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A B S T R A C T Normal human volunteers were intubated with either aspiration tubes or a biopsy capsule placed in the small intestine. The subjects were then fed a test meal containing 50 g of purified bovine serum albumin which served as the model dietary protein.Electrophoretic analysis of intestinal fluids showed that for at least 4 h the fed albumin was detectable in jejunal and ileal fluids. On separate occasions, subjects were fed the same meal without the protein. No protein was detected in intestinal fluids when the protein-free meal was fed. After the protein-rich meal, total concentrations of measured free and peptide amino acids rose from 3.21 to 29.29, and 15.94 to 117.97 ,umol/ml, respectively, (P values <0.02) in the jejunum. Similarly, total concentrations of measured free and peptide amino acids rose from 5.45 to 19.74, and 13.59 to 65.39, respectively, (P values < 0.05) in the ileum. In contrast, concentrations of free and peptide amino acids in intestinal fluids did not increase after the protein-free meal. While intracellular concentrations of amino acids in the jejunal mucosa did not show significant changes, plasma concentrations of each individual free amino acid were increased after the protein-rich meal and were either decreased or unaltered after the proteinfree meal. The amino acid composition of the fed protein was reflected in the increases in intraluminal and plasma concentrations of individual amino acids after the protein-rich meal. It is concluded that after the ingestion of a test meal containing a substantial amount of protein which is within the usual range of dietary

A 30 cm segment of the duodenum, jejunum, or ileum of normal human volunteers was perfused, on separate occasions, with test solutions containing either glycylglycine, free glycine, glycylleucine, or equimolar amounts of free glycine and free leucine. Luminal fluid contained no hydrolytic activity against glycylglycine and minimal activity against glycylleucine. In each intestinal segment, amino acid absorption rates were significantly greater from the test solutions containing the same amount of amino acids in dipeptide than in free form (as high as 185% increase). Perfusion of each intestinal segment with a test solution containing the equimolar mixture of free glycine and free leucine always resulted in a greater leucine than glycine absorption rate. This preferential absorption of leucine, however, was either diminished (jejunum) or almost abolished (duodenum and ileum) when the glycylleucine solution instead of the equimolar mixture was presented to the intestinal mucosa. Among the three segments, the duodenum exhibited the least potential for the disappearance of dipeptides. The jejunal and ileal dipeptide disappearance rates were either similar for glycylleucine (94% vs. 92%) or slightly different for glycylglycine (92% vs. 79%). Despite lack of a remarkable difference in the disappearance rates, absorption rates of constituent amino acids were markedly greater in the jejunum than in the ileum. This reduced amino acid absorption was brought about by a greater accumulation of free amino acids in the lumen of the ileal segment in the lumen than when the glycylglycine test solution did not contain free leucine. Similarly, inhibition of free glycine and free leucine absorption by isoleucine was not accompanied by any remarkable alteration of absorption rates of the constituent amino acids of glycylleucine. The results of these studies suggest that: (a) dipeptide disappearance in the gut lumen is principally accomplished by intact absorption and not by hydrolysis; (b) intracellular hydrolysis of dipeptides is markedly greater in the ileum than in the jejunum, while dipeptide absorption rates are either similar or only slightly different in these two segments; (c) there is no appreciable hydrolysis of glycylglycine by the membrane-bound enzymes and only a small fraction of glycylleucine is hydrolyzed by these enzymes.

The abundance of the oligopeptide transporter (Pept-1) in the brush-border membrane of the intestinal epithelium is the central mechanism for regulation of transport of products of protein digestion (dipeptides and tripeptides) and peptidomimetic drugs (for example, β-lactam antibiotics). Within the past few years, there has been substantial progress in identifying the factors controlling this regulation and the mechanisms of their actions. The purpose of this report is to review this progress. The studies of individual substrates and hormones in a human intestinal cell line (Caco-2) have shown that dipeptides, certain amino acids, insulin, and leptin increase and epidermal growth factor and triiodothyronine decrease the membrane population of Pept-1. In the case of dipeptides, epidermal growth factor, and thyroid hormone, there are parallel changes in the gene expression brought about by alteration of transcription and/or stability of Pept-1 mRNA. In contrast, the treatment with insulin and leptin does not induce any alteration in the Pept-1 gene expression, and the mechanism of increased protein expression appears to be increased trafficking from a preformed cytoplasmic pool to the apical membrane. In vivo studies in rats have shown modulation of protein and gene expressions of the intestinal oligopeptide transporter during the day and during development and in nutritional and metabolic alterations, such as high-protein diet, fasting, and diabetes. Patients with intestinal diseases, such as ulcerative colitis, Crohn's disease, and short-bowel syndrome, may have induction of the Pept-1 expression in their colon. Finally, pharmacological studies have shown that the expression of Pept-1 can be upregulated by agents such as 5 fluorouracil and downregulated by agents such as cyclosporine. In conclusion, the above studies have produced a wealth of new information on regulation of a key transporter in the intestine. This information may have useful applications in nutritional and pharmacological treatments, for example, in diabetic patients needing enteral nutrition or in ulcerative colitis patients needing the suppression of the intestinal inflammation.

The intestinal oligopeptide transporter (cloned as Pept-1) has major roles in protein nutrition and drug therapy. A key unstudied question is whether expression of Pept-1 is hormonally regulated. In this experiment, we investigated whether insulin has such a role. We used a human intestinal cell monolayer (Caco-2) as the in vitro model of human small intestine and glycylglutamine (Gly-Gln) as the model substrate for Pept-1. Results showed that addition of insulin at a physiological concentration (5 nM) to incubation medium greatly stimulates Gly-Gln uptake by Caco-2 cells. This stimulation was blocked when genistein, an inhibitor of tyrosine kinase, was added to incubation medium. Studies of the mechanism of insulin stimulation showed the following. 1) Stimulation occurred promptly (30–60 min) after exposure to insulin. 2) There was no significant change in the Michaelis-Menten constant of Gly-Gln transport, but there was a nearly twofold increase in its maximal velocity. 3) Insulin effect persisted even when Golgi apparatus, which is involved in trafficking of newly synthesized Pept-1, was dismantled. 4) However, there was complete elimination of insulin effect by disruption of microtubules involved in trafficking of preformed Pept-1. 5) Finally, with insulin treatment, there was no change in Pept-1 gene expression, but the amount of Pept-1 protein in the apical membrane was increased. In conclusion, the results show that insulin, when it binds to its receptor, stimulates Gly-Gln uptake by Caco-2 cells by increasing the membrane population of Pept-1. The mechanism appears to be increased translocation of this transporter from a preformed cytoplasmic pool.