Protective Effects of Ethanol Extract of Brazilian Green Propolis and Apigenin against Weak Ultraviolet Ray-B-Induced Barrier Dysfunction via Suppressing Nitric Oxide Production and Mislocalization of Claudin-1 in HaCaT Cells.

Once weak ultraviolet ray-B (UVB) irradiates the skin cells, the generation of reactive nitrogen species (RNS), but not reactive oxygen species (ROS), is stimulated for the mislocalization of claudin-1 (CLDN1), an essential protein for forming tight junctions (TJs). Since our skin is constantly exposed to sunlight throughout our lives, an effective protection strategy is needed to maintain the skin barrier against weak UVB. In the present study, we investigated whether an ethanol extract of Brazilian green propolis (EBGP) and flavonoids had a protective effect against weak UVB irradiation-induced barrier dysfunction in human keratinocyte-derived HaCaT cells. A pretreatment with EBGP suppressed TJ permeability, RNS production, and the nitration level of CLDN1 in the weak UVB-exposed cells. Among the propolis components, apigenin and apigenin-like flavonoids have potent protective effects against NO production and the mislocalization of CLDN1 induced by UVB. The analyses between structures and biological function revealed that the chemically and structurally characteristic flavonoids with a hydroxyl group at the 4' position on the B-ring might contribute to its protective effect on barrier dysfunction caused by weak UVB irradiation. In conclusion, EBGP and its component apigenin protect HaCaT cells from weak UVB irradiation-induced TJ barrier dysfunction mediated by suppressing NO production.

Malvidin Protects against and Repairs Peptic Ulcers in Mice by Alleviating Oxidative Stress and Inflammation.

Peptic ulcer episodes cause damage to the stomach and intestine, with inflammatory cell infiltration and oxidative stress as the main players. In this study, we investigated the potential of anthocyanidin malvidin for preventive and curative peptic ulcer treatment. The anthocyanidin effects were examined in gastric ulcer mouse models induced by ethanol, non-steroidal anti-inflammatory drugs (NSAIDs), ischemia-reperfusion (IR), acetic acid and duodenal ulcer induced by polypharmacy. Expression levels of oxidative and inflammatory genes were measured to investigate the mechanism of anthocyanin activity. At a dose of 5 mg·kg-1, Malvidin prevented gastric ulcer induction by ethanol, NSAID and repaired the tissue after 6 days of IR. Moreover, the anthocyanidin accelerated the healing of acetic acid-induced ulcer, increased the gene expression of EGF and COX-1, and downregulated MMP-9. Anthocyanin treatment mitigated the effect of polypharmacy on inflammation and oxidative stress observed in the intestine. Additionally, the compound downregulated cytokine expression and TLR4 and upregulated HMOX-1 and IL-10, exhibiting protective activity in the mouse gut. Malvidin thus prevented gastric and duodenal ulcers due to prominent anti-inflammatory and antioxidative effects on the gastrointestinal tract that were related to gene expression modulation and an increase in endogenous defense mechanisms.

Prophylactic and therapeutic supplementation using fructo-oligosaccharide improves the intestinal homeostasis after mucositis induced by 5- fluorouracil.

The beneficial effects of prebiotic, such as fructo-oligosaccharides (FOS), in intestinal inflammation have been demonstrated in several studies. Herein, we evaluate whether joint treatment with FOS, both before and during mucositis, had additional beneficial effects and investigated the mechanisms underlying in the action of FOS on the intestinal barrier. BALB/c mice were randomly divided into five groups: CTR (without mucositis + saline solution), FOS (without mucositis + 6 % FOS), MUC (mucositis + saline solution), PT (mucositis + 6 % FOS supplementation before disease induction), and TT (mucositis + 6 % FOS supplementation before and during disease induction). Mucositis was induced by intraperitoneal injection (300 mg/kg) of 5-fluorouracil (5-FU). After 72 h, the animals were euthanized and intestinal permeability (IP), tight junction, bacterial translocation (BT), histology and morphometry, and immunoglobulin A secretory (sIgA), inflammatory infiltrate, and production of short-chain fatty acids (acetate, butyrate and propionate) were evaluated. The MUC group showed an increase in the IP, BT, and inflammatory infiltrate but a decrease in the tight junction expression and butyrate and propionate levels (P < 0.05). In the PT and TT groups, FOS supplementation maintained the IP, tight junction expression, and propionate concentration within physiologic levels, increased butyrate levels, and reduced BT and inflammatory infiltrate (P < 0.05). Total treatment with FOS (TT group) was more effective in maintaining histological score, morphometric parameters, and sIgA production. Thus, total treatment (prophylactic and therapeutic supplementation) with FOS was more effective than pretreatment alone, in reducing 5-FU-induced damage to the intestinal barrier.

High-Fat Diet Induces Disruption of the Tight Junction-Mediated Paracellular Barrier in the Proximal Small Intestine Before the Onset of Type 2 Diabetes and Endotoxemia.

BACKGROUND/AIM: A link between an impaired intestinal barrier, endotoxemia, and the pathogenesis of metabolic diseases, such as type 2 diabetes mellitus (T2DM), has been proposed. In previous work, we have demonstrated that the tight junction (TJ)-mediated intestinal barrier in ileum/colon was marginally changed in prediabetic mice; therefore, it does not seem to mainly contribute to the T2DM onset. In this study, the TJ-mediated epithelial barrier in the duodenum and jejunum was evaluated in mice during the development of type 2 prediabetes. METHODS/RESULTS: HF diet induced prediabetes after 60 days associated with a significant rise in intestinal permeability to the small-sized marker Lucifer yellow in these mice, with no histological signs of mucosal inflammation or rupture of the proximal intestine epithelium. As revealed by immunofluorescence, TJ proteins, such as claudins-1, -2, -3, and ZO-1, showed a significant decrease in junctional content in duodenum and jejunum epithelia, already after 15 days of treatment, suggesting a rearrangement of the TJ structure. However, no significant change in total cell content of these proteins was observed in intestinal epithelium homogenates, as assessed by immunoblotting. Despite the changes in intestinal permeability and TJ structure, the prediabetic mice showed similar LPS, zonulin, and TNF-α levels in plasma or adipose tissue, and in intestinal segments as compared to the controls. CONCLUSION: Disruption of the TJ-mediated paracellular barrier in the duodenum and jejunum is an early event in prediabetes development, which occurs in the absence of detectable endotoxemia/inflammation and may contribute to the HF diet-induced increase in intestinal permeability.

Cholecystokinin Modulates the Mucosal Inflammatory Response and Prevents the Lipopolysaccharide-Induced Intestinal Epithelial Barrier Dysfunction.

The intestinal mucosa plays a critical role in the organism, acting as an interface between the lamina propria and the harmful antigens in the lumen. Sepsis is associated with primary injury to the intestinal mucosa, which in turn induces bacterial translocation and hyperpermeability. Cholecystokinin (CCK) is a peptide synthesized by several cell types, whose immunomodulatory activity has been reported in experimental models of inflammation. We hypothesized that the CCK treatment could modulate the inflammatory response and protect the integrity of the intestinal barrier in endotoxemic rats. Ten minutes before the endotoxemia induction by lipopolysaccharide (LPS) administration, rats were pretreated with CCK at two doses (0.4 µg/kg or 40 µg/kg). Mucosal permeability, bacterial translocation, cytokines production, histology injury, and expression of tight junction (TJ) proteins were the parameters assessed. In the early phase of endotoxemia, rats exhibited impaired intestinal barrier function, increased mucosal permeability, bacterial translocation, and also hyperactivation of the inflammatory response. On the other hand, the pretreatment with CCK modulated the mucosal production of pro-inflammatory cytokines and increased the expression of seal-forming TJ proteins (occludin, claudin-1 and junctional adhesion molecule (JAM-A)) only in the colon and also, reduced the bacterial counts in the mesenteric lymph nodes. However, CCK has a site-specific mechanism of action in the colon via CCK-1R, which is upregulated by the CCK treatment. In synergy with previous findings from our research group, the present results demonstrated that CCK preserves the integrity of the intestinal mucosa and might be a promising hormonal adjuvant therapy for the treatment of sepsis.

Brazilian propolis extract reduces intestinal barrier defects and inflammation in a colitic mouse model.

Brazilian propolis is rich in cinnamic acid derivatives and reportedly reduces intestinal inflammation in rodents; however, the underlying mechanisms remain unclear. We hypothesized that the regulation of tight junction (TJ) barrier, Th17 cell differentiation, and/or, macrophage activation by cinnamic acid derivatives were involved in the propolis-mediated anti-inflammatory effect. Mice were orally administered 2% dextran sodium sulfate (DSS) in combination with either the feeding control or a diet containing 0.3% ethanol extract of Brazilian propolis for 9 days. DSS administration induced acute colitis in mice, whereas the propolis extract mitigated DSS-induced weight loss; colon shortening; increased plasma levels of lipopolysaccharide-binding protein; reduced expression of TJ proteins, such as zonula occludens, junctional adhesion molecule-A, occludin, and claudins; and increased expression of inflammatory cytokines, such as tumor necrosis factor (TNF) α, interleukin (IL) 6, and IL-17a. Cinnamic acid derivatives, such as artepillin C and caffeic acid phenethyl ester, present in the propolis extract suppressed the IL-17 production from cultured murine splenocytes through decreased retinoic acid-related orphan receptor gT expression. Baccharin, drupanin, and culifolin, which are also present in Brazilian propolis, reduced the TNF-α and/or IL-6 production by suppressing inflammatory signaling in murine RAW 264.7 macrophages. Taken together, the regulation of Th17 differentiation and macrophage activation by cinnamic acid derivatives, at least in part, contribute to the anti-inflammatory effect mediated by Brazilian propolis.

Beneficial effects of nutritional supplements on intestinal epithelial barrier functions in experimental colitis models in vivo.

Acute and chronic colitis affect a huge proportion of the population world-wide. The etiology of colitis cases can be manifold, and diet can significantly affect onset and outcome of colitis. While many forms of acute colitis are easily treatable, chronic forms of colitis such as ulcerative colitis and Crohn's disease (summarized as inflammatory bowel diseases) are multifactorial with poorly understood pathogenesis. Inflammatory bowel diseases are characterized by exacerbated immune responses causing epithelial dysfunction and bacterial translocation. There is no cure and therapies aim at reducing inflammation and restoring intestinal barrier function. Unfortunately, most drugs can have severe side effects. Changes in diet and inclusion of nutritional supplements have been extensively studied in cell culture and animal models, and some supplements have shown promising results in clinical studies. Most of these nutritional supplements including vitamins, fatty acids and phytochemicals reduce oxidative stress and inflammation and have shown beneficial effects during experimental colitis in rodents induced by dextran sulphate sodium or 2,4,6-trinitrobenzene sulfonic acid, which remain the gold standard in pre-clinical colitis research. Here, we summarize the mechanisms through which such nutritional supplements contribute to epithelial barrier stabilization.

Gold nanoparticles regulate tight junctions and improve cetuximab effect in colon cancer cells.

Aim: Colon cancer (CC) is the second cause of cancer death worldwide. The use of nanoparticles for drug delivery has been increasing in cancer clinical trials over recent years. Materials & methods: We evaluated cytotoxicity of citrate-capped gold nanoparticles (GNPs) and the role they play on cell-cell adhesion. We also used GNP for delivery of cetuximab into different CC cell lines. Results: CC cells with well-formed tight junctions impair GNP uptake. Noncytotoxic concentration of GNP increases paracellular permeability in Caco-2 cells in a reversible way, concomitantly to tight junctions proteins CLDN1 and ZO-1 redistribution. GNP functionalized with cetuximab increases death of invasive HCT-116 CC cells. Conclusion: GNP can be used for drug delivery and can improve efficiency of CC therapy.

Homoectoine Protects Against Colitis by Preventing a Claudin Switch in Epithelial Tight Junctions.

BACKGROUND: Inflammatory bowel diseases (IBD) are multifactorial disorders affecting millions of people worldwide with alarmingly increasing incidences every year. Dysfunction of the intestinal epithelial barrier is associated with IBD pathogenesis, and therapies include anti-inflammatory drugs that enhance intestinal barrier function. However, these drugs often have adverse side effects thus warranting the search for alternatives. Compatible solutes such as bacterial ectoines stabilize cell membranes and proteins. AIM: To unravel whether ectoine (1,4,5,6-tetrahydro-2-methyl-4-pyrimidinecarboxylic acid) and homoectoine (4,5,6,7-tetrahydro-2-methyl-1H-(1,3)-diazepine-4-carboxylic acid), a synthetic derivative of ectoine, have beneficial effects during dextran sulfate sodium (DSS)-induced colitis in mice. METHODS/RESULTS: We found that the disease activity index was significantly reduced by both ectoines. DSS-induced edema formation, epithelial permeability, leukocyte recruitment and tissue damage were reduced by ectoine and homoectoine, with the latter having stronger effects. Interestingly, the claudin switch usually observed during colitis (decreased expression of claudin-1 and increased expression of the leaky claudin-2) was completely prevented by homoectoine, whereas ectoine only reduced claudin-2 expression. Concomitantly, only homoectoine ameliorated the drop in transepithelial electrical resistance induced by IFN-γ and TNF-α in Caco-2 cells. Both ectoines inhibited loss of ZO-1 and occludin and prevented IFN-γ/TNF-α-induced increased paracellular flux of 4 kDa FITC-dextran in vitro. Moreover, both ectoines reduced expression of pro-inflammatory cytokines and oxidative stress during colitis. CONCLUSION: While both ectoine and homoectoine have protective effects on the epithelial barrier during inflammation, only homoectoine completely prevented the inflammatory claudin switch in tight junctions. Thus, homoectoine may serve as diet supplement in IBD patients to reach or extend remission.

The organophosphate pesticide methamidophos opens the blood-testis barrier and covalently binds to ZO-2 in mice.

Methamidophos (MET) is an organophosphate (OP) pesticide widely used in agriculture in developing countries. MET causes adverse effects in male reproductive function in humans and experimental animals, but the underlying mechanisms remain largely unknown. We explored the effect of MET on mice testes (5 mg/kg/day/4 days), finding that this pesticide opens the blood-testis barrier and perturbs spermatogenesis, generating the appearance of immature germ cells in the epididymis. In the seminiferous tubules, MET treatment changed the level of expression or modified the stage-specific localization of tight junction (TJ) proteins ZO-1, ZO-2, occludin, and claudin-3. In contrast, claudin-11 was barely altered. MET also modified the shape of claudin-11, and ZO-2 at the cell border, from a zigzag to a more linear pattern. In addition, MET diminished the expression of ZO-2 in spermatids present in seminiferous tubules, induced the phosphorylation of ZO-2 and occludin in testes and reduced the interaction between these proteins assessed by co-immunoprecipitation. MET formed covalent bonds with ZO-2 in serine, tyrosine and lysine residues. The covalent modifications formed on ZO-2 at putative phosphorylation sites might interfere with ZO-2 interaction with regulatory molecules and other TJ proteins. MET bonds formed at ZO-2 ubiquitination sites likely interfere with ZO-2 degradation and TJ sealing, based on results obtained in cultured epithelial cells transfected with ZO-2 mutated at a MET target lysine residue. Our results shed light on MET male reproductive toxicity and are important to improve regulations regarding the use of OP pesticides and to protect the health of agricultural workers.

Artificial sweetener saccharin disrupts intestinal epithelial cells' barrier function in vitro.

SCOPE: Consumption of non-nutritive sweeteners (NNS) is a dietary practice used by those who wish to lose weight or by patients on a sugar-restricted diet such as those with DM2. Although these substances are safe, possible biological interactions with the digestive tract, particularly in relation to intestinal permeability, have not been studied. Thus, the current work sought to investigate the action of different NNS on intestinal permeability using an in vitro Caco-2 cell model. METHODS AND RESULTS: Caco-2 cells were incubated with acesulfame K, aspartame, saccharin, or sucralose at equimolar concentrations. Acesulfame K, aspartame, and sucralose did not disrupt monolayer integrity in the cells. However, saccharin increased paracellular permeability and decreased transepithelial electrical resistance (TEER) via a non-cytotoxic mechanism. The levels of the tight junction protein claudin-1 were reduced in Caco-2 cells that had previously been exposed to saccharin. The inhibition of nuclear factor-κB (NF-κB) was able to prevent the reduction in TEER induced by saccharin treatment. Thalidomide, as an inhibitor of ubiquitin ligase, was able to prevent the decrease in claudin-1 protein expression and the TEER reduction in Caco-2 cells. CONCLUSIONS: Saccharin disrupts monolayer integrity and alters paracellular permeability in a Caco-2 cell monolayer model, via a mechanism involving NF-κB activation, resulting in the ubiquitination of the tight junction protein claudin-1. Saccharin consumption may potentially alter the intestinal integrity in humans.

Dietas altas en grasas y sus efectos sobre el intestino / Diets high in fats and their effects on the intestine

El objetivo de esta revisión fue exponer el conocimiento actual sobre la relación existente entre dietas altas en grasa (DAG), alteraciones morfológicas de la mucosa intestinal, efectos inflamatorios y cáncer intestinal. Las DAG inicialmente producen aumento de la microbiota patógena, lo que reduce la cantidad y calidad de la secreción de los exocrinocitos caliciformes, disminuyendo la efectividad de la barrera intestinal. Las bacterias y sus lipopolisacaridos (LPS) promueven la secreción de citoquinas proinflamatorias activando vías de inflamación, que a su vez afectan la integridad de las uniones intercelulares alterando la barrera intestinal. Lo anterior, permite que los LPS ingresen a la lámina propia y circulación sanguínea produciendo inflamación local y sistémica. Así mismo, las DAG generan efectos nocivos en la morfología y función de la mucosa gastrointestinal lo que podría favorecer el desarrollo de cáncer. Lo anterior, podría deberse a que el consumo de DAG es capaz de aumentar la proliferación de células de la mucosa y el número y proliferación de células madres tumorales en el intestino.

The aim of this review was to present current knowledge about the relationship between high fat diets (HFD), morphological alterations of intestinal mucosa, inflammatory effects and intestinal cancer. The HFD initially produces an increase in the pathogenic microbiota, which reduces quantity and quality of secretion of goblet cells, decreasing the effectiveness of intestinal barrier. Bacteria and their lipopolysaccharides (LPS) stimulate the secretion of proinflammatory cytokines by activating inflammation pathways, which in turn affect the integrity of intercellular junctions by changing intestinal barrier. The above allows the LPS enter to lamina propria and blood circulation producing local and systemic inflammation. Likewise, HFD generate deleterious effects on morphology and function of gastrointestinal mucosa, which could favor the development of cancer. This could be due to the fact that consumption of HFD is capable of increasing proliferation of mucosal cells and number and proliferation of tumor stem cells in the intestine.

Acute and chronic exposure to high levels of glucose modulates tight junction-associated epithelial barrier function in a renal tubular cell line.

AIMS: Type 2 diabetes mellitus (T2DM) is one of the most prevalent diseases worldwide. Diabetic nephropathy (DN) is a complication of diabetes and the mechanisms underlying onset and progression of this disease are not fully understood. It has been shown that hyperglycemia is an independent factor to predict the development of DN in individuals with T2DM, however, a link between high plasma glucose levels and renal tubular injuries in DN remains unknown. In this study, we investigated the effect of high levels of glucose (i.e. 180 or 360mg/dL) for up to 24h (acute) or over 72h (chronic) upon tight junction (TJ)-mediated epithelial barrier integrity of the kidney tubular cell line, MDCK. METHODS/KEY FINDINGS: High levels of glucose (180 and 360mg/dL) induced a decrease in transepithelial electrical resistance associated with an increase in TJ cation selectivity at 24h or in TJ permeability to a paracellular marker, Lucifer Yellow, at 72h-exposure when compared to control group (exposed to 100mg/dL glucose). Immunofluorescence analyses showed that glucose treatment induced a significant decrease in the tight junctional content of claudins-1 and -3 as well as a significant increase in claudin-2 (particularly at 24h-exposure) and a time-dependent change in occludin/ZO-1 junctional content. The analyses of total cell content of these junctional proteins by Western blot did not reveal significant changes, except in claudin-2 expression. SIGNIFICANCE: Our data suggest that high levels of glucose induce time-dependence changes in TJ structure in MDCK monolayers, suggesting a possible link between hyperglycemia-induced tubular epithelial barrier disruption and diabetic nephropathy.

Entamoeba histolytica EhCP112 Dislocates and Degrades Claudin-1 and Claudin-2 at Tight Junctions of the Intestinal Epithelium.

During intestinal invasion, Entamoeba histolytica opens tight junctions (TJs) reflected by transepithelial electrical resistance (TEER) dropping. To explore the molecular mechanisms underlying this, we studied in vitro and in vivo the damage produced by the recombinant E. histolytica cysteine protease (rEhCP112) on TJ functions and proteins. rEhCP112 reduced TEER in Caco-2 cells in a dose- and time-dependent manner; and EhCP112-overexpressing trophozoites provoked major epithelial injury compared to control trophozoites. rEhCP112 penetrated through the intercellular space, and consequently the ion flux increased and the TJs fence function was disturbed. However, macromolecular flux was not altered. Functional in vitro assays revealed specific association of rEhCP112 with claudin-1 and claudin-2, that are both involved in regulating ion flux and fence function. Of note, rEhCP112 did not interact with occludin that is responsible for regulating macromolecular flux. Moreover, rEhCP112 degraded and delocalized claudin-1, thus affecting interepithelial adhesion. Concomitantly, expression of the leaky claudin-2 at TJ, first increased and then it was degraded. In vivo, rEhCP112 increased intestinal epithelial permeability in the mouse colon, likely due to apical erosion and claudin-1 and claudin-2 degradation. In conclusion, we provide evidence that EhCP112 causes epithelial dysfunction by specifically altering claudins at TJ. Thus, EhCP112 could be a potential target for therapeutic approaches against amoebiasis.

Combustion-derived nanoparticles, the neuroenteric system, cervical vagus, hyperphosphorylated alpha synuclein and tau in young Mexico City residents.

Mexico City (MC) young residents are exposed to high levels of fine particulate matter (PM2.5), have high frontal concentrations of combustion-derived nanoparticles (CDNPs), accumulation of hyperphosphorylated aggregated α-synuclein (α-Syn) and early Parkinson's disease (PD). Swallowed CDNPs have easy access to epithelium and submucosa, damaging gastrointestinal (GI) barrier integrity and accessing the enteric nervous system (ENS). This study is focused on the ENS, vagus nerves and GI barrier in young MC v clean air controls. Electron microscopy of epithelial, endothelial and neural cells and immunoreactivity of stomach and vagus to phosphorylated ɑ-synuclein Ser129 and Hyperphosphorylated-Tau (Htau) were evaluated and CDNPs measured in ENS. CDNPs were abundant in erythrocytes, unmyelinated submucosal, perivascular and intramuscular nerve fibers, ganglionic neurons and vagus nerves and associated with organelle pathology. ɑSyn and Htau were present in 25/27 MC gastric,15/26 vagus and 18/27 gastric and 2/26 vagus samples respectively. We strongly suggest CDNPs are penetrating and damaging the GI barrier and reaching preganglionic parasympathetic fibers and the vagus nerve. This work highlights the potential role of CDNPs in the neuroenteric hyperphosphorylated ɑ-Syn and tau pathology as seen in Parkinson and Alzheimer's diseases. Highly oxidative, ubiquitous CDNPs constitute a biologically plausible path into Parkinson's and Alzheimer's pathogenesis.

Butyrate reduces high-fat diet-induced metabolic alterations, hepatic steatosis and pancreatic beta cell and intestinal barrier dysfunctions in prediabetic mice.

In this study, we investigated the effect of diet supplementation with sodium butyrate (5% w/w), a short-chain fatty acid produced by the intestinal microbiota, on metabolic parameters, body adiposity, hepatic and pancreatic lipid accumulation, beta cell function/mass as well as on the structure and function of the tight junction-mediated intestinal epithelial barrier in both normal and obese/prediabetic C57 mice fed a regular (control) or high-fat diet for 60 days, respectively. Butyrate treatment significantly inhibited all the high-fat-induced metabolic dysfunctions evaluated, i.e. significantly reduced the weight gain and body adiposity as well as the insulin resistant state, hyperglycemia and hyperinsulinemia, without changing food intake. In addition, high-fat-fed mice treated with this short-chain fatty acid displayed no compensatory hyperplasia of pancreatic beta cells nor marked hepatic steatosis as seen in prediabetic mice after high-fat diet only. Isolated pancreatic islets from high-fat-fed mice treated with butyrate showed improvement of the insulin secretion, which was associated with a significant decrease in lipid accumulation within the pancreas. Butyrate enhanced the intestinal epithelial barrier, as revealed by the FITC-Dextran permeability assay, which was accompanied by a significant increase in the junctional content of the tight junction-associated claudin-1 in intestinal epithelia of jejunum, ileum, and colon of both control and high-fat mice. In conclusion, our results showed that diet supplementation with butyrate inhibits the deleterious effects of high-fat diet intake on metabolic parameters and structure/function of several tissues/organs associated with type 2 diabetes mellitus in a mouse model, suggesting a potential use of this short-chain fatty acid in the treatment of this endocrine-metabolic disorder. Impact statement Butyrate is a short-chain fatty acid produced by the intestinal microbiota through the fermentation of non-absorbable carbohydrates and proteins (e.g. fibers). Sodium butyrate incorporated into the diet displayed a protective action on metabolic, hepatic, pancreatic and intestinal alterations induced by high-fat diet in mice, resulting in significant inhibition of the development of a prediabetic state. Thus, our data suggest that butyrate may have a potential therapeutic use in the treatment of type 2 diabetes and related disorders.

Exendin-4 Reverses Biochemical and Functional Alterations in the Blood-Brain and Blood-CSF Barriers in Diabetic Rats.

Diabetes mellitus (DM) is a metabolic disorder associated with micro- and macrovascular alterations that contribute to the cognitive impairment observed in diabetic patients. Signs of breakdown of the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB) have been found in patients and animal models of DM. Breakdown of the BBB and BCSFB can lead to disruptions in cerebral homeostasis and eventually neural dysfunction and degeneration. However, our understanding of the biochemistry underlying barrier protein modifications is incomplete. Herein, we evaluated changes in the levels of specific proteins in the BBB (occludin, claudin-5, ZO-1, and aquaporin-4) and BCSFB (claudin-2 and aquaporin-1) in the hippocampus of diabetic rats, and we also investigated the functional alterations in these barriers. In addition, we evaluated the ability of exendin-4 (EX-4), a glucagon-like peptide-1 agonist that can cross the BBB to reverse the functional and biochemical modifications observed in these animals. We observed a decrease in BBB proteins (except ZO-1) in diabetic rats, whereas the EX-4 treatment recovered the occludin and aquaporin-4 levels. Similarly, we observed a decrease in BCSFB proteins in diabetic rats, whereas EX-4 reversed such changes. EX-4 also reversed alterations in the permeability of the BBB and BCSFB in diabetic rats. Additionally, altered cognitive parameters in diabetic rats were improved by EX-4. These data further our understanding of the alterations in the central nervous system caused by DM, particularly changes in the proteins and permeability of the brain barriers, as well as cognitive dysfunction. Furthermore, these data suggest a role for EX-4 in therapeutic strategies for cognitive dysfunction in DM.

Experimental Colitis Is Attenuated by Cardioprotective Diet Supplementation That Reduces Oxidative Stress, Inflammation, and Mucosal Damage.

Inflammatory bowel diseases (IBD) such as ulcerative colitis (UC) and Crohn's disease (CD) are multifactorial, relapsing disorders of the gastrointestinal tract. However, the etiology is still poorly understood but involves altered immune responses, epithelial dysfunction, environmental factors, and nutrition. Recently, we have shown that the diet supplement corabion has cardioprotective effects due to reduction of oxidative stress and inflammation. Since oxidative stress and inflammation are also prominent risk factors in IBD, we speculated that corabion also has beneficial effects on experimental colitis. Colitis was induced in male mice by administration of 3.5% (w/v) dextran sulfate sodium (DSS) in drinking water for a period of 3 or 7 days with or without daily gavage feeding of corabion consisting of vitamin C, vitamin E, L-arginine, and eicosapentaenoic and docosahexaenoic acid. We found that corabion administration attenuated DSS-induced colon shortening, tissue damage, and disease activity index during the onset of colitis. Mechanistically, these effects could be explained by reduced neutrophil recruitment, oxidative stress, production of proinflammatory cytokines, and internalization of the junctional proteins ZO-1 and E-cadherin leading to less edema formation. Thus, corabion may be a useful diet supplement for the management of chronic inflammatory intestinal disorders such as IBD.

Polyglucosan Molecules Induce Mitochondrial Impairment and Apoptosis in Germ Cells Without Affecting the Integrity and Functionality of Sertoli Cells.

Glycogen is the main storage form of glucose; however, the accumulation of glycogen-like glucose polymers can lead to degeneration and cellular death. Previously, we reported that the accumulation of glycogen in testis of transgenic animals overexpressing a constitutively active form of glycogen synthase enhances the apoptosis of pre-meiotic male germ cells and a complete disorganization of the seminiferous tubules. Here we sought to further identify the effects of glycogen storage in cells from the seminiferous tubules and the mechanism behind the pro-apoptotic activity induced by its accumulation. Using an in vitro culture of Sertoli cells (line 42GPA9) and spermatocyte-like cells (line GC-1) expressing a superactive form of glycogen synthase or the Protein Targeting to Glycogen (PTG), we found that glycogen synthesized in both cell lines is poorly branched. In addition, the immunodetection of key molecules of apoptotic events suggests that cellular death induced by polyglucosan molecules affects GC-1 cells, but not 42GPA9 cells by mitochondrial impairment and activation of an intrinsic apoptotic pathway. Furthermore, we analyzed the effects of glycogen deposition during the establishment of an in vitro blood-testis barrier. The results using a non-permeable fluorescent molecule showed that, in conditions of over-synthesis of glycogen, 42GPA9 cells do not lose their capacity to generate an impermeable barrier and the levels of connexin43, occludin, and ZO1 proteins were not affected. These results suggest that the accumulation of polyglucosan molecules has a selective effect-triggered by the intrinsic activation of the apoptotic pathway-in germ cells without directly affecting Sertoli cells. J. Cell. Physiol. 231: 2142-2152, 2016. © 2016 Wiley Periodicals, Inc.

Inhibition of angiopoietin-1 (ANGPT1) affects vascular integrity in ovarian hyperstimulation syndrome (OHSS).

Ovarian hyperstimulation syndrome (OHSS) is a complication of ovarian stimulation with gonadotrophins following human chorionic gonadotrophin (hCG) administration. The relationship between hCG and OHSS is partly mediated via the production of angiogenic factors, such as vascular endothelial growth factor A (VEGFA) and angiopoietins (ANGPTs). Here, we investigated the effect of ANGPT1 inhibition on ovarian angiogenesis in follicular fluid (FF) from women at risk of OHSS, using the chorioallantoic membrane (CAM) of quail embryos as an experimental model. We also analysed cytoskeletal changes and endothelial junction protein expression induced by this FF in the presence or absence of an ANGPT1-neutralising antibody in endothelial cell cultures. The presence of this antibody restored the number of vascular branch points and integrin αvß3 levels in the CAMs to control values. ANGPT1 inhibition in FF from OHSS patients also restored the levels of claudin-5, vascular endothelial cadherin and phosphorylated ß-catenin and partially reversed actin redistribution in endothelial cells. Our findings suggest that ANGPT1 increases pathophysiological angiogenesis in patients at risk of OHSS by acting on tight and adherens junction proteins. Elucidating the mechanisms by which ANGPT1 regulates vascular development and cell-cell junctions in OHSS will contribute to identifying new therapeutic targets for the treatment of human diseases with aberrant vascular leakage.