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The objective of this work was to investigate the potential effect of cereal α-amylase/trypsin inhibitors (ATIs) on growth parameters and selective digestive enzymes of Tenebrio molitor L. larvae. The approach consisted of feeding the larvae with wheat, sorghum and rice meals containing different levels and composition of α-amylase/trypsin inhibitors. The developmental and biochemical characteristics of the larvae were assessed over feeding periods of 5 h, 5 days and 10 days, and the relative abundance of α-amylase and selected proteases in larvae were determined using liquid chromatography tandem mass spectrometry. Overall, weight gains ranged from 21% to 42% after five days of feeding. The larval death rate significantly increased in all groups after 10 days of feeding (p < 0.05), whereas the pupation rate was about 25% among larvae fed with rice (Oryza sativa L.) and Siyazan/Esperya wheat meals, and only 8% and 14% among those fed with Damougari and S35 sorghum meals. As determined using the Lowry method, the protein contents of the sodium phosphate extracts ranged from 7.80 ± 0.09 to 9.42 ± 0.19 mg/mL and those of the ammonium bicarbonate/urea reached 19.78 ± 0.16 to 37.47 ± 1.38 mg/mL. The total protein contents of the larvae according to the Kjeldahl method ranged from 44.0 and 49.9 g/100 g. The relative abundance of α-amylase, CLIP domain-containing serine protease, modular serine protease zymogen and C1 family cathepsin significantly decreased in the larvae, whereas dipeptidylpeptidase I and chymotrypsin increased within the first hours after feeding (p < 0.05). Trypsin content was found to be constant independently of time or feed material. Finally, based on the results we obtained, it was difficult to substantively draw conclusions on the likely effects of meal ATI composition on larval developmental characteristics, but their effects on the digestive enzyme expression remain relevant.
The detection and quantification of nut allergens remains a major challenge. The liquid chroma-tography tandem mass spectrometry (LC-MS/MS) is emerging as one of the most widely used methods, but sample preparation prior to the analysis is still a key issue. The objective of this work was to establish optimized protocols for extraction, tryptic digestion and LC-MS analysis of almond, cashew, hazelnut, peanut, pistachio and walnut samples. Ammonium bicar-bonate/urea extraction (Ambi/urea), SDS buffer extraction (SDS), polyvinylpolypyrroli-done (PVPP) extraction, trichloroacetic acid/acetone extraction (TCA/acetone) and chloro-form/methanol/sodium chloride precipitation (CM/NaCl) as well as the performances of con-ventional tryptic digestion and microwave-assisted breakdown were investigated. Overall, the protein extraction yields ranged from 14.9 ± 0.5 (almond extract from CM/NaCl) to 76.5 ± 1.3% (hazelnut extract from Ambi/urea). Electrophoretic profiling showed that the SDS extraction method clearly presented a high amount of extracted proteins in the range of 0–15 kDa, 15–35 kDa, 35–70 kDa and 70–250 kDa compared to the other methods. The linearity of the LC-MS methods in the range of 0 to 0.4 µg equivalent defatted nut flour was assessed and recovery of internal standards GWGG and DPLNV(d8)LKPR ranged from 80 to 120%. The identified bi-omarkers peptides were used to relatively quantifier selected allergenic protein form the inves-tigated nut samples. Considering the overall results, it can be concluded that SDS buffer allows a better protein extraction from almond, peanut and walnut samples while PVPP buffer is more appropriate for cashew, pistachio and hazelnut samples. It was also found that conventional overnight digestion is indicated for cashew, pistachio and hazelnut samples, while microwave assisted tryptic digestion is recommended for almond, hazelnut and peanut extracts.
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.
Over the last few decades, the prevalence of obesity has risen to epidemic proportions worldwide. Consequently, the number of obesity in pregnancy has risen drastically. Gestational overweight and obesity are associated with impaired outcomes for mother and child. Furthermore, studies show that maternal obesity can lead to long-term consequences in the offspring, increasing the risk for obesity and cardiometabolic disease in later life. In addition to genetic mechanisms, mounting evidence demonstrates the induction of epigenetic alterations by maternal obesity, which can affect the offspring's phenotype, thereby influencing the later risk of obesity and cardiometabolic disease. Clear evidence in this regard comes from various animal models of maternal obesity. Evidence derived from clinical studies remains limited. The current article gives an overview of pathophysiological changes associated with maternal obesity and their consequences on placental structure and function. Furthermore, a short excurse is given on epigenetic mechanisms and emerging data regarding a putative interaction between metabolism and epigenetics. Finally, a summary of important findings of animal and clinical studies investigating maternal obesity-related epigenetic effects is presented also addressing current limitations of clinical studies.
The suitability of a newly developed cell-based functional assay was tested for the detection of the activity of a range of neurotoxins and neuroactive pharmaceuticals which act by stimulation or inhibition of calcium-dependent neurotransmitter release. In this functional assay, a reporter enzyme is released concomitantly with the neurotransmitter from neurosecretory vesicles. The current study showed that the release of a luciferase from a differentiated human neuroblastoma-based reporter cell line (SIMA-hPOMC1-26-GLuc cells) can be stimulated by a carbachol-mediated activation of the Gq-coupled muscarinic-acetylcholine receptor and by the Ca2+-channel forming spider toxin α-latrotoxin. Carbachol-stimulated luciferase release was completely inhibited by the muscarinic acetylcholine receptor antagonist atropine and α-latrotoxin-mediated release by the Ca2+-chelator EGTA, demonstrating the specificity of luciferase-release stimulation. SIMA-hPOMC1-26-GLuc cells express mainly L- and N-type and to a lesser extent T-type VGCC on the mRNA and protein level. In accordance with the expression profile a depolarization-stimulated luciferase release by a high K+-buffer was effectively and dose-dependently inhibited by L-type VGCC inhibitors and to a lesser extent by N-type and T-type inhibitors. P/Q- and R-type inhibitors did not affect the K+-stimulated luciferase release. In summary, the newly established cell-based assay may represent a versatile tool to analyze the biological efficiency of a range of neurotoxins and neuroactive pharmaceuticals which mediate their activity by the modulation of calcium-dependent neurotransmitter release.
A fast high performance liquid chromatography tandem mass spectrometry multi-method based on an ACN-precipitation extraction was developed for the analysis of 41 (modified) mycotoxins in beer. Validation according to the performance criteria defined by the European Commission (EC) in Commission Decision no. 657/2002 revealed good linearity (R2 > 0.99), repeatability (RSDr < 15%), reproducibility (RSDR < 15%), and recovery (79–100%). Limits of quantification ranging from 0.04 to 75 µg/L were obtained. Matrix effects varied from −67 to +319% and were compensated for using standard addition. In total, 87 beer samples, produced worldwide, were analyzed for the presence of mycotoxins with a focus on modified mycotoxins, whereof 76% of the samples were contaminated with at least one mycotoxin. The most prevalent mycotoxins were deoxynivalenol-3-glucoside (63%), HT-2 toxin (15%), and tenuazonic acid (13%). Exposure estimates of deoxynivalenol and its metabolites for German beer revealed no significant contribution to intake of deoxynivalenol.
Odd-chain fatty acids (OCFA) are inversely associated with type-2-diabetes in epidemiological studies. They are considered as a biomarker for dairy intake because fermentation in ruminants yields high amounts of propionate, which is used as the primer for lipogenesis. Recently, we demonstrated endogenous OCFA synthesis from propionate in humans and mice, but how this is affected by microbial colonization is still unexplored. Here, we investigated the effect of increasing microbiota complexity on hepatic lipid metabolism and OCFA levels in different dietary settings. Germ-free (GF), gnotobiotic (SIH, simplified human microbiota) or conventional (CONV) C3H/HeOuJ-mice were fed a CHOW or high-fat diet with inulin (HFI) to induce microbial fermentation. We found that hepatic lipogenesis was increased with increasing microbiota complexity, independently of diet. In contrast, OCFA formation was affected by diet as well as microbiota. On CHOW, hepatic OCFA and intestinal gluconeogenesis decreased with increasing microbiota complexity (GF > SIH > CONV), while cecal propionate showed a negative correlation with hepatic OCFA. On HFI, OCFA levels were highest in SIH and positively correlated with cecal propionate. The propionate content in the CHOW diet was 10 times higher than that of HFI. We conclude that bacterial propionate production affects hepatic OCFA formation, unless this effect is masked by dietary propionate intake.
Macrophages in pathologically expanded dysfunctional white adipose tissue are exposed to a mix of potential modulators of inflammatory response, including fatty acids released from insulin-resistant adipocytes, increased levels of insulin produced to compensate insulin resistance, and prostaglandin E₂ (PGE₂) released from activated macrophages. The current study addressed the question of how palmitate might interact with insulin or PGE₂ to induce the formation of the chemotactic pro-inflammatory cytokine interleukin-8 (IL-8). Human THP-1 cells were differentiated into macrophages. In these macrophages, palmitate induced IL-8 formation. Insulin enhanced the induction of IL-8 formation by palmitate as well as the palmitate-dependent stimulation of PGE₂ synthesis. PGE₂ in turn elicited IL-8 formation on its own and enhanced the induction of IL-8 release by palmitate, most likely by activating the EP4 receptor. Since IL-8 causes insulin resistance and fosters inflammation, the increase in palmitate-induced IL-8 formation that is caused by hyperinsulinemia and locally produced PGE₂ in chronically inflamed adipose tissue might favor disease progression in a vicious feed-forward cycle.
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.
Mycotoxins and pesticides regularly co-occur in agricultural products worldwide. Thus, humans can be exposed to both toxic contaminants and pesticides simultaneously, and multi-methods assessing the occurrence of various food contaminants and residues in a single method are necessary. A two-dimensional high performance liquid chromatography tandem mass spectrometry method for the analysis of 40 (modified) mycotoxins, two plant growth regulators, two tropane alkaloids, and 334 pesticides in cereals was developed. After an acetonitrile/water/formic acid (79:20:1, v/v/v) multi-analyte extraction procedure, extracts were injected into the two-dimensional setup, and an online clean-up was performed. The method was validated according to Commission Decision (EC) no. 657/2002 and document N° SANTE/12682/2019. Good linearity (R2 > 0.96), recovery data between 70-120%, repeatability and reproducibility values < 20%, and expanded measurement uncertainties < 50% were obtained for a wide range of analytes, including very polar substances like deoxynivalenol-3-glucoside and methamidophos. However, results for fumonisins, zearalenone-14,16-disulfate, acid-labile pesticides, and carbamates were unsatisfying. Limits of quantification meeting maximum (residue) limits were achieved for most analytes. Matrix effects varied highly (−85 to +1574%) and were mainly observed for analytes eluting in the first dimension and early-eluting analytes in the second dimension. The application of the method demonstrated the co-occurrence of different types of cereals with 28 toxins and pesticides. Overall, 86% of the samples showed positive findings with at least one mycotoxin, plant growth regulator, or pesticide.
Adipose tissue is central to the regulation of energy balance. While white adipose tissue (WAT) is responsible for triglyceride storage, brown adipose tissue specializes in energy expenditure. Deterioration of brown adipocyte function contributes to the development of metabolic complications like obesity and diabetes. These disorders are also leading symptoms of the Bardet-Biedl syndrome (BBS), a hereditary disorder in humans which is caused by dysfunctions of the primary cilium and which therefore belongs to the group of ciliopathies. The cilium is a hair-like organelle involved in cellular signal transduction. The BBSome, a supercomplex of several Bbs gene products, localizes to the basal body of cilia and is thought to be involved in protein sorting to and from the ciliary membrane. The effects of a functional BBSome on energy metabolism and lipid mobilization in brown and white adipocytes were tested in whole-body Bbs4 knockout mice that were subjected to metabolic challenges. Chronic cold exposure reveals cold-intolerance of knockout mice but also ameliorates the markers of metabolic pathology detected in knockouts prior to cold. Hepatic triglyceride content is markedly reduced in knockout mice while circulating lipids are elevated, altogether suggesting that defective lipid metabolism in adipose tissue creates increased demand for systemic lipid mobilization to meet energetic demands of reduced body temperatures. These findings taken together suggest that Bbs4 is essential for the regulation of adipose tissue lipid metabolism, representing a potential target to treat metabolic disorders.
Arsenic can occur in foods as inorganic and organic forms. Inorganic arsenic is more toxic than most watersoluble organic arsenic compounds such as arsenobetaine, which is presumed to be harmless for humans. Within the first German total diet study, total arsenic, inorganic arsenic, arsenobetaine, dimethylarsinic acid and monomethylarsonic acid were analyzed in various foods. Highest levels of total arsenic were found in fish, fish products and seafood (mean: 1.43 mg kg(-1); n = 39; min-max: 0.01-6.15 mg kg(-1)), with arsenobetaine confirmed as the predominant arsenic species (1.233 mg kg 1; n = 39; min-max: 0.01-6.23 mg kg (1)). In contrast, inorganic arsenic was determined as prevalent arsenic species in terrestrial foods (0.02 mg kg (1); n = 38; min-max: 0-0.11 mg kg (1)). However, the toxicity of arsenic species varies and measurements are necessary to gain information about the composition and changes of arsenic species in foods due to household processing of foods.
Background: Being an essential trace element, copper is involved in diverse physiological processes. However, excess levels might lead to adverse effects. Disrupted copper homeostasis, particularly in the brain, has been associated with human diseases including the neurodegenerative disorders Wilson and Alzheimer?s disease. In this context, astrocytes play an important role in the regulation of the copper homeostasis in the brain and likely in the prevention against neuronal toxicity, consequently pointing them out as a potential target for the neurotoxicity of copper. Major toxic mechanisms are discussed to be directed against mitochondria probably via oxidative stress. However, the toxic potential and mode of action of copper in astrocytes is poorly understood, so far. Methods: In this study, excess copper levels affecting human astrocytic cell model and their involvement in the neurotoxic mode of action of copper, as well as, effects on the homeostasis of other trace elements (Mn, Fe, Ca and Mg) were investigated. Results: Copper induced substantial cytotoxic effects in the human astrocytic cell line following 48 h incubation (EC30: 250 ?M) and affected mitochondrial function, as observed via reduction of mitochondrial membrane potential and increased ROS production, likely originating from mitochondria. Moreover, cellular GSH metabolism was altered as well. Interestingly, not only cellular copper levels were affected, but also the homeostasis of other elements (Ca, Fe and Mn) were disrupted. Conclusion: One potential toxic mode of action of copper seems to be effects on the mitochondria along with induction of oxidative stress in the human astrocytic cell model. Moreover, excess copper levels seem to interact with the homeostasis of other essential elements such as Ca, Fe and Mn. Disrupted element homeostasis might also contribute to the induction of oxidative stress, likely involved in the onset and progression of neurodegenerative disorders. These insights in the toxic mechanisms will help to develop ideas and approaches for therapeutic strategies against copper-mediated diseases.
Background Advanced glycation end-products are proteins that become glycated after contact with sugars and are implicated in endothelial dysfunction and arterial stiffening. We aimed to investigate the relationships between advanced glycation end-products, measured as skin autofluorescence, and vascular stiffness in various glycemic strata. Methods We performed a cross-sectional analysis within the European Prospective Investigation into Cancer and Nutrition (EPIC)-Potsdam cohort, comprising n = 3535 participants (median age 67 years, 60% women). Advanced glycation end-products were measured as skin autofluorescence with AGE-Reader (TM), vascular stiffness was measured as pulse wave velocity, augmentation index and ankle-brachial index with Vascular Explorer (TM). A subset of 1348 participants underwent an oral glucose tolerance test. Participants were sub-phenotyped into normoglycemic, prediabetes and diabetes groups. Associations between skin autofluorescence and various indices of vascular stiffness were assessed by multivariable regression analyses and were adjusted for age, sex, measures of adiposity and lifestyle, blood pressure, prevalent conditions, medication use and blood biomarkers. Results Skin autofluorescence associated with pulse wave velocity, augmentation index and ankle-brachial index, adjusted beta coefficients (95% CI) per unit skin autofluorescence increase: 0.38 (0.21; 0.55) for carotid-femoral pulse wave velocity, 0.25 (0.14; 0.37) for aortic pulse wave velocity, 1.00 (0.29; 1.70) for aortic augmentation index, 4.12 (2.24; 6.00) for brachial augmentation index and - 0.04 (- 0.05; - 0.02) for ankle-brachial index. The associations were strongest in men, younger individuals and were consistent across all glycemic strata: for carotid-femoral pulse wave velocity 0.36 (0.12; 0.60) in normoglycemic, 0.33 (- 0.01; 0.67) in prediabetes and 0.45 (0.09; 0.80) in diabetes groups; with similar estimates for aortic pulse wave velocity. Augmentation index was associated with skin autofluorescence only in normoglycemic and diabetes groups. Ankle-brachial index inversely associated with skin autofluorescence across all sex, age and glycemic strata. Conclusions Our findings indicate that advanced glycation end-products measured as skin autofluorescence might be involved in vascular stiffening independent of age and other cardiometabolic risk factors not only in individuals with diabetes but also in normoglycemic and prediabetic conditions. Skin autofluorescence might prove as a rapid and non-invasive method for assessment of macrovascular disease progression across all glycemic strata.
Dietary linoleic acid: will modifying dietary fat quality reduce the risk of type 2 diabetes?
(2021)
The valorization of coffee wastes through modification to activated carbon has been considered as a low-cost adsorbent with prospective to compete with commercial carbons. So far, very few studies have referred to the valorization of coffee parchment into activated carbon. Moreover, low-cost and efficient activation methods need to be more investigated. The aim of this work was to prepare activated carbon from spent coffee grounds and parchment, and to assess their adsorption performance. The co-calcination processing with calcium carbonate was used to prepare the activated carbons, and their adsorption capacity for organic acids, phenolic compounds and proteins was evaluated. Both spent coffee grounds and parchment showed yields after the calcination and washing treatments of around 9.0%. The adsorption of lactic acid was found to be optimal at pH 2. The maximum adsorption capacity of lactic acid with standard commercial granular activated carbon was 73.78 mg/g, while the values of 32.33 and 14.73 mg/g were registered for the parchment and spent coffee grounds activated carbons, respectively. The Langmuir isotherm showed that lactic acid was adsorbed as a monolayer and distributed homogeneously on the surface. Around 50% of total phenols and protein content from coffee wastewater were adsorbed after treatment with the prepared activated carbons, while 44, 43, and up to 84% of hydrophobic compounds were removed using parchment, spent coffee grounds and commercial activated carbon, respectively; the adsorption efficiencies of hydrophilic compounds ranged between 13 and 48%. Finally, these results illustrate the potential valorization of coffee by-products parchment and spent coffee grounds into activated carbon and their use as low-cost adsorbent for the removal of organic compounds from aqueous solutions.
The interplay between diet, intestinal microbiota and host is a major factor impacting health. A diet rich in unsaturated fatty acids has been reported to stimulate the growth of Bilophila wadsworthia by increasing the proportion of the sulfonated bile acid taurocholate (TC). The taurine-induced overgrowth of B. wadsworthia promoted the development of colitis in interleukin-10-deficient (IL-10(-/-)) mice. This study aimed to investigate whether intake of the sulfonates sulfoquinovosyl diacylglycerols (SQDG) with a dietary supplement or their degradation product sulfoquinovose (SQ), stimulate the growth of B. wadsworthia in a similar manner and, thereby, cause intestinal inflammation. Conventional IL-10(-/-) mice were fed a diet supplemented with the SQDG-rich cyanobacterium Arthrospira platensis (Spirulina). SQ or TC were orally applied to conventional IL-10(-/-) mice and gnotobiotic IL-10(-/-) mice harboring a simplified human intestinal microbiota with or without B. wadsworthia. Analyses of inflammatory parameters revealed that none of the sulfonates induced severe colitis, but both, Spirulina and TC, induced expression of pro-inflammatory cytokines in cecal mucosa. Cell numbers of B. wadsworthia decreased almost two orders of magnitude by Spirulina feeding but slightly increased in gnotobiotic SQ and conventional TC mice. Changes in microbiota composition were observed in feces as a result of Spirulina or TC feeding in conventional mice. In conclusion, the dietary sulfonates SQDG and their metabolite SQ did not elicit bacteria-induced intestinal inflammation in IL-10(-/-) mice and, thus, do not promote colitis.
Here were report the combination of biocompatible click chemistry of omega-azidosphinganine with fluorescence microscopy and mass spectrometry as a powerful tool to elaborate the sphingolipid metabolism. The azide probe was efficiently synthesized over 13 steps starting from l-serine in an overall yield of 20% and was used for live-cell fluorescence imaging of the endoplasmic reticulum in living cells by bioorthogonal click reaction with a DBCO-labeled fluorophore revealing that the incorporated analogue is mainly localized in the endoplasmic membrane like the endogenous species. A LC-MS(/MS)-based microsomal in vitro assay confirmed that omega-azidosphinganine mimics the natural species enabling the identification and analysis of metabolic breakdown products of sphinganine as a key starting intermediate in the complex sphingolipid biosynthetic pathways. Furthermore, the sphinganine-fluorophore conjugate after click reaction was enzymatically tolerated to form its dihydroceramide and ceramide metabolites. Thus, omega-azidosphinganine represents a useful biofunctional tool for metabolic investigations both by in vivo fluorescence imaging of the sphingolipid subcellular localization in the ER and by in vitro high-resolution mass spectrometry analysis. This should reveal novel insights of the molecular mechanisms sphingolipids and their processing enzymes have e.g. in infection.
Inhibition of acid sphingomyelinase (ASM), a lysosomal enzyme that catalyzes the hydrolysis of sphingomyelin into ceramide and phosphorylcholine, may serve as an investigational tool or a therapeutic intervention to control many diseases. Specific ASM inhibitors are currently not sufficiently characterized. Here, we found that 1-aminodecylidene bis-phosphonic acid (ARC39) specifically and efficiently (>90%) inhibits both lysosomal and secretory ASM in vitro. Results from investigating sphingomyelin phosphodiesterase 1 (SMPD1/Smpd1) mRNA and ASM protein levels suggested that ARC39 directly inhibits ASM's catalytic activity in cultured cells, a mechanism that differs from that of functional inhibitors of ASM. We further provide evidence that ARC39 dose- and time-dependently inhibits lysosomal ASM in intact cells, and we show that ARC39 also reduces platelet- and ASM-promoted adhesion of tumor cells. The observed toxicity of ARC39 is low at concentrations relevant for ASM inhibition in vitro, and it does not strongly alter the lysosomal compartment or induce phospholipidosis in vitro. When applied intraperitoneally in vivo, even subtoxic high doses administered short-term induced sphingomyelin accumulation only locally in the peritoneal lavage without significant accumulation in plasma, liver, spleen, or brain. These findings require further investigation with other possible chemical modifications. In conclusion, our results indicate that ARC39 potently and selectively inhibits ASM in vitro and highlight the need for developing compounds that can reach tissue concentrations sufficient for ASM inhibition in vivo.
Epigenetic DNA methylation of EBI3 modulates human interleukin-35 formation via NFkB signaling
(2021)
Ulcerative colitis (UC), a severe chronic disease with unclear etiology that is associated with increased risk for colorectal cancer, is accompanied by dysregulation of cytokines. Epstein-Barr virus-induced gene 3 (EBI3) encodes a subunit in the unique heterodimeric IL-12 cytokine family of either pro- or anti-inflammatory function. After having recently demonstrated that upregulation of EBI3 by histone acetylation alleviates disease symptoms in a dextran sulfate sodium (DSS)-treated mouse model of chronic colitis, we now aimed to examine a possible further epigenetic regulation of EBI3 by DNA methylation under inflammatory conditions. Treatment with the DNA methyltransferase inhibitor (DNMTi) decitabine (DAC) and TNF alpha led to synergistic upregulation of EBI3 in human colon epithelial cells (HCEC). Use of different signaling pathway inhibitors indicated NF kappa B signaling was necessary and proportional to the synergistic EBI3 induction. MALDI-TOF/MS and HPLC-ESIMS/MS analysis of DAC/TNF alpha-treated HCEC identified IL-12p35 as the most probable binding partner to form a functional protein. EBI3/IL-12p35 heterodimers (IL-35) induce their own gene upregulation, something that was indeed observed in HCEC cultured with media from previously DAC/TNF alpha-treated HCEC. These results suggest that under inflammatory and demethylating conditions the upregulation of EBI3 results in the formation of anti-inflammatory IL-35, which might be considered as a therapeutic target in colitis.
Mitochondria are critical for hypothalamic function and regulators of metabolism. Hypothalamic mitochondrial dysfunction with decreased mitochondrial chaperone expression is present in type 2 diabetes (T2D). Recently, we demonstrated that a dysregulated mitochondrial stress response (MSR) with reduced chaperone expression in the hypothalamus is an early event in obesity development due to insufficient insulin signaling. Although insulin activates this response and improves metabolism, the metabolic impact of one of its members, the mitochondrial chaperone heat shock protein 10 (Hsp10), is unknown. Thus, we hypothesized that a reduction of Hsp10 in hypothalamic neurons will impair mitochondrial function and impact brain insulin action. Therefore, we investigated the role of chaperone Hsp10 by introducing a lentiviral-mediated Hsp10 knockdown (KD) in the hypothalamic cell line CLU-183 and in the arcuate nucleus (ARC) of C57BL/6N male mice. We analyzed mitochondrial function and insulin signaling utilizing qPCR, Western blot, XF96 Analyzer, immunohistochemistry, and microscopy techniques. We show that Hsp10 expression is reduced in T2D mice brains and regulated by leptin in vitro. Hsp10 KD in hypothalamic cells induced mitochondrial dysfunction with altered fatty acid metabolism and increased mitochondria-specific oxidative stress resulting in neuronal insulin resistance. Consequently, the reduction of Hsp10 in the ARC of C57BL/6N mice caused hypothalamic insulin resistance with acute liver insulin resistance.
The regulation of energy homeostasis is controlled by the brain and, besides requiring high amounts of energy, it relies on functional insulin/insulin-like growth factor (IGF)-1 signalling in the central nervous system. This energy is mainly provided by mitochondria in form of ATP. Thus, there is an intricate interplay between mitochondrial function and insulin/IGF-1 action to enable functional brain signalling and, accordingly, propagate a healthy metabolism. To adapt to different nutritional conditions, the brain is able to sense the current energy status via mitochondrial and insulin signalling-dependent pathways and exerts an appropriate metabolic response. However, regional, cell type and receptor-specific consequences of this interaction occur and are linked to diverse outcomes such as altered nutrient sensing, body weight regulation or even cognitive function. Impairments of this cross-talk can lead to obesity and glucose intolerance and are linked to neurodegenerative diseases, yet they also induce a self-sustainable, dysfunctional 'metabolic triangle' characterised by insulin resistance, mitochondrial dysfunction and inflammation in the brain. The identification of causal factors deteriorating insulin action, mitochondrial function and concomitantly a signature of metabolic stress in the brain is of utter importance to offer novel mechanistic insights into development of the continuously rising prevalence of non-communicable diseases such as type 2 diabetes and neurodegeneration. This review aims to determine the effect of insulin action on brain mitochondrial function and energy metabolism. It precisely outlines the interaction and differences between insulin action, insulin-like growth factor (IGF)-1 signalling and mitochondrial function; distinguishes between causality and association; and reveals its consequences for metabolism and cognition. We hypothesise that an improvement of at least one signalling pathway can overcome the vicious cycle of a self-perpetuating metabolic dysfunction in the brain present in metabolic and neurodegenerative diseases.
Background & aims: Fibroblast growth factor 21 (FGF21) plays a pivotal role in glucose and lipid metabolism and has been proposed as a longevity hormone. However, elevated plasma FGF21 concentrations are paradoxically associated with mortality in higher age and little is known about the postprandial regulation of FGF21 in older adults. In this parallel group study, we investigated postprandial FGF21 dynamics and response in older (65-85 years) compared to younger (18-35 years) adults following test meals with varying macronutrient composition.
Methods: Participants (n = 60 older; n = 60 younger) were randomized to one of four test meals: dextrose, high carbohydrate (HC), high fat (HF) or high protein (HP). Blood was drawn before and 15, 30, 60, 120, 240 min after meal ingestion. Postprandial dynamics were evaluated using repeated measures ANCOVA. FGF21 response was assessed by incremental area under the curve.
Results: Fasting FGF21 concentrations were significantly higher in older adults. FGF21 dynamics were affected by test meal (p < 0.001) and age (p = 0.013), when adjusted for BMI and fasting FGF21. Postprandial FGF21 concentrations steadily declined over 240 min in both age groups after HF and HP, but not after dextrose or HC ingestion. At 240 min, FGF21 concentrations were significantly higher in older than in younger adults following dextrose (133 pg/mL, 95%CI: 103, 172 versus 91.2 pg/mL, 95%CI: 70.4, 118; p = 0.044), HC (109 pg/mL, 95%CI: 85.1, 141 versus 70.3 pg/mL, 95%CI: 55.2, 89.6; p = 0.014) and HP ingestion (45.4 pg/mL, 95%CI: 34.4, 59.9 versus 27.9 pg/mL 95%CI: 20.9, 37.1; p = 0.018). FGF21 dynamics and response to HF were similar for both age groups.
Conclusions: The age-specific differences in postprandial FGF21 dynamics and response in healthy adults, potentially explain higher FGF21 concentrations in older age. Furthermore, there appears to be a significant impact of acute and recent protein intake on FGF21 secretion.
Insulin is the main anabolic hormone secreted by 13-cells of the pancreas stimulating the assimilation and storage of glucose in muscle and fat cells. It modulates the postprandial balance of carbohydrates, lipids and proteins via enhancing lipogenesis, glycogen and protein synthesis and suppressing glucose generation and its release from the liver. Resistance to insulin is a severe metabolic disorder related to a diminished response of peripheral tissues to the insulin action and signaling. This leads to a disturbed glucose homeostasis that precedes the onset of type 2 diabetes (T2D), a disease reaching epidemic proportions. A large number of studies reported an association between elevated circulating fatty acids and the development of insulin resistance. The increased fatty acid lipid flux results in the accumulation of lipid droplets in a variety of tissues. However, lipid intermediates such as diacylglycerols and ceramides are also formed in response to elevated fatty acid levels. These bioactive lipids have been associated with the pathogenesis of insulin resistance. More recently, sphingosine 1-phosphate (S1P), another bioactive sphingolipid derivative, has also been shown to increase in T2D and obesity. Although many studies propose a protective role of S1P metabolism on insulin signaling in peripheral tissues, other studies suggest a causal role of S1P on insulin resistance. In this review, we critically summarize the current state of knowledge of S1P metabolism and its modulating role on insulin resistance. A particular emphasis is placed on S1P and insulin signaling in hepatocytes, skeletal muscle cells, adipocytes and pancreatic 13-cells. In particular, modulation of receptors and enzymes that regulate S1P metabolism can be considered as a new therapeutic option for the treatment of insulin resistance and T2D.
The existence of cancer stem cells (CSCs) poses a major obstacle for the success of current cancer therapies, especially the fact that non-CSCs can spontaneously turn into CSCs, which lead to the failure of the treatment and tumor relapse. Therefore, it is very important to develop effective strategies for the eradication of the CSCs. In this work, we have developed a CSCs-specific targeted, retinoic acid (RA)-loaded gold nanostars-dendritic polyglycerol (GNSs-dPG) nanoplatform for the efficient eradication of CSCs. The nanocomposites possess good biocompatibility and exhibit effective CSCs-specific multivalent targeted capability due to hyaluronic acid (HA) decorated on the multiple attachment sites of the bioinert dendritic polyglycerol (dPG). With the help of CSCs differentiation induced by RA, the self-renewal of breast CSCs and tumor growth were suppressed by the high therapeutic efficacy of photothermal therapy (PTT) in a synergistic inhibitory manner. Moreover, the stemness gene expression and CSC-driven tumorsphere formation were significantly diminished. In addition, the in vivo tumor growth and CSCs were also effectively eliminated, which indicated superior anticancer activity, effective CSCs suppression, and prevention of relapse. Taken together, we developed a CSCs-specific targeted, RA-loaded GNSs-dPG nanoplatform for the targeted eradication of CSCs and for preventing the relapse.
Background:
Epidemiological evidence indicates that diets rich in plant foods are associated with a lower risk of ischaemic heart disease (IHD), but there is sparse information on fruit and vegetable subtypes and sources of dietary fibre. This study examined the associations of major plant foods, their subtypes and dietary fibre with risk of IHD in the European Prospective Investigation into Cancer and Nutrition (EPIC).
Methods:
We conducted a prospective analysis of 490 311 men and women without a history of myocardial infarction or stroke at recruitment (12.6 years of follow-up, n cases = 8504), in 10 European countries. Dietary intake was assessed using validated questionnaires, calibrated with 24-h recalls. Multivariable Cox regressions were used to estimate hazard ratios (HR) of IHD.
Results:
There was a lower risk of IHD with a higher intake of fruit and vegetables combined [HR per 200 g/day higher intake 0.94, 95% confidence interval (CI): 0.90-0.99, P-trend = 0.009], and with total fruits (per 100 g/day 0.97, 0.95-1.00, P-trend = 0.021). There was no evidence for a reduced risk for fruit subtypes, except for bananas. Risk was lower with higher intakes of nuts and seeds (per 10 g/day 0.90, 0.82-0.98, Ptrend = 0.020), total fibre (per 10 g/day 0.91, 0.85-0.98, P-trend = 0.015), fruit and vegetable fibre (per 4 g/day 0.95, 0.91-0.99, P-trend = 0.022) and fruit fibre (per 2 g/day 0.97, 0.95-1.00, P-trend = 0.045). No associations were observed between vegetables, vegetables subtypes, legumes, cereals and IHD risk.
Conclusions:
In this large prospective study, we found some small inverse associations between plant foods and IHD risk, with fruit and vegetables combined being the most strongly inversely associated with risk. Whether these small associations are causal remains unclear.
The mammalian system of energy balance regulation is intrinsically rhythmic with diurnal oscillations of behavioral and metabolic traits according to the 24 h day/night cycle, driven by cellular circadian clocks and synchronized by environmental or internal cues such as metabolites and hormones associated with feeding rhythms. Mitochondria are crucial organelles for cellular energy generation and their biology is largely under the control of the circadian system. Whether mitochondrial status might also feed-back on the circadian system, possibly via mitokines that are induced by mitochondrial stress as endocrine-acting molecules, remains poorly understood. Here, we describe our current understanding of the diurnal regulation of systemic energy balance, with focus on fibroblast growth factor 21 (FGF21) and growth differentiation factor 15 (GDF15), two well-known endocrine-acting metabolic mediators. FGF21 shows a diurnal oscillation and directly affects the output of the brain master clock. Moreover, recent data demonstrated that mitochondrial stress-induced GDF15 promotes a day-time restricted anorexia and systemic metabolic remodeling as shown in UCP1-transgenic mice, where both FGF21 and GDF15 are induced as myomitokines. In this mouse model of slightly uncoupled skeletal muscle mitochondria GDF15 proved responsible for an increased metabolic flexibility and a number of beneficial metabolic adaptations. However, the molecular mechanisms underlying energy balance regulation by mitokines are just starting to emerge, and more data on diurnal patterns in mouse and man are required. This will open new perspectives into the diurnal nature of mitokines and action both in health and disease.
Uruguay River is the most important river in western Rio Grande do Sul, separating Brazil from Argentina and Uruguay. However, its pollution is of great concern due to agricultural activities in the region and the extensive use of pesticides. In a long term, this practice leads to environmental pollution, especially to the aquatic system. The objective of this study was to analyze the physicochemical characteristics, metals and pesticides levels in water samples obtained before and after the planting and pesticides' application season from three sites: Uruguay River and two minor affluents, Mezomo Dam and Salso Stream. For biomonitoring, the free-living nematode Caenorhabditis elegans was used, which were exposed for 24 h. We did not find any significant alteration in physicochemical parameters. In the pre- and post-pesticides' samples we observed a residual presence of three pesticides (tebuconazole, imazethapyr, and clomazone) and metals which levels were above the recommended (As, Hg, Fe, and Mn). Exposure to both pre- and post-pesticides' samples impaired C. elegans reproduction and post-pesticides samples reduced worms' survival rate and lifespan. PCA analysis indicated that the presence of metals and pesticides are important variables that impacted C. elegans biological endpoints. Our data demonstrates that Uruguay River and two affluents are contaminated independent whether before or after pesticides' application season. In addition, it reinforces the usefulness of biological indicators, since simple physicochemical analyses are not sufficient to attest water quality and ecological safety.
Scaling agriculture to the globally rising population demands new approaches for future crop production such as multilayer and multitrophic indoor farming. Moreover, there is a current trend towards sustainable local solutions for aquaculture and saline agriculture. In this context, halophytes are becoming increasingly important for research and the food industry. As Salicornia europaea is a highly salt-tolerant obligate halophyte that can be used as a food crop, indoor cultivation with saline water is of particular interest. Therefore, finding a sustainable alternative to the use of seawater in non-coastal regions is crucial. Our goal was to determine whether natural brines, which are widely distributed and often available in inland areas, provide an alternative water source for the cultivation of saline organisms. This case study investigated the potential use of natural brines for the production of S. europaea. In the control group, which reflects the optimal growth conditions, fresh weight was increased, but there was no significant difference between the treatment groups comparing natural brines with artificial sea water. A similar pattern was observed for carotenoids and chlorophylls. Individual components showed significant differences. However, within treatments, there were mostly no changes. In summary, we showed that the influence of the different chloride concentrations was higher than the salt composition. Moreover, nutrient-enriched natural brine was demonstrated to be a suitable alternative for cultivation of S. europaea in terms of yield and nutritional quality. Thus, the present study provides the first evidence for the future potential of natural brine waters for the further development of aquaculture systems and saline agriculture in inland regions.
The aim of this study was to assess the ability of the FFQ to describe reliable and valid dietary pattern (DP) scores. In a total of 134 participants of the European Prospective Investigation into Cancer and Nutrition-Potsdam study aged 35-67 years, the FFQ was applied twice (baseline and after 1 year) to assess its reliability. Between November 1995 and March 1997, twelve 24-h dietary recalls (24HDR) as reference instrument were applied to assess the validity of the FFQ. Exploratory DP were derived by principal component analyses. Investigated predefined DP were the Alternative Healthy Eating Index (AHEI) and two Mediterranean diet indices. From dietary data of each FFQ, two exploratory DP were retained, but differed in highly loading food groups, resulting in moderate correlations (r 0 center dot 45-0 center dot 58). The predefined indices showed higher correlations between the FFQ (r(AHEI) 0 center dot 62, r(Mediterranean Diet Pyramid Index (MedPyr)) 0 center dot 62 and r(traditional Mediterranean Diet Score (tMDS)) 0 center dot 51). From 24HDR dietary data, one exploratory DP retained differed in composition to the first FFQ-based DP, but showed similarities to the second DP, reflected by a good correlation (r 0 center dot 70). The predefined DP correlated moderately (r 0 center dot 40-0 center dot 60). To conclude, long-term analyses on exploratory DP should be interpreted with caution, due to only moderate reliability. The validity differed extensively for the two exploratory DP. The investigated predefined DP showed a better reliability and a moderate validity, comparable to other studies. Within the two Mediterranean diet indices, the MedPyr performed better than the tMDs in this middle-aged, semi-urban German study population.
Comprehensive untargeted and targeted analysis of root exudate composition has advanced our understanding of rhizosphere processes. However, little is known about exudate spatial distribution and regulation. We studied the specific metabolite signatures of asparagus root exudates, root outer (epidermis and exodermis), and root inner tissues (cortex and vasculature). The greatest differences were found between exudates and root tissues. In total, 263 non-redundant metabolites were identified as significantly differentially abundant between the three root fractions, with the majority being enriched in the root exudate and/or outer tissue and annotated as 'lipids and lipid-like molecules' or 'phenylpropanoids and polyketides'. Spatial distribution was verified for three selected compounds using MALDI-TOF mass spectrometry imaging. Tissue-specific proteome analysis related root tissue-specific metabolite distributions and rhizodeposition with underlying biosynthetic pathways and transport mechanisms. The proteomes of root outer and inner tissues were spatially very distinct, in agreement with the fundamental differences between their functions and structures. According to KEGG pathway analysis, the outer tissue proteome was characterized by a high abundance of proteins related to 'lipid metabolism', 'biosynthesis of other secondary metabolites' and 'transport and catabolism', reflecting its main functions of providing a hydrophobic barrier, secreting secondary metabolites, and mediating water and nutrient uptake. Proteins more abundant in the inner tissue related to 'transcription', 'translation' and 'folding, sorting and degradation', in accord with the high activity of cortical and vasculature cell layers in growth- and development-related processes. In summary, asparagus root fractions accumulate specific metabolites. This expands our knowledge of tissue-specific plant cell function.
Manganese (Mn) and zinc (Zn) are not only essential trace elements, but also potential exogenous risk factors for various diseases. Since the disturbed homeostasis of single metals can result in detrimental health effects, concerns have emerged regarding the consequences of excessive exposures to multiple metals, either via nutritional supplementation or parenteral nutrition. This study focuses on Mn-Zn-interactions in the nematode Caenorhabditis elegans (C. elegans) model, taking into account aspects related to aging and age-dependent neurodegeneration.