Disease models MOLECULAR MEDICINE TODAY, NOVEMBER 1999 (VOL. 5)

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Helicobacter pylori is a major global pathogen,causing up to 90% of duodenal ulcers, 70% ofgastric ulcers and, if appropriate co-factors arepresent, most cases of gastric adenocarcinoma.Yet the pathogenicity of these diseases remainspoorly understood1 and there is no animal modelthat completely mimics them. However, a widerange of models are available which have allowedinvestigation of some key areas of pathogenesis,such as colonization2 (Table 1).

It is seldom appreciated that, although the colo-nization patterns of Helicobacter species in animalmodels might approximate those seen in humans,the pathologies and asymptomatic disease are,with a few exceptions, not the same. Gastritis is theessential precursor lesion to serious disease inhumans. In general, active on chronic gastritis oc-curs in the antrum of the stomach and, in some pop-ulations, extends into the corpus. The principal in-dicator of severe disease is the degree ofinflammation, as shown by the presence of neu-trophils. This is greater in people infected withstrains of H. pylori that carry a pathogenicity island(PAI) called cag, which induces high levels of theproinflammatory cytokine IL-8 (Ref. 3). Neutrophilactivity is rarely seen in the animal models and cor-

relation with the marker gene for the PAI, cagA, isnot seen. Indeed, the most-severe gastritis in themouse is seen with the cagA2 bacterium Helico-bacter felis and this is restricted to the body mucosaaway from areas of maximum bacterial density,suggesting an indirect effect4. The same is seenwith the mouse-adapted Sydney strain of H. pylori(Fig. 1), although inflammation is much milder andtakes longer to develop5. In most of the other ani-mal models, the inflammation is more chronic thanactive, and mononuclear cells rather than neu-trophils predominate. Exceptions are the guineapig, which has an IL-8 homologue6, and theMongolian gerbil, which exhibits an extremely se-vere active and chronic destructive gastritis in thecorpus and antrum of the stomach7. Indeed, gas-tritis in the gerbil model is unlike that seen in thehuman in that it is much more extreme.

UlcersUlcers are rarely seen in the animal models andare not consistently introduced by infection alone.Gastric ulcers have been reported in ferrets,where they are assumed to be caused by Helico-bacter mustelae. In their Helicobacter heilmanniimouse model, Eaton reported gastric ulcers if an

abrasive diet was given8. The one animal modelthat does develop gastric ulcers consistently isthe H. pylori-infected Mongolian gerbil7,9 (Fig. 2).Duodenal ulcers have not been reported in any ofthe animal models of Helicobacter infection.

Gastric cancerOne of the more negative aspects of the importantreport on H. pylori and gastric cancer from the Inter-national Agency on Cancer Research was that thepanel had to rely on epidemiological data. They con-cluded that the evidence in animal models was notsufficient to conclude that H. pylori was a causeof gastric adenocarcinoma10. Since then, mousemodels have provided little evidence, despite sig-nificant long-term pathology. It has been suggestedthat the H. mustelae ferret is more prone to cancerinduction, but the study was uncontrolled11. How-ever, once again, the Mongolian gerbil has providedthe critical evidence to prove that H. pylori infectionalone could induce tumour formation. 37% of in-fected animals developed adenocarcinoma by 62weeks after infection, compared with none of the un-infected controls12. It has recently been reported thatinfection of the liver in mice by the lower-bowel-colonizing species Helicobacter hepatices induced

Animal models of Helicobacter infectionAdrian Lee

Table 1. Animal models of Helicobacter infection a

Animal Colonized by Advantages Disadvantages Refs

Primates H. pylori Closest animal species to human, endoscopy Expensive, colonized by endemic strains, presence 22possible, gastric physiology similar to human of Helicobacter heilmannii-like (HHLO) bacteria

Gnotobiotic piglets H. pylori Colonization pattern similar to human, gastric Chronic gastritis only, expensive short-term 23physiology similar to human, ulcers observed experiments only

Ferrets H. mustelae Natural infection, useful for vaccine studies, Pattern of colonization varies from human, 24gastric physiology similar to human predominantly chronic gastritis only

Cats and dogs H. pylori H. pylori only colonized colony available, gastric Gnotobiotic and SPF animals expensive, 25H. felis physiology similar to human HHLO present in normal cats

Guinea pig IL-8 homologue, active chronic gastritis Little used, data limited 6

Mice H. pylori Economical good for testing vaccines/ Doesn’t mimic human pathology 5, 26, 8H. felis antimicrobials, good colonizing strains of H. pyloriH. heilmannii (e.g. Sydney strain) available

Colonization by isogenic mutants easily tested, immunological reagents available, transgenic/knockout strains available

Gerbils H. pylori Chronic/active antral gastritis, gastric ulcers, Lack of immunological reagents, lack of 7adenocarcinoma induced with infection alone transgenic/knockout strains

aAbbreviations: SPF, specific pathogen free; IL-8, interleukin 8; HHLO, Helicobacter heilmannii-like bacteria.

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not only hepatitis, but also hepatocellular carcinoma.This should convince the sceptics that infection withHelicobacter species alone can induce cancer as aconsequence of long-term severe inflammation13.Equally convincing is the observation that low-grademucosa-associated lymphoid-tissue (MALT) lymph-omas that are almost identical to human tumourscan be induced in mice infected long-term with H. felis, H. heilmannii and H. pylori; these lesionsregress with anti-Helicobacter therapy14.

What the animal models have told usDespite the limitations of the animal models ofHelicobacter infection, they have increased ourknowledge of the human disease. The importanceof local acid to the microbial ecology of the stomachhas begun to explain why different diseases occurin different populations and why ulcers occur onlyat particular sites15. Host factors have been shownto be important in studies on different mouse

strains16. Several bacterial factors involved in thepathogenesis have been identified, for instanceurease, motility and Lewis b adhesins17–19. Animalexperiments have shown that immunization ispossible, and experiments in knockout mice haveeven revealed much about the mechanism of pro-tection20. Finally, these models provide a first stepto the validation of potential new antimicrobials2.

Future directionsThe possession of two complete genome se-quences of H. pylori opens up a completely newapproach to understanding the mechanisms ofcolonization and protection via studies of animalmodels with isogenic mutants21. In vivo expres-sion of key genes can be studied via differentialdisplay and other new gene technologies. Themouse and gerbil models will reveal much aboutthis important gastric pathogen in the years to come.

References1 Lee, A. (1997) The pathogenesis of Helicobacter

pylori infection , Bailliere’s Clin. Infect. Dis. 4, 341–3652 Lee, A. (1998) Animal models for host–pathogen

interaction studies , Br. Med. Bull. 54, 163–1733 Covacci, A. et al. (1999) Helicobacter pylori viru-

lence and genetic geography , Science 284,1328–1333

4 Lee, A. et al. (1993) Long term infection of thegastric mucosa with Helicobacter species doesinduce atrophic gastritis in an animal model ofHelicobacter pylori infection , Zbl Bakt (Int. J.Med. Microbiol.) 280, 38–50

5 Lee, A. et al. (1997) A standardized mouse modelof Helicobacter pylori infection – introducing theSydney strain , Gastroenterology 112, 1386–1397

6 Shomer, N.H. et al. (1998) Experimental Helico-bacter pylori infection induces antral gastritisand gastric mucosa-associated lymphoid tissuein guinea pigs , Infect. Immun. 66, 2614–2618

7 Ikeno, T. et al. (1999) Helicobacter pylori -inducedchronic active gastritis, intestinal metaplasia,and gastric ulcer in Mongolian gerbils , Am. J.Pathol. 154, 951–960

8 Eaton, K.A. et al. (1995) An animal model of gas-tric ulcer due to bacterial gastritis in mice , Vet.Pathol. 32, 489–497

9 Matsumoto, S. et al. (1997) Induction of ulcerationand severe gastritis in Mongolian gerbil by Helico-bacter pylori infection , J. Med. Microbiol. 46, 391–397

10 IARC (1994) IARC monographs on the evaluationof carcinogenic risks to humans , pp. 177–240,World Health Organization

11 Fox, J.G. et al. (1993) MNNG-induced gastric car-cinoma in ferrets infected with Helicobactermustelae , Carcinogenesis 14, 1957–1961

12 Watanabe, T. et al. (1998) Helicobacter pylori in-fection induces gastric cancer in mongoliangerbils, Gastroenterology 115, 642–648

13 Fox, J.G. et al. (1996) Persistent hepatitis andenterocolitis in germfree mice infected with Helico-bacter hepaticus , Infect. Immun. 64, 3673–3681

14 Enno, A. et al. (1998) Antigen-dependent pro-gression of mucosa-associated lymphoid tissue(MALT)-type lymphoma in the stomach – effectsof antimicrobial therapy on gastric MALT lym-phoma in mice , Am. J. Pathol. 152, 1625–1632

15 Van Zanten, S.J.O.V. et al. (1999) The gastric tran-sitional zones: neglected links between gastro-duodenal pathology and Helicobacter ecology ,Gastroenterology 116, 1217–1229

16 Sakagami, T. et al. (1996) Atrophic gastric changesin both Helicobacter felis and Helicobacter pyloriinfected mice are host dependent and separatefrom antral gastritis , Gut 39, 639–648

17 Eaton, K.A. and Krakowka, S. (1994) Effect of gas-tric pH on urease-dependent colonization ofgnotobiotic piglets by Helicobacter pylori , Infect.Immun. 62, 3604–3607

18 Eaton, K.A. et al. (1992) Motility as a factor in thecolonisation of gnotobiotic piglets by Helico-bacter pylori , J. Med. Microbiol. 37, 123–127

19 Guruge, J.L. et al. (1998) Epithelial attachment al-ters the outcome of Helicobacter pylori infec-tion , Proc. Natl. Acad. Sci. U. S. A. 95, 3925–3930

20 Nedrud, J.G. (1999) Animal models for gastricHelicobacter immunology and vaccine studies ,FEMS Immunol. Med. Microbiol. 24, 243–250

21 Alm, R.A. et al. (1999) Genomic–sequencecomparison of two unrelated isolates of thehuman gastric pathogen Helicobacter pylori ,Nature 397, 176–180

22 Dubois, A. (1998) Animal models of Helicobacterinfection , Lab. Anim. Sci. 48, 596–603

23 Krakowka, S. et al. (1998) Antimicrobial therapiesfor Helicobacter pylori infection in gnotobiotic pig-lets , Antimicrob. Agents Chemother. 42, 1549–1554

24 Erdman, S.E. et al. (1997) Helicobacter mustelae -associated gastric MALT lymphoma in ferrets ,Am. J. Pathol. 151, 273–280

25 Fox, J.G. et al. (1996) Local immune response inHelicobacter pylori -infected cats and identifi-cation of H. pylori in saliva, gastric fluid and fae-ces , Immunology 88, 400–406

26 Lee, A. and O’Rourke, J. (1996) The Helicobacterfelis mouse model , in Helicobacter pylori: Tech-niques for Clinical and Basic Research (Lee, A. andMegraud, F., eds), pp. 188–203, WB Saunders

Adrian Lee PhDProfessor

School of Microbiology and Immunology, The University of New South Wales,

Sydney 2502, Australia.Tel: 161 29 385 2101

Fax: 161 29 385 1591e-mail: a.lee@unsw.edu.au

Figure 1. The gastric mucosa of C57BL/6 miceinfected with Helicobacter pylori (the Sydneystrain SS1) showing large numbers of bacteria inthe antral crypts (Steiner stain).

Figure 2. The stomach of Mongolian gerbilsinfected for 12 months with Helicobacter pylori.(a) Gastric ulcer. (b) Severely inflamed antral mu-cosa. Photographs courtesy of Professor TakashiShimoyama and Dr Takashi Sakagami.

Disease mode l s