In vivo insulin action and resistance. The hyperinsulinaemic, euglycaemic clamp technique in conscious rats

After almost 75 years of research and four Nobel prizes awarded for insulin-related investigations, we still do not understand how insulin works. Of course, substantial progress has been made and there are some clues. We review in vivo insulin action from the moment of insulin biosynthesis in the pancreatic β-cell to insulin action on effector systems in target tissues. The mechanism of insulin action at the whole body, tissue and cellular level is discussed. In addition, factors and conditions influencing insulin action and events leading to insulin resistance are summarized. Finally, the reader is introduced into the principles of the hyperinsulinaemic, euglycaemic clamp technique which is considered the "golden standard" for measurement of insulin action and resistance in vivo. An example is given how to clamp conscious rats in a proper way.Biomedical Reviews 1996; 5: 31-45

The existence of an insulin-stimulated glucose and non-essential but not essential amino acid substrate interaction in diabetic pigs

<p>Abstract</p> <p>Background</p> <p>The generation of energy from glucose is impaired in diabetes and can be compensated by other substrates like fatty acids (Randle cycle). Little information is available on amino acids (AA) as alternative energy-source in diabetes. To study the interaction between insulin-stimulated glucose and AA utilization in normal and diabetic subjects, intraportal hyperinsulinaemic euglycaemic euaminoacidaemic clamp studies were performed in normal (n = 8) and streptozotocin (120 mg/kg) induced diabetic (n = 7) pigs of ~40-45 kg.</p> <p>Results</p> <p>Diabetic vs normal pigs showed basal hyperglycaemia (19.0 ± 2.0 vs 4.7 ± 0.1 mmol/L, <it>P </it>< .001) and at the level of individual AA, basal concentrations of valine and histidine were increased (<it>P </it>< .05) whereas tyrosine, alanine, asparagine, glutamine, glutamate, glycine and serine were decreased (<it>P </it>< .05). During the clamp, diabetic vs normal pigs showed reduced insulin-stimulated glucose clearance (4.4 ± 1.6 vs 16.0 ± 3.0 mL/kg·min, <it>P </it>< .001) but increased AA clearance (166 ± 22 vs 110 ± 13 mL/kg· min, <it>P </it>< .05) at matched arterial euglycaemia (5-7 mmol/L) and euaminoacidaemia (2.8-3.5 mmol/L). The increase in AA clearance was mainly caused by an increase in non-essential AA clearance (93.6 ± 13.8 vs 46.6 ± 5.4 mL/kg·min, <it>P </it>< .01), in particular alanine (14.2 ± 2.4 vs 3.2 ± 0.4 mL/kg·min, <it>P </it>< .001)<b/>. Essential AA clearance was largely unchanged (72.9 ± 8.5 vs 63.3 ± 8.5 mL/kg· min), however clearances of threonine (<it>P </it>< .05) and tyrosine (<it>P </it>< .01) were increased in diabetic vs normal pigs (8.1 ± 1.3 vs 5.2 ± 0.5, and 14.3 ± 2.5 vs 6.4 ± 0.7 mL/kg· min, respectively).</p> <p>Conclusions</p> <p>The ratio of insulin-stimulated glucose versus AA clearance was decreased 5.4-fold in diabetic pigs, which was caused by a 3.6-fold decrease in glucose clearance and a 2.0-fold increase in non-essential AA clearance. In parallel with the Randle concept (glucose - fatty acid cycle), the present data suggest the existence of a glucose and non-essential AA substrate interaction in diabetic pigs whereby reduced insulin-stimulated glucose clearance seems to be partly compensated by an increase in non-essential AA clearance whereas essential AA are preferentially spared from an increase in clearance.</p

Cross-Species Comparison of Genes Related to Nutrient Sensing Mechanisms Expressed along the Intestine

Introduction Intestinal chemosensory receptors and transporters are able to detect food-derived molecules and are involved in the modulation of gut hormone release. Gut hormones play an important role in the regulation of food intake and the control of gastrointestinal functioning. This mechanism is often referred to as “nutrient sensing”. Knowledge of the distribution of chemosensors along the intestinal tract is important to gain insight in nutrient detection and sensing, both pivotal processes for the regulation of food intake. However, most knowledge is derived from rodents, whereas studies in man and pig are limited, and cross-species comparisons are lacking. Aim To characterize and compare intestinal expression patterns of genes related to nutrient sensing in mice, pigs and humans. Methods Mucosal biopsy samples taken at six locations in human intestine (n = 40) were analyzed by qPCR. Intestinal scrapings from 14 locations in pigs (n = 6) and from 10 locations in mice (n = 4) were analyzed by qPCR and microarray, respectively. The gene expression of glucagon, cholecystokinin, peptide YY, glucagon-like peptide-1 receptor, taste receptor T1R3, sodium/glucose cotransporter, peptide transporter-1, GPR120, taste receptor T1R1, GPR119 and GPR93 was investigated. Partial least squares (PLS) modeling was used to compare the intestinal expression pattern between the three species. Results and conclusion The studied genes were found to display specific expression patterns along the intestinal tract. PLS analysis showed a high similarity between human, pig and mouse in the expression of genes related to nutrient sensing in the distal ileum, and between human and pig in the colon. The gene expression pattern was most deviating between the species in the proximal intestine. Our results give new insights in interspecies similarities and provide new leads for translational research and models aiming to modulate food intake processes in man

Changes in Plasma Protein Expression Indicative of Early Diet-induced Metabolic Disease in Male Pigs (Sus scrofa).

Recognition of the preclinical stages of metabolic diseases such as diabetes helps to prevent full development of the disease. In our research, we alter the diet composition of pigs to create a model of human metabolic disease. The objective of the current study was to identify plasma proteins and biologic mechanisms that differed in expression between pigs fed a 'cafeteria diet' (considered unhealthy; high in saturated fats) and those fed a 'Mediterranean diet' (considered healthy; high in unsaturated fats). Pigs fed the cafeteria diet showed increased plasma levels of proteins related to LDL ('bad cholesterol'), immune processes, blood clotting, and metal binding. The Mediterranean diet was associated with increased plasma quantities of proteins associated HDL particles ('good cholesterol'), binding of LDL particles, regulation of immune processes, and glycolysis. Pigs fed a cafeteria diet showed molecular signs of diabetes and atherosclerosis-even in the absence of clinical symptoms-which seemed to protect against the development of metabolic disorders. The current results suggest potential biomarkers of the early onset of metabolic syndromes. These biomarkers can help to reveal specific metabolic changes that precede the onset of diabetes, thus enabling the initiation of patient-specific interventions early during pathophysiologic development

Effect of Feeding 0.8% Dried Powdered Chlorella vulgaris Biomass on Growth Performance, Immune Response, and Intestinal Morphology during Grower Phase in Broiler Chickens

An experiment was performed to study the effects of a low inclusion level of Chlorella vulgaris (CV) biomass in broiler diets on performance, immune response related to inflammatory status, and the intestinal histomorphology. The study was performed with 120 Ross 308 male broiler chickens from 0–35 days of age. The broilers were housed in 12 floor pens (1.5 m2) bedded with wood shavings. The broilers received a three phase diet program, either with 0.8% CV biomass (CV) or without CV (CON). Each diet program was replicated in six pens. The final body weight increased (p = 0.053), and the feed conversion ratio (FCR), corrected for body weight, was reduced (p = 0.02) in birds fed CV compared to birds fed CON. In addition, decreased haptoglobins (p = 0.02) and interleukin-13 (p < 0.01) responses were observed during the grower phase of birds fed CV compared to the birds fed CON. A strong correlation (r = 0.82, p < 0.01) was observed between haptoglobin response and FCR. Histomorphology parameters of the jejunum were not different between the groups. It was concluded that the inclusion of 0.8% CV biomass in broiler diets is effective in influencing immune responses related to inflammatory status and promoting broiler growth

Molecular networks affected by neonatal microbial colonization in porcine jejunum, luminally perfused with enterotoxigenic Escherichia coli, F4ac fimbria or Lactobacillus amylovorus

The development of an early complex gut microbiota may play an important role in the protection against intestinal dysbiosis later in life. The significance of the developed microbiota for gut barrier functionality upon interaction with pathogenic or beneficial bacteria is largely unknown. The transcriptome of differently perfused jejunal loops of 12 caesarian-derived pigs, neonatally associated with microbiota of different complexity, was studied. Piglets received pasteurized sow colostrum at birth (d0), a starter microbiota (Lactobacillus amylovorus (LAM), Clostridium glycolicum, and Parabacteroides) on d1-d3, and a placebo inoculant (simple association, SA) or an inoculant consisting of sow's diluted feces (complex association, CA) on d3-d4. On d 26-37, jejunal loops were perfused for 8 h with either enterotoxigenic Escherichia coli F4 (ETEC), purified F4 fimbriae, LAM or saline control (CTRL). Gene expression of each intestinal loop was analyzed by Affymetrix Porcine Gene 1.1_ST array strips. Gene Set Enrichment Analysis was performed on expression values. Compared to CTRL, 184 and 74; 2 and 139; 2 and 48 gene sets, were up- and down-regulated by ETEC, F4 and LAM, respectively. ETEC up-regulated networks related to inflammatory and immune responses, RNA processing, and mitosis. There was a limited overlap in up-regulated gene sets between ETEC and F4 fimbriae. LAM down-regulated genes related to inflammatory and immune responses, as well as to cellular compound metabolism. In CA pigs, 57 gene sets were up-regulated by CA, while 73 were down-regulated compared to SA. CA up-regulated gene sets related to lymphocyte modulation and to cellular defense in all loop perfusions. In CA pigs, compared to SA pigs, genes for chemokine and cytokine activity and for response to external stimuli were down-regulated in ETEC-perfused loops and up-regulated in CTRL. The results highlight the importance of the nature of neonatal microbial colonization in the response to microbial stimuli later in life.Peer reviewe

The effects of starter microbiota and the early life feeding of medium chain triglycerides on the gastric transcriptome profile of 2- or 3-week-old cesarean delivered piglets

Background: The stomach is an underestimated key interface between the ingesta and the digestive system, affecting the digestion and playing an important role in several endocrine functions. The quality of starter microbiota and the early life feeding of medium chain triglycerides may affect porcine gastric maturation. Two trials (T1, T2) were carried out on 12 and 24 cesarean-delivered piglets (birth, d0), divided over two microbiota treatments, but slaughtered and sampled at two or three weeks of age, respectively. All piglets were fed orally: sow serum (T1) or pasteurized sow colostrum (T2) on d0; simple starter microbiota (Lactobacillus amylovorus, Clostridium glycolicum and Parabacteroides spp.) (d1-d3); complex microbiota inoculum (sow diluted feces, CA) or a placebo (simple association, SA) (d3-d4) and milk replacer ad libitum (d0-d4). The The T1 piglets and half of the T2 piglets were then fed a moist diet (CTRL); the remaining half of the T2 piglets were fed the CTRL diet fortified with medium chain triglycerides and 7% coconut oil (MCT). Total mRNA from the oxyntic mucosa was analyzed using Affymetrix</p

Critical review evaluating the pig as a model for human nutritional physiology

The present review examines the pig as a model for physiological studies in human subjects related to nutrient sensing, appetite regulation, gut barrier function, intestinal microbiota and nutritional neuroscience. The nutrient-sensing mechanisms regarding acids (sour), carbohydrates (sweet), glutamic acid (umami) and fatty acids are conserved between humans and pigs. In contrast, pigs show limited perception of high-intensity sweeteners and NaCl and sense a wider array of amino acids than humans. Differences on bitter taste may reflect the adaptation to ecosystems. In relation to appetite regulation, plasma concentrations of cholecystokinin and glucagon-like peptide-1 are similar in pigs and humans, while peptide YY in pigs is ten to twenty times higher and ghrelin two to five times lower than in humans. Pigs are an excellent model for human studies for vagal nerve function related to the hormonal regulation of food intake. Similarly, the study of gut barrier functions reveals conserved defence mechanisms between the two species particularly in functional permeability. However, human data are scant for some of the defence systems and nutritional programming. The pig model has been valuable for studying the changes in human microbiota following nutritional interventions. In particular, the use of human flora-associated pigs is a useful model for infants, but the long-term stability of the implanted human microbiota in pigs remains to be investigated. The similarity of the pig and human brain anatomy and development is paradigmatic. Brain explorations and therapies described in pig, when compared with available human data, highlight their value in nutritional neuroscience, particularly regarding functional neuroimaging techniques

Correlations between plasma proteome profiles and physiological parameters in control pigs fed the two diets.

<p>Number of peaks showing correlation >0.9 between protein peak expression levels and physiological parameters in control animals fed with the Mediterranean or cafeteria diets.a: Non Esterified Fatty Acids; b: Very Low Density Lipoproteins; c: Low Density Lipoproteins; d: High Density Lipoproteins; e: Triglycerides.</p

Plasma proteome profiles comparisons.

<p>Number of protein peaks for each of the protein arrays showing differential expression between the four groups of pigs. It should be noted that some of the peaks may be similar in subsequent fractions.</p