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. 2000 Sep;47(3):423–428. doi: 10.1136/gut.47.3.423

Expression of interleukin 8 (IL-8) and substance P in human chronic pancreatitis

S Di 1, M di 1, F Di 1, G Baccante 1, E Porreca 1, P Innocenti 1, H Friess 1, M Buchler 1
PMCID: PMC1728055  PMID: 10940282

Abstract

BACKGROUND—Changes in substance P content and a relationship between the degree of perineural inflammation and pain has been demonstrated in chronic pancreatitis. Whether a relationship exists between neural alteration and pancreatic inflammation (neurogenic inflammation) is not known.
AIMS—In the present study we evaluated gene expression of preprotachykinin A (PPT-A), the gene encoding substance P, and interleukin 8, a proinflammatory and hyperalgesic mediator whose release is co-regulated by substance P.
PATIENTS—Pancreatic tissue specimens obtained from 21 patients (16 male, five female) with chronic pancreatitis and 18 healthy organ donors (nine male, nine female) were analysed.
METHODS—Gene expression of PPT-A and interleukin 8 was studied by northern blot analysis. Respective proteins were localised using immunohistochemistry.
RESULTS—Northern blot analysis showed that PTT-A mRNA expression levels were present at comparable levels in normal and chronic pancreatitis tissue samples. In contrast, interleukin 8 mRNA was expressed at very low levels in normal controls but was increased 41-fold (p<0.001) in chronic pancreatitis tissue samples. Using immunohistochemistry, interleukin 8 protein was localised mainly in immune cells often found around enlarged pancreatic nerves. In addition, in chronic pancreatitis, intense interleukin 8 immunostaining was present in metaplastic ductal cells of the atrophic pancreatic parenchyma. In chronic pancreatitis samples there was a positive relationship between interleukin 8 mRNA levels and the presence of ductal metaplasia (r=0.795; p<0.001) and the inflammation score (r=0.713; p<0.001).
CONCLUSIONS—Our data indicate that in chronic pancreatitis, the increase in substance P in enlarged pancreatic nerves is not caused by enhanced intrapancreatic PTT-A mRNA expression, suggesting that the location of substance P synthesis is outside of the pancreas. In addition, localisation of interleukin 8 positive immune cells around pancreatic nerves further supports the existence of neuroimmune interactions as a pathophysiological mechanism in chronic pancreatitis.


Keywords: chronic pancreatitis; interleukin 8; neurogenic inflammation; preprotachykinin A; substance P; immunohistochemistry

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Figure 1  .

Figure 1  

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Northern blot analysis. Preprotachykinin A (PPT-A) and interleukin 8 (IL-8) mRNA expression in the normal pancreas (first four lanes) and in tissue samples from patients with chronic pancreatitis (CP) (latter four lanes). PPT-A mRNA was detectable in comparable levels in normal and CP tissue samples. In contrast, enhanced levels of IL-8 mRNA were present in CP samples. The filters were rehybridised with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) cDNA to verify equivalent RNA loading and RNA transfer. PPT-A mRNA migrates as a 1.2 kb band and IL-8 as a 1.8 kb band.

Figure 2  .

Figure 2  

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Densitometric analysis of northern blots. Preprotachykinin A (PPT-A), interleukin 8 (IL-8), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA levels were analysed by laser densitometry in the normal pancreas and in chronic pancreatitis. The ratio of the optical density between PPT-A, IL-8, and the corresponding GAPDH signal was calculated. Bars represent the median with upper and lower quartiles. *p<0.001.

Figure 3  .

Figure 3  

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Immunohistochemical analysis. Interleukin 8 (IL-8) immunohistochemical analysis in the normal pancreas (A) and in chronic pancreatitis (CP) (B-D, F, H). In the normal pancreas (A), no IL-8 immunostaining was detectable. In contrast, in CP tissue sections (B-D, F, H), moderate to intense IL-8 immunoreactivity was found, mainly in the inflammatory cells (D, F, arrows) surrounding enlarged pancreatic nerves. In some cases those nerves were substance P immunoreactive (E). Panels (G) and (H) (arrowheads) show staining of consecutive tissue sections of CP for CD-68 (G) and IL-8 (H). The inserts in (G) and (H) show that IL-8 immunoreactive cells (H) are most probably also stained CD-68 positive (G). In addition, in CP tissue samples, moderate to strong IL-8 immunoreactivity was also found in metaplastic ductal cells (B, C, arrows). n=nerve; d=duct. (A) IL-8 immunostaining; (B- D, F, H) IL-8 immunostaining (patients Nos 14, 16, see table 1); (E) substance P immunostaining (patient No 14); (G) CD-68 immunostaining (patient No 16). Original magnification ×200.

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