I-FABP Expression Research Articles

Experimental study on the expression and diagnostic significance of I-FABP in acute intestinal ischemia

Objective: To detect the expression and distribution of I-FABP in intestinal tissue and the changes of serum concentrations at different time of acute intestinal ischemia, and explore the significance and mechanism of I-FABP in early diagnosis of acute ischemic bowel disease. Methods: The selected 96 healthy adult SD rats were randomly divided into the experimental group and control group; 48 in each group. Each group was randomly subdivided into 6 groups with 8 rats in each group. The superior mesenteric artery was ligated in the experimental group and the peritoneal switch operation was performed in the control group. The venous blood samples were extracted from each group rats’ right ventricle at 0.5 h, 1 h, 2 h, 4 h, 8 h, 12 h after the operation and the concentration of I-FABP was tested respectively. Then the rats were killed, and the diseased intestinal tubes were cut out for paraffin sections. The I-FABP in intestinal tissue was stained by routine HE staining and direct immunofluorescence staining. Results: The I-FABP was mainly expressed in the epithelial villi of intestinal mucosa, and there was a small amount of expression in the intestinal submucosa and even the muscularis. Within 1 hour of intestinal ischemia, the number of I-FABP positive granules in the intestine and intestinal cavity increased gradually, and then gradually decreased after 1 hour. The difference has statistically significant between the experimental group and the control group (P < 0.05). The serum I-FABP: In the experimental group, the serum I-FABP concentration began to increase at 0.5 h, and reached a peak at 1 h (290. 24 ± 156.69) μg·L–1, then gradually decreased. Compared with the control group, the difference was statistically significant (P < 0.05). Conclusion: I-FABP usually mainly exists in the epithelial cells of intestinal mucosa. When acute intestinal ischemia occurs, the epithelial cells of intestinal mucosa permeability changes; I-FABP expression rapidly releases to intestinal tissue and intestinal cavity, and is absorbed into the blood. Therefore, I-FABP has certain clinical significance in early diagnosis and treatment of acute intestinal ischemia.

Protective effects of heparin on early septic shock of intestinal mucosal injury in dogs

Objective Determination of plasma markers of D-lactic acid and I-FABP and analyzed to verify the early septic shock in dogs and intestinal fatty acid binding protein expression levels of D-lactate changes and improvements to septic shock heparin early jejunal villus microcirculation in dogs and intestinal fatty acid binding protein and the expression of D-lactic acid levels, confirmed the protective effect of heparin on intestinal mucosal injury. Methods Beagles early septic shock by intravenous injection of LPS Methods. The 40 animals were randomly random number divided into control group, LPS group, the basic treatment group, heparin group. Stable after 1 hour, i. e. the point T0, to modeling LPS 1 hour, i. e. at T1, after 5 hours was observed sequential T2, T3, T4, T5, T6 point. Observed at the corresponding time points (T1, T3, T6 time points) canine blood collected by 1 500 rpm, 15 minutes after centrifugation, the plasma to the EP tube into aliquots stored at -80 ℃, measured by ELISA intestinal fatty acid binding protein (IFABP) and D-lactate (D-lac). Results heparin group IFABP (21.6±2.3) pg / mL LPS comparison group IFABP (100.48±8.42) pg / mL will be significantly decreased, with statistical significance, P 0.05, on the basis of adequate volume resuscitation on heparin therapy, I-FABP expression levels can be significantly reduced. Conclusions The presence of septic shock early intestinal mucosal microcirculation and intestinal mucosa. Under the premise of its full capacity of recovery, heparin early septic shock in dogs with intestinal mucosal microcirculation improvement, thereby protecting the intestinal mucosal barrier function. Key words: Sepsis / Septic shock; Heparin; Intestinal mucosa; Microcirculation; Lipopolysaccharides; Intestinal binding protein; D-lactic acid; Orthogonal polarization spectral imager

Commensal Streptococcus salivarius Modulates PPARγ Transcriptional Activity in Human Intestinal Epithelial Cells.

The impact of commensal bacteria in eukaryotic transcriptional regulation has increasingly been demonstrated over the last decades. A multitude of studies have shown direct effects of commensal bacteria from local transcriptional activity to systemic impact. The commensal bacterium Streptococcus salivarius is one of the early bacteria colonizing the oral and gut mucosal surfaces. It has been shown to down-regulate nuclear transcription factor (NF-кB) in human intestinal cells, a central regulator of the host mucosal immune system response to the microbiota. In order to evaluate its impact on a further important transcription factor shown to link metabolism and inflammation in the intestine, namely PPARγ (peroxisome proliferator-activated receptor), we used human intestinal epithelial cell-lines engineered to monitor PPARγ transcriptional activity in response to a wide range of S. salivarius strains. We demonstrated that different strains from this bacterial group share the property to inhibit PPARγ activation independently of the ligand used. First attempts to identify the nature of the active compounds showed that it is a low-molecular-weight, DNase-, proteases- and heat-resistant metabolite secreted by S. salivarius strains. Among PPARγ-targeted metabolic genes, I-FABP and Angptl4 expression levels were dramatically reduced in intestinal epithelial cells exposed to S. salivarius supernatant. Both gene products modulate lipid accumulation in cells and down-regulating their expression might consequently affect host health. Our study shows that species belonging to the salivarius group of streptococci impact both host inflammatory and metabolic regulation suggesting a possible role in the host homeostasis and health.

Evaluation of the expression of I-FABP and L-FABP in a necrotizing enterocolitis model after the use of Lactobacillus acidophilus

Background/purposeNecrotizing enterocolitis (NEC) is a severe intestinal disease that primarily affects premature babies, leading to high mortality and morbidity. Probiotics represent an important alternative prophylaxis for NEC but its mechanism of action is poorly understood. Moreover, intestinal and liver-type fatty acid-binding proteins (I-FABP and L-FABP) may be utilized because markers of intestinal injury, including NEC. We aimed to evaluate the protection induced by the Lactobacillus acidophilus on the intestines of newborn rats submitted to experimental NEC using I-FABP and L-FABP as biomarkers. MethodsSprague–Dawley newborn rats were divided into three groups: (1) C (control)—breast-fed; (2) NEC—subjected to NEC protocol and (3) NECP—NEC+probiotic. Morphometric, intestinal lesion, immunohistochemistry and Western blotting analysis were performed. Statistical significant differences were considered when p<0.05. ResultsIntestinal weight was lower in NEC and NECP compared to C (p<0.05). Intestinal injury was lower in NECP compared to NEC. Prophylactic probiotic recovered mucosa and muscular layers' thickness to C levels (p<0.05). I-FABP and L-FABP expressions in NECP showed intermediate values between C and NEC. ConclusionL. acidophilus had a protective effect on the development of NEC and FABPs could demonstrate the degree of tissular damage of the intestine.

L-FABP and I-FABP expression in newborn rats changes inversely in the model of necrotizing enterocolitis.

To determine the expression of hepatic L-FABP and intestinal I-FABP in an experimental model of necrotizing enterocolitis (NEC) in neonatal rats. Newborn Sprague-Dawley rats were divided into four groups: Control (C1) - exclusive breastfeeding at the first and sixth procedures (C6), NEC1 - fed formula milk and submitted to hypoxia and hypothermia at the first and sixth procedures (NEC6). The newborn pups were fed twice a day for three days, for a total of six procedures. Samples were collected for morphometric evaluation (body weight, liver weight, liver weight/body weight ratio, intestinal weight and intestinal/body weight ratio) and for immunohistochemical and Western blotting analysis. The values obtained were analyzed statistically, with the level of significance set at p<0.05. Morphometric measurements showed reduction of body and liver weights in the NEC group (p<0.05). Both immunohistochemistry and western blotting revealed that L-FABP expression in the liver was decreased and I-FABP expression in the ileum was increased in the NEC group (p<0.05). L-FABP and I-FABP expression changed inversely in the rat NEC model. These findings can contribute to a better diagnosis of NEC in human newborns.

Protective mechanism of omega-3 fish oil on mucosal barrier with severe acute panereatitis

Objective To evaluate the protective effect of omega-3 fish oil on the intestinal mucosal barrier with severe acute pancreatitis (SAP).Methods Forty Six SAP patients were enrolled (23 patients in each group) and received total parenteral nutrition (TPN) for 6 days,providing 0.8 g/(kg· d)soybean oil +0.2 g/(kg·d) ω-3 FAs based fat (FO group) or 1.0 g/(kg·d) standard soybean-oil based fat (SO group).The levels of plasma endotoxin,D-lactate and serum I-FABP expression were measured before and 7 days day after TPN.Results Forty-six patients included had significantly higher plasma level of endotoxin and D-lactate,and higher serum I-FABP concentration in the first 72 h (P ＞ 0.05).At 7th day aforementionged indexes were significantly decreased in all SAP patients.Gastrointestinal function score,plasma endotoxin and D-lactate,and serum I-FABP concentrations in FO group were significantly lower than in SO group (P ＜ 0.05,1.61 ± 0.79 vs.2.02 ± 0.68 ; 0.24 ± 0.15 vs.0.42 ± 0.11 ; 2.89 ±1.23 vs.3.95 ± 1.17 ; 77.5 ± 38.4 vs.100.5 ± 42.8).Conclusion Omega-3 fish oil supplementation reduces plasma endotoxin and D-lactate,and serum I-FABP concentrations,protects the intestinal mucosal barrier,and thereby decreases intestinal mucosal permeability in SAP. Key words: Acute pancreatitis; Omega-3 fish oil; Intestinal mucosal barrier

Novel Potential Interacting Partners of Fibronectin in Spontaneous Animal Model of Interstitial Cystitis

Feline idiopathic cystitis (FIC) is the only spontaneous animal model for human interstitial cystitis (IC), as both possess a distinctive chronical and relapsing character. Underlying pathomechanisms of both diseases are not clearly established yet. We recently detected increased urine fibronectin levels in FIC cases. The purpose of this study was to gain further insight into the pathogenesis by assessing interacting partners of fibronectin in urine of FIC affected cats. Several candidate proteins were identified via immunoprecipitation and mass spectrometry. Considerable changes in FIC conditions compared to physiological expression of co-purified proteins were detected by Western blot and immunohistochemistry. Compared to controls, complement C4a and thioredoxin were present in higher levels in urine of FIC patients whereas loss of signal intensity was detected in FIC affected tissue. Galectin-7 was exclusively detected in urine of FIC cats, pointing to an important role of this molecule in FIC pathogenesis. Moderate physiological signal intensity of galectin-7 in transitional epithelium shifted to distinct expression in transitional epithelium under pathophysiological conditions. I-FABP expression was reduced in urine and urinary bladder tissue of FIC cats. Additionally, transduction molecules of thioredoxin, NF-κB p65 and p38 MAPK, were examined. In FIC affected tissue, colocalization of thioredoxin and NF-κB p65 could be demonstrated compared to absent coexpression of thioredoxin and p38 MAPK. These considerable changes in expression level and pattern point to an important role for co-purified proteins of fibronectin and thioredoxin-regulated signal transduction pathways in FIC pathogenesis. These results could provide a promising starting point for novel therapeutic approaches in the future.

Fatty acid‐binding protein expression in the gastrointestinal tract of calves and cows

Fatty acid-binding protein (FABP) has high affinity for long-chain fatty acids and appears to participate in the metabolism and intracellular transport of lipids. Liver- and intestinal-type FABP (L-FABP and I-FABP, respectively) are expressed in the small intestine. However, in the gastrointestinal tract of ruminants, expression and localization of FABPs are unknown. In this study, we investigated the expression of I-FABP and L-FABP in the gastrointestinal tract of cattle. I- and L-FABP had higher messenger RNA (mRNA) and protein expression levels in the duodenum and jejunum relatively to other gastrointestinal regions in both calves and cows. Furthermore, L-FABP mRNA and protein expression were high in the colon. Both these protein types were confirmed to be in the cytosol of jejunal epithelial cells, where they were found in the villi rather than in the crypts. We concluded that duodenal and jejunal FABPs might be involved in the metabolism of fatty acids mainly in epithelial cells in cattle.

Molecular regulation of both dietary vitamin A and fatty acid absorption and metabolism associated with larval morphogenesis of Senegalese sole ( Solea senegalensis)

The present study aimed to deepen the understanding of molecular mechanisms governing the absorption and metabolism of some nutrients, growth and development in larvae of Senegalese sole ( Solea senegalensis) fed with Artemia enriched with Easy Selco© (ES, INVE) or Aquagrow Gold© (AGG, ABN), which mainly differed in their vitamin A (VA) content and fatty acid composition. The expression profile of genes involved in VA metabolism ( crbp2, rbp, crabp1), lipid transport ( i-fabp, l-fabp), nuclear receptors for VA and fatty acids ( rarα1, rxrα, pparβ), growth ( igf1, igf2 and their receptor igf1r) and development ( bgp) was analyzed at 22, 30 and 38 days post hatching. The main results suggested that the amount of VA absorbed by larvae is controlled at the intestinal level by crbp2 in both groups, preventing excessive accumulation of this vitamin in the target tissues. The stable expression of i-fabp in the ES group with age could cause an excessive fat accumulation in the intestine inducing, in turn, the steatosis found in the liver and vascular system of these specimens. In liver, the regulation of rbp and fabp expression reflected the status of the physiological functions demanding VA and lipids. The findings revealed that dietary composition induced different strategies for VA and lipid absorption and metabolism affecting, in turn, larval development, growth and health.

Proanthocyanidins modulate triglyceride secretion by repressing the expression of long chain acyl-CoA synthetases in Caco2 intestinal cells

In this work we determined the ability of grape seed proanthocyanidins (GSPE) to modulate triglyceride (TG) secretion by enterocytes in post-prandial and fasting states, using Caco2 cell line. GSPE did not modify TG secretion in the post-prandial state, even though it modulated the long chain acyl-CoA synthetase (ACSL) 5 and the carnitine-palmitoyl-transferase-1a gene expression. On the contrary, GSPE decreased TG secretion in the fasting state, significantly repressing ACSL3, ACSL5, I-FABP and PPARalpha gene expression. Intestinal cells can use two different pathways to carry out TG synthesis. Fatty acids (FA) delivered by ACSL3 and by ACSL5 would be directed towards the monoacylglycerol and glycerol-3-phosphate pathways, respectively. Therefore, proanthocyanidins repress the supply of FA towards the monoacylglycerol pathway in the post-prandial state, whereas they repress the supply of FAs towards both pathways in the fasting state, suggesting that the feeding state is a key factor regarding the effectiveness of proanthocyanidins to reduce triglyceridaemia.

OP011 N-3 PUFAS PREVENT THE DESTRUCTION OF INTESTINAL BARRIER AFTER ISCHEMIA/REPERFUSION INJURY BY l-FABP INDUCING THE PPARγ PATHWAY ACTIVATION

Rationale: This study was designed to investigate whether altered expression of tight junctions (TJs) proteins in intestinal membrane in ischemia/reperfusion injury was prevented by n-3 PUFAs, and the molecular mechanism underlying this beneficial effect. Methods: 24 Male SD rats were assigned to 4 groups: control group (group 1), I/R injury group (group 2), pretreated with n-3PUFAs for 7 days before I/R model was performed (group 3), pretreated with PPARg agonist (15dPGJ2) 30 minutes before I/R injury (group 4). The serum samples were collected for ELISA test, and intestinal mucosa samples were collected for TEM, western blotting and EMSA analysis. Results: After I/R injury, the structure of TJs was disrupted and the expression of occludin decreased. The serum intestinal fatty acid binding protein (IFABP) elevated and the expression of intracellular I-FABP decreased. The expression of activated PPARg in nucleus was attenuated. Pretreatment with n-3PUFAs and 15dPGJ2 reduced the serum level of I-FABP and kept TJs morphology intact comparing with group 2. The expression of occludin and the activated PPARg were both improved in group 3 and 4. Moreover, the expression of cytoplasmic I-FABP was increased in group 3. Conclusion: The intestinal barrier is severely destroyed after I/R injury, which is related to the redistribution of I-FABP. Our findings for the first time indicate that n-3PUFAs can protect the intestinal barrier by modifying intracellular I-FABP, transporting n-3 PUFAs from cytoplasm to nuclei, activating the PPARg pathway in nuclei, and then up regulating the expression of TJ proteins of the intestinal barrier.

Functional Analysis of Pegasus: The ‘Atypical' Member of the Ikaros Gene Family.

Abstract 3645Poster Board III-581The Ikaros family of zinc finger transcription factors are an important class of hematopoietic regulators involved in the development of the immune system. However, the in vivo role of its most divergent ‘atypical' member, Pegasus, remains unknown. Therefore, we have utilised zebrafish to explore the function of Pegasus. Bioinformatics analysis identified a single pegasus orthologue in zebrafish. Wholemount in situ hybridisation revealed an initial broad distribution of maternally-derived pegasus transcripts during early zebrafish embryogenesis, which was replaced by specific expression in the brain, eye, thymus, gut and pancreas during late embryogenesis. Morpholino-mediated gene knockdown of zebrafish pegasus resulted in a reduction in cells expressing markers for hematopoietic stem cells (lmo2), granulocytes (mmp9), macrophages (fms), as well as early (ikaros) and late (rag1) lymphoid cells while, conversely, those expressing early (spi1) and late (l-plastin) myeloid markers were increased. No difference was noted in the early erythroid (gata1) compartment, although mature red blood cells appeared reduced, as judged by o-dianisidine staining. During emergency hematopoiesis (elicited by LPS injection), numbers of granulocytes (mpo+) and macrophages (lyz+) were both significantly further reduced. In addition, morphants also showed anterior defects, including reduced forebrain, smaller eyes and otoliths, which corresponded with a reduction of expression of the brain markers pax2.1, krox20 and dlx1. Gut expression of ifabp also appeared affected. Collectively, these experiments suggest a clear role for pegasus in hematopoiesis, as well as novel functions in brain and gut development. Disclosures:No relevant conflicts of interest to declare.

New Insight in Loss of Gut Barrier during Major Non-Abdominal Surgery

BackgroundGut barrier loss has been implicated as a critical event in the occurrence of postoperative complications. We aimed to study the development of gut barrier loss in patients undergoing major non-abdominal surgery.Methodology/Principal FindingsTwenty consecutive children undergoing spinal fusion surgery were included. This kind of surgery is characterized by long operation time, significant blood loss, prolonged systemic hypotension, without directly leading to compromise of the intestines by intestinal manipulation or use of extracorporeal circulation. Blood was collected preoperatively, every two hours during surgery and 2, 4, 15 and 24 hours postoperatively. Gut mucosal barrier was assessed by plasma markers for enterocyte damage (I-FABP, I-BABP) and urinary presence of tight junction protein claudin-3. Intestinal mucosal perfusion was measured by gastric tonometry (PrCO2, Pr-aCO2-gap). Plasma concentration of I-FABP, I-BABP and urinary expression of claudin-3 increased rapidly and significantly after the onset of surgery in most children. Postoperatively, all markers decreased promptly towards baseline values together with normalisation of MAP. Plasma levels of I-FABP, I-BABP were significantly negatively correlated with MAP at ½ hour before blood sampling (−0.726 (p<0.001), −0.483 (P<0.001), respectively). Furthermore, circulating I-FABP correlated with gastric mucosal PrCO2, Pr-aCO2-gap measured at the same time points (0.553 (p = 0.040), 0.585 (p = 0.028), respectively).Conclusions/SignificanceThis study shows the development of gut barrier loss in children undergoing major non-abdominal surgery, which is related to preceding hypotension and mesenterial hypoperfusion. These data shed new light on the potential role of peroperative circulatory perturbation and intestinal barrier loss.

Dairy protein and leucine alter GLP-1 release and mRNA of genes involved in intestinal lipid metabolism in vitro

Objective A growing body of evidence supports an antiobesity effect of dairy products; however, the mechanisms remain unclear. The objective of this study was to explore possible intestinal mechanisms by which dairy delivers an antiobesity effect. The human intestinal cell line, NCI-H716, was used to test the hypothesis that branched-chain amino acids and dairy proteins regulate satiety hormone secretion and modulate genes involved in fatty acid and cholesterol metabolism. Methods In dose–response (0.5%, 1.0%, 2.0%, and 3.0%) studies, the effect of leucine, isoleucine, valine, skim milk, casein, and whey on glucagon-like peptide-1 release and the expression of selected genes were tested. Results Leucine, isoleucine, skim milk, and casein stimulated glucagon-like peptide-1 release ( P < 0.05). Isoleucine and whey downregulated the expression of intestinal-type fatty acid binding protein ( i-FABP), fatty acid transport protein 4 ( FATP4), Niemann-Pick C-1–like-1 protein ( NPC1L1), acetyl-coenzyme A carboxylase ( ACC), fatty acid synthase ( FAS), sterol regulatory element-binding protein-2 ( SREBP-2), and 3-hydroxy-3-methylglutaryl-CoA reductase ( HMGCR; P < 0.05). Leucine and valine downregulated the expression of NPC1L1, ACC, FAS, SREBP-2, and HMGCR ( P < 0.05). Casein downregulated the expression of i-FABP, FATP4, ACC, FAS, SREBP-2, and HMGCR ( P < 0.05). Skim milk downregulated the expression of ACC, FAS, and SREBP-2, but not i-FABP, FATP4, and NPC1L1. Conclusion This work suggests that the antiobesity effect of dairy may be mediated, at least in part, by integration of events that promote glucagon-like peptide-1 secretion and inhibit expression of genes involved in intestinal fatty acid and cholesterol absorption and synthesis.

An Acute and Coincident Increase in FABP Expression and Lymphatic Lipid and Drug Transport Occurs During Intestinal Infusion of Lipid-Based Drug Formulations to Rats

To determine a) whether administration of lipid-based formulations can acutely up-regulate the intestinal expression of I-FABP and L-FABP and b) whether this occurs coincidentally with an increase in intestinal lymphatic lipid and drug transport. The expression of I-FABP and L-FABP mRNA (using q-PCR) and protein (using immunohistochemistry and Western blotting) in enterocytes was compared with data describing transport of lipid and drug into intestinal lymph following infusion of a set of lipid-based formulations. Administration of relatively small amounts of oleic acid (5-20 mg/h) over a 5 h period to rats acutely up-regulated the expression, and altered the intracellular distribution of, I-FABP and L-FABP in the enterocytes of the small intestinal epithelia. The increase in expression of I-FABP and L-FABP correlated well with previous data describing the transport of lipid and drug into intestinal lymph following infusion of the same formulations. The expression and intracellular distribution of I-FABP and L-FABP are acutely influenced by lipid infusion over a time period relevant to feeding or the administration of pharmaceutical lipidic formulations, and these changes occur coincidentally with increased drug transport into the lymphatics.

Dairy proteins and branched‐chain amino acids stimulate GLP‐1 release and downregulate genes involved in fatty acid and cholesterol metabolism in the human NCI‐H716 cell line

The possibility that increased dairy intake can reduce body weight or exaggerate weight loss is attracting an increasing amount of attention. The objective of this work is to test the hypothesis that dairy proteins and branched-chain amino acids (BCAA) regulate satiety hormone secretion, and modulate selective genes involved in fatty acid and cholesterol metabolism. A human intestinal cell line was used to determine the dose effects of skim milk, casein, whey, leucine, isoleucine and valine on glucagon-like peptide-1 (GLP-1) release and the mRNA expression of i-FABP (intestinal type fatty acid binding protein), FATP4 (fatty acid transport protein), NPC1L1 (Niemann-Pick C1 like protein), ACC (acetyl CoA carboxylase), FAS (fatty acid synthase), SREBP-2 (sterol regulatory element-binding protein-2) and HMGCR [3-hydroxy-3-methylglutary-CoA (HMG-CoA) reductase]. The results showed that skim milk, casein, leucine and isoleucine stimulated GLP-1 release. Whey and isoleucine down-regulated the expressions of i-FABP, FATP4, NPC1L1, ACC, FAS, SREBP-2 and HMGCR. Skim milk and casein down-regulated the expression of ACC, FAS, and SREBP-2, but not i-FABP, FATP4 and NPC1L1. Leucine and valine down-regulated the expressions of NPC1L1, ACC, FAS, SREBP-2 and HMGCR, but not i-FABP, FATP4. The findings suggested that the anti-obesity effect of dairy may be mediated, in part, by integration of events that promote GLP-1 secretion and inhibit expression of genes involved in fatty acid and cholesterol absorption and synthesis. (Supported by CIHR and NSERC)

Dietary exposure to soy or whey proteins alters colonic global gene expression profiles during rat colon tumorigenesis

BackgroundWe previously reported that lifetime consumption of soy proteins or whey proteins reduced the incidence of azoxymethane (AOM)-induced colon tumors in rats. To obtain insights into these effects, global gene expression profiles of colons from rats with lifetime ingestion of casein (CAS, control diet), soy protein isolate (SPI), and whey protein hydrolysate (WPH) diets were determined.ResultsMale Sprague Dawley rats, fed one of the three purified diets, were studied at 40 weeks after AOM injection and when tumors had developed in some animals of each group. Total RNA, purified from non-tumor tissue within the proximal half of each colon, was used to prepare biotinylated probes, which were hybridized to Affymetrix RG_U34A rat microarrays containing probes sets for 8799 rat genes. Microarray data were analyzed using DMT (Affymetrix), SAM (Stanford) and pair-wise comparisons. Differentially expressed genes (SPI and/or WPH vs. CAS) were found. We identified 31 induced and 49 repressed genes in the proximal colons of the SPI-fed group and 44 induced and 119 repressed genes in the proximal colons of the WPH-fed group, relative to CAS. Hierarchical clustering identified the co-induction or co-repression of multiple genes by SPI and WPH. The differential expression of I-FABP (2.92-, 3.97-fold down-regulated in SPI and WPH fed rats; P = 0.023, P = 0.01, respectively), cyclin D1 (1.61-, 2.42-fold down-regulated in SPI and WPH fed rats; P = 0.033, P = 0.001, respectively), and the c-neu proto-oncogene (2.46-, 4.10-fold down-regulated in SPI and WPH fed rats; P < 0.001, P < 0.001, respectively) mRNAs were confirmed by real-time quantitative RT-PCR. SPI and WPH affected colonic neuro-endocrine gene expression: peptide YY (PYY) and glucagon mRNAs were down-regulated in WPH fed rats, whereas somatostatin mRNA and corresponding circulating protein levels, were enhanced by SPI and WPH.ConclusionsThe identification of transcripts co- or differentially-regulated by SPI and WPH diets suggests common as well as unique anti-tumorigenesis mechanisms of action which may involve growth factor, neuroendocrine and immune system genes. SPI and WPH induction of somatostatin, a known anti-proliferative agent for colon cancer cells, would inhibit tumorigenesis.

An isocaloric PUFA diet enhances lipid uptake and weight gain in aging rats.

Aging is associated with a change in the morphology and absorptive capacity of the small intestine. In young rats, feeding a semisynthetic diet containing saturated FA (SFA) increases nutrient uptake, as compared with an isocaloric diet containing polyunsaturated FA (PUFA). We tested the hypotheses that (i) aging is associated with a decline in lipid absorption in the Fischer 344 rat; (ii) this decline can be corrected by manipulating the fat composition of the diet; and (iii) the age- and diet-associated variations in lipid uptake are associated with changes in the ileal lipid-binding protein (ILBP) or the intestinal or liver FA-binding proteins (I- or L-FABP, respectively) in the cytosol of the enterocyte. In rats fed SFA or PUFA, aging was associated with a decline in the in vitro uptake of stearic acid (18:0) when expressed on the basis of intestinal or mucosal weight. In contrast, age had no effect on lipid uptake when expressed on the basis of serosal surface area, whereas lipid uptake increased with age when expressed on the basis of mucosal surface area. The age-associated variations in lipid uptake were not associated with changes in protein abundance and/or expression of ILBP, I-FABP, or L-FABP. In 24-mon-old rats, when uptake of lipids was expressed on the basis of mucosal surface area, feeding PUFA enhanced lipid uptake and body weight gain as compared with rats fed SFA. Future studies must determine whether the enhanced lipid uptake and body weight gain observed in older animals fed PUFA have any therapeutic benefit.

Distinct epithelial responses in experimental colitis: implications for ion uptake and mucosal protection.

In the present study, we aimed to investigate enterocyte- and goblet cell-specific functions during the different phases of acute colitis induced with dextran sulfate sodium (DSS). Rats were treated with DSS for 7 days, followed by a 7-day recovery period. Colonic tissue was excised on days 2 (onset of disease), 7 (active disease), and 14 (regenerative phase). Enterocyte functions were studied by the expression of carbonic anhydrases (CAs), sodium/hydrogen exchangers (NHEs) and intestinal fatty acid-binding protein (iFABP) and by alkaline phosphatase (AP) activity. The expression and secretion of the mucin Muc2 and trefoil factor family peptide-3 (TFF3) were used as parameters for goblet cell function. DSS induced a downregulation of the CAs, NHEs, and iFABP in some normal-appearing surface enterocytes and in most of the flattened-surface enterocytes during disease onset and active disease. During the regenerative phase most enterocytes expressed these genes again. Quantitative analysis revealed a significant decrease in CAs, NHEs, and iFABP expression levels during onset and active disease. During the regenerative phase, the expression levels of the CAs were restored, whereas the expression levels of the NHEs and iFABP remained decreased. In contrast, enterocyte-specific AP activity was maintained in normal and flattened enterocytes during DSS-induced colitis. Goblet cells continued to express MUC2 and TFF3 during and after DSS treatment. Moreover, Muc2 and TFF3 expression and secretion levels were maintained or even increased during each of the DSS-induced disease phases. In conclusion, DSS-induced colitis was associated with decreased expression of CAs, NHEs, and iFABP. The loss of these genes possibly accounts for some of the pathology seen in colitis. The maintenance or upregulation of Muc2 and TFF3 synthesis and secretion levels implies that goblet cells at least maintain their epithelial defense and repair capacity during acute inflammation induced by DSS.

Modulation of intestinal and liver fatty acid-binding proteins in Caco-2 cells by lipids, hormones and cytokines.

Intestinal and liver fatty acid binding proteins (I- and L-FABP) are thought to play a role in enterocyte fatty acid (FA) trafficking. Their modulation by cell differentiation and various potential effectors was investigated in the human Caco-2 cell line. With the acquisition of enterocytic features, Caco-2 cells seeded on plastic progressively increased L-FABP quantities, whereas I-FABP was not detectable even very late in the maturation process. On permeable filters that improved differentiation markers (sucrase, alkaline phosphatase, transepithelial resistance), Caco-2 cells furthered their L-FABP content and expressed I-FABP. Western blot analysis showed a significant increase in I- and L-FABP expression following an 8-hour incubation period with butyric acid, oleic acid, and phosphatidylcholine. However, in all cases, I-FABP levels were higher than L-FABP concentrations regardless of the lipid substrates added. Similarly, hydrocortisone and insulin enhanced the cellular content of I- and L-FABP whereas leptin triggered I-FABP expression only after an 8-hour incubation. Finally, tumor necrosis factor-alpha was more effective in increasing the cytosolic amount of I-FABP levels. In conclusion, our data demonstrate that I-FABP expression is limited to fully differentiated Caco-2 cells and can be more easily regulated than L-FABP by lipids, hormones, and cytokines.