@article{HauffeRathAgyapongetal.2022, author = {Hauffe, Robert and Rath, Michaela and Agyapong, Wilson and Jonas, Wenke and Vogel, Heike and Schulz, Tim Julius and Schwarz, Maria and Kipp, Anna Patricia and Bl{\"u}her, Matthias and Kleinridders, Andr{\´e}}, title = {Obesity Hinders the Protective Effect of Selenite Supplementation on Insulin Signaling}, series = {Antioxidants}, volume = {11}, journal = {Antioxidants}, edition = {5}, publisher = {MDPI}, address = {Basel, Schweiz}, issn = {2076-3921}, doi = {10.3390/antiox11050862}, pages = {1 -- 16}, year = {2022}, abstract = {The intake of high-fat diets (HFDs) containing large amounts of saturated long-chain fatty acids leads to obesity, oxidative stress, inflammation, and insulin resistance. The trace element selenium, as a crucial part of antioxidative selenoproteins, can protect against the development of diet-induced insulin resistance in white adipose tissue (WAT) by increasing glutathione peroxidase 3 (GPx3) and insulin receptor (IR) expression. Whether selenite (Se) can attenuate insulin resistance in established lipotoxic and obese conditions is unclear. We confirm that GPX3 mRNA expression in adipose tissue correlates with BMI in humans. Cultivating 3T3-L1 pre-adipocytes in palmitate-containing medium followed by Se treatment attenuates insulin resistance with enhanced GPx3 and IR expression and adipocyte differentiation. However, feeding obese mice a selenium-enriched high-fat diet (SRHFD) only resulted in a modest increase in overall selenoprotein gene expression in WAT in mice with unaltered body weight development, glucose tolerance, and insulin resistance. While Se supplementation improved adipocyte morphology, it did not alter WAT insulin sensitivity. However, mice fed a SRHFD exhibited increased insulin content in the pancreas. Overall, while selenite protects against palmitate-induced insulin resistance in vitro, obesity impedes the effect of selenite on insulin action and adipose tissue metabolism in vivo.}, language = {en} } @article{dePinhoTavaresLealdaSilvaRochaGomesetal.2021, author = {de Pinho Tavares Leal, Pedro Ernesto and da Silva, Alexandre Alves and Rocha-Gomes, Arthur and Riul, Tania Regina and Cunha, Rennan Augusto and Reichetzeder, Christoph and Villela, Daniel Campos}, title = {High-Salt Diet in the Pre- and Postweaning Periods Leads to Amygdala Oxidative Stress and Changes in Locomotion and Anxiety-Like Behaviors of Male Wistar Rats}, series = {Frontiers in Behavioral Neuroscience}, volume = {15}, journal = {Frontiers in Behavioral Neuroscience}, publisher = {Frontiers Research Foundation}, address = {Lausanne, Schweiz}, issn = {1662-5153}, doi = {10.3389/fnbeh.2021.779080}, pages = {1 -- 12}, year = {2021}, abstract = {High-salt (HS) diets have recently been linked to oxidative stress in the brain, a fact that may be a precursor to behavioral changes, such as those involving anxiety-like behavior. However, to the best of our knowledge, no study has evaluated the amygdala redox status after consuming a HS diet in the pre- or postweaning periods. This study aimed to evaluate the amygdala redox status and anxiety-like behaviors in adulthood, after inclusion of HS diet in two periods: preconception, gestation, and lactation (preweaning); and only after weaning (postweaning). Initially, 18 females and 9 male Wistar rats received a standard (n = 9 females and 4 males) or a HS diet (n = 9 females and 5 males) for 120 days. After mating, females continued to receive the aforementioned diets during gestation and lactation. Weaning occurred at 21-day-old Wistar rats and the male offspring were subdivided: control-control (C-C)—offspring of standard diet fed dams who received a standard diet after weaning (n = 9-11), control-HS (C-HS)—offspring of standard diet fed dams who received a HS diet after weaning (n = 9-11), HS-C—offspring of HS diet fed dams who received a standard diet after weaning (n = 9-11), and HS-HS—offspring of HS diet fed dams who received a HS diet after weaning (n = 9-11). At adulthood, the male offspring performed the elevated plus maze and open field tests. At 152-day-old Wistar rats, the offspring were euthanized and the amygdala was removed for redox state analysis. The HS-HS group showed higher locomotion and rearing frequency in the open field test. These results indicate that this group developed hyperactivity. The C-HS group had a higher ratio of entries and time spent in the open arms of the elevated plus maze test in addition to a higher head-dipping frequency. These results suggest less anxiety-like behaviors. In the analysis of the redox state, less activity of antioxidant enzymes and higher levels of the thiobarbituric acid reactive substances (TBARS) in the amygdala were shown in the amygdala of animals that received a high-salt diet regardless of the period (pre- or postweaning). In conclusion, the high-salt diet promoted hyperactivity when administered in the pre- and postweaning periods. In animals that received only in the postweaning period, the addition of salt induced a reduction in anxiety-like behaviors. Also, regardless of the period, salt provided amygdala oxidative stress, which may be linked to the observed behaviors.}, language = {en} } @phdthesis{Radloff2018, author = {Radloff, Katrin}, title = {The role of the fatty acid profile and its modulation by cytokines in the systemic inflammation in cancer cachexia}, school = {Universit{\"a}t Potsdam}, pages = {156}, year = {2018}, abstract = {Systemic inflammation is a hallmark of cancer cachexia. Among tumor-host interactions, the white adipose tissue (WAT) is an important contributor to inflammation as it suffers morphological reorganization and lipolysis, releasing free fatty acids (FA), bioactive lipid mediators (LM) and pro-inflammatory cytokines, which accentuate the activation of pro-inflammatory signaling pathways and the recruitment of immune cells to the tissue. This project aimed to investigate which inflammatory factors are involved in the local adipose tissue inflammation and what is the influence of such factors upon enzymes involved in FA or LM metabolism in healthy individuals (Control), weight stable gastro-intestinal cancer patients (WSC) and cachectic cancer patients (CC). The results demonstrated that the inflammatory signature of systemic inflammation is different from local adipose tissue inflammation. The systemic inflammation of the cachectic cancer patients was characterized by higher levels of circulating saturated fatty acids (SFA), tumor-necrosis-factor-α (TNF-α), interleukins IL-6, IL-8 and CRP while levels of polyunsaturated fatty acids (PUFAs), especially n3-PUFAs, were lower in CC than in the other groups. In vitro and in adipose tissue explants, pro-inflammatory cytokines and SFAs were shown to increase the chemokines IL-8 and CXCL10 that were found to be augmented in adipose tissue inflammation in CC which was more profound in the visceral adipose tissue (VAT) than in subcutaneous adipose tissue (SAT). Systemic inflammation was negatively associated with the expression of PUFA synthesizing enzymes, though gene and protein expression did hardly differ between groups. The effects of inflammatory factors on enzymes in the whole tissue could have been masked by differentiated modulation of the diverse cell types in the same tissue. In vitro experiments showed that the expression of FA-modifying enzymes such as desaturases and elongases in adipocytes and macrophages was regulated into opposing directions by TNF-α, IL-6, LPS or palmitate. The higher plasma concentration of the pro-resolving LM resolvin D1 in CC cannot compensate the overall inflammatory status and the results indicate that inflammatory cytokines interfere with synthesis pathways of pro-resolving LM. In summary, the data revealed a complex inter-tissue and inter-cellular crosstalk mediated by pro-inflammatory cytokines and lipid compounds enhancing inflammation in cancer cachexia by feed-forward mechanisms.}, language = {en} } @article{PueschelKlauderHenkelOberlaender2022, author = {P{\"u}schel, Gerhard and Klauder, Julia and Henkel-Oberl{\"a}nder, Janin}, title = {Macrophages, low-grade inflammation, insulin resistance and hyperinsulinemia}, series = {Journal of Clinical Medicine : open access journal}, volume = {11}, journal = {Journal of Clinical Medicine : open access journal}, number = {15}, publisher = {MDPI}, address = {Basel, Schweiz}, issn = {2077-0383}, doi = {10.3390/jcm11154358}, pages = {1 -- 30}, year = {2022}, abstract = {Metabolic derangement with poor glycemic control accompanying overweight and obesity is associated with chronic low-grade inflammation and hyperinsulinemia. Macrophages, which present a very heterogeneous population of cells, play a key role in the maintenance of normal tissue homeostasis, but functional alterations in the resident macrophage pool as well as newly recruited monocyte-derived macrophages are important drivers in the development of low-grade inflammation. While metabolic dysfunction, insulin resistance and tissue damage may trigger or advance pro-inflammatory responses in macrophages, the inflammation itself contributes to the development of insulin resistance and the resulting hyperinsulinemia. Macrophages express insulin receptors whose downstream signaling networks share a number of knots with the signaling pathways of pattern recognition and cytokine receptors, which shape macrophage polarity. The shared knots allow insulin to enhance or attenuate both pro-inflammatory and anti-inflammatory macrophage responses. This supposedly physiological function may be impaired by hyperinsulinemia or insulin resistance in macrophages. This review discusses the mutual ambiguous relationship of low-grade inflammation, insulin resistance, hyperinsulinemia and the insulin-dependent modulation of macrophage activity with a focus on adipose tissue and liver.}, language = {en} } @article{HauffeRathSchelletal.2021, author = {Hauffe, Robert and Rath, Michaela and Schell, Mareike and Ritter, Katrin and Kappert, Kai and Deubel, Stefanie and Ott, Christiane and J{\"a}hnert, Markus and Jonas, Wenke and Sch{\"u}rmann, Annette and Kleinridders, Andr{\´e}}, title = {HSP60 reduction protects against diet-induced obesity by modulating energy metabolism in adipose tissue}, series = {Molecular Metabolism}, volume = {53}, journal = {Molecular Metabolism}, publisher = {Elsevier}, address = {Amsterdam, Niederlande}, issn = {2212-8778}, doi = {10.1016/j.molmet.2021.101276}, pages = {1 -- 14}, year = {2021}, abstract = {Objective Insulin regulates mitochondrial function, thereby propagating an efficient metabolism. Conversely, diabetes and insulin resistance are linked to mitochondrial dysfunction with a decreased expression of the mitochondrial chaperone HSP60. The aim of this investigation was to determine the effect of a reduced HSP60 expression on the development of obesity and insulin resistance. Methods Control and heterozygous whole-body HSP60 knockout (Hsp60+/-) mice were fed a high-fat diet (HFD, 60\% calories from fat) for 16 weeks and subjected to extensive metabolic phenotyping. To understand the effect of HSP60 on white adipose tissue, microarray analysis of gonadal WAT was performed, ex vivo experiments were performed, and a lentiviral knockdown of HSP60 in 3T3-L1 cells was conducted to gain detailed insights into the effect of reduced HSP60 levels on adipocyte homeostasis. Results Male Hsp60+/- mice exhibited lower body weight with lower fat mass. These mice exhibited improved insulin sensitivity compared to control, as assessed by Matsuda Index and HOMA-IR. Accordingly, insulin levels were significantly reduced in Hsp60+/- mice in a glucose tolerance test. However, Hsp60+/- mice exhibited an altered adipose tissue metabolism with elevated insulin-independent glucose uptake, adipocyte hyperplasia in the presence of mitochondrial dysfunction, altered autophagy, and local insulin resistance. Conclusions We discovered that the reduction of HSP60 in mice predominantly affects adipose tissue homeostasis, leading to beneficial alterations in body weight, body composition, and adipocyte morphology, albeit exhibiting local insulin resistance.}, language = {en} } @article{KotthoffLisecSchwerdtleetal.2019, author = {Kotthoff, Lisa and Lisec, Jan and Schwerdtle, Tanja and Koch, Matthias}, title = {Prediction of transformation products of monensin by electrochemistry compared to microsomal assay and hydrolysis}, series = {Molecules}, volume = {24}, journal = {Molecules}, number = {15}, publisher = {MDPI}, address = {Basel}, issn = {1420-3049}, doi = {10.3390/molecules24152732}, pages = {12}, year = {2019}, abstract = {The knowledge of transformation pathways and identification of transformation products (TPs) of veterinary drugs is important for animal health, food, and environmental matters. The active agent Monensin (MON) belongs to the ionophore antibiotics and is widely used as a veterinary drug against coccidiosis in broiler farming. However, no electrochemically (EC) generated TPs of MON have been described so far. In this study, the online coupling of EC and mass spectrometry (MS) was used for the generation of oxidative TPs. EC-conditions were optimized with respect to working electrode material, solvent, modifier, and potential polarity. Subsequent LC/HRMS (liquid chromatography/high resolution mass spectrometry) and MS/MS experiments were performed to identify the structures of derived TPs by a suspected target analysis. The obtained EC-results were compared to TPs observed in metabolism tests with microsomes and hydrolysis experiments of MON. Five previously undescribed TPs of MON were identified in our EC/MS based study and one TP, which was already known from literature and found by a microsomal assay, could be confirmed. Two and three further TPs were found as products in microsomal tests and following hydrolysis, respectively. We found decarboxylation, O-demethylation and acid-catalyzed ring-opening reactions to be the major mechanisms of MON transformation.}, language = {en} } @article{SaguTchewonpiHuschekWaldbachBragaetal.2022, author = {Sagu Tchewonpi, Sorel and Huschek, Gerd and Waldbach Braga, Tess and Rackiewicz, Michal and Homann, Thomas and Rawel, Harshadrai Manilal}, title = {Design of Experiment (DoE) for Optimization of HPLC Conditions for the Simultaneous Fractionation of Seven α-Amylase/Trypsin Inhibitors from Wheat (Triticum aestivum L.)}, series = {Processes : open access journal}, volume = {10}, journal = {Processes : open access journal}, edition = {2}, publisher = {MDPI}, address = {Basel, Schweiz}, issn = {2227-9717}, doi = {10.3390/pr10020259}, pages = {1 -- 18}, year = {2022}, abstract = {Wheat alpha-amylase/trypsin inhibitors remain a subject of interest considering the latest findings showing their implication in wheat-related non-celiac sensitivity (NCWS). Understanding their functions in such a disorder is still unclear and for further study, the need for pure ATI molecules is one of the limiting problems. In this work, a simplified approach based on the successive fractionation of ATI extracts by reverse phase and ion exchange chromatography was developed. ATIs were first extracted from wheat flour using a combination of Tris buffer and chloroform/methanol methods. The separation of the extracts on a C18 column generated two main fractions of interest F1 and F2. The response surface methodology with the Doehlert design allowed optimizing the operating parameters of the strong anion exchange chromatography. Finally, the seven major wheat ATIs namely P01083, P17314, P16850, P01085, P16851, P16159, and P83207 were recovered with purity levels (according to the targeted LC-MS/MS analysis) of 98.2 ± 0.7; 98.1 ± 0.8; 97.9 ± 0.5; 95.1 ± 0.8; 98.3 ± 0.4; 96.9 ± 0.5, and 96.2 ± 0.4\%, respectively. MALDI-TOF-MS analysis revealed single peaks in each of the pure fractions and the mass analysis yielded deviations of 0.4, 1.9, 0.1, 0.2, 0.2, 0.9, and 0.1\% between the theoretical and the determined masses of P01083, P17314, P16850, P01085, P16851, P16159, and P83207, respectively. Overall, the study allowed establishing an efficient purification process of the most important wheat ATIs. This paves the way for further in-depth investigation of the ATIs to gain more knowledge related to their involvement in NCWS disease and to allow the absolute quantification in wheat samples.}, language = {en} } @article{FigueroaCamposGKTKruizengaSaguTchewonpietal.2022, author = {Figueroa Campos, Gustavo Adolfo and G. K. T. Kruizenga, Johannes and Sagu Tchewonpi, Sorel and Schwarz, Steffen and Homann, Thomas and Taubert, Andreas and Rawel, Harshadrai Manilal}, title = {Effect of the post-harvest processing on protein modification in green coffee beans by phenolic compounds}, series = {Foods : open access journal}, volume = {11}, journal = {Foods : open access journal}, edition = {2}, publisher = {MDPI}, address = {Basel, Schweiz}, issn = {2304-8158}, doi = {10.3390/foods11020159}, pages = {19}, year = {2022}, abstract = {The protein fraction, important for coffee cup quality, is modified during post-harvest treatment prior to roasting. Proteins may interact with phenolic compounds, which constitute the major metabolites of coffee, where the processing affects these interactions. This allows the hypothesis that the proteins are denatured and modified via enzymatic and/or redox activation steps. The present study was initiated to encompass changes in the protein fraction. The investigations were limited to major storage protein of green coffee beans. Fourteen Coffea arabica samples from various processing methods and countries were used. Different extraction protocols were compared to maintain the status quo of the protein modification. The extracts contained about 4-8 µg of chlorogenic acid derivatives per mg of extracted protein. High-resolution chromatography with multiple reaction monitoring was used to detect lysine modifications in the coffee protein. Marker peptides were allocated for the storage protein of the coffee beans. Among these, the modified peptides K.FFLANGPQQGGK.E and R.LGGK.T of the α-chain and R.ITTVNSQK.I and K.VFDDEVK.Q of β-chain were detected. Results showed a significant increase (p < 0.05) of modified peptides from wet processed green beans as compared to the dry ones. The present study contributes to a better understanding of the influence of the different processing methods on protein quality and its role in the scope of coffee cup quality and aroma. View Full-Text}, language = {en} } @article{SchjeideSchenkeSeegeretal.2022, author = {Schjeide, Brit-Maren and Schenke, Maren and Seeger, Bettina and P{\"u}schel, Gerhard}, title = {Validation of a novel double control quantitative copy number PCR method to quantify off-target transgene integration after CRISPR-induced DNA modification}, series = {Methods and protocols : M\&Ps}, volume = {5}, journal = {Methods and protocols : M\&Ps}, number = {3}, publisher = {MDPI}, address = {Basel, Schweiz}, issn = {2409-9279}, doi = {10.3390/mps5030043}, pages = {1 -- 14}, year = {2022}, abstract = {In order to improve a recently established cell-based assay to assess the potency of botulinum neurotoxin, neuroblastoma-derived SiMa cells and induced pluripotent stem-cells (iPSC) were modified to incorporate the coding sequence of a reporter luciferase into a genetic safe harbor utilizing CRISPR/Cas9. A novel method, the double-control quantitative copy number PCR (dc-qcnPCR), was developed to detect off-target integrations of donor DNA. The donor DNA insertion success rate and targeted insertion success rate were analyzed in clones of each cell type. The dc-qcnPCR reliably quantified the copy number in both cell lines. The probability of incorrect donor DNA integration was significantly increased in SiMa cells in comparison to the iPSCs. This can possibly be explained by the lower bundled relative gene expression of a number of double-strand repair genes (BRCA1, DNA2, EXO1, MCPH1, MRE11, and RAD51) in SiMa clones than in iPSC clones. The dc-qcnPCR offers an efficient and cost-effective method to detect off-target CRISPR/Cas9-induced donor DNA integrations.}, language = {en} } @article{SchaeferKakularamReischetal.2022, author = {Sch{\"a}fer, Marj{\"a}nn Helena and Kakularam, Kumar Reddy and Reisch, Florian and Rothe, Michael and Stehling, Sabine and Heydeck, Dagmar and P{\"u}schel, Gerhard Paul and Kuhn, Hartmut}, title = {Male Knock-in Mice Expressing an Arachidonic Acid Lipoxygenase 15B (Alox15B) with Humanized Reaction Specificity Are Prematurely Growth Arrested When Aging}, series = {Biomedicines}, volume = {10}, journal = {Biomedicines}, edition = {6}, publisher = {MDPI}, address = {Basel, Schweiz}, issn = {2227-9059}, doi = {10.3390/biomedicines10061379}, pages = {1 -- 22}, year = {2022}, abstract = {Mammalian arachidonic acid lipoxygenases (ALOXs) have been implicated in cell differentiation and in the pathogenesis of inflammation. The mouse genome involves seven functional Alox genes and the encoded enzymes share a high degree of amino acid conservation with their human orthologs. There are, however, functional differences between mouse and human ALOX orthologs. Human ALOX15B oxygenates arachidonic acid exclusively to its 15-hydroperoxy derivative (15S-HpETE), whereas 8S-HpETE is dominantly formed by mouse Alox15b. The structural basis for this functional difference has been explored and in vitro mutagenesis humanized the reaction specificity of the mouse enzyme. To explore whether this mutagenesis strategy may also humanize the reaction specificity of mouse Alox15b in vivo, we created Alox15b knock-in mice expressing the arachidonic acid 15-lipoxygenating Tyr603Asp+His604Val double mutant instead of the 8-lipoxygenating wildtype enzyme. These mice are fertile, display slightly modified plasma oxylipidomes and develop normally up to an age of 24 weeks. At later developmental stages, male Alox15b-KI mice gain significantly less body weight than outbred wildtype controls, but this effect was not observed for female individuals. To explore the possible reasons for the observed gender-specific growth arrest, we determined the basic hematological parameters and found that aged male Alox15b-KI mice exhibited significantly attenuated red blood cell parameters (erythrocyte counts, hematocrit, hemoglobin). Here again, these differences were not observed in female individuals. These data suggest that humanization of the reaction specificity of mouse Alox15b impairs the functionality of the hematopoietic system in males, which is paralleled by a premature growth arrest.}, language = {en} } @article{RancanVolkmannGiulbudagianetal.2019, author = {Rancan, Fiorenza and Volkmann, Hildburg and Giulbudagian, Michael and Schumacher, Fabian and Stanko, Jessica Isolde and Kleuser, Burkhard and Blume-Peytavi, Ulrike and Calderon, Marcelo and Vogt, Annika}, title = {Dermal Delivery of the High-Molecular-Weight Drug Tacrolimus by Means of Polyglycerol-Based Nanogels}, series = {Pharmaceutics : Molecular Diversity Preservation International}, volume = {11}, journal = {Pharmaceutics : Molecular Diversity Preservation International}, number = {8}, publisher = {MDPI}, address = {Basel}, issn = {1999-4923}, doi = {10.3390/pharmaceutics11080394}, pages = {14}, year = {2019}, abstract = {Polyglycerol-based thermoresponsive nanogels (tNGs) have been shown to have excellent skin hydration properties and to be valuable delivery systems for sustained release of drugs into skin. In this study, we compared the skin penetration of tacrolimus formulated in tNGs with a commercial 0.1\% tacrolimus ointment. The penetration of the drug was investigated in ex vivo abdominal and breast skin, while different methods for skin barrier disruption were investigated to improve skin permeability or simulate inflammatory conditions with compromised skin barrier. The amount of penetrated tacrolimus was measured in skin extracts by liquid chromatography tandem-mass spectrometry (LC-MS/MS), whereas the inflammatory markers IL-6 and IL-8 were detected by enzyme-linked immunosorbent assay (ELISA). Higher amounts of tacrolimus penetrated in breast as compared to abdominal skin or in barrier-disrupted as compared to intact skin, confirming that the stratum corneum is the main barrier for tacrolimus skin penetration. The anti-proliferative effect of the penetrated drug was measured in skin tissue/Jurkat cells co-cultures. Interestingly, tNGs exhibited similar anti-proliferative effects as the 0.1\% tacrolimus ointment. We conclude that polyglycerol-based nanogels represent an interesting alternative to paraffin-based formulations for the treatment of inflammatory skin conditions.}, language = {en} }