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. 1997 Aug;41(2):187–194. doi: 10.1136/gut.41.2.187

Gastric ulcer healing in the rat: kinetics and localisation of de novo procollagen synthesis

M Shahin 1, A Gillessen 1, T Pohle 1, C Weber 1, D Schuppan 1, H Herbst 1, W Domschke 1
PMCID: PMC1891472  PMID: 9301497

Abstract

Background and aims—To gain further insight into the role of the extracellular matrix during healing of peptic ulcers, sequential changes of procollagen expression were studied over 30 days of ulcer healing. 
Materials and methods—Procollagens α1(I), α1(III), and α1(IV) RNA and their polypeptides were assessed in acetic acid induced rat gastric ulcers by in situ hybridisation and immunohistochemistry. 
Results—Three days after ulcer induction, intense hybridisation signals were obtained with all probes, with procollagen α1(I) showing the highest transcript levels. Procollagen gene expression remained elevated up to day 15, but was reduced to initial low levels on day 30. Immunohistochemical staining documented increased deposition of the three procollagen types parallel to their respective transcript levels, again with type I showing the earliest and the most prominent deposits. The highest procollagen transcript levels were found in the intact submucosa surrounding the ulcer margins, followed by the muscularis propria and the serosa, with the lamina propria exhibiting the lowest transcript levels. 
Conclusion—The procollagens studied are regulated differentially at the transcriptional and post-transcriptional levels. The early onset and long duration of procollagen expression as well as the involvement of all layers of the gastric wall points to their central structural and functional role in gastric ulcer healing. 



Keywords: gastric ulcer; in situ hybridisation; procollagen RNA

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Figure 1 .

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: (A) Fully developed rat antral ulcer with overhanging edges three days after induction with acetic acid. The ulcer penetrates the muscularis mucosae as well as the muscularis propria. The ulcer floor is occupied by necrotic debris and a round cell infiltrate. (B) Complete mucosal re-epithelialisation on day 30 with marked cystic dilatation of the regenerated glands and the muscularis mucosae still disrupted. (A) Original magnification ×20; (B) ×100. 


Figure 2 .

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: In situ hybridisation with the 35S labelled α1(I) procollagen antisense RNA probe. The gastric mucosa of a sham operated rat (A) shows only minimal signal; in the gastric tissue contralateral to the ulcer (B) the autoradiographic signals are obviously increased. Original magnification ×240. 


Figure 3 .

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: In situ hybridisation with the 35S labelled α1(I) procollagen antisense RNA probe on rat gastric ulcer. There is strong cellular labelling predominantly in the submucosa (A) as well as around the muscle bundles at the ulcer margin indicating a remarkable up regulation of collagen synthesis as early as day 3 after ulcer induction. (B) Magnification shows clustering of the autoradiographic signals in the submucosa. (C) A slightly weaker signal is observed at the ulcer base. (D) Parallel section hybridised with a 35S labelled sense RNA probe for α1(I) procollagen showing only a few background signals. Original magnification: (A) ×60; (B,C,D) ×240. 


Figure 4 .

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: In situ hybridisation with the 35S labelled α1(I) procollagen antisense RNA probe on a rat gastric ulcer (12 days after induction). There is ubiquitous distribution of cells expressing RNA in cellular sheets along the muscle fibres of the muscularis propria (A) or within the interglandular stroma of the lamina propria (B). Original magnification: (A) ×100; (B) ×280. 


Figure 5 .

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: In situ hybridisation with the 35S labelled α1(III) (A) and α1(IV) procollagen (B,C) antisense RNA probes on a rat gastric ulcer three days after induction. (A) Compared with α1(I) procollagen the density of the silver grains in the submucosal cells is slightly weaker. α1(IV) procollagen gene transcripts are enhanced within mesenchymal cells of the submucosa (B), and in the endomysial vascular connective tissue (C). Original magnification: ×280. 


Figure 6 .

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: Immunostaining for collagens type I and III (APAAP technique). (A) Intense collagen specific staining (red colour) is already observed on day 3 after ulcer induction both in the submucosa and the base (b) of the ulcer margin (m). (B) On day 7 substantial deposits of collagen type III are detected in the lamina propria and at the ulcer base (b). Original magnification: ×60. 


Figure 7 .

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: Immunostaining for type IV procollagen (APAAP technique). (A) On day 3 after ulcer induction there is deposition around blood vessels (b.v.) and in the basement membranes of the gastric glands and around the inflammatory cellular infiltrate at the ulcer rim. (B) With progression of ulcer healing dense procollagen IV deposition is observed in the stromal tissue of the lamina propria and of the submucosa. Original magnification: ×60. 


Figure 8 .

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: Immunostaining for type I collagen (APAAP technique) on day 7 after ulcer induction. Only faint collagen type I deposition is found in the muscularis mucosae (m.m.) or propria (m.p.); (b) indicates ulcer base. Original magnification: ×60. 


Figure 9 .

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: Immunostaining (APAAP technique) with non-immune serum in a three day old negative control ulcer specimen. Original magnification: ×60. 


Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Becker J., Schuppan D., Benzian H., Bals T., Hahn E. G., Cantaluppi C., Reichart P. Immunohistochemical distribution of collagens types IV, V, and VI and of pro-collagens types I and III in human alveolar bone and dentine. J Histochem Cytochem. 1986 Nov;34(11):1417–1429. doi: 10.1177/34.11.3772076. [DOI] [PubMed] [Google Scholar]
  2. Ernst H., Grunert S., Schneider H. T., Beck W. S., Brune K., Hahn E. G. Distribution of extracellular matrix proteins in indomethacin-induced lesions in the rat stomach. Scand J Gastroenterol. 1995 Sep;30(9):847–853. doi: 10.3109/00365529509101590. [DOI] [PubMed] [Google Scholar]
  3. Genovese C., Rowe D., Kream B. Construction of DNA sequences complementary to rat alpha 1 and alpha 2 collagen mRNA and their use in studying the regulation of type I collagen synthesis by 1,25-dihydroxyvitamin D. Biochemistry. 1984 Dec 4;23(25):6210–6216. doi: 10.1021/bi00320a049. [DOI] [PubMed] [Google Scholar]
  4. Gillessen A., Shahin M., Pohle T., Foerster E., Domschke W. Collagen mRNA and fibronectin are increased in healing gastric ulcers in man. J Physiol Pharmacol. 1995 Mar;46(1):57–62. [PubMed] [Google Scholar]
  5. Gillessen A., Shahin M., Pohle T., Foerster E., Krieg T. H., Domschke W. Evidence of de novo collagen synthesis in healing human gastric ulcers. Scand J Gastroenterol. 1995 Jun;30(6):515–518. doi: 10.3109/00365529509089782. [DOI] [PubMed] [Google Scholar]
  6. Gillessen A., Voss B., Rauterberg J., Domschke W. Distribution of collagen types I, III, and IV in peptic ulcer and normal gastric mucosa in man. Scand J Gastroenterol. 1993 Aug;28(8):688–689. doi: 10.3109/00365529309098273. [DOI] [PubMed] [Google Scholar]
  7. Golik A., Modai D., Gluskin I., Schechter I., Cohen N., Eshchar J. Aeromonas in adult diarrhea: an enteropathogen or an innocent bystander? J Clin Gastroenterol. 1990 Apr;12(2):148–152. doi: 10.1097/00004836-199004000-00007. [DOI] [PubMed] [Google Scholar]
  8. Ishizaki M., Zhu G., Haseba T., Shafer S. S., Kao W. W. Expression of collagen I, smooth muscle alpha-actin, and vimentin during the healing of alkali-burned and lacerated corneas. Invest Ophthalmol Vis Sci. 1993 Nov;34(12):3320–3328. [PubMed] [Google Scholar]
  9. Liau G., Yamada Y., de Crombrugghe B. Coordinate regulation of the levels of type III and type I collagen mRNA in most but not all mouse fibroblasts. J Biol Chem. 1985 Jan 10;260(1):531–536. [PubMed] [Google Scholar]
  10. Martens M. F., Huyben C. M., Hendriks T. Collagen synthesis in fibroblasts from human colon: regulatory aspects and differences with skin fibroblasts. Gut. 1992 Dec;33(12):1664–1670. doi: 10.1136/gut.33.12.1664. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Maruoka A., Fujishima H., Misawa T., Chijiiwa Y., Nawata H. Evaluation of acetic acid-induced gastric ulcers in dogs by endoscopic ultrasonography. Scand J Gastroenterol. 1993 Dec;28(12):1055–1061. doi: 10.3109/00365529309098309. [DOI] [PubMed] [Google Scholar]
  12. Matthes H., Herbst H., Schuppan D., Stallmach A., Milani S., Stein H., Riecken E. O. Cellular localization of procollagen gene transcripts in inflammatory bowel diseases. Gastroenterology. 1992 Feb;102(2):431–442. doi: 10.1016/0016-5085(92)90087-f. [DOI] [PubMed] [Google Scholar]
  13. Mauch C., Hatamochi A., Scharffetter K., Krieg T. Regulation of collagen synthesis in fibroblasts within a three-dimensional collagen gel. Exp Cell Res. 1988 Oct;178(2):493–503. doi: 10.1016/0014-4827(88)90417-x. [DOI] [PubMed] [Google Scholar]
  14. Mikami H., Watanabe S., Hirose M., Sato N. Role of extracellular matrix in wound repair by cultured gastric mucosal cells. Biochem Biophys Res Commun. 1994 Jul 15;202(1):285–292. doi: 10.1006/bbrc.1994.1925. [DOI] [PubMed] [Google Scholar]
  15. Milani S., Herbst H., Schuppan D., Hahn E. G., Stein H. In situ hybridization for procollagen types I, III and IV mRNA in normal and fibrotic rat liver: evidence for predominant expression in nonparenchymal liver cells. Hepatology. 1989 Jul;10(1):84–92. doi: 10.1002/hep.1840100117. [DOI] [PubMed] [Google Scholar]
  16. Milani S., Herbst H., Schuppan D., Kim K. Y., Riecken E. O., Stein H. Procollagen expression by nonparenchymal rat liver cells in experimental biliary fibrosis. Gastroenterology. 1990 Jan;98(1):175–184. doi: 10.1016/0016-5085(90)91307-r. [DOI] [PubMed] [Google Scholar]
  17. Nath P., Laurent M., Horn E., Sobel M. E., Zon G., Vogeli G. Isolation of an alpha 1 type-IV collagen cDNA clone using a synthetic oligodeoxynucleotide. Gene. 1986;43(3):301–304. doi: 10.1016/0378-1119(86)90220-9. [DOI] [PubMed] [Google Scholar]
  18. Nusgens B., Merrill C., Lapiere C., Bell E. Collagen biosynthesis by cells in a tissue equivalent matrix in vitro. Coll Relat Res. 1984 Oct;4(5):351–363. doi: 10.1016/s0174-173x(84)80003-5. [DOI] [PubMed] [Google Scholar]
  19. Okabe S., Roth J. L., Pfeiffer C. J. A method for experimental, penetrating gastric and duodenal ulcers in rats. Observations on normal healing. Am J Dig Dis. 1971 Mar;16(3):277–284. doi: 10.1007/BF02235252. [DOI] [PubMed] [Google Scholar]
  20. Reubi J. C., Waser B., Schmassmann A., Halter F. Persistent lack of somatostatin receptors in gastric mucosa of healing ulcers in rats. Gastroenterology. 1994 Aug;107(2):339–346. doi: 10.1016/0016-5085(94)90157-0. [DOI] [PubMed] [Google Scholar]
  21. Schuppan D., Cantaluppi M. C., Becker J., Veit A., Bunte T., Troyer D., Schuppan F., Schmid M., Ackermann R., Hahn E. G. Undulin, an extracellular matrix glycoprotein associated with collagen fibrils. J Biol Chem. 1990 May 25;265(15):8823–8832. [PubMed] [Google Scholar]
  22. Takemoto T., Namiki M., Ishikawa M., Tsuneoka K., Oshiba S., Kawai K., Ogawa N. Ranitidine and sucralfate as maintenance therapy for gastric ulcer disease: endoscopic control and assessment of scarring. Gut. 1989 Dec;30(12):1692–1697. doi: 10.1136/gut.30.12.1692. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Tarnawski A., Douglass T. G., Stachura J., Krause W. J. Quality of gastric ulcer healing: histological and ultrastructural assessment. Aliment Pharmacol Ther. 1991;5 (Suppl 1):79–90. doi: 10.1111/j.1365-2036.1991.tb00751.x. [DOI] [PubMed] [Google Scholar]
  24. Völker W., Böhm A., Schmidt A., Svahn C. M., Gellerbring A. K., Mattsson C., Ekvärn S., Robenek H., Buddecke E. Inhibition of smooth muscle cell proliferation and neointimal growth by low-anticoagulant heparin. Arzneimittelforschung. 1995 May;45(5):546–550. [PubMed] [Google Scholar]

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