Exosomes derived from mesenchymal stem cells in diabetes and diabetic complications.

Diabetes, a group of metabolic disorders, constitutes an important global health problem. Diabetes and its complications place a heavy financial strain on both patients and the global healthcare establishment. The lack of effective treatments contributes to this pessimistic situation and negative outlook. Exosomes released from mesenchymal stromal cells (MSCs) have emerged as the most likely new breakthrough and advancement in treating of diabetes and diabetes-associated complication due to its capacity of intercellular communication, modulating the local microenvironment, and regulating cellular processes. In the present review, we briefly outlined the properties of MSCs-derived exosomes, provided a thorough summary of their biological functions and potential uses in diabetes and its related complications.

Triclocarban and triclosan promote breast cancer progression in vitro and in vivo via activating G protein-coupled estrogen receptor signaling pathways.

Triclocarban (TCC) and triclosan (TCS) have been detected ubiquitously in human body and evoked increasing concerns. This study aimed to reveal the induction risks of TCC and TCS on triple negative breast cancer through non-genomic GPER-mediated signaling pathways. Molecular simulation indicated that TCC exhibited higher GPER binding affinity than TCS theoretically. Calcium mobilization assay displayed that TCC/TCS activated GPER signaling pathway with the lowest observed effective concentrations (LOEC) of 10 nM/100 nM. TCC and TCS also upregulated MMP-2/9, EGFR, MAPK3 but downregulated MAPK8 via GPER-mediated signaling pathway. Proliferation assay showed that TCC/TCS induced 4 T1 breast cancer cells proliferation with the LOEC of 100 nM/1000 nM. Wound-healing and transwell assays showed that TCC/TCS promoted 4 T1 cells migration in a concentration-dependent manner with the LOEC of 10 nM. The effects of TCC on breast cancer cells proliferation and migration were stronger than TCS and both were regulated by GPER. TCC/TCS induced migratory effects were more significantly than proliferative effect. Mechanism study showed that TCC/TCS downregulated the expression of epithelial marker (E-cadherin) but upregulated mesenchymal markers (snail and N-cadherin), which was reversed by GPER inhibitor G15. These biomarkers results indicated that TCC/TCS-induced 4 T1 cells migration was a classic epithelial to mesenchymal transition mechanism regulated by GPER signaling pathway. Orthotopic tumor model verified that TCC promoted breast cancer in-situ tumor growth and distal tissue metastasis via GPER-mediated signaling pathway at human-exposure level of 10 mg/kg/d. TCC-induced tissue metastasis of breast cancer was more significantly than in-situ tumor growth. Overall, we demonstrated for the first time that TCC/TCS could activate the GPER signaling pathways to induce breast cancer progression.

Triclocarban and triclosan exacerbate high-fat diet-induced hepatic lipid accumulation at environmental related levels: The potential roles of estrogen-related receptors pathways.

Triclosan (TCS) and triclocarban (TCC) have become ubiquitous pollutants detected in human body with concentrations up to hundreds of nanomolar levels. Previous studies about the hepatic lipid accumulation induced by TCS and TCC were focused on pollutant itself, which showed weak or no effects. High-fat diet (HFD), as a known environmental factor contributing to lipid metabolism-related disorders, its synergistic action with environmental pollutants deserves concern. The present study aimed to demonstrate the combined effects and potential molecular mechanisms of TCS and TCC with HFD at cellular and animal levels. The in vitro studies showed that TCC and TCS alone had negligible impact on lipid accumulation in HepG2 cells but induced lipid deposition at nanomolar levels when co-exposure with fatty acid. TCC exhibited much higher induction effects than TCS, which was related to their differential regulatory roles in adipogenic-related genes expression. The in vivo studies showed that TCC had little influence on hepatic lipid accumulation in mice fed with normal diet (ND) but could exacerbate the lipid accumulation in mice fed with HFD. Meanwhile, TCC-induced dyslipidemia in mice fed with HFD was more significant than that fed with ND. Therefore, we speculated that TCC might increase the risk of nonalcoholic fatty liver disease (NAFLD) and atherosclerosis in HFD humans. Molecular mechanism studies showed that TCC and TCS could bind to and activate estrogen-related receptor α (ERRα) and ERRγ as well as regulate their expression. TCC had higher activity on ERRα and ERRγ than TCS, which explained partly the differential regulatory roles of two receptors in the lipid accumulation induced by TCC and TCS. This work revealed synergistic effects and molecular mechanisms of TCC and TCS with excessive fatty acid on the hepatic lipid metabolism, which provided a novel insight into the toxic mechanism of pollutants from the perspective of dietary habits.

Neonicotinoid insecticides promote breast cancer progression via G protein-coupled estrogen receptor: In vivo, in vitro and in silico studies.

Neonicotinoid insecticides (NIs) have been widely detected in environmental media and human body with concentrations reaching hundreds of nanomolar to micromolar levels. However, the information about their human health toxicology and mechanism is deficient. Previous studies have implied that NIs might exert estrogenic disruption and promote breast cancer progression, but the molecular mechanism is unclear, especially the molecular initiating event. G protein-coupled estrogen receptor (GPER), as a candidate therapeutic target, plays vital roles in the development of breast cancer. This work aimed to reveal the potential mechanism through GPER pathway. Firstly, we screened the activities of seven most common NIs on GPER signal pathway by calcium mobilization assay. Clothianidin, acetamiprid (ACE), and dinotefuran activated GPER most potently and ACE displayed the highest agonistic activity with the lowest observed effective concentration (LOEC) of 1 µM. The molecular docking and dynamics simulation showed favored interaction trend between the NIs and GPER. The three NIs with GPER activity induced 4T1 breast cancer cells migration and ACE showed the highest potency with LOEC of 100 nM. ACE also induced 4T1 cells proliferation at high concentration of 50 µM and up-regulated GPER expression in a dose-dependent manner. We speculated that both the induction effects of ACE on 4T1 cells proliferation and migration might be owing to the activation and up-regulation of GPER. By using 4T1-Luc cells injected orthotopic tumor model, we found that ACE also promoted in-situ breast cancer growth and lung metastasis in normal mouse dependent on GPER. However, ACE only promoted in-situ breast cancer growth through GPER but not lung metastasis in ovariectomized mice, implying that the ACE-induced lung metastasis should be related to endogenous estrogen from ovary. Overall, we demonstrated that NIs promoted breast cancer progression via GPER pathway at human related exposure levels and their female health risks need urgent concerns.

Benzotriazole Ultraviolet Stabilizers Promote Breast Cancer Cell Proliferation via Activating Estrogen-Related Receptors α and γ at Human-Relevant Levels.

Benzotriazole ultraviolet stabilizers (BUVSs) are ubiquitous emerging pollutants that have been reported to show estrogenic disruption effects through interaction with the classic estrogen receptors (ERs) in the fashion of low activity. The present study aims at revealing the potential disruption mechanism via estrogen-related receptors α and γ (ERRα and ERRγ) pathways. By the competitive binding assay, we first found that BUVSs bond to ERRγ ligand binding domain (ERRγ-LBD) with Kd ranging from 0.66 to 19.27 µM. According to the results of reporter gene assays, the transcriptional activities of ERRα and ERRγ were promoted by most tested BUVSs with the lowest observed effective concentrations (LOEC) from 10 to 100 nM, which are in the range of human exposure levels. At 1 µM, most tested BUVSs showed higher agonistic activity toward ERRγ than ERRα. The most effective two BUVSs promoted the MCF-7 proliferation dependent on ERRα and ERRγ with a LOEC of 100 nM. The molecular dynamics simulation showed that most studied BUVSs had lower binding free energy with ERRγ than with ERRα. The structure-activity relationship analysis revealed that molecular polarizability, electron-donating ability, ionization potential, and softness were the main structural factors impacting the binding of BUVSs with ERRγ. Overall, our results provide novel insights into the estrogenic disruption effects of BUVSs.

Effects of supplementing sow diets during late gestation with Pennisetum purpureum on antioxidant indices, immune parameters and faecal microbiota.

The purpose of this study was to investigate the effects of adding Pennisetum purpureum (P. purpureum, also known as Napier grass or elephant grass) to the diets of late gestation on the antioxidant indexes, immune indexes and faecal microbiota of sows. At the 90 days of gestation, 300 healthy sows were randomly divided into three groups, and they received the basic commercial diet or added 5% P. purpureum and 10% P. purpureum, respectively. The experiment started from 90 days of gestation to parturition. The results showed that the total antioxidant capacity, immunoglobulins and serum equol concentrations of sows on 100 days of gestation and at parturition increased linearly (p < .05) with the increase of the content of P. purpureum in the gestation diet. The 5% P. purpureum increased the relative abundance of Bacteroidetes (p = .027) and Actinobacteria (p < .001) at phylum level, Coriobacteriaceae (p < .001) at family level and Prevotellaceae_UCG_001 (p = .004) at genus level, and decreased the relative abundance of Escherichia_Shigella (p < .001) at genus level. In summary, this study shows that the additive of P. purpureum can increase the concentration of serum equol, improve the antioxidant capacity and immune function of sow in late gestation. In addition, the additive of 5% P. purpureum in the diet might change the composition of intestinal microbiota of sows, particularly the relative abundance of Coriobacteriaceae (p < .001) increased.

Effect of glucose, soya oil and glutamine on protein expression and mammalian target of rapamycin complex 1 pathway of jejunal crypt enterocytes in weaned piglets.

The present study was conducted to evaluate the effects of glucose, soya oil or glutamine on jejunal morphology, protein metabolism and protein expression of the mammalian target of rapamycin complex 1 (mTORC1) signalling pathway in jejunal villus or crypt compartment of piglets. Forty-two 21 d-weaned piglets were randomly allotted to one of the three isoenergetic diets formulated with glucose, soya oil or glutamine for 28 d. On day 14 or 28, the proteins in crypt enterocytes were analysed with isobaric tags for relative and absolute quantification and proteins involved in mTORC1 signalling pathway in villus or crypt compartment cells were determined by Western blotting. Our results showed no significant differences (P > 0·05) in jejunal morphology among the three treatments on day 14 or 28. The differentially expressed proteins mainly took part in a few network pathways, including antimicrobial or inflammatory response, cell death and survival, digestive system development and function and carbohydrate metabolism. On day 14 or 28, there were higher protein expression of eukaryotic initiation factor-4E binding protein-1 in jejunal crypt compartment of piglets supplemented with glucose or glutamine compared with soya oil. On day 28, higher protein expression of phosphor-mTOR in crypt compartment was observed in piglets supplemented with glucose compared with the soya oil. In conclusion, the isoenergetic glucose, soya oil or glutamine did not affect the jejunal morphology of piglets; however, they had different effects on the protein metabolism in crypt compartment. Compared with soya oil, glucose or glutamine may be better energy supplies for enterocytes in jejunal crypt compartment.

The effects of dietary reduced mineral elements and coated cysteamine supplementation on bacterial diversity in the ileum of finishing pigs.

This study was conducted to investigate the effects of different levels of dietary partial MEs and coated cysteamine (CC) supplementation on gut microbiota in finishing pigs. Results showed that whittling down dietary partial MEs (Cu, Fe, Zn, Mn) by 20% and 40% had little effect on the microbial diversity, community structure, and bacterial relative abundance in the ileum of finishing pigs. Supplementation with 1,600 mg/kg CC also had no obvious effect on the microbial diversity, community structure, and bacterial relative abundance in the finishing pig ileum when fed diets with a normal MEs level. However, the abundance of Peptostreptococcaceae, Pasteurella, and Pasteurella_aerogenes was higher, and the abundance of Actinobacillus_minor was lower in the 20% ME reduction diet treatment than that in the 20% ME reduction with 1,600 mg/kg CC diet group (p < 0.05). In conclusion, our results suggested that there is no obvious effect on gut microbiota when dietary partial MEs are reduced by 20% or 40%, which indicates the feasibility of reducing dietary partial MEs by 20% or 40% in finishing pigs. Supplementation with CC changed the relative abundance of some bacteria related to opportunistic pathogenicity in the finishing pig ileum when were fed a 20% ME reduction diet.

Mini-gut: a promising model for drug development.

Until recently, major advances in drug development have been hampered by a lack of proper cell and tissue models; but the introduction of organoid technology has revolutionized this field. At the level of the gastrointestinal tract, the so-called mini-gut comprises all major cell types of native intestine and recapitulates the composition and function of native intestinal epithelium. The mini-gut can be classified as an intestinal organoid (IO), derived from pluripotent stem cells, or as an enteroid, consisting only of epithelial cells and generated from adult stem cells. Both classifications have been used as models to develop drugs against cystic fibrosis, cancer and infectious disease, as well as for drug screening, personalized medicine and the development of new medical tools. In this review, we highlight and discuss the importance of mini-guts for drug development and point out their limitations and future prospects.

Enteroids: Promising in Vitro Models for Studies of Intestinal Physiology and Nutrition in Farm Animals.

The lack of sophisticated in vitro models limits our current understanding of gastrointestinal functions in farm animals. Conventional 2D cell lines or primary cells fail to recapitulate the physiology of in vivo intestinal epithelium. In contrast stem cell-derived, nontransformed 3D enteroids partially recreate the villus-crypt anatomy of the native intestine and comprise most if not all intestinal cell types including enterocytes, enteroendocrine cells, goblet cells, Paneth cells, and stem cells. This review summarizes the techniques used for generating and culturing enteroids of various farm animal species, focuses on important factors influencing the longevity of enteroids, and provides an overview of their current applications in modeling veterinary pathogens and in developing chemicals and bioactives for treating animal disease and improving production performance. It also mentions current limitations of enteroid models and potential solutions and highlights the opportunities for using these enteroids as a platform in studies regarding veterinary sciences and animal nutrition.

Effects of yeast products on the intestinal morphology, barrier function, cytokine expression, and antioxidant system of weaned piglets.

The goal of this study was to evaluate the effects of a mixture of yeast culture, cell wall hydrolysates, and yeast extracts (collectively "yeast products," YP) on the performance, intestinal physiology, and health of weaned piglets. A total of 90 piglets weaned at 21 d of age were blocked by body weight, sex, and litter and randomly assigned to one of three treatments for a 14-d feeding experiment, including (1) a basal diet (control), (2) 1.2 g/kg of YP, and (3) 20 mg/kg of colistin sulfate (CSE). No statistically significant differences were observed in average daily feed intake, average daily weight gain, or gain-to-feed ratio among CSE, YP, and control piglets. Increased prevalence of diarrhea was observed among piglets fed the YP diet, whereas diarrhea was less prevalent among those fed CSE. Duodenal and jejunal villus height and duodenal crypt depth were greater in the control group than they were in the YP or CSE groups. Intraepithelial lymphocytes (IEL) in the duodenal and jejunal villi were enhanced by YP, whereas IEL in the ileal villi were reduced in weaned piglets fed YP. Secretion of jejunal and ileal interleukin-10 (IL-10) was higher and intestinal and serum antioxidant indexes were affected by YP and CSE. In YP- and CSE-supplemented animals, serum D-lactate concentration and diamine oxidase (DAO) activity were both increased, and intestinal mRNA expressions of occludin and ZO-1 were reduced as compared to the control animals. In conclusion, YP supplementation in the diets of weaned piglets appears to increase the incidence of diarrhea and has adverse effects on intestinal morphology and barrier function.

Dietary supplementation with soybean oligosaccharides increases short-chain fatty acids but decreases protein-derived catabolites in the intestinal luminal content of weaned Huanjiang mini-piglets.

The improvement of gut health and function with prebiotic supplements after weaning is an active area of research in pig nutrition. The present study was conducted to test the working hypothesis that medium-term dietary supplementation with soybean oligosaccharides (SBOS) can affect the gut ecosystem in terms of microbiota composition, luminal bacterial short-chain fatty acid and ammonia concentrations, and intestinal expression of genes related to intestinal immunity and barrier function. Ten Huanjiang mini-piglets, weaned at 21 days of age, were randomly assigned to 2 groups. Each group received a standard diet containing either dietary supplementation with 0.5% corn starch (control group) or 0.5% SBOS (experimental group). The results showed that dietary supplementation with SBOS increased the diversity of intestinal microflora and elevated (P < .05) the numbers of some presumably beneficial intestinal bacteria (e.g., Bifidobacterium sp, Faecalibacterium prausnitzii, Fusobacterium prausnitzii, and Roseburia). Soybean oligosaccharide supplementation also increased the concentration of short-chain fatty acid in the intestinal lumen, and it reduced (P < .05) the numbers of bacteria with pathogenic potential (e.g., Escherichia coli, Clostridium, and Streptococcus) and the concentration of several protein-derived catabolites (e.g., isobutyrate, isovalerate, and ammonia). In addition, SBOS supplementation increased (P < .05) expression of zonula occludens 1 messenger RNA, and it decreased (P < .05) expression of tumor necrosis factor α, interleukin 1ß, and interleukin 8 messenger RNA in the ileum and colon. These findings suggest that SBOS supplementation modifies the intestinal ecosystem in weaned Huanjiang mini-piglets and has potentially beneficial effects on the gut.

The acute and chronic effects of monosodium L-glutamate on serum iron and total iron-binding capacity in the jugular artery and vein of pigs.

We analyzed the effects of acute and chronic oral administration of monosodium L-glutamate (MSG) on serum iron (Fe) levels and total iron-binding capacity (TIBC) in piglets. In the first experiment, 12 piglets were randomly assigned to two groups: one fed a standard diet (SD) and the other fed an SD containing MSG (10 g/kg). On day 30, serum, liver, kidney, and spleen samples were collected to determine the Fe levels. In the second experiment, six pigs were surgically fitted with a catheter in the jugular artery and vein to investigate the dynamic changes of serum Fe and TIBC. Blood samples were taken from each pig via the catheter every 30 min, for a period of 4 h. The results show that MSG increases Fe levels in the spleen (P < 0.05) and in serum obtained from the jugular artery (P < 0.01). In addition, TIBC in serum obtained from the jugular artery demonstrated an increasing trend in pigs fed the MSG diet; however, this trend was not observed in the jugular vein. In conclusion, MSG increases Fe retention by enhancing TIBC in serum.

Up-regulated expression of peroxisome proliferator-activated receptor gamma in the hypothalamic-pituitary-adrenal axis of weaned pigs after Escherichia coli lipopolysaccharide challenge.

The expression of peroxisome proliferator-activated receptor gamma (PPARgamma) was investigated in the hypothalamic-pituitary-adrenal (HPA) axis of weaned pigs after injection with 100 microg/kg bodyweight Escherichia coli lipopolysaccharide (LPS) (n=6) and control pigs injected with sterile saline (n=6). LPS increased PPARgamma mRNA and protein expression in the hypothalamus (23.8 and 3.1-fold relative to controls, respectively), pituitary gland (9.2 and 2.0-fold, respectively) and adrenal gland (3.5 and 2.3-fold, respectively) (P<0.05). LPS also induced an increase in PPARgamma immunohistochemical staining in the hypothalamus (1.3-fold), adenohypophysis (1.3-fold), adrenal cortex (1.4-fold) and adrenal medulla (1.6-fold) (P<0.05). Concurrent with up-regulated expression of PPARgamma, LPS increased the concentrations of plasma corticotrophin-releasing hormone (2.1-fold) and adrenocorticotrophin (1.4-fold) (P<0.05). LPS also induced elevations of interleukin 6 and tumour necrosis factor alpha mRNA levels in the hypothalamus (4.0 and 3.2-fold, respectively), pituitary gland (20.7 and 5.1-fold, respectively) and adrenal gland (3.9 and 3.3-fold, respectively) (P<0.05). PPARgamma may play a role in the regulation of neuroendocrine responses associated with immunological stress in pigs.

Regulation of ornithine aminotransferase gene expression and activity by all-transretinoic acid in Caco-2 intestinal epithelial cells.

Ornithine aminotransferase (OAT) is a crucial enzyme in the synthesis of citrulline and arginine from glutamine/glutamate and proline by enterocytes of the small intestine. However, a role for OAT in intestinal polyamine synthesis and cell growth is not known. All-transretinoic acid (RA), an active metabolite of vitamin A, regulates the activity of several metabolic enzymes related to OAT, including ornithine decarboxylase and arginase, which may influence the function of OAT through effects on substrate (ornithine) availability. The objective of the present study was to test the hypothesis that RA regulates OAT mRNA expression and enzymatic activity in intestinal epithelial cells. Caco-2 cells were cultured for 12-72 h in the presence of 0, 0.01 and 1 microM RA and then used for measurements of OAT mRNA levels and enzyme activity as well as ornithine and polyamines. Treatment with RA induced increases in OAT gene expression and enzymatic activity, which resulted in decreased intracellular concentrations of ornithine and polyamines (putrescine, spermidine and spermine) in a dose-dependent manner. These changes occurred concomitantly with a decrease in the total number of cells, and the increase in OAT activity was due to increased OAT mRNA expression. In cells treated with 1 microM RA, addition of 10 microM putrescine to culture medium restored both cellular levels of polyamines and cell numbers to the values for the control group (without addition of RA). We conclude that exposure of Caco-2 cells to RA induces OAT expression for increasing ornithine catabolism. This leads to a reduced availability of intracellular ornithine for polyamine synthesis, thereby decreasing cell proliferation. These novel findings indicate a functional role for OAT in regulating intestinal polyamine synthesis and growth.

Amino acids and immune function.

A deficiency of dietary protein or amino acids has long been known to impair immune function and increase the susceptibility of animals and humans to infectious disease. However, only in the past 15 years have the underlying cellular and molecular mechanisms begun to unfold. Protein malnutrition reduces concentrations of most amino acids in plasma. Findings from recent studies indicate an important role for amino acids in immune responses by regulating: (1) the activation of T lymphocytes, B lymphocytes, natural killer cells and macrophages; (2) cellular redox state, gene expression and lymphocyte proliferation; and (3) the production of antibodies, cytokines and other cytotoxic substances. Increasing evidence shows that dietary supplementation of specific amino acids to animals and humans with malnutrition and infectious disease enhances the immune status, thereby reducing morbidity and mortality. Arginine, glutamine and cysteine precursors are the best prototypes. Because of a negative impact of imbalance and antagonism among amino acids on nutrient intake and utilisation, care should be exercised in developing effective strategies of enteral or parenteral provision for maximum health benefits. Such measures should be based on knowledge about the biochemistry and physiology of amino acids, their roles in immune responses, nutritional and pathological states of individuals and expected treatment outcomes. New knowledge about the metabolism of amino acids in leucocytes is critical for the development of effective means to prevent and treat immunodeficient diseases. These nutrients hold great promise in improving health and preventing infectious diseases in animals and humans.