Introduction
The possibility that a novel stomach pentadecapeptide BPC 157, (thought to be essential for beneficial activity of an organoprotective protein isolated from human gastric juice (named BPC)) and which has noted beneficial effects (Sikiric et al., 1993b, Sikiric et al., 1994, Sikiric et al., 1996a, Sikiric et al., 1996b, Sikiric et al., 1997a, Sikiric et al., 1997b; Seiwerth et al., 1997; Grabarevic et al., 1997) could interact with NO was investigated.
Based on its salutary effect on ethanol-induced lesions and on other lesions gastrointestinal lesions induced by various challengers (i.e., prolonged stress, nonsteroidal anti-inflammatory analgetics, cystamine, trinitrobenzene sulfonic acid, dinitrofluorbenzene sulfonic acid, acute pancreatitis, somatosensory neurons depletion) (Sikiric et al., 1993b, Sikiric et al., 1994, Sikiric et al., 1996a, Sikiric et al., 1996b, Sikiric et al., 1997a, Sikiric et al., 1997b; Veljaca et al., 1994a, Veljaca et al., 1994b, Veljaca et al., 1995b, Paré and Klucyznski, 1994; Sandor et al., 1996) BPC was recently claimed to have a `cytoprotective' effect, particularly on the gastrointestinal intestinal mucosa, along with a particular beneficial effect on endothelium integrity (Sikiric et al., 1993b, Sikiric et al., 1994). Considering the existence of BPC-157 in gastric juice and its unusual stability (Sikiric et al., 1993b) (i.e., incubated in human gastric juice or in water, this pentadecapeptide is not subjected to degradation at least for 24 h (Veljaca et al., 1995a)) it is likely that this peptide has some biological actions. It is suggested that BPC, which is formed constitutively in the gastric mucosa (body) and which is present in gastric juice (Sikiric et al., 1993a, Sikiric et al., 1993b, Sikiric et al., 1993c, Sikiric et al., 1994, Sikiric et al., 1996a, Sikiric et al., 1996b, Sikiric et al., 1997a, Sikiric et al., 1997b, Sikiric et al., 1997c), protect the stomach against injury. Evidence shows that this pentadecapeptide has a strong anti-inflammatory activity in both acute and chronic inflammation models (Sikiric et al., 1993a, Sikiric et al., 1997c), reduces the release of inflammatory mediators (i.e., myeloperoxidase, leukotriene B4, tromboxane B2) in vitro and in vivo (Veljaca et al., 1994a, Veljaca et al., 1994b, Veljaca et al., 1995b), markedly attenuates various experimental lesions in other organs (i.e., liver (Sikiric et al., 1993c), pancreas (Sikiric et al., 1996b), heart (Sikiric et al., 1993b; Bosnjak et al., 1994; Grabarevic et al., 1997)) and promotes wound and fracture healing in rats (Sikiric et al., 1993b, Seiwerth et al., 1997). Interestingly, an effect on disturbed blood pressure (e.g., Goldblatt's hypertension) has also been noted (Sikiric et al., 1993b). However, the mechanism remains elusive. A complex interaction with adrenergic and dopaminergic systems has been reported (Sikiric et al., 1997a). The release of this pentadecapeptide from somatosensory neurones is suggested, based on experiments with neurotoxin capsaicin, in a nociception assay (Sikiric et al., 1993a) and nasal (Kalogjera et al., 1997) and intestinal (Sikiric et al., 1996a) lesions studies. Since neuropeptides originating from the afferent sensory neurones in the vicinity of the microvessels are involved in the regulation of the release of the endothelium-derived mediators (Whittle et al., 1992), it was reasonable to investigate whether the effects of this novel stomach pentadecapeptide, BPC, could be a result of its interaction with nitric oxide (NO).
Extension of NO studies (usually limited to blunted generation of NO) and investigation of the influence of this novel gastric pentadecapeptide, under conditions of both a blunted generation of NO and NO substrate application, appear to be a rather logical approach in light of the widely suggested importance of NO as an essential signalling system in the gastrointestinal and cardiovascular systems (for review, see Moncada et al., 1991; Whittle et al., 1992). Bearing in mind the recognized dual role of NO, both an inhibition and an uncontrolled excess of NO would lead to significant damage (Lopez-Belmonte et al., 1993; Whittle et al., 1992). A beneficial effect was suggested for l-arginine, the precursor of NO (for review see Whittle et al., 1992), but agents which liberate NO are ulcerogenic when given in the higher doses (Lopez-Belmonte et al., 1993). l-arginine analogues and NO generation inhibitors are consistently claimed to be ulcerogenic (for review see Whittle et al., 1992) but bicarbonate secretion could be enhanced by these agents (Takeuchi et al., 1993; Takeuchi and Okabe, 1994), an effect also associated with an increase in systemic blood pressure (Takeuchi and Okabe, 1994). Finally, besides being essential for blood pressure maintenance, induction of NO synthase seems to be responsible for severe shock (Moncada et al., 1991). Thus, this investigation could help to further elucidate a variety of NO interactions.
Therefore, in addition to studying the interplay of NO/BPC 157 in the maintenance of stomach mucosal integrity, the effect of BPC 157 application on the blood pressure changes induced by a potent endothelial NO synthesis inhibitor NG-nitro-l-arginine methylester (l-NAME) (Whittle et al., 1992) and a NO precursor, l-arginine (Moncada et al., 1991), are an additional focus in the present report. In addition, using the biochemical techniques described before by Whittle et al. (1992), the influence of this pentadecapeptide (along with l-arginine or l-NAME, given alone, or in combination) on the activity of NO synthase in homogenate supernatants of gastric mucosa from the rat stomach was investigated. To emphasize the experimental rationale for the present study, the NO system has to date not challenged by simultaneously testing the effect of an agent on the changes induced by l-arginine as well as l-NAME, given both separately and together, even though this dual activity of the NO system is widely recognized (for review, see Moncada et al., 1991; Whittle et al., 1992), at least from a theoretical point of view.
Section snippets
Drugs
The pentadecapeptide BPC157 (GlyGluProProProGlyLysProAlaAspAspAlaGlyLeuVal, M.W. 1419), a partial sequence of human gastric juice peptide BPC (Sikiric et al., 1993a, Sikiric et al., 1993b, Sikiric et al., 1993c, Sikiric et al., 1994, Sikiric et al., 1996a, Sikiric et al., 1996b), is freely soluble in water at pH 7.0 and in saline and was prepared as described before (Sikiric et al., 1993a, Sikiric et al., 1993b, Sikiric et al., 1993c, Sikiric et al., 1994, Sikiric et al., 1996a, Sikiric et al.,
Results
Methodologically, given the complex dual activity of NO (Moncada et al., 1991; Whittle et al., 1992), investigation of the various changes induced by BPC 157 added together with agents that affect the NO system in opposite ways, a simple way to assess the efficacy of the test drug. Such a protocol has not been previously applied in NO studies.
Discussion
Theoretically, the use of agents that positively or negatively affect the NO system, given separately or in combination, would be advantageous for a through determination of the possible relation of NO with other agents and systems. Although to date this approach has not been used extensively (for review, see, e.g., Moncada et al., 1991; Whittle et al., 1992), it seemed to us that this approach could be particularly helpful against the background of the suggested dual role of NO.
The known
Acknowledgements
This study was subsidized by grants from the Ministry of Science of Republic of Croatia and Pliva, Zagreb, Croatia.