Helicobacter pylori-induced gastritis in guinea pigs: Model development, diagnostic methods and comparison with mouse protocols

Helicobacter pylori is a gram-negative spiral-shaped bacterium that colonizes the human stomach and causes gastric inflammation, gastric and duodenal peptic ulcers, gastric cancer and MALT-lymphoma. A novel guinea pig model of H. pylori infection was developed. Infection was induced by oral inoculation and was confirmed by cultivation from the stomach mucosa. The infection led to severe gastric inflammatory changes and a detectable serum antibody response similar to that in humans. When infected by H. pylori, guinea pigs were found to develop more severe gastritis than mice. The infection induced an acute-phase response in guinea pigs, as revealed by the elevated serum C3 levels obtained, and appeared to cause elevated serum cholesterol levels, suggesting a possible link between H. pylori infection and cardiovascular disease. The effects on H. pylori-infected guinea pigs of dietary supplements consisting of a combination of antioxidant vitamins and selenium were evaluated. Animals receiving these supplements displayed a lesser density of H. pylori colonization in the stomach than control animals and their gastric inflammation appeared to be suppressed. It was concluded that in order to obtain optimal gastric inflammation in the guinea pig model the antioxidant levels in the feed should be kept low. H. pylori infection in guinea pigs was followed for a five-month period. It was found to persist and to be accompanied by severe gastric inflammation. Control animals tested for Helicobacter by use of culture methods and a genus-specific PCR protocol were found to be free of natural Helicobacter infection. Methods of monitoring H. pylori colonisation were evaluated in both mice and guinea pigs. Of two commercial H. pylori faeces-antigen tests assessed in mice, a test based on monoclonal antibodies displayed 100% accuracy, whereas a polyclonally based test sometimes gave false positive results, probably by cross-reacting with a natural murine Helicobacter infection. Indigenous Helicobacter ganmani infection in the mouse colony was detected by PCR-denaturing gradient gel electrophoresis. The polyclonal antibody test was found to cross-react in vitro with several non-pylori Helicobacter species, including species reported in humans. Analyses of faecal samples from guinea pigs, in contrast to mice, by the use of an antigen test or PCR methods were found unable to detect the H. pylori infection. In conclusion, the investigation considers the development and optimisation of experimental H. pylori infection in guinea pigs, and also explores different methods of monitoring the infection. The study shows the guinea pig to be a viable model for studying H. pylori-related gastric and possible extra-gastric manifestations

High intake of selenium, beta-carotene, and vitamins A, C, and E reduces growth of Helicobacter pylori in the guinea pig

PURPOSE: Helicobacter pylori is a human gastroduodenal pathogen associated with type-B gastritis and gastric cancer. Low gastric tissue antioxidant levels are believed to increase the risk of developing gastric cancer. We investigated whether dietary antioxidant levels protect against infection and type-B gastritis in H. pylori-infected guinea pigs. METHODS: Dunkin-Hartley guinea pigs infected for 6 weeks with H. pylori were fed diets with various antioxidant levels. Stomach specimens were cultured, and gastritis was graded from 0 to 3. RESULTS: Supplementation with vitamins A, C, and E and with selenium yielded H. pylori recovery from 17% of challenged animals, compared with 43% of those fed a control diet. Gastritis was scored at 0.33 and 0.93, respectively. Supplementation with only vitamin C or astaxanthin had less effect on gastritis and recovery rate. In a second experiment, gastritis score in a group given vitamins A, C, E, and selenium and beta-carotene was 2.25 and in a control group, it was 2.57. The H. pylori recovery rate was 75 and 100%, respectively, with fewer colonies from animals given antioxidant supplementation (P < 0.05). CONCLUSIONS: A combination of antioxidants can protect against H. pylori infection in guinea pigs. In animal studies, antioxidant intake should be low to optimize development of H. pylori-associated disease. Furthermore we established that H. pylori causes severe gastritis in guinea pigs

Comparative study of Helicobacter pylori infection in guinea pigs and mice - elevation of acute-phase protein C3 in infected guinea pigs

Eighteen Dunkin-Hartley guinea pigs and 50 NMRI mice were inoculated with Helicobacter pylori and the infection followed by culture, histopathology, antibody response, and plasma levels of the acute-phase proteins albumin, C3, and transferrin for up to 7 weeks. The immune response to H. pylori surface proteins was studied by an enzyme immunoassay (EIA) and Western immunoblot and the plasma levels of albumin, C3, and transferrin were analyzed by single radial immunodiffusion. Guinea pigs had a more severe gastritis and a higher EIA immune response than NMRI mice. Serum C3 levels were elevated in infected guinea pigs after 3 and 7 weeks indicating a systemic inflammatory response and a possible link between H. pylori infection and extragastric manifestations such as vasculitis associated with atherosclerosis. Serum cholesterol levels were analyzed in guinea pigs at 7 weeks and indicated a higher level in H. pylori-infected than in control animals, but this difference was not statistically significant

Lectin histochemistry of the gastric mucosa in normal and Helicobacter pylori infected guinea-pigs

Helicobacter pylori attaches via lectins, carbohydrate binding proteins, to the carbohydrate residues of gastric mucins. Guinea-pigs are a suitable model for a H. pylori infection and thus the carbohydrate composition of normal and H. pylori infected gastric mucosa was investigated by lectin histochemistry. The stomach of all infected animals showed signs of an active chronic gastritis in their mucosa, whereas no inflammation was present in the control animals. The corpus-fundus regions of the controls showed heterogeneous WGA, SNA-I, UEA-I and HPA binding in almost all parts of the gastric glands. While these lectins labelled the superficial mucous cells and chief cells heterogeneously, the staining of the parietal cells was limited to WGA and PHA-L. Mucous neck cells reacted heterogeneously with UEA-I, HPA, WGA and PHA-L. In the antrum, the superficial mucous cells and glands were stained by WGA, UEA-I, HPA, SNA-I or PHA-L. WGA, UEA-I, SNA-I and HPA labelled the surface lining cells strongly. The mucoid glands reacted heterogeneously with WGA, UEA-I, HPA, SNA-I and PHA-L. In both regions, the H. pylori infected animals showed similar lectin binding pattern as the controls. No significant differences in the lectin binding pattern and thus in the carbohydrate composition between normal and H. pylori infected mucosa could be detected, hence H. pylori does not induce any changes in the glycosylation of the mucosa of the guinea-pig. This unaltered glycosylation is of particular relevance for the sialic acid binding lectin SNA-I as H. pylori uses sialic acid binding adhesin for its attachment to the mucosa. As sialic acid binding sites are already expressed in the normal mucosa H. pylori can immediately attach via its sialic acid binding adhesin to the mucosa making the guinea-pig particularly useful as a model organism

The relationship between O-chain expression and colonisation ability of Helicobacter pylori in a mouse model

The influence of lipopolysaccharide (LPS) O-polysaccharide chain production on the colonisation ability of Helicobacter pylori in four mouse models (NMRI, C57BL/6, CBA/Ca, and BALB/cA mice) was studied. H. pylori strains that produced smooth-form LPS (S-LPS) detectable in silver-stained electrophoretic gels colonised mice. In contrast, a laboratory-passaged strain G50 and the culture collection strain CCUG 17874 did not colonise mice; the former strain produced low amounts of O-chains only detectable in immunoblotting but not in silver-stained gels, whereas the latter produced rough-form LPS (R-LPS) without O-chains. Furthermore, a galE isogenic mutant, which produced R-LPS, did not colonise mice. However, after repeated broth culture, strains G50 and CCUG 17874 produced S-LPS detectable in silver-stained gels and were capable of colonising mice. Consistent with the production of O-chains, all colonising strains produced Lewis (Le) antigens, Le(x) and/or Le(y). Except for low expression of Le(y) by non-colonising G50, reflecting low production of O-chains, all other non-colonising strains and the galE mutant lacked expression of Le antigens consistent with their production of R-LPS. Lectin typing of strains supported these findings, and also showed that lectin types did not differ before and after colonisation. The low level of O-chain production and Le antigen expression by the non-colonising G50 may not be sufficient to aid colonisation. Examination of protein profiles of H. pylori strains before inoculation showed that protein expression was not significantly different between colonising and non-colonising strains. These results show that S-LPS production with O-chain expression is required by H. pylori for colonisation in a number of mouse models and that care should be taken with inoculating H. pylori strains that loss of O-chains does not occur during subculturing

Dietary factors influence the recovery rates of Helicobacter pylori in a BALB/cA mouse model

The aim of this study was to assess the ability of different mouse diets to sustain an H. pylori infection in BALB/cA mice. Four commercially available mouse diets were compared. Experiment 1: Mice were fed the four diets for seven days before infection, infected three times at two-day intervals with 0.1 ml of 10(9) colony-forming units/ml H. pylori cells. H. pylori strains (n = 4) were cultured on GAB-Camp agar for 2 days, harvested and suspended in PBS. All animals were sacrificed at 2 and 4 weeks post inoculation. Experiment 2: Mice infected for 8 weeks were fed RM2, changed to the different diets for 10 days and sacrificed. Stomachs were collected, cultured on GAB-Camp agar to estimate H. pylori growth and stomach biopsies were analyzed by PCR. There were significant differences between diets in their ability to sustain growth of H. pylori. The range was from a few hundred colonies to no growth at all on the GAB-Camp agar. PCR signals showed good correlation with the culture results. All H. pylori-infected mice gave a significantly higher inflammation score compared to non-infected mice. The diet RM2, having the highest number of culturable H. pylori in the mouse stomach, also showed the highest inflammation. These results suggest that the dietary factors affect the amounts of H. pylori in an infection of BALB/cA mice

The evolution of epilepsy surgery in tuberous sclerosis in Sweden: A national registry study

PurposeThis study aimed to characterize the Swedish cohort of surgically treated patients with TSC and explore differences in preoperative investigation and outcome over time.MethodsData on patient and seizure characteristics were retrieved from the Swedish National Epilepsy Surgery Register. Two-year follow-up results were compared between the years 1997–2010 and 2011–2018. Preoperative investigations were re-evaluated.ResultsEighteen tuberectomies and seven callosotomies were identified. Seizure freedom after tuberectomy was achieved in 11 % (1/9) 1997–2010 and 56 % (5/9) 2011–2018. The number of tuberectomies increased each decade. Patients operated on in 1997–2010 had higher seizure frequency (median 175 seizures/month vs. 102) and incidence of infantile spasms (4/9 vs. 1/9, none after 2011). There was a trend towards surgery at a younger age (median 86 months 1997–2010 vs. 48 months 2011–2018). None with >200 seizure/month, SEGA, or history of infantile spasms achieved seizure freedom. Two patients underwent anterior callosotomy (1992 and 1994) and became free of drop attacks. Five callosotomies were performed between 2011 and 2013, one patient became free of drop attacks. Two complications with new neurological deficits were reported. The median age at surgery was higher in the callosotomy group (14 years) than in the tuberectomy group (5 years).ConclusionSeizure freedom after tuberectomy in patients with TSC has increased over time in our cohort. Signs of a heavier disease burden were more frequently observed 1997–2010 and associated with worse outcomes. Callosotomy operations were prevalent at the beginning of the 2010s

Infection with cagA- and vacA-positive and -negative strains of Helicobacter pylori in a mouse model

To study the role of cytotoxin-associated protein (cagA) and vacuolating cytotoxin (vacA) in Helicobacter pylori infection in an experimental murine model, mice were infected with seven strains with different cagA and vacA status. Groups of 10 NMRI mice were challenged and were killed 5 weeks later. In a second study, 20 mice were challenged with a mixture of the same seven strains and killed 1, 3, 15 and 17 weeks post-inoculation. All seven strains were found to colonize the mice for the 5-week experimental period. Animals infected with vacA-positive strains, regardless of cagA status, showed an elevation of antibody titers. Two cagA-negative and vacA-positive strains and one cagA- and vacA-positive strain were found to 'take over' in the mixed infection as analyzed by the randomly amplified polymorphic DNA-polymerase chain reaction technique and in one mouse stomach we found coexistence of two of the strains. We found no evidence of the different strains colonizing different parts of the stomach