To the extent possible under law, AOP-Wiki has waived all copyright and related or neighboring rights to KER:2028
Inhibition, trypsin leads to Increased monitor peptide
Key Event Relationship Overview
AOPs Referencing Relationship
|AOP Name||Adjacency||Weight of Evidence||Quantitative Understanding||Point of Contact||Author Status||OECD Status|
|Trypsin inhibition leading to pancreatic acinar cell tumors||adjacent||Moderate||Low||Shigeru Hisada (send email)||Under development: Not open for comment. Do not cite||Under Development|
Life Stage Applicability
|All life stages||High|
Key Event Relationship Description
Pancreatic acinar cells secrete digestive enzymes including trypsin into the small intestine.
In rats, one of the pancreatic soluble trypsin inhibitors (TIs), monitor peptide (MP), is simultaneously secreted in the pancreatic juice. MP forms complexes with trypsin in the empty intestine, which keeps the intestinal level of free MP low. Once the gastric contents are transported to the small intestine, secretion of the pancreatic proteases including trypsin and MP is induced, where trypsin is used for protein hydrolysis, and the level of free MP is subsequently increased. The increased MP level stimulates CCK release from I cells lining the small intestinal mucosa via MP receptors, and the resulting increase in CCK stimulates exocrine secretion including MP from the pancreas. Increased MP further stimulates CCK secretion via a positive feedback loop as long as duodenal contents remain to consume trypsin for proteolysis.
After trypsin inhibitors are ingested, the intestinal content of free MP increases rapidly, especially in an empty intestine, via positive feedback regulation.
Evidence Supporting this KER
Trypsin is a digestive enzyme secreted by pancreatic acinar cells that cleaves peptide bonds at the carboxyl end of basic amino acids (lysine and arginine). Secretion of pancreatic digestive enzymes including trypsin is regulated mainly by cholecystokinin (CCK) released from enteroendocrine I cells located in the duodenal mucosa of the small intestine [Wang BJ and Cui ZJ, 2007], and CCK release is controlled by multiple mechanisms [Caron J et al, 2017]. These mechanisms involve feedback regulation of trypsin-sensitive CCK-releasing peptides, one being positive feedback regulation of MP and the other negative feedback regulation of luminal CCK-releasing factor (LCRF) [Miyasaka K and Funakoshi A, 1998; Wang BJ and Cui ZJ, 2007; Guan D et al, 1990].
MP is one of the PSTIs in rats, which stimulates CCK release from duodenal enteroendocrine I cells as well as inhibition of trypsin activity. MP consists of 61 amino acids and has a molecular weight of approximately 6000. MP was first purified from rat pancreatic juice, and its amino acid sequence was subsequently determined [Iwai K et al, 1987; Lin YZ et al, 1990].
MP is bound to trypsin in the empty intestine. Once gastric contents are transported into the small intestine, secretion of the pancreatic proteases with MP is increased, where trypsin instead hydrolyzes these proteins, leading to an increase in the free MP level [Iwai K et al, 1988; Liddle RA, 1995; Graf R, 2006]. The increased level of MP stimulates CCK release from I cells, and then pancreatic exocrine secretion is stimulated [Liddle RA et al, 1992; Guan D et al, 1990; Cuber JC et al, 1990]. It was shown that MP binds to the surface of CCK-immunoreactive mucosal cells of the small intestine [Yamanishi R et al, 1993a; Yamanishi R et al, 1993b].
Following the increased secretion of pancreatic enzymes, proteolysis decreases intestinal protein contents, which once again decreases the intestinal level of free MP due to the excess of trypsin and in turn CCK release [Liddle RA, 1995; Miyasaka K and Funakoshi A, 1998; Graf R, 2006].
When raw soya flour (RSF), which contains trypsin inhibitory activity, or TIs such as camostat are ingested, trypsin activity is inhibited to increase the intestinal level of free MP especially in the empty intestine, followed by an increase in the blood level of CCK [Liddle RA, 1995; Miyasaka K and Funakoshi A, 1998]. TI ingestion-induced increases in blood levels of CCK leads to further CCK release due to increased pancreatic secretion of proteins including MP in a positive feedback manner. On the other hand, TIs may elevate the luminal concentration of LCRF to stimulate CCK release; however, this increase might not be as exaggerated as that of MP, because increased blood level of CCK does not induce further secretion of LCRF.
Uncertainties and Inconsistencies
In normal rats, positive regulation of CCK release by MP seems to require some level of pancreatic secretion before to be effective. In the presence of nutritional protein in the duodenum, trypsin is used for digestion of protein and increased levels of MP stimulates CCK release. On the other hand, after most of the protein is digested, increased free MP might be inactivated with excess of trypsin or other proteases, as follows [Foltz M, 2008]:
1) MP is degraded by trypsin and other proteases.
2) MP forms a complex with trypsin as other PSTIs.
3) MP forms a complex with trypsin, thereafter degraded by proteases.
No study has shown a direct quantitative relationship between MIE and KE1.
No study has reported the time from trypsin inhibition to alteration of intestinal MP content. However, as mentioned above, treatment with trypsin inhibitors or MP increased the plasma concentration of CCK within 30 min in rats.
Known modulating factors
Raw soya flour and trypsin inhibitors such as camostat inhibit trypsin activity, leading to an increase in CCK release from the upper intestine into the bloodstream, where the increased CCK released seems to be mediated by increased luminal concentration of MP due to trypsin inhibition [Green GM and Miyasaka K, 1983; Liddle RA et al, 1984; Goke B et al, 1986; Douglas BR et al, 1989; Cuber JC et al, 1990; Playford RJ et al, 1993; Obourn JD et al, 1997; Tashiro M et al, 2004; Komarnytsky S et al, 2011; Calam J et al, 1987] .
Known Feedforward/Feedback loops influencing this KER
MP stimulates CCK release from intestinal I cells, and the increased CCK level in turn promotes pancreatic acinar cells to secrete pancreatic enzymes including CCK-stimulating MP. Therefore, MP-mediated CCK release is under positive feedback regulation [Liddle RA, 1995; Wang BJ and Cui ZJ, 2007; Chey WY and Chang T, 2001], and the effects of trypsin inhibitors seem robust. As discussed previously, trypsin-sensitive LCRF released from intestinal mucosal cells also stimulate duodenal I cells to release CCK with negative feedback loop.
Domain of Applicability
Isoforms of trypsin are found in many species, for example, cationic and anionic trypsins (trypsins 1 and 2) and mesotrypsin in humans, cationic and anionic trypsins in cows, and anionic trypsin and P23 in rats [Chen JM and Claude Férec C, 2013; Fukuoka S and Nyaruhucha CM, 2002] . Despite differences among species, the three-dimensional structures of the isoforms are highly conserved among species, and the natural substrates for the enzymes are generally any peptide that contains Lys or Arg [Baird Jr TT, 2017]. The active site of trypsin has a specific catalytic triad structure composed of serine, histidine, and aspartate, and the flanking amino acid sequences are entirely conserved [Baird Jr TT and Craik CS, 2013; Baird Jr TT, 2017]. Therefore, trypsin inhibitors have comparable effects on the enzymatic activity of trypsin isoforms among animal species including humans and rats [Savage GP and Morrison SC, 2003].
MP secreted from rat pancreatic acinar cells into the small intestine stimulates I cells of the small intestinal mucosa to release CCK.
MP-like peptides are also found in rats and other mammalian species [Eddeland A and Ohlsson K, 1976]. Rat soluble trypsin inhibitor [Tsuzuki S et al, 1992; Tsuzuki S et al, 1991], human soluble trypsin inhibitor [Pubols MH et al, 1974; Kikuchi N et al, 1985], and bovine soluble trypsin inhibitor [Greene LJ and Giordano JS Jr, 1969; Guy O et al, 1971] are homologous peptides, all of which show trypsin inhibitory activity but no CCK-stimulatory activity [Miyasaka K et al, 1989a; Miyasaka K et al, 1989b; Marchbank T et al, 1998; Voet D and Voet JG, 1995].
1. Baird Jr TT, Craik CS: Trypsin. Academic Press, Cambridge, Massachusetts (pp)2594-2600,2013
2. Baird Jr TT: Trypsin. Elsevier,2017
3. Calam J, Bojarski JC, Springer CJ: Raw soya-bean flour increases cholecystokinin release in man. Br J Nutr 58:175-179,1987
4. Caron J, Domenger D, Dhulster P, Ravallec R, Cudennec B: Protein digestion-derived peptides and the peripheral regulation of food intake. Front Endocrinol (Lausanne) 8:85,2017
5. Chen J-M, Claude Férec C: Human trypsins. Academic Press, Cambridge, Massachusetts (pp) 2600-2609,2013
6. Chey WY, Chang T: Neural hormonal regulation of exocrine pancreatic secretion. Pancreatology 1:320-335,2001
7. Cuber JC, Bernard G, Fushiki T, Bernard C, Yamanishi R, Sugimoto E, Chayvialle JA: Luminal CCK-releasing factors in the isolated vascularly perfused rat duodenojejunum. Am J Physiol 259:G191-197,1990
8. Douglas BR, Woutersen RA, Jansen JB, de Jong AJ, Rovati LC, Lamers CB: Modulation by CR-1409 (lorglumide), a cholecystokinin receptor antagonist, of trypsin inhibitor-enhanced growth of azaserine-induced putative preneoplastic lesions in rat pancreas. Cancer Res 49:2438-2441,1989
9. Eddeland A, Ohlsson K: Purification of canine pancreatic secretory trypsin inhibitor and interaction in vitro with complexes of trypsin-alpha-macroglobulin. Scand J Clin Lab Invest 36:815-820,1976
10. Foltz M, Ansems P, Schwarz J, Tasker MC, Lourbakos A, Gerhardt CC: Protein hydrolysates induce CCK release from enteroendocrine cells and act as partial agonists of the CCK1 receptor. J Agric Food Chem 56:837-843,2008
11. Fukuoka S, Nyaruhucha CM: Expression and functional analysis of rat P23, a gut hormone-inducible isoform of trypsin, reveals its resistance to proteinaceous trypsin inhibitors. Biochim Biophys Acta 1588:106-112,2002
12. Fushiki T, Kajiura H, Fukuoka S, Kido K, Semba T, Iwai K: Evidence for an intraluminal mediator in rat pancreatic enzyme secretion: reconstitution of the pancreatic response with dietary protein, trypsin and the monitor peptide. J Nutr 119:622-627,1989
13. Goke B, Printz H, Koop I, Rausch U, Richter G, Arnold R, Adler G: Endogenous CCK release and pancreatic growth in rats after feeding a proteinase inhibitor (camostate). Pancreas 1:509-515,1986
14. Graf R, Bimmler D: Biochemistry and biology of SPINK-PSTI and monitor peptide.. Endocrinol Metab Clin North Am 35:333-43, ix,2006
15. Green GM, Miyasaka K: Rat pancreatic response to intestinal infusion of intact and hydrolyzed protein. Am J Physiol 245:G394-8,1983
16. Greene LJ, Giordano JS Jr: The structure of the bovine pancreatic secretory trypsin inhibitor--Kazal's inhibitor. I. The isolation and amino acid sequences of the tryptic peptides from reduced aminoethylated inhibitor. J Biol Chem 244:285-298,1969
17. Guan D, Ohta H, Tawil T, Liddle RA, Green GM: CCK-releasing activity of rat intestinal secretion: effect of atropine and comparison with monitor peptide. Pancreas 5:677-684,1990
18. Guy O, Shapanka R, Greene LJ: The structure of the bovine pancreatic secretory trypsin inhibitor--Kazal's inhibitor. 3. Determination of the disulfide bonds and proteolysis by thermolysin. J Biol Chem 246:7740-7747,1971
19. Iwai K, Fukuoka S, Fushiki T, Tsujikawa M, Hirose M, Tsunasawa S, Sakiyama F: Purification and sequencing of a trypsin-sensitive cholecystokinin-releasing peptide from rat pancreatic juice. Its homology with pancreatic secretory trypsin inhibitor. J Biol Chem 262:8956-8959,1987
20. Iwai K, Fushiki T, Fukuoka S: Pancreatic enzyme secretion mediated by novel peptide: monitor peptide hypothesis. Pancreas 3:720-728,1988
21. Kikuchi N, Nagata K, Yoshida N, Ogawa M: The multiplicity of human pancreatic secretory trypsin inhibitor. J Biochem 98:687-694,1985
22. Komarnytsky S, Cook A, Raskin I: Potato protease inhibitors inhibit food intake and increase circulating cholecystokinin levels by a trypsin-dependent mechanism. Int J Obes (Lond) 35:236-243,2011
23. Liddle RA, Goldfine ID, Williams JA: Bioassay of plasma cholecystokinin in rats: effects of food, trypsin inhibitor, and alcohol. Gastroenterology 87:542-549,1984
24. Liddle RA, Misukonis MA, Pacy L, Balber AE: Cholecystokinin cells purified by fluorescence-activated cell sorting respond to monitor peptide with an increase in intracellular calcium.. Proc Natl Acad Sci U S A 89:5147-5151,1992
25. Liddle RA: Regulation of cholecystokinin secretion by intraluminal releasing factors. Am J Physiol 269:G319-27,1995
26. Lin YZ, Isaac DD, Tam JP: Synthesis and properties of cholecystokinin-releasing peptide (monitor peptide), a 61-residue trypsin inhibitor. Int J Pept Protein Res 36:433-439,1990
27. Marchbank T, Freeman TC, Playford RJ: Human pancreatic secretory trypsin inhibitor. Distribution, actions and possible role in mucosal integrity and repair. Digestion 59:167-174,1998
28. Miyasaka K, Nakamura R, Funakoshi A, Kitani K: Stimulatory effect of monitor peptide and human pancreatic secretory trypsin inhibitor on pancreatic secretion and cholecystokinin release in conscious rats. Pancreas 4:139-144,1989a
29. Miyasaka K, Funakoshi A, Nakamura R, Kitani K, Uda K, Murata A, Ogawa M: Differences in stimulatory effects between rat pancreatic secretory trypsin inhibitor-61 and -56 on rat pancreas. Jpn J Physiol 39:891-899,1989b
30. Miyasaka K, Funakoshi A: Luminal feedback regulation, monitor peptide, CCK-releasing peptide, and CCK receptors. Pancreas 16:277-283,1998
31. Obourn JD, Frame SR, Chiu T, Solomn TE, Cook JC: Evidence that A8947 enhances pancreas growth via a trypsin inhibitor mechanism.Toxicol Appl Pharmacol 146:116-126,1997
32. Playford RJ, King AW, Deprez PH, De-Belleroche J, Freeman TC, Calam J: Effects of diet and the cholecystokinin antagonist; devazepide (L364,718) on CCK mRNA, and tissue and plasma CCK concentrations. Eur J Clin Invest 23:641-647,1993
33. Pubols MH, Bartelt DC, Greene LJ: Trypsin inhibitor from human pancreas and pancreatic juice. J Biol Chem 249:2235-2242,1974
34. Savage GP, Morrison SC: Trypsin inhibitors. Elsevier (pp) 5878-5884,2003
35. Tashiro M, Samuelson LC, Liddle RA, Williams JA: Calcineurin mediates pancreatic growth in protease inhibitor-treated mice. Am J Physiol Gastrointest Liver Physiol 286:G784-790,2004
36. Tsuzuki S, Fushiki T, Kondo A, Murayama H, Sugimoto E: Effect of a high-protein diet on the gene expression of a trypsin-sensitive, cholecystokinin-releasing peptide (monitor peptide) in the pancreas. Eur J Biochem 199:245-252,1991
37. Tsuzuki S, Miura Y, Fushiki T, Oomori T, Satoh T, Natori Y, Sugimoto E: Molecular cloning and characterization of genes encoding rat pancreatic cholecystokinin (CCK)-releasing peptide (monitor peptide) and pancreatic secretory trypsin inhibitor (PSTI). Biochim Biophys Acta 1132:199-202,1992
38. Voet D, Voet JG: Biochemistry (2nd ed.). John Wiley & Sons (pp) 396-400,1995
39. Wang BJ, Cui ZJ: How does cholecystokinin stimulate exocrine pancreatic secretion? From birds, rodents, to human.. Am J Physiol Regul Integr Comp Physiol 292:R666-78,2007
40. Yamanishi R, Kotera J, Fushiki T, Soneda T, Iwanaga T, Sugimoto E: Characteristic and localization of the monitor peptide receptor. Biosci Biotechnol Biochem 57:1153-1156,1993a
41. Yamanishi R, Kotera J, Fushiki T, Soneda T, Saitoh T, Oomori T, Satoh T, Sugimoto E: A specific binding of the cholecystokinin-releasing peptide (monitor peptide) to isolated rat small-intestinal cells.. Biochem J 291 ( Pt 1):57-63,1993b