Helicobacter pylori infection delays healing of ischaemia-reperfusion induced gastric ulcerations: new animal model for studying pathogenesis and therapy of H. pylori infection.
OBJECTIVE
Helicobacter pylori (Hp) infection is a major risk factor of peptic ulcerations but studies on its pathogenicity are limited due to the lack of an adequate animal model. In this study we developed the new model of gastric Hp infection in rat gastric mucosa, with acute gastric erosions progressing into ulcers in animals subjected initially to ischaemia-reperfusion (I/R).


METHODS
I/R lesions were produced in rats by clamping the coeliac artery for 0.5 h followed by 1 h reperfusion and gastric inoculation with type I Hp strain (CagA and VacA positive) or type II Hp strain (CagA and VacA negative), obtained from fresh clinical isolates, or with vehicle (saline). Gastric secretion during recovery from I/R lesions was determined in a separate group of rats equipped with chronic gastric fistula to inoculate the animals with Hp and then to collect gastric juice for determination of gastric acidity and pepsin outputs as well as luminal content of somatostatin.


METHODS
The animals were killed at 0, 3, 12 or 24 h and 3, 5, 10 or 15 days after Hp inoculation and the area of gastric lesions was determined planimetrically and gastric blood flow (GBF) was measured by the H2-gas clearance technique. The venous blood was withdrawn for measurement of plasma interleukin (IL)-1beta and tumour necrosis factor alpha (TNFalpha) by ELISA and plasma gastrin, luminal somatostatin by RIA and the mucosal expression of transforming growth factor (TGFalpha) was analysed using RT-PCR with specific primers. Gastric Hp infection was assessed by histology, rapid urease test and Hp culture. The effect of triple therapy with omeprazole, amoxycillin and tinidazole on Hp infection and ulcer healing was also determined.


RESULTS
Ischaemia alone resulted in an immediate fall in GBF and almost complete suppression of gastric secretion but without any gastric lesions. When ischaemia was followed by 1 h of reperfusion, acute gastric erosive lesions immediately occurred, reaching a maximum at 12 h after I/R and progressing after 3 days to deeper gastric ulcers that disappeared after 15 days. In Hp-inoculated rats, the number of viable Hp colonies gradually increased, reaching maximum at day 10 with infection with type I and at day 15 with infection with type II Hp. At day 15 the difference in Hp colonization was not significantly different between the stomachs infected by type I and type II Hp. Inoculation, especially with the type I Hp strain, significantly delayed healing of I/R-induced acute lesions and accelerated their progression into deeper chronic ulcers. This effect was accompanied by a significant fall in GBF and a higher increment in plasma IL-1beta and TNFalpha levels. Gastric acid secretion, which was completely inhibited up to 12 h after I/R, returned to the control value 24 h upon completion of the I/R procedure. This return was delayed in Hp-infected rats and accompanied by a significant elevation of plasma gastrin and a decrease in luminal somatostatin. The immunoreactivity of TGFalpha and expression of TGFalpha mRNA determined by RT-PCR were well defined in intact gastric mucosa but were significantly decreased, especially in mucosa infected with type I Hp strain, at day 15 after I/R. The triple therapy which cured Hp infection completely abolished the delay in ulcer healing caused by Hp.


CONCLUSIONS
(1) Gastric infection with the Hp strain expressing cagA and vacA encoded cytotoxins delays healing of I/R-induced acute gastric lesions due to an impairment of gastric microcirculation, and excessive proinflammatory cytokine release and suppression of anti-ulcer TGFalpha expression. (2) The I/R induced suppression of gastric acid secretion may contribute to the hypergastrinaemia as a secondary phenomenon and may account for the spread of Hp infection observed during the progression of acute erosions into chronic ulcers. (3) I/R induced gastric ulcers may be a useful model for studying the action of Hp infection on gastric ulcerogenesis in ischaemic stomach and for testing anti-Hp therapy.
Konturek
S J
SJ
Department of Physiology and Pathomorphology, Jagiellonian University School of Medicine, Cracow, Poland. pogonow@cyf-kr.edu.pl
Brzozowski
T
T
Konturek
P C
PC
Kwiecien
S
S
Karczewska
E
E
Drozdowicz
D
D
Stachura
J
J
Hahn
E G
EG
D000818:Animals; D001483:Base Sequence; D001707:Biopsy, Needle; D004195:Disease Models, Animal; D004797:Enzyme-Linked Immunosorbent Assay; D005753:Gastric Mucosa / Q000098:blood supply* / Q000378:metabolism / Q000473:pathology*; D016481:Helicobacter Infections / Q000188:drug therapy / Q000473:pathology*; D016480:Helicobacter pylori / Q000302:isolation & purification* / Q000472:pathogenicity; D007150:Immunohistochemistry; D007375:Interleukin-1 / Q000032:analysis; D007511:Ischemia / Q000473:pathology*; D008297:Male; D008969:Molecular Sequence Data; D011336:Probability; D011863:Radioimmunoassay; D051381:Rats; D015427:Reperfusion Injury; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018570:Risk Assessment; D012680:Sensitivity and Specificity; D018709:Statistics, Nonparametric; D013276:Stomach Ulcer / Q000473:pathology* / Q000503:physiopathology; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis; D014945:Wound Healing
D007375:Interleukin-1; D014409:Tumor Necrosis Factor-alpha