INTRODUCTION

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In recent years, studies of the host response to Helicobacter pylori infection have been primarily directed at a protective effect. Most studies have been directed towards the development of vaccination strategies (3, 14-17, 28, 35); only a few studies have examined the role of the host in tissue damage associated with infection by H. pylori (30, 31). In spite of this emphasis on vaccination schemes, a body of evidence regarding the pathogenesis of gastritis is accumulating. Marked differences in the severity of disease in different host species and, more recently, in different strains of the same host species have led to the suggestion that host immune response is important in determining the severity of gastritis. In some hosts, such as humans (26), rodents (12, 19, 27), and cheetahs (13), infection with some Helicobacter species results in severe gastritis with attendant epithelial changes ranging from erosion to ulceration and from metaplasia to neoplasia. In other hosts, such as dogs and cats (9, 21, 33, 34, 41), gastric Helicobacter infection results in little or no inflammation and no epithelial changes. In addition to species differences, host strain differences determine the outcome of infection. C57BL/6 mice are readily susceptible to some species and strains of Helicobacter and develop gastritis in response to infection, whereas BALB/c mice are more resistant to both colonization and inflammation (32, 38). These differences suggest that host response is important in determining if an infected individual develops severe gastritis, ulcers, neoplasia, only mild chronic gastritis, or no disease.

In addition to host response, a large body of data has been generated regarding the potential role of bacterial factors in the damage of host gastric mucosa. A number of bacterial virulence factors, including vacA (1), cagA and the related cag pathogenicity island (37), and iceA1 (36), have been shown to be associated with the increased severity of disease in humans, and there is clear evidence that in vitro H. pylori gene products can cause epithelial damage (6) or induce cytokine secretion potentially leading to inflammation (7). In spite of these correlations, however, studies evaluating the roles of bacterial factors in vivo in experimental models have been limited, and a direct effect of bacterial products causing host epithelial cell damage in vivo is controversial (8, 40). Thus, it is not clear if bacterial products damage host cells directly in vivo, if damage occurs as a by-product of host inflammation, or if both effects are important.

The overall goals of this study were (i) to demonstrate that H. pylori-associated gastritis and gastric epithelial damage are dependent on an intact adaptive immune response, (ii) to determine if such gastritis can be transferred to immunodeficient hosts by splenocytes from infected or uninfected donor mice, and (iii) to determine if gastritis is accompanied by in vivo evidence of a T-cell-mediated immune response (delayed-type hypersensitivity [DTH] response).

	MATERIALS AND METHODS

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Mice. Four- to six-week-old female helicobacter-free C57BL/6J mice and C57BL/6J scid/scid (severe combined immunodeficient [SCID]) mice were purchased from Jackson Laboratory (Bar Harbor, Maine). SCID mice are deficient in a DNA-dependent kinase necessary for recombination of both T-cell receptors and antibodies (22). Thus, neither T nor B cells mature in these mice, and they have no adaptive immunity. Mice were housed in microisolator cages and offered sterile Teklad lab chow and water ad libitum. One week following arrival, SCID mice were anesthetized with methoxyflurane, and 100 µl of blood was collected from the periorbital sinus. Serum was separated and stored at 20°C until used for the determination of the immunoglobulin G (IgG) level by enzyme-linked immunosorbent assay (ELISA) (see below). SCID phenotype was verified by the absence of detectable mouse IgG (optical density at 405 nm, <0.1) and/or the absence of a detectable serologic response to antigenic challenge.

DTH response. For the preparation of bacterial sonicate, broth cultures containing approximately 10⁹ CFU of H. pylori per ml were washed twice in phosphate-buffered saline (PBS), resuspended in 1/10 of the original volume of PBS, and sonicated with a Bradford sonifier at 4°C for two cycles of four 30-s bursts with 30-s rests. Intact bacteria were removed by centrifugation (5,000 rpm in a Sorvall RC-5B refrigerated superspeed centrifuge for 10 min), and sterility was ascertained by culture. The protein concentration was determined by a microprotein assay (Bio-Rad Laboratories, Hercules, Calif.), and aliquots were stored at 20°C until used. For mouse footpad injection, sonicates were adjusted to 10 µg/30 µl, and each mouse was given 30 µl of sonicate by injection into one hind footpad. Previous evaluation determined that 10 µg of antigen elicited a detectable and specific response (present only in sensitized mice [data not shown]). The opposite footpad received sterile PBS. Footpad thickness was measured with a dial thickness gauge (Swiss Precision Instruments, Los Angeles, Calif.) before injection, 24 h after injection, and, in some cases, 48 h after injection.

Bacterial strains. H. pylori SS1 (25) was kindly supplied by Adrian Lee. This strain is a cagA- and VacA-positive human isolate which is adapted for the colonization of mice. Bacteria were grown in brucella broth with 10% fetal calf serum or on blood agar plates under microaerobic conditions at 37°C. Broth cultures were enumerated by a hemacytometer and/or plate dilution.

Adoptive transfer. For the isolation of splenocytes, spleens were disaggregated in cold Hanks balanced salt solution, clumps were allowed to settle out, and the supernatant was sedimented at 1,000 rpm in an IEC PR-600 centrifuge for 10 min. Erythrocytes were removed by hypotonic lysis, and the splenocytes were resuspended in Dulbecco's PBS (D-PBS). Splenocytes were examined for viability by trypan blue exclusion and adjusted to 10⁷ viable cells/ml, and recipient SCID mice were given 0.1 ml by intraperitoneal injection 4 weeks after bacterial inoculation.

Flow cytometry. Splenocytes from C57BL/6 and SCID mice were isolated as described above, and 1 × 10⁶ to 2 × 10⁶ cells were resuspended in 100 µl of Fc Block (Pharmingen, San Diego, Calif.) and incubated for 10 min at 4°C. Fluorescein isothiocyanate- or phycoerythrin-labeled anti-CD4, anti-CD8, or anti-CD45R/B220 (Pharmingen) was added, and cells were incubated for 45 min at 4°C. Cells were washed three times in D-PBS with bovine serum albumin and 0.1% NaN₃ and resuspended in 100 µl of D-PBS, and 100 µl of 4% paraformaldehyde was added. Cell suspensions were evaluated on an EPICS flow cytometer.

ELISA. Murine IgG levels in SCID mice and anti-H. pylori IgG levels in all mouse sera taken at termination (herein referred to as terminal sera) were determined by ELISA. ELISA plates (96 wells; Corning) were coated either with 4 µg of unconjugated goat anti-mouse IgG (Pierce) per ml for the determination of circulating IgG in naive SCID mice or with 100 µg of H. pylori SS1 sonicate per ml for the determination of the serologic response to H. pylori. For the determination of mouse IgG levels, plates were coated with IgG for 12 h at 4°C, washed, blocked with 200 µl of 0.1% bovine serum albumin per well, washed, and incubated for 2 h at room temperature with SCID mouse sera diluted 1:20. Plates were washed, incubated with peroxidase-conjugated goat anti-mouse IgG for 90 min at room temperature, and washed again, and peroxidase was detected by incubation with azino-bisthiosulfonate in the dark for 5 to 10 min. The optical density at 405 nm was recorded.

Immunohistochemistry for expression of PCNA. To determine the labeling index, formalin-fixed sections were deparaffinized, antigens were exposed with antigen retrieval solution (Bio Genex), and the presence of proliferating cell nuclear antigen (PCNA) was determined by histochemical staining. Primary antibody was from Pharmingen (PC-10, mouse anti-PCNA), and secondary antibody was from Vector Laboratories (biotinylated goat anti-mouse antibody). Antibody staining was detected with an ABC detection kit from Vector Laboratories, and slides were counterstained with hematoxylin. The labeled and nonlabeled nuclei in three well-oriented glands in three different locations in the gastric fundus were enumerated, and the labeling index was calculated by dividing the number of labeled nuclei by the total number of nuclei counted and then multiplying by 100.

Histologic evaluation. The severity of gastritis in hematoxylin-and-eosin-stained sections was determined by enumerating the number of 20× microscopic fields containing the following pathologic changes: (i) neutrophilic inflammation (polymorphonuclear leukocytes [PMNs]), (ii) inflammatory infiltrate (lymphocytic, plasmacytic, histiocytic, neutrophilic, or, most commonly, mixed) which was severe enough to displace glands (gastritis), (iii) the destruction of gastric glands or neutrophilic gland abscesses by inflammatory infiltrate (adenitis), and (iv) the loss of normal fundic gland morphology with a replacement of specialized cells by mucus-type epithelium (metaplasia) and epithelial erosions. Genta-stained sections were evaluated to determine the number of fields containing Alcian blue-positive (intestinal-type mucus-containing) epithelial cells (intestinal metaplasia). Each of the categories was expressed as percent affected mucosa (the number of fields containing the lesion divided by the total number of fields). Because mild antral inflammation was present nonspecifically in all mouse groups, only fundic and cardiac mucosae were scored. All sections collected were examined in their entirety, and slides were read blind, i.e., without knowledge of their source.

Statistics. All experimental groups contained five to eight mice. Group means were compared by nonparametric methods (Mann-Whitney U test). A P value of <0.05 was considered statistically significant.

	RESULTS

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Adoptive transfer. Flow cytometric analysis revealed that isolated C57BL/6 splenocytes were 32% T cells (17% CD4 and 14% CD8) and 59% B cells (CD45R/B220) (Table 1). Flow cytometric analysis of cell markers in SCID mice was complicated by low-level staining of large, undifferentiated cells. However, peak populations revealed that cells from recipient SCID mice were enriched with T cells (39%) and depleted of B cells (23%). The CD4-to-CD8 ratio of donor splenocytes (1:1.18) did not differ significantly from that of recipient SCID mice (1:1.22, P > 0.5). Nonrecipient SCID mice did not have a detectable peak in any of the cell marker categories examined.

Gastritis in C57BL/6 and SCID mice. Histologic evaluation of gastritis included four categories of inflammation: PMN, adenitis, gastritis, and mucus metaplasia (see definitions above). These changes are illustrated in Fig. 1. Neutrophils were present in all groups of C57BL/6 mice, infected and uninfected, although in infected mice they increased late in the course of infection, their levels becoming significantly elevated by 16 weeks after inoculation (Fig. 2A). Thus, neutrophils alone correlated only weakly with infection status. In contrast, the levels of inflammation of the other three categories increased markedly in response to bacterial infection. Adenitis, gastritis, and metaplasia were not present in uninfected mice, but their levels significantly increased in infected mice by 6 or 8 weeks after infection. By 16 weeks, all levels were 10- to 20-fold greater than levels in uninfected C57BL/6 mice (Fig. 2A).

View larger version (134K): [in this window] [in a new window]   FIG. 1.   Histologic appearance of gastritis in an infected C57BL/6 mouse killed 16 weeks after bacterial inoculation. (A) Junction of normal and abnormal fundic mucosae. The affected area (to the right of the arrow) is characterized by widespread mixed inflammation and the loss of parietal and chief cells, which were replaced by undifferentiated mucus cells (metaplasia). The area to the left of the arrow is normal, characterized by gastric glands containing mostly parietal and chief cells. Bar = 50 µm. (B) Higher magnification demonstrating neutrophilic adenitis with the destruction of glands (large arrow), mixed inflammation including lymphocytes and plasma cells (arrowhead), and the almost complete replacement of gastric glands by undifferentiated mucus cells. A single remaining parietal cell is evident (small arrow). Bar = 14 µm.

View larger version (31K): [in this window] [in a new window]   FIG. 2.   Change in gastric inflammation and bacterial colonization in infected C57BL/6 and congenic SCID mice between 5 and 16 weeks postinoculation (PI). (A) In C57BL/6 mice, all four indicators of inflammation increased over the course of infection, but bacterial colonization did not change. (B) Variable numbers of neutrophils were present in infected and uninfected SCID mice at all experimental time intervals. Otherwise, inflammation remained low and did not change over the course of infection. Bacterial colonization was somewhat greater than that in C57BL/6 mice and did not change over the course of infection. *, P < 0.05 compared to result for uninfected mice (week 0).

Gastritis in recipient SCID mice. In contrast to nonrecipient SCID mice, H. pylori-infected SCID mice that received splenocytes developed severe gastritis rapidly after adoptive transfer (Fig. 3A and B). Within 2 weeks of the transfer of splenocytes (i.e., 6 weeks after infection) from either infected (Fig. 3A) or uninfected (Fig. 3B) C57BL/6 donors, SCID mice developed widespread neutrophilic infiltrates (up to 100% of the gastric mucosa), adenitis (up to 94%), gastritis (up to 90%), and metaplasia (up to 50%). Mean inflammation of all four categories continued to increase over time both in SCID mice receiving splenocytes from infected donors and in those receiving splenocytes from uninfected donors.

View larger version (43K): [in this window] [in a new window]   FIG. 3.   Changes in inflammation and colonization in infected and uninfected SCID recipients of splenocytes from infected or uninfected donors. (A) In infected recipients of splenocytes from infected donors, levels of neutrophilic inflammation, adenitis, gastritis, and metaplasia were significantly elevated 2 weeks posttransfer (PT) and continued to increase over the course of the experiment. (B) Infected recipients of cells from uninfected donors responded similarly, but inflammation in all categories was somewhat more pronounced than in the recipients of cells from infected donors. Bacterial colonization decreased precipitously in both groups of mice. Uninfected recipients of splenocytes from infected (C) or uninfected (D) donors developed only minimal, inconsistent gastric inflammation. Slight increases in levels of adenitis and gastritis in recipients of cells from infected donors may have been due to cross-reaction with enteric flora. *, P < 0.05 compared to results for nonrecipients (week 0). Note that since adoptive transfer was performed 4 weeks after bacterial infection, the postinoculation interval (Fig. 2) was 4 weeks longer than the PT interval. Thus, recipient SCID mice killed 12 weeks PT had been colonized for as long as nonrecipient SCID mice killed 16 weeks postinoculation (see Materials and Methods).

View larger version (44K): [in this window] [in a new window]   FIG. 4.   Effects of host and donor strains on gastric inflammation and bacterial colonization. SCID mice were killed 4 weeks after transfer (8 weeks after infection), and C57BL/6 mice were killed 16 weeks after infection (the most severely affected C57BL/6 group). Levels of neutrophilic inflammation, adenitis, gastritis, and metaplasia were significantly higher in infected recipient SCID mice than in infected C57BL/6 mice, even though C57BL/6 mice had been infected 8 weeks longer. In SCID recipients of naive cells, inflammation was the greatest and bacterial colonization was the least of any of the groups. Infected nonrecipient SCID mice did not develop inflammation, and bacterial colonization was highest in that group. *, P < 0.05 compared to results for C57BL/6 mice.

View larger version (91K): [in this window] [in a new window]   FIG. 5.   (A) Severe inflammation in an infected recipient SCID mouse infected with H. pylori, given splenocytes from an uninfected donor, and killed 4 weeks after transfer (8 weeks after bacterial inoculation). Note the marked, widespread infiltration of inflammatory cells, loss of parietal cells, destruction of glands, and overall thickening of the mucosa compared to panel B. (B) Moderate, multifocal inflammation in an infected C57BL/6 mouse killed 8 weeks after bacterial inoculation. Inflammation is less extensive and areas of normal mucosa are retained (arrows). Bars = 60 µm.

Epithelial lesions. In mice killed 3 months after transfer, four morphologic indicators of epithelial damage were evaluated. Mucus metaplasia was defined as described above. The percentages of gastric mucosae with epithelial erosions and intestinal metaplasia were evaluated in Genta-stained sections. Epithelial erosions were defined as the loss of necrosis of surface epithelium accompanied by the flattening or proliferation of adjacent epithelial cells. Intestinal metaplasia was defined as the presence of intestinal-type (Alcian blue-positive) mucus in gastric epithelial cells. Finally, the labeling index was determined in PCNA-stained sections. All four epithelial changes correlated significantly with gastritis and adaptive immune response (Table 2). Mucus metaplasia and erosions were present only in mice with gastritis (infected C57BL/6) or severe gastritis (infected recipient SCID). The group with the most severe gastritis (recipients of splenocytes from uninfected donors) had the most extensive mucus metaplasia, intestinal metaplasia, and epithelial erosions and the highest rate of epithelial proliferation (labeling index). In contrast, the highest degree of bacterial colonization was present in the group with no gastritis and no epithelial lesions (infected nonrecipient SCID mice). In addition to significant differences in mouse groups, there was a statistically significant correlation between the severity of gastritis and epithelial changes. Slopes, significance levels (P), and correlation coefficients (r²) of linear correlations between gastritis and all four epithelial changes are given in Table 3. Gastritis was positively correlated with epithelial changes.

Bacterial colonization. Concentrations of H. pylori organisms colonizing C57BL/6 mice ranged from 2 × 10⁷ to 3 × 10⁷ CFU/g of gastric mucosa and did not change significantly over the course of the experiment despite increasing inflammation (Fig. 2A). In SCID mice, however, bacterial colonization was inversely correlated with the severity of gastritis. In nonrecipient SCID mice with no gastritis, concentrations ranged from 2 × 10⁷ to 6 × 10⁷ CFU/g and did not change with time (Fig. 2B). In contrast, in recipient SCID mice, colonization decreased as the severity of inflammation increased (Fig. 3A and B). In SCID mice that received cells from infected donors, the concentration decreased more than 200-fold, from >5 × 10⁷ CFU/g 1 week after transfer to 2 × 10⁵ CFU/g 12 weeks after transfer; in those receiving cells from uninfected donors, the concentration decreased more than 300-fold, from >3 × 10⁷ CFU/g 1 week after transfer to fewer than 9 × 10⁴ CFU/g 8 weeks after transfer (P = 0.055 compared to value for mice given cells from infected donors). Three of five recipient mice given cells from uninfected donors were culture negative 8 weeks after transfer.

DTH response. A DTH response, as measured by a marked, statistically significant increase in footpad thickness 24 h after antigen injection, was present only in infected recipient SCID mice. DTH response was absent in both infected C57BL/6 (Fig. 6A) and infected nonrecipient SCID (Fig. 6B) mice. For infected recipient SCID mice, however, mean footpad swelling increased markedly both in groups receiving cells from infected donors (Fig. 7A) and in those receiving cells from uninfected donors (not shown). Slight increases in mean footpad thickness were present in uninfected SCID mice which received cells from infected donors and were killed 2, 4, and 8 weeks after transfer, but these differences were small, were not statistically significant, and did not persist to 12 weeks (Fig. 7B).

View larger version (15K): [in this window] [in a new window]   FIG. 6.   Change in DTH response (footpad swelling) and serum immune response to H. pylori antigens in infected C57BL/6 and SCID mice. (A) C57BL/6 mice did not develop a DTH response to H. pylori antigens even 16 weeks postinoculation (PI) but did develop a strong serum IgG response by 5 weeks PI (DTH was not measured 12 weeks PI). *, P < 0.05 compared to results for uninfected mice. (B) Infected SCID mice developed neither DTH nor serum immune responses (serum IgG was undetectable). OD405, optical density at 405 nm.

View larger version (15K): [in this window] [in a new window]   FIG. 7.   Change in DTH response (footpad swelling) and serum immune response to H. pylori antigens in infected and uninfected recipient SCID mice. (A) Infected SCID recipients of splenocytes from infected donors developed strong DTH responses by 2 weeks posttransfer (PT). Serum IgG responses developed later and remained consistently lower than those in C57BL/6 mice. Responses of SCID recipients of splenocytes from uninfected donors were similar (not shown). *, P < 0.05 compared to results for nonrecipients. (B) DTH and serum immune responses were low and inconsistent in uninfected SCID recipients of splenocytes from infected donors. OD405, optical density at 405 nm.

Immunoglobulin response. Infected C57BL/6 mice responded to H. pylori infection with strong systemic anti-H. pylori IgG responses that were detectable within 5 weeks of inoculation and reached maximal levels (indicated by optical density, as determined by ELISA) by 6 weeks (Fig. 6A). In contrast, infected SCID mice did not produce any anti-H. pylori IgG (Fig. 6B), and IgG levels in uninfected C57BL/6 mice remained low. Infected recipient SCID mice developed variable levels of anti-H. pylori IgG which gradually increased after transfer but did not reach the levels found in C57BL/6 mice (Fig. 7).

	DISCUSSION

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This study revealed several novel and important findings. First, we have shown that gastritis and epithelial lesions due to H. pylori are dependent on an intact adaptive immune response and do not develop in T- and B-cell-deficient SCID mice. These results differ from those of a previous study, in which the responses of SCID and wild-type mice to Helicobacter felis did not differ (4). There were a number of differences between that study and the present one which likely explain the different outcomes. Most likely to have contributed to the differences in these studies is the difference in the mouse strains used. Blanchard et al. (4) used C.B-17 mice and congenic SCID mice. These mice are essentially BALB/c mice with a single genetic difference (an IgG isotype) (5). BALB/c mice have been shown in several studies to develop only minimal gastritis in response to both H. felis (32, 38) and H. pylori (38). Based on reported results of these studies as well as our own observations (not shown), BALB/c mice would be expected to have minimal gastritis at 8 weeks after infection (the sacrifice time used in the Blanchard study). Thus, the lack of difference in inflammation and colonization in the Blanchard study may be largely attributable to the failure of the wild-type mice to respond rather than to any significant response by the SCID mice.

A second difference between C.B-17 and C57BL/6 SCID mice is the demonstrated "leakiness" of the former strain. C.B-17 SCID mice are known to produce antibody in increasing concentrations as they age (5, 39), and the possibility that some adaptive immune function remains in these mice must be considered when using this strain. Thus, it is conceivable that the SCID mice in the Blanchard study did develop some specific immune response to H. pylori, resulting in inflammation that approached the gastritis (albeit mild) described for the wild-type mice. In contrast, C57BL/6 mice are not known to be leaky. Furthermore, in the present study, all SCID mice were tested for circulating mouse IgG and shown to be negative at the onset of the study. Mice which did not receive splenocytes remained IgG negative, did not develop H. pylori-specific antibody in response to infection, and did not develop DTH responses, suggesting that they remained immunodeficient throughout the study.

The second important finding of this study is the severity of gastritis in recipient SCID mice. The lesions are notable for two reasons. The most intriguing aspect is the difference between recipient SCID and donor C57BL/6 mice. The marked increases in the severity and rapidity of the onset of gastritis in recipient SCID mice (within 2 weeks of transfer), as well as the induction of DTH not seen in donor mice, indicate a loss of regulation resulting in the derepression of inflammation in recipient SCID mice. It is likely that regulatory cells, cytokines, or both suppress inflammation in normal mice, dampening and delaying gastritis. The absence of these regulatory influences in SCID mice results in the lesions that were produced in this study. While the effector cells are likely T cells (as evidenced by the correlation with DTH and the T-cell enrichment of the engrafted cells), the identification of the regulatory population in donor mice was not addressed by this study.

In addition to providing evidence suggesting the regulation of inflammation due to H. pylori in normal mice, the severe gastritis in recipient SCID mice represents an important addition to the growing number of animal models of disease due to H. pylori. The overall severity and rapidity of the onset of the lesions described here are more extensive and acute than those described for other animals colonized with H. pylori. Lesions of comparable severity and rapid onset (within 4 weeks) have been described for interleukin-10-deficient mice colonized with H. felis (2), and several studies have described severe lesions after prolonged (several months to years) colonization by H. felis (24, 25, 38) or H. pylori (27). However, in most studies, colonization by H. pylori resulted in relatively mild gastritis unaccompanied by epithelial lesions (9, 10, 18, 23).

The third important finding of this study is that epithelial lesions depend on the presence of gastritis and that by themselves, even large numbers of bacteria in close contact with the gastric epithelium do not cause epithelial erosions, metaplasia, or increased epithelial proliferation in vivo in the absence of a host response. These results indicate that if bacterial virulence factors are important in determining the outcome of infection, it is likely that their roles are in the induction of host response rather than in having a direct deleterious effect on host epithelial cells.

In addition to the three important findings described above, several other characteristics of H. pylori gastritis are addressed in this study. This study is the first to describe the effect of adoptive transfer in H. pylori-infected immunodeficient mice. A previous investigation by Mohammadi et al. examined the effect of adoptive transfer between groups of H. felis-infected C57BL/6 mice (31). In that study, findings were similar to those of the present study in that adoptive transfer exacerbated gastritis and suppressed colonization, but the differences in the severity of gastritis between nonrecipient and recipient mice were not as marked as they were in the present study. One interesting difference between the results of Mohammadi et al. and our own was the effect of prior exposure to antigen in donor mice. Mohammadi et al. found that splenocytes from naturally infected or immunized donors induced more severe gastritis than did splenocytes from uninfected donors. In our study, splenocytes from uninfected donors induced more severe gastritis and suppressed bacterial colonization more than splenocytes from infected donors did.

Differences between the study by Mohammadi et al. and the present study are likely due to differences in mouse strain, bacterial strain, and experimental protocol. Probably the most significant difference was that adoptive transfer in that study was done between groups of C57BL/6 mice rather than between C57BL/6 and SCID mice, as was done in the present study. Thus, the recipient mice were immunocompetent, and the contribution of the immune response of the recipients to the outcome could not be determined. Because the recipients in the present study were immunodeficient, endogenous adaptive immunity did not contribute to their responses. Other differences included the number of splenocytes transferred; the bacterial strain (H. felis was used in the study by Mohammadi et al.); and the infection, transfer, and sacrifice times. Finally, Mohammadi et al. did not evaluate DTH responses in their mice.

Other findings of this study are of interest. The strong inverse correlation between gastritis and bacterial colonization is evidence that the host immune response is effective in the suppression of bacterial colonization. This is compatible with other studies showing that increased immunity (e.g., due to vaccination) is accompanied by increased gastritis (postimmunization gastritis) as well as suppressed bacterial colonization (15). The correlation of DTH with severe gastritis is also compatible with previous studies suggesting that T-cell-mediated immunity is necessary for bacterial clearance (and, by extension, for the induction of gastritis) (3, 14, 31, 35). Finally, the development of DTH in infected recipient SCID mice but not infected donors is further evidence of immune regulation in donor mice.

The overall results of this study confirm that host immunity is necessary for both gastritis and the suppression of H. pylori colonization in infected mice. In addition, we demonstrate that epithelial damage is dependent on the host immune response rather than resulting from a direct bacterial effect, and we provide evidence that T-cell-dependent immunity is necessary for both gastritis and bacterial suppression. Finally, we demonstrate that gastritis is derepressed in SCID recipients, suggesting that gastritis is regulated and thus suppressed in normal infected C57BL/6 mice.