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Inflammatory bowel diseases (IBD) are chronic disabling conditions with a complex and multifactorial etiology, which is still not completely understood. In the last 20 years, anti-TNF-α antagonists have revolutionized the treatment of IBD, but many patients still do not respond or experience adverse events. Therefore, new biological therapies and small molecules, targeting several different pathways of gut inflammation, have been developed of which some have already been introduced in clinical practice while many others are currently investigated. Moreover, therapeutic procedures such as leukocytapheresis, fecal microbiota transplant and stem cell transplantation are currently being investigated for treating IBD. Lastly, complementary and alternative medicine has become a field of interest for gastroenterologist to reduce symptom burden in IBD patients. In this comprehensive and updated review, a novel classification of current and developing drugs is provided.

Inflammatory bowel diseases (IBD) are chronic and relapsing inflammatory conditions of the gut that include Crohn's disease and ulcerative colitis. The pathogenesis of IBD is not completely unraveled, IBD are multi-factorial diseases with reported alterations in the gut microbiota, activation of different immune cell types, changes in the vascular endothelium, and alterations in the tight junctions' structure of the colonic epithelial cells. Proteomics represents a useful tool to enhance our biological understanding and to discover biomarkers in blood and intestinal specimens. It is expected to provide reproducible and quantitative data that can support clinical assessments and help clinicians in the diagnosis and treatment of IBD. Sometimes a differential diagnosis of Crohn's disease and ulcerative colitis and the prediction of treatment response can be deducted by finding meaningful biomarkers. Although some non-invasive biomarkers have been described, none can be considered as the "gold standard" for IBD diagnosis, disease activity and therapy outcome. For these reason new studies have proposed an "IBD signature", which consists in a panel of biomarkers used to assess IBD. The above described approach characterizes "omics" and in this review we will focus on proteomics.

Background: Interleukin (IL)-33 is a member of interleukin (IL)-1 family of cytokines which has been linked to the development of inflammatory conditions and cancer in the gastrointestinal tract. This study is designed to investigate whether IL-33 has direct effect on human gastric epithelial cells (GES-1) and on human gastric adenocarcinoma cell line (AGS) and gastric carcinoma cell line (NCI-N87), assessing its role in regulation of cell proliferation and cell cycle, apoptosis and necrosis. Cell cycle regulation was also determined in ex vivo gastric cancer samples obtained during endoscopy and surgical procedures. Methods: cell lines and tissue samples underwent stimulation with rhIL-33. Proliferation was assessed by XTT and CFSE assay, we also evaluated apoptosis by Caspase 3/8 Activity assay and Annexin V assays. Cell cycle were analyzed by means of Propidium Iodine assay and gene expression regulation was assessed by RT-PCR Profiling. Results: we found that IL-33 has an antiproliferative and proapoptotic effect on cancer cell line, while it can stimulate proliferation and reduce apoptosis in normal epithelial cell line. These effects are also confirmed by the analysis of cell cycle gene expression which showed a reduced expression of proproliferative genes in cancer cells, in particular genes involved in G0/G1 and G2/M checkpoint. These results are confirmed by the gene expression analysis on surgical and bioptic specimens. Conclusions: the aforementioned results indicate that IL-33 may be involved in cell proliferation in an environment- and cell type-dependent fashion.

Background Inflammatory bowel diseases (IBD) are characterized by an increased thrombosis risk of uncertain etiology. Coagulation derangement arising from inflammation may be a triggering factor. We hypothesized that strong inflammation inhibitors (eg, anti-tumor necrosis factor-α drugs) may affect coagulation. Methods Forty patients with IBD were compared with 57 control patients for coagulation factors and endogenous thrombin potential (ETP), the latter being the most sensitive marker of in vivo pro- and anticoagulation balance. We measured ETP in the presence and absence of thrombomodulin (the physiologic protein C [PC] activator). Coagulation at different timepoints was also assessed for 28 of these patients during infliximab treatment. Results The median ETP (nM thrombin × minutes) and range (minimum-maximum) were each higher in patients at baseline than in control patients in both the absence (2120 [1611-3041] vs 1865 [1270-2337]) and the presence (1453 [464-2522] vs 831 [104-1741]) of thrombomodulin. The ETP ratio (with/without thrombomodulin) was high at baseline (0.73 [0.21-0.90] vs 0.45 [0.07-0.85]). The ETP and ETP ratio declined during treatment and were significantly lower at the end than at baseline. Factor (F) VIII and fibrinogen, which were high at baseline, decreased during treatment and at the end were significantly lower than at baseline. The FVIII/PC ratio, which was high in patients at baseline, declined during treatment and at the end was lower than at baseline. C-reactive protein recorded at the end of treatment was lower than at baseline. Conclusions Patients with IBD have a procoagulant imbalance as shown by increased ETP at baseline. The ETP decreases during treatment with infliximab, which is related to decreased FVIII and FVIII/PC ratio. This effect is also related to the improvement of inflammation as shown by decreased fibrinogen and C-reactive protein.

The Multimatrix® (MMX®) preparation MMX® is a recently obtained drug formulation developed to facilitate release of high concentrations of active drugs into the colon, with a homogeneous distribution along all colonic segments, particularly the most distal ones; the distal colonic tracts, indeed, are the most difficult to reach in significant amounts when a drug is given orally. The MMX® formulation is characterized by a lipophilic matrix dispersed in a hydrophilic structure. Indeed, in the last few years, MMX® technology has been widely used in the development of various drugs for the treatment of inflammatory and infectious gastrointestinal diseases localized in the colon. In particular, MMX® mesalamine, budesonide and parnaparin formulations have been investigated in patients with ulcerative colitis, and the first two have reached worldwide registration for the treatment of this disease. Moreover, MMX®-rifamycin is being positively tested in the treatment of colonic bacterial infections, including traveler’s diarrhea. MMX® technology is, thus, proving to be a very effective formulation for the treatment of various colonic diseases. This effectiveness has been related not only to specific colonic delivery, but also to its ability to act in a once-daily dosage, thus favouring patients’ adherence to prescribed schedules of treatment. The effective delivery of the active molecule to the site of need in the colon is also associated with very low systemic absorption and very low rates of adverse events (AEs). In this paper, we have reviewed all clinical trials performed with an MMX®-bound drug and all possible real-life reports, in order to give an overall evaluation of MMX®.

Interleukin (IL)-33 is a member of the interleukin (IL)-1 family of cytokines linked to the development of inflammatory conditions and cancer in the gastrointestinal tract. This study is designed to investigate whether IL-33 has a direct effect on human gastric epithelial cells (GES-1), the human gastric adenocarcinoma cell line (AGS), and the gastric carcinoma cell line (NCI-N87) by assessing its role in the regulation of cell proliferation, migration, cell cycle, and apoptosis. Cell cycle regulation was also determined in ex vivo gastric cancer samples obtained during endoscopy and surgical procedures. Cell lines and tissue samples underwent stimulation with rhIL-33. Proliferation was assessed by XTT and CFSE assays, migration by wound healing assay, and apoptosis by caspase 3/7 activity assay and annexin V assay. Cell cycle was analyzed by means of propidium iodine assay, and gene expression regulation was assessed by RT-PCR profiling. We found that IL-33 has an antiproliferative and proapoptotic effect on cancer cell lines, and it can stimulate proliferation and reduce apoptosis in normal epithelial cell lines. These effects were also confirmed by the analysis of cell cycle gene expression, which showed a reduced expression of pro-proliferative genes in cancer cells, particularly in genes involved in G0/G1 and G2/M checkpoints. These results were confirmed by gene expression analysis on bioptic and surgical specimens. The aforementioned results indicate that IL-33 may be involved in cell proliferation in an environment- and cell-type-dependent manner.

Biochemical and histological approaches were used to study the metabolic adaptations of transition dairy goats to dietary supplementation with saturated and unsaturated fatty acids. Twenty-three Alpine dairy goats were divided into three groups and fed a basal pre-kidding and lactation diet (C) or the same diet supplemented with fish oil (FO) or stearic acid (ST) starting 1 week before kidding until 21 days in milk (DIM). No differences were observed in milk production and composition. However, the serum non-esterified fatty acid (NEFA) and beta-hydroxybutyrate (BOHB) concentrations were changed over time by the treatments. The mean adipocyte area, measured on a subset of 12 goats, which included four subjects from each experimental group, decreased constantly in the C and ST groups from −7 to 21 days, while the FO group did not change between days 7 to 21. These results support the idea that FO is able to limit lipolysis, although the energy balance is still negative. No inflammatory processes were observed in the liver in accordance with the blood leukocytes trend, even if moderate to severe fatty changes in the liver were observed in the experimental goats. In the FO group, however, fatty infiltration appeared more severe and it occurred more gradually compared with the other diets. Overall, these results suggest an interesting ability of dietary lipid supplements to affect the fat mobilizing machinery; FO in particular seems able to reduce/delay fat mobilization and could improve hepatocyte adaptation to fatty infiltration, allowing the cells to better maintain their function.

Magnesium (Mg(2+)) is the second most copious element inside human cells and the fourth most abundant positively charged ion in the human body. It is of central importance for a broad variety of physiological processes, including intracellular signaling, neuronal excitability, muscle contraction, bone formation and enzyme activation. Its overall balance is tightly regulated by the concerted actions of the intestine, bones and kidneys. Disturbance of this balance can have serious consequences. Symptoms of hypomagnesaemia include tetany, seizures and cardiac arrhythmias, whereas hypermagnesaemia may cause cardiovascular and neuromuscular abnormalities. Drugs can interfere with Mg(2+) homoeostasis in several ways, and proton-pump inhibitors (PPIs) have been associated with hypomagnesaemia. A better understanding of the molecular mechanisms underlying the adverse effects of these medications on Mg(2+) balance will isuggest ideas for prevention and treatment, and might provide greater insight into Mg(2+) homoeostasis. This review gives an overview of the influence of PPIs on Mg(2+) homoeostasis and provides some understanding of the underlying physiological mechanisms. Moreover, we will discuss the potential link between PPI-induced changes in Mg(2+) homeostasis, and the reported cardiovascular risk observed in long-term PPI users.

Flexible biphenyls have been applied as efficient and practical chiroptical probes for absolute configuration assignment to chiral primary amines. The mechanism of the central-to-axial chirality transfer from the amine moiety to the conformationally flexible biphenyl system has been determined by NMR and computational studies. This allowed proposing a general non-empirical rule in order to establish, simply by looking at the sign of the 250 nm A band in the ECD spectrum of the biphenyl derivative, the torsion of the biphenyl and thus the absolute configuration of the amine. The method proved to be very reliable and sensitive, allowing treatment of samples on the μmol scale and permitting the simultaneous determination of the amine sample's absolute configuration and enantiopurity.

The Multimatrix® (MMX®) preparation MMX® is a recently obtained drug formulation developed to facilitate release of high concentrations of active drugs into the colon, with a homogeneous distribution along all colonic segments, particularly the most distal ones; the distal colonic tracts, indeed, are the most difficult to reach in significant amounts when a drug is given orally. The MMX® formulation is characterized by a lipophilic matrix dispersed in a hydrophilic structure. Indeed, in the last few years, MMX® technology has been widely used in the development of various drugs for the treatment of inflammatory and infectious gastrointestinal diseases localized in the colon. In particular, MMX® mesalamine, budesonide and parnaparin formulations have been investigated in patients with ulcerative colitis, and the first two have reached worldwide registration for the treatment of this disease. Moreover, MMX®-rifamycin is being positively tested in the treatment of colonic bacterial infections, including traveler’s diarrhea. MMX® technology is, thus, proving to be a very effective formulation for the treatment of various colonic diseases. This effectiveness has been related not only to specific colonic delivery, but also to its ability to act in a once-daily dosage, thus favouring patients’ adherence to prescribed schedules of treatment. The effective delivery of the active molecule to the site of need in the colon is also associated with very low systemic absorption and very low rates of adverse events (AEs). In this paper, we have reviewed all clinical trials performed with an MMX®-bound drug and all possible real-life reports, in order to give an overall evaluation of MMX®.

Background. Anaemia is common in inflammatory bowel disease (IBD), frequently resulting from a combination of iron deficiency and of anaemia of chronic disease (ACD). ACD is characterized by macrophage iron retention induced by proinflammatory cytokines. Hepcidin is the master inducer of iron accumulation during ACD, and its production is mainly regulated by IL-6 and the novel erythroid hormone erythroferrone (ERFE). This study evaluates whether anti-TNF monoclonal antibodies therapy modurates hepcidin production and the levels of its main regulators, leading to a restoration of iron homeostasis. Methods. Sera were collected from 21 IBD patients, before each anti-TNF administration, for the first 6 weeks of therapy. Prohepcidin, erythropoietin, erythroferrone, C reactive protein, interleukin-6, iron markers, and haemoglobin levels were measured and clinical activity indexes were evaluated. Results. Serum prohepcidin, IL-6, CRP, and ferritin were significantly reduced after 6-week treatment; an increase in serum iron and total transferrin was observed. No changes in the EPO-ERFE axis were found. Remarkably, haemoglobin was significantly increased. Conclusions. Anti-TNF therapy improves iron metabolism and, subsequently, anaemia in IBD. This effect appears to be related to the modulation of the cytokine network and specifically IL-6 leading to a relevant decrease of hepcidin, a master regulator of ACD.

Background Inflammatory bowel disease (IBD) is characterized by a chronic inflammation of the gut, partly driven by defects in the expression and function of pattern recognition receptors, including the IFI16 protein. Because this protein is a target for autoantibodies and its aberrant expression was reported in colonic mucosa from active patients with ulcerative colitis, we studied its expression and specific seroresponse in patients with IBD before and after infliximab (IFX) therapy. Methods Anti-IFI16 antibodies (IgG and IgA subtypes) were measured in the sera of 74 patients with IBD: 48 patients with Crohn's disease (CD) and 26 patients with ulcerative colitis, prospectively harvested before and after IFX therapy. Anti-GP2 antibodies (both IgG and IgA subtypes) were also tested for comparison. The patient antibody statuses were qualitatively and quantitatively associated with disease phenotype and response to IFX therapy. Results Significantly higher titers of anti-IFI16 IgG were found in both CD and ulcerative colitis patients compared with healthy controls. Anti-IFI16 IgA titers were also present in patients with CD. Anti-GP2 IgG subtype titers were significantly increased in patients with CD, as were IgA subtype titers. Significant changes in anti-IFI16 IgG subtype titers were observed after IFX in patients with CD who correlated with clinical remission or response. Conclusions Our results highlight the importance of IFI16 in IBD pathogenesis showing that its de novo overexpression in the gut epithelial cells leads to a breakdown in immune tolerance and the subsequent development of specific autoantibodies. Anti-IFI16 IgG antibodies hold the potential to serve as a biomarker of response to IFX therapy.

Stem cells represent an important tool in veterinary therapeutic field such as tissue engineering. In the present study, equine amnion-derived mesenchymal stromal cells were investigated for applications in veterinary science as an alternative source to bone marrow mesenchymal stem cells and adipose stem cells. Amnion stromal cells isolation and characterization protocol is described; the in vitro cell growth rate was calculated by measuring viable cell number over 20 days. The expression of stem cell markers such as Oct-4, Nanog, Sox-2 and CD105 was assessed by retrotranscription quantitative PCR (RT-qPCR) and differentiation into adipocytes, osteocytes and chondrocytes precursors was analyzed by cytochemical staining. This study showed that amnion stromal cells expressing stem cell markers can differentiate into mesoderm lineage and may be an alternative source to mesenchymal stem cells derived from adipose tissue and bone marrow for the use in tissue repair.

In the pig ovary, the transition from primordial to primary and secondary ovarian follicles begins before birth, but antral follicles can be observed, for the first time, at approximately 60-90 d of age. At approximately the same time, secondary follicles become responsive to gonadotropins, leading to the formation of antral follicles. Placing pieces of ovarian tissue under the kidney capsule of immunodeficient (SCID) mice allows the requirements for follicular recruitment and development to be studied. The objective of this study was to investigate if primordial follicles contained in ovarian fragments isolated from newborn piglets (36 +/- 12 h old) and immediately transplanted under the kidney capsule of SCID mice, are able to become responsive to gonadotropins after 60 d (as in an unaltered animal). Ovarian fragments were transplanted under the kidney capsule of three groups of four female and four male SCID mice. The first group did not receive any hormonal treatment for 12 wk. The second group was treated from the 9th week with 1 IU of FSH/LH on alternating days for 3 wk, and the third group was treated with 5 IU Pregnant Mare Serum Ganadotropin (PMSG) 48 h before euthanasia. Primordial follicles contained in ovarian fragments isolated from newborn piglets developed only to the secondary stage. Therefore, development of gonadotropin responsiveness in ovarian fragments xenotransplanted in SCID mice was delayed compared to what occurs in the unaltered animal, and there was minimal response to exogenous gonadotropins.

An in vitro study was carried out to examine the influence of two fish-oil-derived long chain omega-3 polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on goat polymorphonuclear leukocytes (PMN). Twelve Saanen healthy goats were used as blood donors. Neutrophils were isolated from blood and incubated with increasing concentration of EPA and DHA (25, 50, 100, 200muM). Control samples were incubated in the absence of omega-3 PUFAs. Phagocytosis was evaluated by fluorescein-labeled Escherichia coli incorporation, while extracellular Reactive Oxygen Species (ROS) production was determined by cytochrome c reduction assay, which was selected among the others due to its specificity for extracellular superoxide anion release. Phagocytic activity was significantly increased by EPA (P

Microscopic colitis (MC) is a common cause of chronic diarrhea. The 2 most frequent forms of MC are collagenous colitis and lymphocytic colitis. Over the past years, the incidence and prevalence of microscopic colitis are rising and this is largely attributed to a greater awareness, and concomitantly an increasing number of diagnoses. Patients with microscopic colitis report watery, nonbloody diarrhea of chronic course, abdominal pain, weight loss, and fatigue that may impair patient's health-related quality of life. The underlying mechanisms involved in the pathogenesis of microscopic colitis remain unspecified but is probably multifactorial. Collagenous colitis and lymphocytic colitis may represent specific mucosal responses to different luminal agents in predisposed individuals, resulting in an uncontrolled immune response. Genetic predisposition, altered modulation of cytokines and miRNAs, and aberrant response to drugs seem to be involved in the development of MC. Despite the progress of knowledge, still many questions remain unsolved regarding the etiology, pathophysiology, and optimal management of MC. This review gives an update on the immunological aspects of collagenous colitis and lymphocytic colitis.

MicroRNAs (miRNAs) are a family of regulatory molecules involved in many physiological processes, including activation of cells of the immune system. This study investigated the effect of Escherichia coli lipopolysaccharide (LPS) and Staphylococcus aureus enterotoxin B (SEB) on the expression of five miRNAs involved in the inflammatory response, including miR-9, miR-125 b, miR-155, miR-146 a and miR-223, in bovine CD14(+) cells (monocytes). Incubation of monocytes with SEB induced down-regulation of miR-155, miR-223 and miR-125 b, but not the anti-inflammatory miRNA miR-146 a. Conversely, incubation with LPS upregulated both miR-155 and miR-146 a. In vitro incubation of isolated CD14(+) bovine monocytes with LPS and SEB elicited different and opposite expression of miRNAs reportedly involved in inflammatory reactions.

Most of the acute intestinal diseases are caused by foodborne pathogens with infants and elderly people being at major risk. The aim of this study was to develop a procedure to simultaneously detect 20 foodborne pathogens in complex alimentary matrices such as milk, cheese and meat. The list of targets include, among the others, Listeria spp., Salmonella spp., Shigella spp., Escherichia coli spp., Campylobacter spp., Clostridium spp. and Staphylococcus aureus. The accuracy of detection was determined by using ATCC strains as positive and negative controls. The achieved sensitivity of each of assays was 1 pg of genomic DNA, which was equivalent to ∼1 cfu. The working ranges of the TaqMan(®) Real-time PCR assays, when used quantitatively on cheese and meat samples inoculated with serial dilution of Listeria spp., Listeria monocytogenes, S. aureus, Salmonella enterica, Shigella boydii, E. coli O157:H7, Bacillus cereus, Campylobacter coli, Yersinia enterocolitica, Enterobacter sakazakii and Pseudomonas aeruginosa was 10(8) cfu/g to 10(4) cfu/g. No matrix interferences were observed.

Six new phosphite ligands were prepared and their efficiency as chiral ligands was investigated in the copper-catalyzed asymmetric conjugate addition of dimethylzinc to diethyl 3-phenylpropylidenmalonate (5), affording the addition product with ees up to 74%. Moreover, a simple and straightforward route to floral fragrances Phenoxanol, Citralis, and Nitrile Citralis in optically active form through the above cited reaction was proposed. Chirality, 2011. © 2011 Wiley-Liss, Inc.

A new family of umpolung sulfoxide reagents for the synthesis of α-benzyl alcohols, chlorides, and acetamides is developed.

Acute phase proteins (APPs) are produced mainly by the liver and their concentration is increased during the systemic inflammatory response. Expression of haptoglobin (Hp), serum amyloid A (SAA), lipopolysaccharide-binding protein (LBP) and α-1 acid glycoprotein (AGP) was determined in the mucosa of the normal bovine forestomachs and abomasum by qualitative and quantitative reverse transcriptase-PCR for mRNA and by Western blot analysis and immunohistochemistry for proteins. Although expression of SAA mRNA was evident in the forestomachs and abomasum, SAA protein was identified only in the abomasum. Expression of Hp protein was high in the forestomachs and abomasum, even though expression of Hp mRNA was negligible. The main site of expression of LBP mRNA was the omasum, whereas the highest protein expression was evident in the abomasum. AGP was expressed at low levels in the bovine forestomachs. Western blot analysis revealed a heterogeneous electrophoretic pattern for AGP, LBP and Hp, indicating that different stomach compartments produce isoforms that are different to those expressed by the liver. Expression of APPs by the bovine forestomachs and abomasum may contribute to regulation of the innate immune response against pathogens.

The in vitro effects of the ω-3 polyunsaturated fatty acids (PUFAs) eicosapentanoic acid (EPA) and docosahexanoic acid (DHA) on phagocytosis and the extracellular respiratory burst in caprine monocytes were assessed. Blood monocytes incubated with increasing concentrations of EPA or DHA (25-200 μM) demonstrated increased phagocytosis compared to unexposed monocytes. Generation of reactive oxygen species (ROS) was not markedly affected in the presence of EPA and DHA, except at 200 μM, at which concentrations monocyte viability was also reduced.

Maternal effect genes produce mRNA or proteins that accumulate in the egg during oogenesis and control the developmental program until embryonic genome activation takes place. NLRP5 (NLR family, Pyrin domain containing 5), also called MATER (Maternal Antigen That Embryos Require) is one of the genes required for normal early embryonic development, although its precise function remains to be elucidated. The aim of the present study was to analyze the NLRP5 gene expression pattern and protein distribution in somatic tissues and germ cells in the pig. Reverse transcription was performed on mRNA from germinal vescicle (GV) oocytes and total RNA from spermatozoa and tissues from different organs. The transcript for NLRP5 gene was identified only in ovaries and oocytes. The presence of NLRP5 protein was detected only in ovaries by western blot analysis and immunohistochemistry.

Background Mesenchymal stem cells (MSCs) have been recently investigated for their potential use in regenerative medicine. MSCs, in particular, have great potential, as in various reports they have shown pluripotency for differentiating into many different cell types. However, the ability of MSCs to differentiate into tendon cells in vitro has not been fully investigated. Results In this study, we show that equine bone marrow mesenchymal stem cells (BM-MSCs), defined by their expression of markers such as Oct4, Sox-2 and Nanog, have the capability to differentiate in tenocytes. These differentiated cells express tendon-related markers including tenomodulin and decorin. Moreover we show that the same BM-MSCs can differentiate in osteocytes, as confirmed by alkaline phosphatase and von Kossa staining. Conclusion As MSCs represent an attractive tool for tendon tissue repair strategies, our data suggest that bone marrow should be considered the preferred MSC source for therapeutic approaches.

The present study was designed to investigate the relationship between pre-mating nutrition and the relative amounts of a panel of developmentally relevant genes in ovine oocytes and granulosa cells. Cast age ewes were fed a ration providing 0.5× (0.5 M) or 1.5× (1.5 M) live weight maintenance requirements for 2 weeks before slaughter. The ewes were synchronized and superovulated with FSH and pregnant mares serum gonadotropin. At slaughter, oocytes and granulosa cells were aspirated from follicles >2 mm in diameter and the relative abundance of 8 and 17 transcripts in oocytes and granulosa cells respectively were analyzed by semi-quantitative RT-PCR. In the oocytes, no differences between groups were observed for five transcripts (GDF9,BMP15, c-kit, glucose transporter 1 (SLC2A1), and hexokinase 1), but a lower amount of glucose transporter 3 (SLC2A3), sodium/glucose cotransporter 1 (SLC5A1), and Na+/K+ATPase mRNAs was detected in the 0.5 M group. Increased expression ofPTGS2,HAS2, and the leptin receptor long form was observed in granulosa cells from the 0.5 M group. No differences between groups were observed for the other transcripts (early growth response factor-1, estrogen receptor-α, LH and FSH receptors, gremlin 1, pentraxin 3, KIT ligand, glucose transporters 1, 3, and 8,IGF1, IGF1 receptor, leptin receptor, and tumor necrosis factor-stimulated gene 6). Expression of leptin and sodium/glucose cotransporter 1 was not detected in both groups. The present data indicate that pre-mating nutrition is associated with alteration in the mRNA content in oocytes and surrounding follicle cells in ewes, which may account for the reduced reproductive performance typical of ewes that are fed a restricted ration for a short period of time before mating.

Background and aims: Gastric cancer is still a relevant cause of death. Interleukin (IL)-1 family cytokines play key roles in promoting gastric cancer growth. IL-33, a novel member of the IL-1 family, binds to the receptor ST2 and induces prominent epithelial hyperplasia in the gut. Aim of this study is to characterize IL-33 and ST2 expression modifications throughout the progression from healthy gastric mucosa to cancer and obtain insights on the potential involvement of the IL-33/ST2 axis in regulating gastric epithelial proliferation. Methods: Biopsies from normal gastric mucosa (CONT) (N=7), Helicobacter pylori (Hp)(N=8) and non Hp-related gastritis (N=8), gastric intestinal metaplasia (IM) (N=4), gastric cancer (GC) (N=11) and normal tissue close to cancer lesions (N=11) were collected from patients undergoing upper gastrointestinal endoscopies. IL-33 and ST2 expression was evaluated by WB and qRT-PCR. AGS gastric epithelial cell were cultured with growing concentration of rIL-33 (0 ng/ml-10 ng/ml). Cell proliferation was assessed by means of XTT and CFSE assay using 0, 6 and 24h, or 4, 6 and 8 days' timepoints, respectively. Wound-healing scratch test was performed on AGS monolayers. Caspase-Glo 3/7 luminescent assay evaluated AGS apoptosis. Results: WB for IL-33 showed both full-length (30 kD) and cleaved (22 kD) forms in all the experimental groups; IL-33 mRNA transcripts were increased in non Hprelated gastritis (9.1±3.2 fold increase; p=0.04), compared to CONT (set as 1), not significantly changed in gastric cancer, whereas a trend towards reduction was detected in Hp-related gastritis and IM. WB for ST2 demonstrated both the soluble sST2 and the transmembrane ST2L variants in all the experimental groups; qRT-PCR for sST2 showed no significant changes in the different experimental groups; ST2L mRNA was more abundant in the normal tissue close to cancer lesions (34.2±4.3 fold increase, p=0.05) and in gastric cancer (9.7±4.6 fold increase, p=0.14), compared to CONT (set as 1). The XTT proliferation assay, wound healing experiments and CFSE assay showed an inhibitory effect of IL-33 on AGS in a dose-dependent way. In addition, apoptosis increased consensually to IL-33 concentrations. Conclusions: These data suggest that IL-33/ST2 axis is dysregulated during the progression from gastritis to gastric cancer and may inhibit pathologic gastric epithelial growth.

Previous studies on sheep have demonstrated effects of maternal nutrition on fetal development with effects being observed on reproductive function (Rae et al. 2002 Anim. Reprod. Sci. 72, 63–71), blood pressure (Gopalakrishnan et al. 2004 Am. J. Physiol. Regul. Integr. Comp. Physiol. 287, R12–R20), and behavior (Erhard et al. 2004 Behav. Brain Res. 151, 25–35) in the adult offspring. It is known that gene expression in developing oocytes and very early embryos is influenced by their environment and some of the effects of nutrition on reproductive and other systems are likely to be expressed through such early changes (Hyttel 2000 Anim. Reprod. Sci. 60–61, 49–60). Effects of nutrition on oocyte function (i.e. before ovulation and fertilization) and resultant embryo quality have been observed (Gonzalez-Bulnes et al. 2004 Reprod. Fertil. Dev. 16, 421–435), but effects on oocyte gene expression have not been investigated. The aim of the present study was to investigate the effects of premating nutrition on the expression of selected genes in sheep cumulus cells and oocytes. For this purpose, 16 cast age ewes of proven fertility were used. At the time of manipulation and study, all ewes had a condition score of approximately 2.5. During the 2 weeks before slaughter, ewes of one group were fed a ration providing 0.5-live weight (n = 10) maintenance requirements and the other group 1.5-live weight (n = 6) maintenence requirements. This treatment ensured that all animals had similar numbers of potentially ovulatory follicles but different patterns of ovarian follicle maturation and, accordingly, different numbers of follicles and oocytes undergoing the final stages of maturation. The ewes's reproductive cycles were synchronized using intravaginal pessaries inserted 2 weeks before slaughter. At slaughter, ovaries were recovered and oocytes and associated cumulus cells were aspirated from all follicles >2 mm in diameter. For each ovary, oocytes derived from follicles with a diameter 4 mm were pooled separately, and stored for further analysis. Associated cumulus masses were collected and stored singly for subsequent investigations. As expected from previous observations, the number of visible follicles, the ratio of small-to-large antral follicles, and the total numbers of oocytes per ewe were similar for control and underfed ewes. Furthermore, the expression levels of the markers for oocyte growth, GDF-9, BMP-15, and c-kit, were investigated. Data were analyzed by ANOVA followed by Duncan test. Results showed no major differences between groups, independent of follicle diameter. These data suggest that the nutritional status of donor ewes did not significantly affect the expression profiles of these genes. However, further analyses on a greater panel of genes is required to rule out premating nutrition effects on oocyte gene expression. Furthermore, effects on the cumulus cell gene expression profile remain to be investigated. This work was supported by the University of Milan, FIRST 2006; Scottish Executive Environment and Rural Affairs Department.