components of the Wnt signaling pathway were decreased. VAd- and VDd-fed mice, compared to control mice, exhibited increased expression of pro-inflammatory markers and β-defensins in the colon. Organoid cell culture confirmed that vitamins A and D regulate AMP expression, likely through the Jak/STAT5 signaling pathway. In conclusion, our data show that vitamin A and D regulate intestinal antimicrobial peptide defense through Wnt and STAT5 signaling pathways.
Editorial: Updates on RIG-I-like receptor-mediated innate immune responses
(2023) Kishore, Uday; Kufer, Thomas A.
Inflammation and nutrition: friend or foe?
(2023) Stumpf, Franziska; Keller, Bettina; Gressies, Carla; Schuetz, Philipp
The importance of the interplay between inflammation and nutrition has generated much interest in recent times. Inflammation has been identified as a key driver for disease-related malnutrition, leading to anorexia, reduced food intake, muscle catabolism, and insulin resistance, which are stimulating a catabolic state. Interesting recent data suggest that inflammation also modulates the response to nutritional treatment. Studies have demonstrated that patients with high inflammation show no response to nutritional interventions, while patients with lower levels of inflammation do. This may explain the contradictory results of nutritional trials to date. Several studies of heterogeneous patient populations, or in the critically ill or advanced cancer patients, have not found significant benefits on clinical outcome. Vice versa, several dietary patterns and nutrients with pro- or anti-inflammatory properties have been identified, demonstrating that nutrition influences inflammation. Within this review, we summarize and discuss recent advances in both the role of inflammation in malnutrition and the effect of nutrition on inflammation.
Benefits of fiber-enriched foods on satiety and parameters of human well-being in adults with and without cardiometabolic risk
(2023) Ehret, Janine; Brandl, Beate; Schweikert, Karsten; Rennekamp, Rachel; Ströbele-Benschop, Nanette; Skurk, Thomas; Hauner, Hans
Consumption of fiber-rich foods is linked to beneficial effects on chronic diseases and gut health, while implications towards improving satiety and parameters of well-being remain unclear. A randomized placebo-controlled intervention study was conducted to compare the effects of fiber-enriched foods to their non-enriched counterparts in adults over a 12-week period on selected clinical parameters—satiety, quality of life, body sensation, and life satisfaction—subjective health status, and importance of diet for well-being. Quality of life (QOL) differed significantly between intervention and control groups at baseline, throughout, and at the end of the study. No effects on satiety, satisfaction with life, or the importance of diet for well-being could be shown between groups. With higher fiber intake, body sensation ratings increased. A higher BMI was significantly associated with lower-body sensation, subjective health status and quality of life. Fiber-enriched foods do not seem to affect feeling of satiety or parameters of well-being. Larger samples and additional methods are necessary to fully explore the effect of increased fiber intake on patient-related outcomes in more detail.
The influence of knowledge about the environmental impact of foods on consumers' decisions and perceptions
(2024) Bschaden, Andreas; Ströbele-Benschop, Nanette
Food production substantially impacts the environment, with large differences between foods, dishes, and dietary styles. Therefore, consumer behaviour is an important issue in sustainability research. Several theories attempt to explain consumer behaviour, such as the Theory of Planned Behaviour or the transtheoretical model. In both models, attitudes play a central role in explaining behaviour. Attitudes are in part influenced by knowledge, and by information individuals are exposed to. This dissertation project aimed to enhance the understanding of the interplay between consumers’ knowledge and dietary behaviour, with the focus on ecological sustainability. Three studies were conducted, each focusing on diverse aspects. First, the project investigated how aware the participants are about the environmental impact of meat consumption, and how two informative videos could influence that knowledge, as well as participants’ meat consumption. Second, the hedonic perception of a snack that was offered under different sustainability information conditions was explored. Third, the impact of carbon footprint information of dishes on food choices, as well as participants’ perception of that information was examined. The first study assessed participants’ knowledge regarding the environmental benefits of various dietary strategies, such as purchasing regional, seasonal, and organic foods, avoiding plastic packaging and foods imported by airplane, and eating less meat. Additionally, an intervention using three kinds of informative videos was conducted to investigate the impact of the provided information on the participants’ knowledge. Data were collected through a questionnaire before and after the intervention. The second study examined consumers’ hedonic perception of a novel savoury tortilla-chips shaped snack made from by-products of the food industry. The snack was offered to a sample of participants under two varying information conditions to investigate how information about the snack’s ingredients, its sustainability with general information about food losses compared to tasting the snack without such information, influenced the hedonic experience. The third study investigated the influence of displaying the carbon footprint of dishes on dish choices across various gastronomic settings and with visually different presentation methods. In addition to sales figures, customer perceptions of the provided information were also evaluated, gathered through a survey. Results suggest that consumers still underestimate the environmental benefit of eating less meat, both in general and compared to other strategies. While this result applies for all investigated groups, there were significant differences between gender and educational groups. The documentary about the environmental impact of meat consumption led to higher assessments of the environmental benefit of eating less meat a week later, but no effect was observed a year later. No such effect was observed for the educational video about the same topic, nor for the control video. Regarding the perception of a savoury snack under different information conditions, the MANOVA model showed significant effects for study condition on perceived saltiness, for gender on willingness to pay and odour, and for ecological awareness on health perception. An interaction-effect was observed for study condition and gender for perceived taste and saltiness, suggesting that women liked the snack better when sustainability information was provided, and men liked it better with no such information. Investigating the impact of carbon footprint information on food choice, in the participating university canteen, the average carbon footprint of all chosen dishes was 1.5 percent lower when carbon footprint information was provided, while there were no significant effects in company canteens, food trucks, and a restaurant. The feedback from customers in the questionnaire was predominantly positive overall. In line with other research, the results have found a lack of knowledge regarding the environmental impact of food production and consumption, particularly among male participants and individuals with lower educational levels. While the documentary appeared to be effective as an educational tool in the short term, no impact was observed after a year. The presentation of the carbon footprint on menus showed inconsistent results regarding the food choices. Since this is not in line with other research findings, the topic should be further investigated in various gastronomic settings and with various ways of presenting the information. As it seems challenging to translate knowledge into action, beliefs, or attitudes, further research should prioritise that aspect. The project found consumers being open to sustainable nutrition, and desiring more information. Sustainability information about reducing food losses processing by-products did not consistently evoke a better hedonic experience in a snack tasting. However, as other studies reported such effects investigating different products and information, further research is needed to explore the effects for various information, such as climate-friendliness of products. In practice, food producers, retailers and caterers, consumers, and policymakers can adopt various strategies to address the challenge of supporting the food system to become more sustainable.
Study protocol to investigate the efficacy of confocal laser endomicroscopy-based selective single-elimination diet over standard fivefold elimination diet in patients with endomicroscopically proven food intolerance: app-assisted, monocentric, double-blind, randomised and controlled trial in Germany
(2023) Heßler, Nicole; Kordowski, Anna; Sasse, Jill; Ahlemann, Greta; Schulz, Franziska; Schröder, Torsten; Exner, Anna; Jablonski, Lennart; Jappe, Uta; Bischoff, Stephan C; Grzegorzek, Marcin; König, Inke R; Sina, Christian
Introduction: Imprecise nutritional recommendations due to a lack of diagnostic test accuracy are a frequent problem for individuals with adverse reactions to foods but no precise diagnosis. Consequently, patients follow very broad and strict elimination diets to avoid uncontrolled symptoms such as diarrhoea and abdominal pain. Dietary limitations and the uncertainty of developing gastrointestinal symptoms after the inadvertent ingestion of food have been demonstrated to reduce the quality of life (QoL) of affected individuals and subsequently might increase the risk of malnutrition and intestinal dysbiosis. This trial aims to investigate the effects of a tailored diet based on the confocal laser endoscopy (CLE) examination result to limit the side effects of unspecific and broad elimination diets and to increase the patient’s QoL. Methods and analysis: The study is designed as a prospective, double-blind, monocentric, randomised and controlled trial conducted at the University Hospital of Schleswig-Holstein, Campus Lübeck, Germany. One hundred seventy-two patients with non-IgE-related food allergies and positive CLE results will be randomised to either a tailored diet or a standard fivefold elimination diet. The primary endpoints are the difference between the end and the start of the intervention in health-related QoL and the sum score of the severity of symptoms after 12 weeks. Key secondary endpoints are changes in the severity of symptoms, further QoL measurements, self-assessed state of health and number of days with a pathologically altered stool. Microbiome diversity and metabolome of stool, urine and blood will also be investigated. Safety endpoints are body composition, body mass index and adverse events. Ethics and dissemination: The study protocol was accepted by the ethical committee of the University of Lübeck (AZ: 22-111) on 4 May2022. Results of the study will be published in peer-reviewed journals and presented at scientific meetings. Trial registration number: German Clinical Trials Register (DRKS00029323).
NOD1 cooperates with HAX‐1 to promote cell migration in a RIPK2‐ and NF‐ĸB‐independent manner
(2023) Hezinger, Lucy; Bauer, Sarah; Ellwanger, Kornelia; Piotrowsky, Alban; Biber, Felix; Venturelli, Sascha; Kufer, Thomas A.
The human Nod-like receptor protein NOD1 is a well-described pattern-recognition receptor (PRR) with diverse functions. NOD1 associates with F-actin and its protein levels are upregulated in metastatic cancer cells. A hallmark of cancer cells is their ability to migrate, which involves actin remodelling. Using chemotaxis and wound healing assays, we show that NOD1 expression correlated with the migration rate and chemotactic index in the cervical carcinoma cell line HeLa. The effect of NOD1 in cell migration was independent of the downstream kinase RIPK2 and NF-ĸB activity. Additionally, NOD1 negatively regulated the phosphorylation status of cofilin, which inhibits actin turnover. Co-immunoprecipitation assays identified HCLS1-associated protein X-1 (HAX-1) as a previously unknown interaction partner of NOD1. Silencing of HAX-1 expression reduced the migration behaviour to similar levels as NOD1 knockdown, and simultaneous knockdown of NOD1 and HAX-1 showed no additive effect, suggesting that both proteins act in the same pathway. In conclusion, our data revealed an important role of the PRR NOD1 in regulating cell migration as well as chemotaxis in human cervical cancer cells and identified HAX-1 as a protein that interacts with NOD1 and is involved in this signalling pathway.
The role of NLRC5 in obesity
(2024) Bauer, Sarah Katharina; Kufer, Thomas
Obesity and its associated morbidities are major global health problems. It has become evident in the last decades that the state of obesity is intimately linked with our immune system. Pattern recognition receptors (PRRs), the main sensor molecules of the innate immune system, were shown to play an essential role in the pathology of obesity and its associated morbidities. Among others, members of the nucleotide-binding and oligomerization domain (NOD) -like receptors (NLRs), a family of cytosolic PRRs, were associated with the obesity-accompanying low-grade inflammatory response contributing to obesity-associated morbidities. NLRC5 is a NLR protein functioning as key transcriptional regulator of major histocompatibility complex (MHC) class I genes responsible for antigen presentation. Recent observations now suggest novel roles of NLRC5 in metabolic trades, but so far, no confirmation of these singular observations is available, and the underlying mechanisms remain elusive. The aim of this thesis was to characterize the role of the NLR protein NLRC5 in obesity. To this end, two Nlrc5 deficient mouse lines (Nlrc5 deltaExon4-7 and Nlrc5 deltaExon4) were subjected to high-fat diet (HFD) feeding and phenotypic, morphological, and biochemical analyses were performed. Female Nlrc5 deltaExon4-7 mice presented with higher body and adipose tissue (AT) weight gain and larger adipocytes compared to wildtype (WT) animals. This phenotype, however, could not be recapitulated in the Nlrc5 deltaExon4 mouse line. Microbiome analysis revealed subtle alterations of the faecal microbiome by diet:genotype interactions. To further characterize the effect of NLRC5 deficiency on adipocyte differentiation, the CRISPR/Cas9 gene editing system was used to modify Nlrc5 expression in the 3T3-L1 preadipocyte cell line. Using inducible HeLa cell lines with stable GFP-NLRC5 expression we showed NLRC5 to interact with the master regulator of adipogenesis peroxisome proliferator-activated receptor y (PPARy) and to enhance the expression of PPARy target genes. In addition, a contribution of NLRC5 to PPARy’s anti-inflammatory actions was revealed using NLRC5 deficient THP-1 macrophage-like cells and bone marrow-derived macrophages from Nlrc5 deltaExon4-7 mice. To elucidate the mechanism behind the synergy between NLRC5 and PPARy, reporter gene and chromatin immunoprecipitation (ChIP) assays were performed. Lastly, the expression of multiple NLR family members was correlated with body mass index (BMI) in obese human patients and investigated in the adipose tissue and liver of HFD-fed mice, the latter revealing Nlrp10 to be highly upregulated by HFD feeding. Taken together, this thesis provides a comprehensive characterization of Nlrc5 deficient mice on HFD and reveals a function of NLRC5 as transcriptional co-regulator of PPARy targets and its anti-inflammatory properties. In addition, this work provides first insights into the potential mechanism behind the synergistic transcriptional regulation by NLRC5 and PPARy and extends the knowledge on the regulation of NLR expression by HFD feeding.
Die Mikroalge Phaeodactylum tricornutum : Bioverfügbarkeit, Sicherheit und potenzieller gesundheitlicher Nutzen für die humane Ernährung
(2023) Kopp, Lena Janine; Bischoff, Stephan C.
The dissertation by Lena Kopp investigated the suitability of the microalga Phaeodactylum tricornutum (PT) for human nutrition. PT contains essential nutrients such as the long-chain omega-3 fatty acid eicosapentaenoic acid (EPA), which is otherwise found mainly in fish. In addition, PT contains a high content of other nutrients such as proteins, carotenoids (in particular fucoxanthin), vitamins and β-glucans, which have nutritive and therapeutic potential. Clinical and animal studies have shown that the PT biomass ingestion is safe and has potential health effects, such as anti-inflammatory and prebiotic effects. The results suggest that PT can be used as a food for human nutrition with possible health-promoting effects.
Immunomodulatory effects of resveratrol on human intestinal mast cell signaling in vitro and mast cell associated enteritis and colitis in mice
(2023) Bilotta, Sabrina; Lorentz, Axel
By releasing their pre-stored or de novo synthesized mediators, mast cells (MC) are important immunoregulatory cells responsible for a variety of inflammatory reactions. Although known to be major effector cells in immunoglobuline (Ig) E dependent allergic reactions, MC have been widely shown to play a role in various inflammations of the gut. Diseases of the gastrointestinal tract (GIT) are widespread and multicausal. Those affected suffer from the sometimes severe symptoms and may experience restrictions on their daily life. Even if conventional medication is applied routinely, aim of the past and current research is to establish supportive and/or alternative medication that is based on natural substances. These may be on the basis of small natural components like resveratrol, a stilbene mostly found in grapes. Numerous positive properties are attributed to resveratrol. These are anti-inflammatory, anti-cancerogenic, anti-oxidative, as well as neuroprotective effects. The use of substances of natural origin as so-called nutraceuticals can help to increase the acceptance of medication by those affected, but also to reduce and overcome the side effects associated with conventional treatment. Effects of resveratrol were examined on the reactivity of MC isolated from patients’ tissue undergoing bowel resection. The results of this work show that resveratrol exhibited potent inhibitory effects on high affinity IgE receptor mediated activation of MC, strongly inhibiting not only MC degranulation, but also gene expression of the pro-inflammatory cytokines C-X-C motif chemokine ligand (CXCL) 8, C-C motif chemokine ligand (CCL) 2, CCL3, CCL4 and tumor necrosis factor (TNF-) α. Ultimately, the intracellular signaling cascade triggered during MC activation via IgE receptor leads to mediator release. Following IgE receptor mediated activation, phosphorylation of signaling molecules like extracellular signal-regulated kinase (ERK) 1/2 and signal transducer and activator of transcription (STAT) 3, occurs. ERK1/2 was found to be responsible for phosphorylation of mitochondrial STAT3, which contributes significantly to MC degranulation. Treatment with resveratrol was able to inhibit the phosphorylation of STAT3 by more than 50 % and that of ERK1/2 by almost 100 %. Furthermore, the experiments performed succeeded in isolating the mitochondrial fraction from relatively low human intestinal MC (hiMC) numbers. Also, in this fraction we could detect phosphorylation of STAT3 and ERK1/2 after MC activation, which was reduced after treatment with resveratrol. Having shown the strong inhibitory effects in vitro, we set out to examine immunomodulatory effects of resveratrol in vivo. Presence and activity of MC are closely related to intestinal inflammations in consequence of food allergy (FA) and inflammatory bowel disease (IBD). In mice, FA can be studied using the ovalbumin (OVA)-induced allergic enteritis model and colitis can be studied using the IL-10 knockout (-/-) mice, which develop a spontaneous form of chronic colitis. We could show that the oral application of resveratrol inhibited the increase of MC numbers in the colon and duodenum of affected animals in both experimental settings. Less pronounced but still visible effects of resveratrol administration were observed in the colon with regard to epithelial damage, cell infiltration and reduction of goblet cell numbers. In all cases, based on a scoring system, the damage decreased to the level of the corresponding controls receiving no additive and in which no allergic enteritis was induced or nor colitis developed. Overall, allergic enteritis resulted in a weaker symptomatology, and IL-10-/- animals showed a delayed appearance of the typical symptoms. The results of this thesis show a strong inhibitory effect of resveratrol on hiMC. This could be detected for mediator release as well as on gene expression levels and in the phosphorylation of the signaling molecules ERK1/2 and STAT3, which we could also identify in the mitochondria of hiMC. We observed positive influences on MC-associated parameters in the OVA enteritis and IL-10-/- colitis mouse models. With regard to its use as nutraceutical, resveratrol could therefore come more of a focus in the future.
Einfluss der mediterranen Ernährung auf das Fettsäuremuster von Erythrozytenmembranen sowie auf Darmmikrobiota- und Darmbarriere-assoziierte Biomarker : Mechanismen und klinische Anwendungen
(2022) Seethaler, Benjamin; Bischoff, Stephan C.
Dissertation from Benjamin Seethaler: "Effect of the Mediterranean diet on the fatty acid pattern of erythrocyte membranes and on gut microbiota- and gut barrier-associated biomarkers - mechanisms and clinical applications". In summary, the results of the PhD project offer new insights into the biomedical mechanisms of action and health effects of the Mediterranean diet. Of particular importance is the relationship we have shown between dietary fiber from the Mediterranean diet, its fermentation to short-chain fatty acids, and its beneficial influence on impaired intestinal barrier function. In the future, our studies may provide the basis for personalized nutritional therapy to improve impaired gut barrier function in high-risk breast cancer patients. Furthermore, we were able to establish LBP and zonulin as biomarkers to detect gut barrier function and to determine and assess gut barrier disorders. This method validation simplifies or enables the assessment of intestinal barrier function in clinical practice or clinical trials.
Einfluss verschiedener Getreidearten und Herstellungsverfahren auf den Gehalt immunogener Substanzen in Brot sowie in vivo auf die Verträglichkeit an der Maus und im Menschen
(2022) Zimmermann, Julia; Bischoff, Stephan C.
There are three medical conditions that are triggered by consumption of cereals. Celiac disease, wheat allergy and non-celiac wheat sensitivity (NCWS). While the underlying triggers and mechanisms of the first two entities have been extensively studied, there is still uncertainty in this regard for NCWS. Symptoms are nonspecific and diagnostic markers are lacking. Besides bacterial fermentable carbohydrates (FODMAPs), selected cereal proteins such as gluten or α-amylase trypsin inhibitors (ATIs) are in the focus of research as triggers. The aim of the present work was firstly to investigate the influence of the choice of cereal (common wheat, spelt, rye) and the production of bread (degree of milling and choice between yeast and sourdough) on the presence of potentially immunogenic proteins based on proteomic analysis. In a second step, the tolerability of selected breads should be investigated in a transgenic mouse model with intestinal inflammation and in a human study in patients with NCWS and subjective spelt tolerance. This was to narrow down possible triggers of NCWS and to investigate underlying mechanisms. Within the project, protein composition of bread and flour samples was analyzed based on a quantitative proteomics method (nano-UHPLC-ESI-MS based). In addition, a list of known and potentially immunogenic cereal proteins was generated based on Pfam annotation, which was used for the analysis of allergens in flour and bread. This showed that neither the absolute number nor the abundance of these allergenic proteins were dependent on the degree of milling of the flour or the fermentation process of the dough, which means that they are not selectively degraded during bread production. However, such proteins were identified in higher numbers and higher relative amounts in spelt and wheat samples compared to rye samples. Furthermore, different bread types from the proteome analysis were investigated in a mouse model with intestinal inflammation. This did not demonstrate better tolerability of rye bread compared to spelt and wheat bread. Instead, there was a trend for sourdough bread to have less negative effects on intestinal inflammation compared to yeast dough bread. It also turned out that inflammation was increased independently of gluten. No differences were found between wheat and spelt in either the proteomic analysis or the animal studies in this project. However, in a subgroup of NCWS patients, spelt bread is subjectively better tolerated than wheat bread, which could be due to both genetics and the different production of wheat and spelt bread. In order to verify the phenomenon and identify underlying mechanisms, if any, a clinical study was conducted in patients of this subgroup. The aim of the blinded study was to investigate whether spelt bread is actually better tolerated than wheat bread and whether the production process (16h dough or 1h dough + baking agent) has an influence. After each bread (4 days each + 3 days washout), gastrointestinal symptoms were assessed using the Irritable Bowel Syndrome Severity Scoring System (IBS-SSS) questionnaire. Extraintestinal symptoms and various blood and stool parameters were also analyzed. It was found that spelt bread was not better tolerated compared to wheat bread after blinded consumption and that FODMAP-rich bread was not worse tolerated compared to FODMAP-poor bread.
Characterization of the role of the NLR proteins NLRC5 and NLRP11 in the immune response
(2021) Kienes, Ioannis; Kufer, Thomas
Recognition of conserved molecular patterns by pattern recognition receptors (PRRs) is crucial for the initiation of an innate immune response. Within PRRs, the NOD-like receptor (NLR) family is, in humans, a group of 22 cytosolic proteins, which function as PRRs of the innate immune system and as regulators of adaptive immune responses. However, it has become evident, that several NLR proteins also function as regulators of innate immune responses. In this thesis the function of human NLR proteins NLRC5 and NLRP11 in immune responses was further characterized. The first part of this thesis was focused on the NOD-like receptor NLRC5. NLRC5 and major histocompatibility complex (MHC) class II transcriptional activator (CIITA) are the master regulators of MHC class I and II transcription, respectively. Both proteins can translocate into the nucleus, where they induce transcription of MHC class I and class II, respectively. As NLRC5 and CIITA do not possess intrinsic DNA binding capacities, they exert their function by binding to a common multiprotein complex, termed MHC enhanceosome and through recruitment of further transcriptional regulators. Although MHC enhanceosome components are, as known thus far, identical, NLRC5 and CIITA are specific for their respective transcriptional targets. In this work we employed several techniques to identify novel interaction partners of NLRC5 to understand the mechanisms behind this specificity. As the N-terminal domain death-domain like fold (DD) of NLRC5 has previously been shown to be involved in conferring specificity, we adapted a protocol for proximal ligation by fusion of the NLRC5 DD to biotin ligase from Aquifex aeolicus (BioID2) to unravel the interactome of this NLRC5 domain. By enrichment of biotinylated proteins through streptavidin-biotin precipitation and analysis of the proteins by LC-MS/MS, we aimed to identify novel putative interactors with functions in transcriptional regulation. Additionally, we used yeast 2 hybrid screening of the NLRC5 DD against a library of human CD4+ and CD8+ T cells for the identification of novel interaction partners. This led to the identification of the paired amphipathic helix protein Sin3A (Sin3A) and the negative elongation factor B (NELFB) as interactors of NLRC5 DD. Characterization of their role in transcriptional regulation of MHC class I revealed an inhibitory role of both proteins. However, as we also observed repression of CIITA-mediated MHC class II transcription, both proteins are likely not involved in determination of target specificity of NLRC5. Translocation of NLRC5 into the nucleus is essential for the induction of MHC class I transcription. It has however previously been shown, that forced nuclear localization of NLRC5 strongly diminishes its transcriptional activity. We therefore employed co-immunoprecipitation of differentially localized NLRC5 constructs to identify interaction partners which might be involved in post translational regulation of NLRC5. Further, to advance our understanding of the NLRC5 DD, we aimed to elucidate its crystal structure. For this, we established a protocol for large-scale recombinant expression and purification of the NLRC5 DD for subsequent crystallization of the recombinant protein. The second part of this thesis was focused on NLRP11. Tight regulation of inflammatory cytokine and interferon (IFN) production in innate immunity is pivotal for optimal control of pathogens and avoidance of immunopathology. NLRP11 has previously been shown to regulate type I IFN and other pro-inflammatory responses. To gain a deeper understanding of the underlying mechanism, we aimed to identify novel NLRP11 interactors, through which the inhibition is conferred. Here we generated cell lines stably expressing NLRP11-eGFP as novel tools to elucidate the functions of NLRP11. We identified the ATP-dependent RNA helicase DDX3X as a novel binding partner of NLRP11 by co immunoprecipitation and LC-MS/MS. DDX3X is known to enhance type I IFN responses and NLRP3 inflammasome activation. We demonstrate that NLRP11 can abolish IKKe-mediated phosphorylation of DDX3X, resulting in lower type I IFN induction upon viral infection. These effects were dependent on the leucine-rich repeat (LRR) domain of NLRP11 that we mapped as the interaction domain for DDX3X. In addition, NLRP11 also suppressed NLRP3-mediated caspase-1 activation in an LRR domain-dependent manner, suggesting, that NLRP11 might sequester DDX3X and prevent it from promoting NLRP3-induced inflammasome activation. Taken together, our data revealed DDX3X as a central target of NLRP11, which can mediate the effects of NLRP11 on type I IFN induction, as well as NLRP3 inflammasome activation. This expands our knowledge of the molecular mechanisms underlying NLRP11 function in innate immunity and suggests that both NLRP11 and DDX3X might be promising targets for modulation of innate immune responses.
Effekte der Proteinzufuhr während einer Gewichtsabnahme auf fettfreie Masse, Ruheenergieumsatz und physische Funktion bei übergewichtigen postmenopausalen Frauen - eine randomisierte, kontrollierte Studie
(2020) Englert, Isabell; Kohlenberg-Müller, Kathrin
Aim Weight loss in old age increases the risk of sarcopenia caused by the age-related reduction of fat-free mass (FFM). Due to the strong correlation between FFM and resting energy expenditure (REE), the maintenance of this must also be considered. In addition, the physical function (PF) must be maintained. The objective was to investigate the impact of protein intake on changes in fat-free mass (FFM), resting energy expenditure (REE), and physical function (PF) during weight loss. Methods 54 postmenopausal women (BMI 30.9 ± 3.4 kg/m²; 59 ± 7 years of age) were randomized into two groups with 0.8 g (K) or 1.5 g protein/kg body weight/d (P) energy-restricted diets (- 750 kcal of individual energy requirements) for 12 weeks, followed by six months weight maintenance with ad libitum food intake. The protein dose was evenly distributed to two liquid and one solid meal. The shakes of the P group were additionally enriched with pure whey protein. Four seminars were held to provide information on the course of the study and in particular on healthy nutrition. At the beginning and at the end of the study, the subjects kept a 7-day nutrition protocol. FFM (by bioelectrical impedance analysis), REE (by indirect calorimetry), PF (strength, endurance, and balance by short physical performance battery test (SPPB), 400 m walking speed and handgrip strength by hand dynamometer), blood parameters (lipid and carbohydrate profile, urea, vitamin D, calcium, magnesium, liver and kidney values from serum) and blood pressure were measured at baseline, after weight loss, and after follow up. The evaluation was primarily based on an intention to treat analysis with correlation and regression analysis, paired and unpaired t-tests, whereby the significance level was set ≤ at 0.05. The values given are continuous mean values ± standard deviation. Results 46 women completed the weight loss intervention and 29 were followed up after weight maintenance. Weight loss was -4.6 ± 3.6 kg (P) and -5.2 ± 3.4 kg (K) (both p < 0.001) and weight regain during follow up was 1.3 ± 2.8 kg (P, p = 0.028) and 0.4 ± 2.5 kg (K) (p = 0.392) with no differences between protein groups. Similar losses in FFM (-0.9 ± 1.1 kg (P) vs. -1.0 ± 1.3 kg (K)) and REE (-206 ± 136 kcal/d (P) vs. -239 ± 134 kcal/d (K), both p < 0.001) were observed in both groups. At follow-up, no changes in FFM were detected in both groups whereas in the NP group the REE increased again (138 ± 296, p = 0.02). The main determinants of the FFM loss were the energy deficit and the speed of weight loss. In the NP group, SPPB score improved with weight loss (0.6 ± 0.8, p < 0.001) and handgrip strength decreased (-1.7 ± 3.4 kg, p < 0.001) whereas no changes were observed in the HP group. The blood profile improved, especially regarding the carbohydrate profile, due to weight loss, and blood pressure. Conclusion A high protein weight loss diet without exercise had no impact on preserving FFM and REE but may help maintain muscle strength in postmenopausal women. As the handgrip strength can be a sensitive parameter for incipient sarcopenia even before the muscle mass decreases noticeably, it can be concluded that an increased protein intake during weight loss can counteract the risk of sarcopenia. Energy deficit and speed of weight loss should be considered as confounders in future studies. In addition, further strategies must be pursued to maintain FFM in weight loss in old age.
The dietary quality of food pantry users from a socio-ecological perspective
(2019) Simmet, Anja; Ströbele-Benschop, Nanette
In Germany, around 1.5 Mio people with a low income receive food for a small fee from one of around 940 so-called “Tafel”. In high-income countries like Germany, users of food pantries are a particularly vulnerable population group, as they are characterized by cumulative health risks. They often suffer from food insecurity and from chronic diseases such as hypertension, diabetes mellitus type 2 or obesity. Given that these diseases as well as food insecurity both relate to diet, the dietary quality plays a critical role in the health status of food pantry users. To illustrate the dietary quality and the different levels of factors influencing the dietary quality among food pantry users, this thesis has the following aims: 1. to provide a summary of the scientific evidence about the dietary quality of food pantry users in high-income countries; 2. to provide a summary of the scientific evidence about the nutritional quality of food provided by food pantries in high-income countries; 3. to examine the distribution of Tafel food pantries and food banks and to provide a representative picture of their resources (e.g. food, volunteers etc.), activities (e.g. provided programs) and users in Germany; 4. to examine the distribution of Tafel food pantries and to identify compositional and physical environmental correlates of food pantry use in Berlin. To reach these aims, two systematic reviews were conducted (first and second publication). In addition, an explorative cross-sectional study consisting of an analysis of secondary data and a comprehensive survey among all Tafel belonging to the federal association “Tafel Deutschland” was performed (third publication). Finally, an ecological study was conducted by analyzing and mapping food pantry use and compositional and physical characteristics of areas in Berlin (fourth publication). The first review revealed that the dietary quality among the reviewed food pantry users tended to be low as reflected by an inadequate intake of energy, fruit and vegetables, dairy products and calcium compared to national recommendations. The reviewed food pantry users had, in particular, a lower consumption of dairy products compared to the general populations. The second review demonstrated that the nutritional quality of reviewed pre-packed food bags provided by food pantries was highly variable within and between included studies. It also showed that the nutritional quality of most of the food bags was low, reflected, in particular, by a low provision of dairy products, vitamins A and C and calcium compared to national recommendations. None of the studies included in the reviews were nationally representative. The third publication showed that the German food bank system Tafel Deutschland provided a comprehensive net of food pantries, social supermarkets, food banks and other services. However, the number of Tafel per 10,000 welfare recipients was lower in eastern Germany (M = 1.37) compared to western Germany ((M = 2.12), t(162.54) = 4.2424, p < 0.0001). In contrast to the results of the studies included in the second review, the Tafel mainly provided perishable food such as fruits and vegetables (41.42% of the amount of food distributed), bakery products (19.85%) and dairy products (13.39%). However, due to the dependence on donors such as retailers, the amount of food distributed varied widely. The assistance of most local Tafel is based on volunteer labor as 89.97% of Tafel’s staff were volunteers. In 79 districts, in which all Tafel participated in the survey, there was an average of 179 Tafel beneficiaries per 1000 welfare recipients and 17 Tafel beneficiaries per 1000 residents overall. An even lower usage was found by the fourth publication which revealed that only two out of 1000 adult inhabitants and six out of 1000 children received assistance from one of the 44 investigated Tafel food pantries in Berlin. Tafel use by adults (A) or by children (B) was related to the percentage of welfare recipients (A, β = 0.17, p < 0.001 und B, β = 0.16, p = 0.002), the percentage of inhabitants with migration background (A, β = - 0.08, p = 0.002), the number of discount grocery percentage stores per 1000 children (B, β = 5.65, p = 0.010), and the number of stops of the public transport within a radius of 500 meters (A, β = - 0.24, p = 0.020). The most important limitation of both studies was the unknown reliability of the data collected from the Tafel. The dietary quality of food pantry users in high-income countries was seen to be low. Although food pantries did not provide a full meal plan for a healthy diet, they may have a positive impact on the dietary quality of its users. However, to better understand the role of food pantries in the complex interplay of individual, social and environmental influences on the diet of food pantry users, multi-level approaches should be used in the future. Moreover, researchers are strongly recommended to investigate the mechanisms by which using a food pantry might impact users’ dietary quality. In addition to these implications for researchers, this thesis makes several recommendations to practitioners at food pantries in Germany and other high-income countries. Following these recommendations could make the position of food pantries in civil society into an important entry point for health promotion among a part of the food insecure population in the future.
Metabolic chamber studies on energy- and macronutrient metabolism : impact of meal skipping and energy flux
(2018) Nas, Alessa; Bosy-Westphal, Anja
The classical concept of body weight regulation attributes the development of obesity to a chronically positive energy balance. There is, however, evidence indicating that beyond this basic concept, the effectiveness of body weight regulation is affected by the circadian regulation of metabolism and the level of energy flux (EF, level of energy balance). Meal skipping affects circadian regulation and might therefore also affect the regulation of body weight. In addition, an asymmetric regulation of body weight is hypothesized with improved effectiveness when EF is high (active lifestyle) and less effectiveness at a low EF (sedentary lifestyle). Metabolic chambers offer the opportunity to acquire short-term parameters of energy and macronutrient balance that precede long-term weight gain and therefore, can help to understand the impact of nutrition and physical activity interventions on body weight regulation. This thesis presents the implementation of a metabolic chamber system (Chapter II) and investigates the acute impact of meal skipping (Chapter III) and energy flux (Chapter IV) on energy and macronutrient metabolism by performing two well-controlled, cross-over intervention studies using metabolic chambers. The implementation of the metabolic chambers revealed, that thorough considerations must be made in terms of the metabolic chamber environment (room ventilation and position of analyzer unit), the additional devices (e.g. air conditioner) used as well as the study protocol, in order to obtain good data quality. The study on meal skipping includes 17 healthy participants who underwent 3 isocaloric 24-h interventions (55%, 30%, and 15% carbohydrate, fat and protein, respectively): a breakfast skipping day (BSD) and a dinner skipping day (DSD) separated by a conventional 3-meal-structure day (control). Energy and macronutrient balance were measured and postprandial glucose and insulin concentrations, as well as 24-h glycemia and 24-h insulin secretion (C-peptide), were analyzed. When compared with the 3-meal control, 24-h energy expenditure was higher on DSD (DSD: +69 kcal/d; p < 0.05), but not on BSD. Whereas, fat oxidation increased on the BSD only (+13 g/d; p < 0.01). Spontaneous physical activity, 24-h glycemia, and 24-h insulin secretion did not differ between intervention days. The postprandial homeostasis model assessment index (+54%) and glucose concentrations after lunch (+46%) were, however, higher on the BSD than on the DSD (both p < 0.05). When compared with 3 meals/d, dinner skipping increased energy expenditure. In contrast, higher postprandial insulin concentrations and increased fat oxidation with breakfast skipping show the development of metabolic inflexibility in response to prolonged fasting that may in the long-term lead to impaired glucose homeostasis. The study on energy flux includes 16 healthy participants who underwent three 24-h interventions with different levels of EF: (i) low EF, physical activity level (PAL) = 1.3 – 1.4 (ii) medium EF, PAL = 1.5 – 1.6 and (iii) high EF, PAL = 1.7 – 1.8 each at energy balance (EB), caloric restriction (CR), and overfeeding (OF) (100%, 75% and 125% of individual energy requirement with 50% carbohydrate, 35% fat, 15% protein). Different levels of EF were accomplished by walking (4 km/h) on a treadmill (0, 165 and 330 min). Sleeping energy expenditure (SEE), 24-h macronutrient oxidation and relative macronutrient balance (oxidation relative to intake) were determined. During EB and OF, 24-h fat oxidation increased with higher EF. This resulted in a higher relative fat balance at medium EF (EB: +17%, OF: +14%) and high EF (EB: +23%, OF: +17%) compared to low EF (all p < 0.05). SEE during EB and OF was higher at medium (EB: +5 kcal/3h and OF: +12 kcal/3h) and high (EB: +7 kcal/3h and OF: +18 kcal/3h) EF compared to low EF (all, p < 0.05). In contrast, during CR 24-h fat oxidation was only higher at high EF compared to low EF and neither relative fat balance nor SEE differed between the EF levels. A higher EF might have beneficial effects on body weight regulation during short-term overfeeding and energy balance because it increased SEE and improved relative fat balance. However, during short-term caloric restriction, a higher EF had no impact on the regulation of energy or fat balance. Therefore, a high EF especially can attenuate the adverse effects of short-term overfeeding. Altogether, this thesis emphasizes the importance of physical activity in daily life and suggests that the adverse metabolic outcome of breakfast skipping (caused by a positive energy balance after lunch with a preceding prolonged fasting period) might be attenuated by a high EF.
Ernährung von sozial benachteiligten Menschen am Beispiel von Tafelkunden : Betrachtung des Ernährungs- und Gesundheitsverhaltens, der Verbreitung von Ernährungsarmut und des Obst- und Gemüsekonsums
(2018) Depa, Julia; Ströbele-Benschop, Nanette
In industrialized countries the distribution of mortality of morbidity follows a social gradient. Hence, among people with a lower socioeconomic status (SES) the nutritional quality is poor and low fruit and vegetable consumption occur more frequently compared to people with a higher SES. A particularly vulnerable group are socially disadvantaged people such as food bank users. Food insecurity (FI) is also more common in this population group. Food banks exist worldwide and distribute donated groceries (e.g. from food retailers) to socially disadvantaged people. In Germany, 1.5 million users are supported by over 900 food banks providing mainly fresh fruits and vegetables (FV). In Germany, little is known about the diet of food bank users. The following research questions were developed for this thesis: 1. Are there differences in health and nutrition status among people using food banks in different types of cities and can differences in these variables be found when comparing food bank users with the as low SES defined German population (chapter 2, first publication)? 2. How widespread is FI among food bank users and which socio-demographic, food bank-related and health variables are associated with FI (chapter 3, second publication)? 3. Are there differences in FV intake between male food bank users and male eligible non-food bank users and can FV intake among this study population be increased by an intervention providing weekly free and personally selected FV (14 portions/ per week) for four weeks (chapter 4, third publication)? For the publications of this thesis data of food bank users regarding socio-demographic, health- and nutrition-related variables were collected in different cities. The questions were taken from the questionnaire of the national study DEGS (German Health Interview and Examination Survey for Adults) and GEDA (German Health Update) and from the FIES (Food Insecurity and Experience Scale). Additionally, questionnaires were adapted to the study population. In all publications cross-sectional study designs were used. Except in the third publication for the second part of the research questions an intervention study using a longitudinal design was conducted. The first publication shows that food bank users from the three examined cities (Berlin n=94, Ludwigsburg n=64, Fulda n=114) are not a homogenous group. Food bank users assess their self-rated health mostly worse than people from the low SES German population (proportion self-rated health as moderate, bad or very bad: men 67.4% vs. 43.5%, women 68.8% vs. 36.7%). Additionally, they consume less fruit daily (proportion of daily fruit consumption: men 39.8% vs. 43.5%, women 56.2% vs. 62.4%). The second publication reveals with 70.2% a high rate of FI among food bank users (Stuttgart n=510, Karlsruhe n=186, Berlin n=337). Especially age (r τ = -0.224, p<0.000) and smoking (V=0.219, p<0.000) are strongly associated with FI. The third publication clarifies that male food banks users (n=24) from Stuttgart did not differ in consumed FV amount (2.2 portions/day vs. 1.8 portions/day) and variety (17 types/month vs. 14.4 types/month) compared to non-food bank users (n=28). Besides, the weekly provision of free fruit and vegetables for four weeks (14 portions/ month) increases the consumed fruit and vegetable amount (difference-IG 1.1 portions/day vs. difference-CG -0.2 portions/day) and variety (difference-IG 2.6 types/month vs. difference-CG -1.2 types/month) among the intervention group (n=25) compared to the control group (n=27). It is important to note the high amount of smokers among food bank users both in the first (46.9%) and the third publication (66.7%). The reported results correspond to research from abroad. Results of the publications are limited by unbalanced standardization and representation procedures (first and second publication) and the cross-sectional design (first and second publication and first research question of the third publication). It can be concluded that food banks are a suitable option to target socially disadvantaged people and to explore their nutrition and health behavior as well as a suitable option to provide intervention opportunities. The extent of health inequality is probably underestimated. FI is widespread among food bank users and smoking as poverty factor among food bank users should be further examined. The free provision of fruit and vegetables seems to be an appropriate possibility to increase the consumption and variety of fruit and vegetables among socially disadvantaged people.
Strategien zur Optimierung der Ernährung in schulischen Einrichtungen mit besonderem Fokus auf die Nährstoffversorgung
(2014) Böhringer, Stefanie; Grimm, Peter
Changes in society, such as growing number of employed women lead to a larger amount of schools in Baden-Württemberg that offer full time school programs. Due to this, the number of all day school pupils has more than tripled over the last 10 years (Landesinstitut für Schulentwicklung et al. 2011). As a result, in 2008, the Federal Ministry of Food, Agriculture and Consumer Protection set up the Vernetzungsstelle Schulverpflegung in Baden-Württemberg in joint financing with the Ministry of Rural Affairs and Consumer Protection. The Vernetzungsstelle Schulverpflegung Baden-Württemberg supports the schools in this new additional task of lunch catering and improves the quality and acceptance of the school meals in Baden-Württemberg. This study aims to evaluate and enhance the methods of the Vernetzungsstelle Schulverpflegung Baden-Württemberg to optimize the nutrition in educational institutions. For this purpose, ten schools with the corresponding seven canteens and 6000 pupils had been reviewed from June 2010 until October 2011 by the Vernetzungsstelle Schulverpflegung Baden-Württemberg. This supervision targets to improve the nutrition consumption of the pupils and thereby increase the amount of pupils with normal body-mass-index In order to reach this aim, both, the quality as well as the general requirements (e.g. menu, reservation) of the school lunch needs to be improved. Furthermore, the satisfaction of the customers as well as the resulting weekly participation in lunch at school canteens should be increased. Additionally, the pupils shall be sensitized for an appropriate diet. The “round table” is the fundament of the supervision of schools by the Vernetzungsstelle Schulverpflegung Baden-Württemberg. Several target groups come together to organize the improvement of the school canteen. For each school canteen, individual important optimizing and advertising strategies take place. Optimizing strategies include the training of caterers and canteen personal. Advertising strategies comprise several events, like pasta day or Santa Clause day, which help to raise the acceptance of the school canteen. To be able to measure the changes caused by intervention, an analyses takes place at the beginning and the end of the the study. For that, the surrounding conditions are evaluated by an expert interview with the responsible person of the school canteen. The satisfaction of the customers, such as pupils, teachers and parents is evaluated by written questionnaire. The quality of the lunch is analyzed by the criteria of the DGE quality standards of school feeding. Additionally, anthropometric data, like body height, body weight and abdominal girth, of six and seventh class pupils are measured. A bioelectric impedance analysis determines the body composition. Furthermore, a food frequency questionnaire and a 24h recall are conducted. Results of the study show, that during the time of intervention, on average 8 optimizing strategies and 15 advertising strategies are executed. 2991 (2966) pupils, 1939 (1785) parents and 143 (114) teachers take part at the first (second) survey. The cooperation of the schools with the Vernetzungsstelle Schulverpflegung Baden-Württemberg leads to an increased acceptance of the scool canteen by pupils, teachers and parents. Pupils and parents upgrade the school canteen judgment (scale 1-5) from 2,8 to 2,4 and teachers upgrade from 2,5 to 2,2. The cooperation doesn´t lead to an improvement of the nutrition composition of the canteen food. The pupils declare that they eat in the city more rarely, however rather bring food from home or eat at home. These changes in lunch habbits, though, do not have any impact on the total nutrition intake of the pupils from the sixth and seven grades (N first evaluation=436, N second evaluation=413). Therefore, neither the antropometroc parameters nor the body composition of the pupils are influenced. The methods of the Vernetzungsstelle Schulverpflegung Baden-Württemberg can be used successfully to raise the acceptance of the school canteen. Reasons, why there have been no changes in the nutrition intake of the pupils, can be discussed versatile. One explanation is that the intervention phase is too short in order to take measurable influence on the nutrition intake. As a result of the cooperation one can say that the school canteen initially needs to be integrated in every day school life, in order to improve the lunch catering. Moreover, the schools should make sure that one person takes responsibility for the catering. In addition to that, all involved parties have to work together on the optimization. Not until these points are settled, optimizing strategies, such as quality improvement of the canteen food, can´t be possible and successful. Finally, the food offer needs to be acceptable for the pupils, in order to be advertising in a good way.
Role of plasminogen activator inhibitor (PAI-1) in the pathogenesis of fructose-induced non-alcoholic fatty liver disease (NAFLD)
(2012) Kanuri, Giridhar; Bischoff, Stephan C.
Non-alcoholic fatty liver disease (NAFLD), a liver disease frequently associated with obesity, type 2 diabetes and dyslipidemia has become a worldwide health problem during the last decades. Results of recent studies suggest that a diet rich in fructose may also be a risk factor for the development of NAFLD. Results of our own group but also other group suggest that TNFα and PAI-1 may be involved in the development of NAFLD in rodents but also humans. Therefore, the aim of the present study was to investigate the role TNFα and PAI-1 in the onset of fructose-induced NAFLD in a mouse model as well as in human NAFLD patients. The specific aims were 1) Are TNFR1-/- mice protected from fructose-induced NAFLD? If yes, what are the molecular mechanisms involved? TNFR1 -/- and wild-type mice were either fed 30% fructose solution or tap water. Chronic fructose feeding caused a significant ~5-fold increase in triglyceride accumulation and neutrophil infiltration in livers of wild-type mice and an ~8-fold increase in plasma alanine aminotransferase (ALT) levels in comparison to control mice. Similar effects of fructose feeding were not found in TNFR 1-/- mice. Indeed, the protective effect of the tumor necrosis factor receptor 1 (TNFR1) deletion against the onset of fructose-induced steatosis was associated with decreased sterol regulatory element-binding protein 1 (SREBP-1), fatty acid synthase (FAS) and plasminogen activator inhibitor 1 (PAI-1) expression in the liver. Furthermore, the protective effect was also associated with protection against alterations markers of insulin signaling cascade (e.g. adenosine monophosphate-activated protein kinase (AMPK), protein kinase B (Akt) levels). However, markers of hepatic lipid peroxidation, inducible nitric oxide synthase (iNOS) protein and adenosine triphosphate (ATP) levels were similar between wild-type and TNFR1 -/- mice fed fructose. Taken together, these data suggest that TNFα plays a casual role in the onset of fructose-induced liver damage as well as insulin resistance in mice through signaling cascades downstream of TNFR1. 2) Are PAI-1-/- mice protected from fructose-induced NAFLD? And if so, what are the molecular mechanisms involved? To address if PAI-1 is also a critical factor in the onset of fructose-induced NAFLD, PAI-1-/- and wild-type mice were either fed a fructose solution or tap water. Chronic fructose feeding in wild-type mice caused a marked increase in hepatic triglycerides, PAI-1 expression and plasma ALT levels in comparison to water controls. A similar effect of fructose feeding was not found in PAI-1-/- mice. PAI-1-/- mice fed fructose were protected from hepatic steatosis despite similar portal endotoxin levels, alterations of markers of insulin resistance and hepatic TNFα protein levels between fructose fed groups. The protective effect of the loss of PAI-1 against the onset of fructose-induced steatosis was associated with a significant increase in phospho-cMet, phospho Akt, expression of apolipoprotein B (ApoB) and activity of microsomal triglyceride transfer protein (MTTP) in livers of PAI-1-/- mice in comparison to fructose fed wild-type mice. Moreover, in PAI-1-/- mice expression of CD1d and markers of CD1d-reactive iNKT cells were markedly higher than in wild-type mice; however, expression of markers of activation of CD1d-reactive iNKT cells (e.g. interleukin 15 (IL-15) and interferon γ (INFγ)) were only found to be increased in livers of fructose fed PAI-1-/- mice. Taken together, these data suggest that PAI-1 plays a causal role in mediating the early phase of fructose-induced liver damage in mice through signalling cascades down-stream of Kupffer cells and TNFα. 3) Are molecular mechanisms identified in mouse studies also relevant to human situation? To determine if the alterations found in livers of animals with NAFLD are also relevant in humans with NAFLD, markers of lipid peroxidation, insulin signaling and number of iNKT cells were determined in 6 controls and 11 NAFLD patients.4-hydroxynonenal (4-HNE) protein adducts levels were significantly higher in livers of NAFLD patients whereas expression of insulin receptor substrate (IRS-1) was reduced by ~80 % in comparison to controls. PAI-1 protein levels primarily found in hepatocytes was significantly higher in NAFLD patients; however, hepatic CD1d and MTTP mRNA expression did not differ between groups. Hepatic c-Met and BCL-2l mRNA expressions were significantly lower in NAFLD patients in comparison to controls and number of CD3ζ positive cells was higher. In contrast, expression of iNKT cell markers (e.g. IL-4 and IL-15) was significantly lower in livers of patients with NAFLD when compared with controls.Taken together, the present study suggests that the molecular mechanism involved in the progression of NAFLD is similar in both rodents and humans. Furthermore, TNFα and PAI-1 may be considered as therapeutic targets for NAFLD.
Einfluss der IgG-Subklassen sowie der immunmodulatorischen Aminosäuren Arginin und Glutamin auf die Aktivierung menschlicher Darmmastzellen
(2011) Lechowski, Sandra; Lorentz, Axel
Mast cells are key effector cells in allergic diseases, like food allergy. Activation of mast cells occurs mainly by crosslinking of immunoglobulin E (IgE) receptor FcεRI. The role of allergen specific immunoglobulin G (IgG) subclasses IgG1, IgG2, IgG3, and IgG4 in activation of intestinal mast cells is not yet fully clarified. On the one hand, mast cells can be activated via IgG1 and by the mechanisms of ?IgG-supercrosslinking?. On the other hand, IgG1 and IgG4 are up regulated during immunotherapy and might be able to attenuate allergic reactions. In this work, we investigated the effect of IgG subclasses on IgE dependent and IgE-independent mediator release of human intestinal mast cells. Mast cells, isolated and purified from human intestinal tissue, were cultured with their growth factor stem cell factor (SCF) in combination with interferon γ (IFN-γ) or with interleukin 4 (IL-4). IFN-γ is known to up regulate expression of the activating IgG receptor FcγRI on mast cells, derived from peripheral blood. IL-4 is known to enhance IgE triggered mediator release from human intestinal mast cells. Expression of FcγRI, FcγRII, FcγRIII and FcεRI was analysed by flow cytometry. IgG subclasses should be isolated from human sera by affinity chromatography. However, the required purity of at least 95 % for each subclass could not be achieved. Thus, intestinal mast cells were stimulated with human myeloma IgG1-4 and their specific anti-IgG1-4 antibodies in combination with IgE/anti-IgE and the release of inflammatory mediators was measured. We could show that human intestinal mast cells cultured with IFN-γ express FcγRI, while FcγRII and FcγRIII were only weakly expressed. Mast cells cultured with IL-4 do not express FcγR. IgG subclasses themselves did not activate intestinal mast cells neither in culture with IFN-γ, nor cultured with IL-4. In combination with IgE/anti-IgE, the IgG subclasses IgG1/anti IgG1, IgG2/anti IgG2 and IgG4/anti IgG4 tended to decrease the release of pre-stored β-hexosaminidase and de novo synthesized leukotriene C4 (LTC4) but the reduction was not significant. To conclude, IFN-γ induced in human intestinal mast cells the expression of FcγRI, but this did not result in stimulation of the cells by IgG. Thus, human intestinal mast cells do not respond or only to a small extend to IgG subclasses and differ from mast cells derived from peripheral blood which could be activated via IgG1. The second part of the work examines the effect of the immunomodulatory amino acids arginine and glutamine on IgE-dependent activation of human intestinal mast cells. Background was the measured anti-inflammatory effect of combined pharmacological doses of both amino acids on intestinal biopsies from patients with Crohn?s disease. Beside allergic diseases, mast cells play a role in chronic intestinal inflammation. Therefore, they could be involved in the described effect. Human intestinal mast cells were incubated over night with combined physiological doses or pharmacological doses of arginine (0.1 mmol/L or 2 mmol/l) and glutamine (0.6 mmol/l or 10 mmol/l). Mast cells were stimulated with IgE/anti-IgE and release of β-hexosaminidase and LTC4, induction of cytokines and activation of signaling molecules was analysed. In human intestinal mast cells, cultured in SCF and IL-4, combined pharmacological concentrations of arginine and glutamine led to decreased release of LTC4 about 40 ± 22 % and to reduced induction of cytokines like CCL2 (monocyte chemotactic protein 1), CCL4 (macrophage inflammatory protein 1 beta), CXCL8 (IL-8), and TNF (tumor growth factor alpha) about an average of 58 ± 35 % compared to physiological doses of arginine and glutamine. The anti-inflammatory effects of arginine and glutamine were associated with decreased activation levels of MAP kinases like ERK1/2, p38, JNK pan und MEK1/2 about 38 ± 17 % as well as serine/threonine kinase Akt about 23 ± 20 %. Therewith, arginine and glutamine reduce activation levels of signaling molecules known to be involved in IgE dependent mast cell cytokine expression. Here, the results show that arginine and glutamine exert anti-inflammatory effects on human intestinal mast cells in vitro. Further investigations are necessary to monitor these findings in vivo prior to use arginine and glutamine as immunomodulatory agents in inflammatory and allergic diseases.

Browse

Recent Submissions