Messenger RNA expression and localization of xenin in the gastrointestinal tract in sheep.

The present study aimed to determine the primary sequence of ovine xenin and clarify the mRNA expression and peptide localization of xenin in the gastrointestinal tract in sheep. The colocalization of xenin and glucose-dependent insulinotropic polypeptide was also compared in the antrum and duodenum. Analysis of the nucleotide sequence of ovine xenin revealed a high degree (97.9%) of sequence homology of the sequence between sheep and cattle, and the amino acids sequence determined for ovine xenin coincided (100%) with that of other mammalian species. Real-time quantitative PCR for ovine xenin did not show regional difference in the mRNA expression ratio of xenin. In contrast to the real-time quantitative PCR results, anti-xenin positive cells were abundantly localized in the abomasal antrum (P < 0.01) and at a lesser amount in the duodenum, but no antixenin positive cells were observed in the other regions. Anti-xenin single-positive cells were in a majority in the abomasal antrum, whereas anti-xenin single-positive cells, and anti-GIP single-positive cells, and double-positive cells were even colocalized in the duodenum. These results suggest that abomasal antrum is a major source of xenin in the ovine gastrointestinal tract.

Establishment of novel specific assay for short-form glucose-dependent insulinotropic polypeptide and evaluation of its secretion in nondiabetic subjects.

The short-form glucose-dependent insulinotropic polypeptide (GIP) (1-30) is released from islet alpha cells and promotes insulin secretion in a paracrine manner in vitro. However, it is not well elucidated how GIP (1-30) is involved in glucose metabolism in vivo, since a specific assay system for GIP (1-30) has not yet been established. We first developed a sandwich enzyme-linked immunosorbent assay (ELISA) specific for GIP (1-30) by combining a novel antibody specific to the GIP (1-30) C terminus with the common antibody against GIP N terminus. Then, we explored cross-reactivities with incretins and glucagon-related peptides in this ELISA. GIP (1-30) amide, but not GIP (1-42), GLP-1, or glucagon increased absorbance in a dose-dependent manner. We next measured plasma GIP (1-30) concentrations in nondiabetic participants (ND) during a 75-g oral glucose tolerance test or cookie meal test (carbohydrates 75 g, lipids 28.5 g, proteins 8.5 g). Both glucose and cookie load increased GIP (1-30) concentrations in ND, but the increases were much lower than those of GIP (1-42). Furthermore, the DPP-4 inhibitor significantly increased GIP (1-30) concentrations similarly to GIP (1-42) in ND. In conclusion, we for the first time developed an ELISA specific for GIP (1-30) and revealed its secretion in ND.

Is gastrointestinal motility related to alkaloids of Charred Semen Arecae?

ETHNOPHARMACOLOGICAL RELEVANCE: Semen Arecae (SA) is one of the most commonly used Traditional Chinese Medicine. Charred Semen Arecae (CSA) is the processed product of SA. Alkaloids are considered as pharmacological mechanisms of SA and CSA on gastrointestinal motility. Recent studies have shown alkaloids decreased quickly after procession. However, the promoting on gastrointestinal motility were not decreased. Is gastrointestinal motility related to alkaloids of CSA? This study explored the effects of SA, CSA, Semen Arecae-Removal (SA-R), and Charred Semen Arecae-Removal (CSA-R) on gastrointestinal motility, Gastric Inhibitory Polypeptide (GIP), Glucagon Like Peptide-1 (GLP-1), gastric juice and bile in rats. MATERIAL AND METHODS: Rats were randomly divided into six groups, including the Control group, SA group, CSA group, SA-R group, CSA-R group, and Positive drug group (Mosapride). Alkaloids of samples were knocked out by using the "target constituent removal" strategy. Gastric residue and intestinal propulsion rate were evaluated in rats. Serum levels of GIP and GLP-1 were measured by Enzyme-Linked Immunosorbent Assay (ELISA). Gastric juice and bile were examined, respectively. RESULTS: CSA-R and SA-R have been investigated by the Preparative Thin-layer Chromatography (PTLC) method. Intestinal propulsion and gastric residue assessments confirmed the effectiveness of CSA and CSA-R. CSA-R was higher than SA-R in the GLP-1, pepsin activity, the secretion of bile, Bilirubin (BIL), and Cholesterol (CHO). The statistical comparison demonstrated that there is no difference between the CSA group and CSA-R group. CONCLUSIONS: After processing, the promoting gastrointestinal motility might be not related to alkaloids. Maillard reaction could be produced to promote the secretion of GLP-1, bile, and CHO for gastrointestinal motility. Our findings provide a pharmacological reference for the clinical application of SA and CSA in the treatment of digestive diseases.

GLP1- and GIP-producing cells rarely overlap and differ by bombesin receptor-2 expression and responsiveness.

The incretin hormones glucagon-like peptide-1 (GLP1) and glucose-dependent insulinotropic polypeptide (GIP) are secreted from intestinal endocrine cells, the so-called L- and K-cells. The cells are derived from a common precursor and are highly related, and co-expression of the two hormones in so-called L/K-cells has been reported. To investigate the relationship between the GLP1- and GIP-producing cells more closely, we generated a transgenic mouse model expressing a fluorescent marker in GIP-positive cells. In combination with a mouse strain with fluorescent GLP1 cells, we were able to estimate the overlap between the two cell types. Furthermore, we used primary cultured intestinal cells and isolated perfused mouse intestine to measure the secretion of GIP and GLP1 in response to different stimuli. Overlapping GLP1 and GIP cells were rare (∼5%). KCl, glucose and forskolin+IBMX increased the secretion of both GLP1 and GIP, whereas bombesin/neuromedin C only stimulated GLP1 secretion. Expression analysis showed high expression of the bombesin 2 receptor in GLP1 positive cells, but no expression in GIP-positive cells. These data indicate both expressional and functional differences between the GLP1-producing 'L-cell' and the GIP-producing 'K-cell'.

Measurement of the incretin hormones: glucagon-like peptide-1 and glucose-dependent insulinotropic peptide.

The two incretin hormones, glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP), are secreted from the gastrointestinal tract in response to meals and contribute to the regulation of glucose homeostasis by increasing insulin secretion. Assessment of plasma concentrations of GLP-1 and GIP is often an important endpoint in both clinical and preclinical studies and, therefore, accurate measurement of these hormones is important. Here, we provide an overview of current approaches for the measurement of the incretin hormones, with particular focus on immunological methods.

Distribution of K and L cells in the feline intestinal tract.

Glucose-dependent insulinotropic peptide (GIP), glucagon-like peptide (GLP)-1 and GLP-2 are hormones secreted from specialized K cells (GIP) and L cells (GLP-1, GLP-2) in the intestinal mucosa. These hormones play major roles in health and disease by modulating insulin secretion, satiety, and multiple intestinal functions. The aim of this study was to describe the distribution of K cells and L cells in the intestines of healthy cats. Samples of duodenum, mid-jejunum, ileum, cecum, and colon were collected from 5 cats that were euthanized for reasons unrelated to this study and had no gross or histologic evidence of gastrointestinal disease. Samples stained with rabbit-anti-porcine GIP, mouse-anti-(all mammals) GLP-1, or rabbit-anti-(all mammals) GLP-2 antibodies were used to determine the number of cells in 15 randomly selected 400× microscopic fields. In contrast to other mammals (eg, dogs) in which K cells are not present in the ileum and aborally, GIP-expressing cells are abundant throughout the intestines in cats (>6/high-power field in the ileum). Cells expressing GLP-1 or GLP-2 were most abundant in the ileum (>9/high-power field) as in other mammals, but, although GLP-1-expressing cells were abundant throughout the intestines, GLP-2-expressing cells were rarely found in the duodenum. In conclusion, the distribution of GIP-secreting K cells in cats is different from the distribution of K cells that is described in other mammals. The difference in distribution of GLP-2- and GLP-1-expressing cells suggests that more than 1 distinct population of L cells is present in cats.

The small intestine of the adult New Hampshire chicken: an immunohistochemical study.

The presence and distribution of glucose-dependent insulinotropic polypeptide or gastric inhibitory polypeptide (GIP), gastric-releasing peptide (GRP) and glucagon immunoreactivity were studied in the small intestine of the New Hampshire chicken using immunohistochemistry. This is the first report of the presence of GIP-immunoreactive (ir) cells in avian small intestine. GIP, GRP and glucagon immunoreactivity was localized in the epithelium of the villi and crypts of the duodenum, jejunum and ileum. In particular, both in the duodenum and in the jejunum immunoreactive endocrine cells to GIP, GRP and glucagon were observed. In the ileum, we noticed GIP-ir and glucagon-ir cells. GRP-ir was found in nerve fibres of all three segments of the small intestine. The distribution of these bioactive agents in the intestinal tract of the chicken suggests that GIP and glucagon may play a role in the enteropancreatic axis in which intestinal peptides modulate pancreas secretion.

Effect of ezetimibe on incretin secretion in response to the intestinal absorption of a mixed meal.

Ezetimibe is a potent inhibitor of cholesterol absorption by enterocytes. Although ezetimibe minimally affects the absorption of triglyceride, it is unknown whether ezetimibe affects the secretion of the incretin hormones glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1). It has been shown that ezetimibe-treated mice are protected from diet-induced insulin resistance. Since GIP and GLP-1 promote the actions of insulin, we hypothesized that ezetimibe may affect the secretion of GIP and GLP-1 by enteroendocrine cells into lymph in response to the intestinal absorption of a mixed meal (Ensure). To test this hypothesis, we used the lymph fistula rat model to determine GIP and GLP-1 concentrations in lymph during the 2 h after the infusion of Ensure. Ezetimibe significantly reduced lymphatic cholesterol output during fasting, without coincident decreases in glucose, protein, and triglyceride outputs. However, ezetimibe did not influence cholesterol output after infusion of Ensure. Interestingly, ezetimibe significantly reduced the secretion of both GIP and GLP-1 into lymph after the infusion of Ensure. Therefore, the inhibitory effect of ezetimibe on GIP and GLP-1 secretion by enteroendocrine cells occurs outside of the effects of glucose, protein, or triglyceride secretion by the intestine.

The effect of duodenal-jejunal bypass on glucose-dependent insulinotropic polypeptide secretion in Wistar rats.

BACKGROUND: Enteroendocrine K cells secrete the incretin hormone glucose-dependent insulinotropic peptide (GIP) and are predominately located in the duodenum. GIP levels should decrease after gastric bypass due to duodenal exclusion; however, studies have found conflicting data regarding the changes in GIP secretion after gastric bypass and duodenal-jejunal bypass (DJB). METHODS: We performed a DJB or Sham surgery on Wistar rats followed by an oral glucose tolerance test on postoperative (post-op) day 12 and superior mesenteric lymphatic cannulation on post-op day 14. We measured meal-stimulated GIP concentrations and small bowel GIP and GLP-1 protein content after DJB or Sham surgery. RESULTS: There was no difference in glucose tolerance by 12 days post-op. We found no difference in lymphatic GIP concentration area under the curve between DJB and Sham rats (15,240 pg/ml min +/- 2,651 vs. 17,201 pg/ml min +/- 2,763, respectively, p = 0.62). GIP and GLP-1 protein contents were both significantly increased only in the midjejunum in DJB rats compared to Sham rats (p = 0.009 and p = 0.01, respectively). CONCLUSIONS: Plasma and lymphatic GIP concentrations did not significantly change after DJB in Wistar rats. DJB increased GIP protein content in the midjejunum at the new site of nutrient absorption, but this was surprisingly not countered by a decrease in GIP protein content in the bypassed duodenum. Further studies are needed to determine the mechanisms that account for the discrepancy in GIP production and subsequent secretion after DJB as well as what role GIP plays in the effect of gastrointestinal surgery on glucose homeostasis.

Immunoassays for the incretin hormones GIP and GLP-1.

The measurement of the incretin hormones, glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), using immunologically based assays is made difficult by the fact that the processing of the precursor molecules gives rise to a number of different peptides which cross-react with antisera raised against the two hormones. For GLP-1, the picture is further complicated because of the necessity to differentiate between the intestinal and pancreatic proglucagon products. Finally, once secreted, both incretins are rapidly degraded by the enzyme dipeptidyl peptidase-4 (DPP-4) to generate metabolites which have lost their insulinotropic activities. These metabolites are the major circulating forms of the incretins, accounting for 60-80% of total immunoreactive GLP-1 and GIP in the peripheral plasma, while concentrations of the intact forms can be very low and, in some cases, barely detectable. The use of highly specific assays using well-characterised antisera and careful sample handling is therefore required for a reliable determination of incretin hormone concentrations.

Las hormonas gastrointestinales en el control de la ingesta de alimentos / Gastrointestinal hormones in food intake control

El descubrimiento de la existencia de hormonas gastrointestinales que modulan la homeostasis energética ha despertado un gran interés. Algunas de estas hormonas, actuando en el hipotálamo o el núcleo del tracto solitario en el tronco encefálico, ejercen efectos moduladores del apetito y la saciedad. En términos generales, las señales endocrinas generadas en el tracto gastrointestinal tienen efecto anorexigénico directo o indirecto a través del sistema nervioso vegetativo. Sólo la ghrelina, hormona producida en el estómago, se ha asociado de manera consistente con el inicio de la ingesta y se la considera una de las principales señales orexigénicas en los modelos animales estudiados y en humanos. En esta revisión, se describen brevemente las principales hormonas de origen gastrointestinal implicadas en la regulación del apetito. Dada la importancia que los trastornos de la ingesta de alimentos, especialmente la obesidad, han adquirido, un mejor conocimiento de los mecanismos de acción de estas señales endocrinas podría contribuir al desarrollo de nuevas moléculas que incrementen y mejoren nuestro arsenal terapéutico para tratar la obesidad y las enfermedades crónicas relacionadas con ella (AU)

The discovery of gut hormones regulating the energy balance has aroused great interest in the scientific community. Some of these hormones modulate appetite and satiety, acting on the hypothalamus or the solitary tract nucleus in the brainstem. In general, the endocrine signals generated in the gut have direct or indirect (through the autonomous nervous system) anorexigenic effects. Only ghrelin, a gastric hormone, has been consistently associated with the initiation of food intake and is regarded as the main orexigenic signal both in animal models and humans. In this review, we provide a brief description of the major gastrointestinal hormones implicated in the regulation of food intake. Given the increased importance of food intake disturbances, especially obesity, a better understanding of the underlying mechanisms of action of the gastrointestinal hormones might contribute to the development of new molecules that could increase the therapeutic arsenal for treating obesity and its associated comorbidities (AU)

Distribution of neuroendocrine cells in the small and large intestines of the one-humped camel (Camelus dromedarius).

The distribution and relative frequency of neuroendocrine cells in the small and large intestines of one-humped camel were studied using antisera against 5-hydroxytryptamine (5-HT), cholecystokinin (CCK-8), somatostatin (SOM), peptide tyrosine tyrosine (PYY), gastric inhibitory polypeptide (GIP), neuronal nitric oxide synthase (nNOS), gastrin releasing peptide (GRP), substance P (SP), and neurokinin A (NKA). Among these cell types, CCK-8 immunoreactive (IR) cells were uniformly distributed in the mucosa, while others showed varied distribution in the villi or crypts of the small intestine. Immunoreactive cells like 5HT, CCK-8, and SOM showed peak density in the villi and crypts of the small intestine and in the colonic glands of the large intestine, while cells containing SP were discerned predominately in the crypts. 5-HT, CCK-8 and SOM cells were mainly flask-shaped and of the open-variety, while PYY and SP immunoreactive cells were mainly rounded or basket-shaped and of the closed variety. Basically the distribution pattern of the endocrine cells in the duodenum, jejunum and colon of the one-humped camel is similar to that of other mammals. Finally, the distribution of these bioactive agents may give clues as to how these agents aid in the function of the intestinal tract of this desert animal.

Helicobacter pylori infection stimulates intestinalization of endocrine cells in glandular stomach of Mongolian gerbils.

Intestinal metaplasia has been investigated extensively as a possible premalignant condition for stomach cancer but its pathogenesis is still not fully understood. In the present study, we examined the relationship between endocrine and mucous cell marker expression periodically after Helicobacter pylori infection in the Mongolian gerbil model. The numbers of chromogranin A (CgA)-positive, gastrin-positive and gastric inhibitory polypeptide (GIP)-positive cells in H. pylori-infected groups was increased significantly compared with the non-infected case. However, CgA-positive and gastrin-positive cells then decreased from 50 through 100 experimental weeks after H. pylori infection, whereas GIP-positive cells increased. Coexistence of gastrin-positive and GIP-positive cells was detected in the same gastric and intestinal mixed phenotypic glandular-type glands. In conclusion, the endocrine cell phenotype is in line with that of the mucous counterpart in the glands of H. pylori-infected Mongolian gerbil stomach, supporting the concept that development of intestinal metaplasia is due to the abnormal differentiation of a stem cell.

Coexistence of gastric- and intestinal-type endocrine cells in gastric and intestinal mixed intestinal metaplasia of the human stomach.

Intestinal metaplasia (IM) in the human stomach has previously been classified into a gastric and intestinal mixed (GI-IM) and a solely intestinal phenotype (I-IM). The phenotypes of mucous and endocrine cells were evaluated in 3034 glandular ducts associated with chronic gastritis. In the pyloric region, the relative expression of gastric endocrine cell markers, such as gastrin and somatostatin, decreased gradually from glandular ducts with only gastric mucous cell phenotype (G type) to GI-IM toward I-IM, while that of the intestinal endocrine cell markers, glicentin, gastric inhibitory polypeptide (GIP), and glucagon-like peptide-1 (GLP-1) was inversely correlated. In the fundic region, gastrin-positive cells emerged in the pseudo-pyloric and GI-IM glands, whereas I-IM glands did not possess any gastrin-positive cells, suggesting the presence of a distinct pathway of intestinalization. Double staining revealed coexistence of gastrin- and GLP-1-positive cells in the same gland and occasionally in the same cell in GI-IM glands. These results suggest that the phenotypes of endocrine cells are in line with those for mucous counterparts and support the concept that all of the different types of mucous and endocrine cells in normal and IM glands might be derived from a single progenitor cell in each gland.

A stereological evaluation of secretin and gastric inhibitory peptide-containing mucosal cells of the perinatal small intestine of the pig.

Stereological methods were used to quantify secretin and gastric inhibitory peptide (GIP)-immunoreactivity (GIP-IR) in paraffin sections of the duodenum, jejunum and ileum of fetal and neonatal piglets. In addition, sections were processed for GLP-1-immunohistochemistry. The volume density of the tunica mucosa increased after birth, giving rise to a decreased volume density of the tela submucosa and tunica muscularis. Generally known region-specific morphological distinctions were reflected in differing volume densities of the various layers. The highest volume density of GIP-IR epithelial cells was observed in the jejunum of the neonate. In contrast, the volume density of secretin-IR epithelial cells was highest in the duodenum of both fetal and neonatal piglets. The volume occupied by GIP-IR and secretin-IR epithelial cells increased in the jejunum after birth. Additionally, ileal secretin-IR epithelial cells were more numerous in the neonatal piglet. In conclusion, the quantitative and qualitative presence of GIP-IR and secretin-IR epithelial cells agree with earlier reports of their presence and co-localization between GIP-IR and GLP-1-IR, in the pig small intestine. Furthermore, the differences suggest that age- and region-related functional demands are temporally and probably causally related with the morphological diversification of the intestine and its endocrine cells.

The early effect of the Roux-en-Y gastric bypass on hormones involved in body weight regulation and glucose metabolism.

OBJECTIVE: To evaluate the early effect of Roux-en-Y (RYGB) gastric bypass on hormones involved in body weight regulation and glucose metabolism. SIGNIFICANT BACKGROUND DATA: The RYGB is an effective bariatric procedure for which the mechanism of action has not been elucidated yet. Reports of hormonal changes after RYGB suggest a possible endocrine effect of the operation; however, it is unknown whether these changes are the cause or rather the effect of surgically induced weight loss. We speculated that if the mechanism of action of the RYGB involves an endocrine effect, then hormonal changes should occur early after surgery, prior to substantial body weight changes. METHODS: Ten patients with a mean preoperative body mass index (BMI) of 46.2 kg/m (40-53 kg/m) underwent laparoscopic RYGB. Six patients had type 2 diabetes treated by oral hypoglycemic agents. Preoperatively and 3 weeks following surgery, all patients were tested for fasting glucose, insulin, glucagon, insulin-like growth factor 1 (IGF-1), leptin, gastric inhibitory polypeptide (GIP), glucagon-like peptide-1 (GLP-1), cholecystokinin (CCK), adrenocorticotropic hormone (ACTH), corticosterone, and neuropeptide Y (NPY). RESULTS: Changes in mean BMI were rather minimal (43.2 kg/m; P = not significant), but there was a significant decrease in blood glucose (P = 0.005), insulin (P = 0.02), IGF-1 (P < 0.05), leptin (P = 0.001), and an increase in ACTH levels (P = 0.01). The other hormones were not significantly changed by surgery. All the 6 diabetic patients had normal glucose and insulin levels and did not require medications after surgery. The RYGB reduced GIP levels in diabetic patients (P < 0.01), whereas no changes in GIP levels were found in nondiabetics. CONCLUSIONS: Roux-en-Y gastric bypass determines considerable hormonal changes before significant BMI changes take place. These results support the hypothesis of an endocrine effect as the possible mechanism of action of RYGB.

Glucose-dependent insulinotropic polypeptide and insulin-like immunoreactivity in saliva following sham-fed and swallowed meals.

Gastrointestinal peptides, including insulin, glucagon and glucose-dependent insulinotropic polypeptide (GIP) have previously been reported in salivary glands. Recent evidence has suggested they might influence postprandial macronutrient metabolism. This study therefore investigated and compared postprandial hormone concentrations in saliva and plasma to determine whether their secretion was influenced by oral food stimuli. In a within-subject randomised cross-over comparison of hormone concentrations in plasma and saliva following a mixed meal, 12 subjects were given two 1708 kJ mixed meals. On one occasion the meal was chewed and swallowed (swallowed meal), on the other it was chewed and expectorated (sham-fed meal). Salivary and plasma levels of immunoreactive insulin, GIP and glucagon-like peptide-1 (GLP-1), total protein, alpha-amylase, glucose and non-esterified fatty acid were measured before and for 90 min following the meals. Saliva total protein and alpha-amylase rose following both meals, indicating that the stimulus for salivary protein release is related to the presence of food in the mouth. GLP-1 was not detected in saliva. Fasting salivary insulin levels were lower in saliva than plasma (28+/-6 vs 40+/-25 pmol/l respectively). Both increased following the swallowed meal but the rise in saliva was slower and less marked than in plasma (peak levels 96+/-18 and 270+/-66 pmol/l for saliva and plasma respectively, P<0.01). Both were unchanged following the sham-fed meal. GIP was detected in saliva. Fasting GIP levels were significantly higher in saliva than plasma (183+/-23 compared with 20+/-7 pmol/l, P<0.01). They decreased in saliva following both swallowed and sham-fed meals to nadirs of 117+/-17 and 71+/-12 pmol/l respectively, but rose following the swallowed meal to peak levels of 268+/-66 pmol/l. These findings are consistent with insulin in saliva being an ultrafiltrate of that circulating in blood, but GIP in saliva being the product of local salivary gland synthesis, whose secretion is influenced, directly or indirectly, by oral stimuli. The function of salivary GIP is unknown, but we speculate that it may play a role in the regulation of gastric acid secretion in the fasting state.

Immunohistochemical study of the distribution of endocrine cells in the gastrointestinal tract of the camel (Camelus bactrianus).

The regional distribution and relative frequency of endocrine cells in the gastrointestinal tract of the camel, Camelus bactrianus, were investigated using immunohistochemical methods. Ten types of immunoreactive (IR) endocrine cells were identified in this study. Among these cell types, only serotonin- and somatostatin-IR cells were detected in almost all regions of the gastrointestinal tract. Most of the cell types showed peak density in the pyloric gland region. The others showed restricted distribution: gastrin, cholecystokinin (CCK), motilin, bovine pancreatic polypeptide (BPP), and (gastric) substance P in the stomach; gastrin, CCK, BPP, gastric inhibitory polypeptide (GIP), glucagon, peptide tyrosine tyrosine (PYY) and substance P in the small intestine; and CCK, motilin, BPP, and PYY in the large intestine. Fundamentally the distribution pattern of endocrine cells in the gastrointestinal tract of the camel is similar to that of cattle. The distribution and frequency of endocrine cells in the glandular sac region are the same as those of the cardiac gland.

Indirect RT-PCR in-situ hybridization: a novel non-radioactive method for detecting glucose-dependent insulinotropic peptide.

To establish indirect in-situ PCR for the detection of intestinal peptide hormones, rat intestine and a murine intestinal tumor cell line, STC 1, were used. The results exhibited intensive staining of GIP-producing K-cells. Paraformaldehyde-fixed cryostat sections yielded the best results in signal to background ratio with RT-PCR in-situ hybridization. Moreover, it was possible to elevate the positive staining signal and to reduce background staining. Digoxigenin-labeled in-situ hybridization served as a control for specificity and sensitivity of GIP (glucose-dependent insulinotropic peptide) mRNA expression on cryostat as well as paraffin sections. In conclusion, this RT-PCR in-situ hybridization protocol proves to be a specific, sensitive and reliable non-radioactive technique for the detection of intestinal peptide hormone mRNA, especially in tissues or tumor cells where the application of ISH is limited.

Detection of glycated gastric inhibitory polypeptide within the intestines of diabetic obese (ob/ob) mice.

Gastric inhibitory polypeptide (GIP) is produced within endocrine cells of the small intestine and released into the circulation upon nutrient ingestion. This study has quantified the levels of this insulinotropic peptide in the intestines of lean and diabetic obese ob/ob mice and estimated the proportion that is glycated. The total intestinal GIP concentration and content of the diabetic mice were significantly greater (p < 0.01) than that of control animals. Affinity chromatographic separation and side-viewing GIP radioimmunoassay demonstrated that approx 20% of the GIP extracted from intestines of ob/ob mice was present in glycated form. Less than 2% of intestinal GIP was glycated in lean mice. In conclusion substantial quantities of glycated GIP exist within the intestines of diabetic ob/ob mice, suggesting that this may be a contributing factor to the physiological disarray of this syndrome.