Printed in U.S.A. Copyright © 1997 by The Endocrine Society Circulating and Tissue Forms of the Intestinal Growth Factor, Glucagon-Like Peptide-2*

Glucagon-like peptide-2 (GLP-2) has recently been identified as a stimulator of intestinal epithelial growth, prompting the development of RIA and HPLC methodologies to study this peptide in more detail. A GLP-2-specific antiserum (UTTH-7) was developed that recognizes amino acids 25-30 of human and rat GLP-2-(1-33). UTTH-7 cross-reacts with N- and C-terminally modified forms of GLP-2, proglucagon, and the major proglucagon fragment. Analysis of rat ileal extracts demonstrated the presence of GLP-2-(1-33) as well as significant amounts of GLP-2-(3-33) (16 +/- 7% of total GLP-2). The level of total immunoreactive GLP-2 in plasma from fasted rats was 700 +/- 71 pg/ml, and this increased 3.6-fold (P < 0.001) in 24-h fed rats. HPLC analysis demonstrated the presence of both GLP-2-(1-33) and GLP-2-(3-33) in plasma from fasted rats, with increments in both peptides in plasma from fed rats. Immunoreactive GLP-2 increased in plasma from human subjects 2 h after a meal, rising from 851 +/- 230 to 1106 +/- 211 pg/ml (P < 0.05); 15 +/- 4% of this immunoreactivity was accounted for by the presence of intact GLP-2. HPLC showed the presence of both GLP-2-(1-33) and GLP-2-(3-33) in plasma from fed humans. Incubation of human GLP-2-(1-33) with the enzyme dipeptidylpeptidase IV resulted in liberation of GLP-2-(3-33), whereas replacement of Ala2 with Gly2 prevented this cleavage. Thus, while GLP-2-(1-33) is a major circulating and tissue form of GLP-2, GLP-2-(3-33) is a significant component ofimmunoreactive GLP-2 in both intestine and plasma.

[1]  D. Drucker,et al.  Regulation of the biological activity of glucagon-like peptide 2 in vivo by dipeptidyl peptidase IV , 1997, Nature Biotechnology.

[2]  P. Brubaker,et al.  Proglucagon processing in islet and intestinal cell lines , 1996, Regulatory Peptides.

[3]  R. Pederson,et al.  Degradation of glucose-dependent insulinotropic polypeptide and truncated glucagon-like peptide 1 in vitro and in vivo by dipeptidyl peptidase IV. , 1995, Endocrinology.

[4]  V. Marks,et al.  Glucagon-like peptide-1 (7-36)amide and glucose-dependent insulinotropic polypeptide secretion in response to nutrient ingestion in man: acute post-prandial and 24-h secretion patterns. , 1993, The Journal of endocrinology.

[5]  J. Holst,et al.  Both Subcutaneously and Intravenously Administered Glucagon-Like Peptide I Are Rapidly Degraded From the NH2-Terminus in Type II Diabetic Patients and in Healthy Subjects , 1995, Diabetes.

[6]  D. Drucker,et al.  Biological determinants of intestinotrophic properties of GLP-2 in vivo. , 1997, The American journal of physiology.

[7]  J. Holst,et al.  Carboxypeptidase‐B‐like processing of the C‐terminus of glucagon‐like peptide‐2 in pig and human small intestine , 1989, FEBS letters.

[8]  G. Géraud,et al.  Dipeptidyl peptidase IV expression in rat jejunal crypt-villus axis is controlled at mRNA level. , 1991, The American journal of physiology.

[9]  O. H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[10]  P. Blache,et al.  Metabolic clearance rates of oxyntomodulin and glucagon in the rat: contribution of the kidney , 1990, Regulatory Peptides.

[11]  L. Frohman,et al.  Rapid enzymatic degradation of growth hormone-releasing hormone by plasma in vitro and in vivo to a biologically inactive product cleaved at the NH2 terminus. , 1986, The Journal of clinical investigation.

[12]  G. Weir,et al.  Insulinotropin: glucagon-like peptide I (7-37) co-encoded in the glucagon gene is a potent stimulator of insulin release in the perfused rat pancreas. , 1987, The Journal of clinical investigation.

[13]  T. Yoshimoto,et al.  Post-proline cleaving enzyme. Synthesis of a new fluorogenic substrate and distribution of the endopeptidase in rat tissues and body fluids of man. , 1979, Biochimica et biophysica acta.

[14]  D. Drucker,et al.  Tissue-specific differences in the levels of proglucagon-derived peptides in streptozotocin-induced diabetes. , 1989, Endocrinology.

[15]  B. Gallwitz,et al.  Dipeptidyl-peptidase IV hydrolyses gastric inhibitory polypeptide, glucagon-like peptide-1(7-36)amide, peptide histidine methionine and is responsible for their degradation in human serum. , 1993, European journal of biochemistry.

[16]  S. Bloom,et al.  GLUCAGON-LIKE PEPTIDE-1 7-36: A PHYSIOLOGICAL INCRETIN IN MAN , 1987, The Lancet.

[17]  J. Fahrenkrug,et al.  Preparation of 125I-labeled synthetic porcine secretin for radioimmunoassay. , 1976, Scandinavian journal of clinical and laboratory investigation.

[18]  D. Swallow,et al.  Regional expression of epithelial dipeptidyl peptidase IV in the human intestines. , 1994, Biochemical and biophysical research communications.

[19]  J. López-Novoa,et al.  Renal catabolism of human glucagon-like peptides 1 and 2. , 1990, Canadian journal of physiology and pharmacology.

[20]  J. Holst,et al.  Degradation of glucagon-like peptide-1 by human plasma in vitro yields an N-terminally truncated peptide that is a major endogenous metabolite in vivo. , 1995, The Journal of clinical endocrinology and metabolism.

[21]  L. Orci,et al.  Preproglucagon gene expression in pancreas and intestine diversifies at the level of post-translational processing. , 1986, The Journal of biological chemistry.

[22]  J. Holst,et al.  Glucagon-like peptides GLP-1 and GLP-2, predicted products of the glucagon gene, are secreted separately from pig small intestine but not pancreas. , 1986, Endocrinology.

[23]  S. Bloom,et al.  Molecular forms of glucagon‐like peptides in man , 1985, FEBS letters.

[24]  N. Seidah,et al.  Role of prohormone convertases in the tissue-specific processing of proglucagon. , 1996, Molecular endocrinology.

[25]  J. Holst,et al.  Radio-immunoassays for glucagon-like peptides 1 and 2 (GLP-1 and GLP-2). , 1987, Scandinavian journal of clinical and laboratory investigation.

[26]  L. Frohman,et al.  Dipeptidylpeptidase IV and trypsin-like enzymatic degradation of human growth hormone-releasing hormone in plasma. , 1989, The Journal of clinical investigation.

[27]  T. Steinmetzer,et al.  Dipeptidyl peptidase IV in the immune system. Effects of specific enzyme inhibitors on activity of dipeptidyl peptidase IV and proliferation of human lymphocytes. , 1991, Biological chemistry Hoppe-Seyler.

[28]  D. Drucker,et al.  Intestinal function in mice with small bowel growth induced by glucagon-like peptide-2. , 1997, The American journal of physiology.

[29]  J. Holst,et al.  Antidiabetogenic effect of glucagon-like peptide-1 (7-36)amide in normal subjects and patients with diabetes mellitus. , 1992, The New England journal of medicine.

[30]  D. Drucker,et al.  Cell-specific post-translational processing of preproglucagon expressed from a metallothionein-glucagon fusion gene. , 1986, The Journal of biological chemistry.

[31]  D. Drucker,et al.  Induction of intestinal epithelial proliferation by glucagon-like peptide 2. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[32]  H. Bennett,et al.  Isolation and characterization of corticotropin- and melanotropin-related peptides from the neurointermediary lobe of the rat pituitary by reversed-phase liquid chromatography. , 1981, Biochemistry.

[33]  J. Holst,et al.  Tissue and Plasma Concentrations of Amidated and Glycine-Extended Glucagon-Like Peptide I in Humans , 1994, Diabetes.

[34]  J. Holst,et al.  Naturally occurring products of proglucagon 111-160 in the porcine and human small intestine. , 1988, The Journal of biological chemistry.