Front. gastrointest. Res., vol. 1, pp.12-48 (Karger, Basel 1975)
Gastritis R.G.
STRICKLAND
Division of Gastroenterology, Department of Medicine, School of Medicine, University of New Mexico, Albuquerque, N. M.
Contents I. Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . II. Gastric Autoimmunity - Historical Aspects. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III. Gastric Autoantibodies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . A. Intrinsic Factor Antibodies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . B. Parietal Cell Antibody. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . C. Autoantibodies in Pernicious Anemia. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. Classical (adult onset) Pernicious Anemia. . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Juvenile Vitamin B12 Deficiency ................................... D. Autoantibodies in Atrophic Gastritis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E. Gastric Morphology and Function in Atrophic Gastritis ................ F. Significance of Gastric Autoantibodies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV. Cell-Mediated Immunity. .. . . .. . . .. . . . .. . . . . . . .. . . .. . . ... . . .. . . . . . . . . . . V. Immune Deficiency ...................................... , . . . . . . . .. . . . VI. Genetic Influences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII. Associated Diseases. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII. Effects of Corticosteroids. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IX. Experimental Autoimmune Gastritis ................. . . . . . . . . . . . . . . . . . . . X. Conclusions ......................................................... Acknowledgments. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
12 14 15 15 17 18 18 20 21 21 26 29 30 31 32 33 34 36 38 38
Several problems are encountered in attempting to classify human gastritis. Accurate diagnosis rests on a histological examination of the mucosa. While single specimens obtained by peroral biopsy are representative in the more severe forms of chronic gastritis, this may not be so when the inflam-
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I. Introduction
STRICKLAND
13
mation or atrophy has a focal distribution. Most previous studies of gastritis have focused on morphological and functional disturbances in the gastric corpus, and little attention has been given to simultaneous investigation of the pyloric antral mucosa which, on the basis of histology and function, must be considered a separate structure (fig. I). Short gastric pits and straight glands lined by parietal or acid-producing cells and chief or pepsinogen-producing cells distinguish the corpus mucosa. The antral mucosa takes on a more villous appearance; the glands are tortuous and lined for the most part by mucussecreting cells. Interspersed between these cells are several types of enterochromaffin cells; the best characterized being the gastrin secreting (G) cells. Many potential pathogenic factors have been identified in gastritis, but few of them are sufficiently well-documented to justify attempts to classify this disease on etiological grounds. Finally it is clear that gastritis is often clinically silent, and its prevalence in the general population is uncertain. This has raised doubts as to its significance once discovered. A generally accepted classification is that based on morphology: (A)
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Fig.I. Normal histological appearance of gastric pyloric antral mucosa (left) and corpus mucosa (right). HE. x 150.
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14
acute; (B) chronic, (1) superficial, (2) atrophic, (a) multifocal, (b) diffuse, (c) gastric atrophy; (C) indeterminate, (1) hypertrophic, (2) eosinophilic. This terminology is generally used to describe changes in the corpus mucosa, but a similar range of histological findings may occur in the antral mucosa. The only form of human gastritis in which substantial evidence favors the pathogenic role of immunologic factors is chronic atrophic gastritis (CAG). This includes the gastric lesion of pernicious anemia (PA) where the essential defect is failure of production of intrinsic factor (IF) to such a degree that absorption of vitamin B12 is severely impaired.
Investigation into the phenomenon of acquired resistance to oral hog IF in patients with PA resulted in the discovery by SCHWARTZ [149] in 1958 of a serum factor which interfered with the action of hog IF in promoting vitamin B12 absorption. In the same year TAYLOR and MORTON [166] reported the induction of antibody to IF in rabbits immunized with human gastric juice and the ability of this antibody to block IF mediated vitamin B12 absorption. TAYLOR [164] and SCHWARTZ [150] then demonstrated independently a similar inhibitory factor in the y-globulin fraction of serum from patients with PA who had never received oral treatment with hog IF. The hypothesis that the inhibitor was an autoantibody to IF was subsequently confirmed in 1962 by the studies of JEFFRIES et al. [97] who showed that the y-globulin fraction ofPA serum could combine with IF-vitamin B12 complex and retard its electrophoretic mobility in starch gel. In the same year complement-fixing antibody to a saline extract of human gastric mucosa was demonstrated in PA sera by IRVINE et al. [89] and MARKSON and MOORE [116], and TAYLOR et al. [167] showed by immunofluorescence that this antibody reacted specifically with gastric parietal cells. These studies were the first to identify a direct link between diffuse chronic inflammatory atrophy of the gastric mucosa and organspecific immunologic abnormalities. They marked the beginning of a new era of investigation into the pathogenesis of this disease. This chapter presents a review of subsequent studies, both human and experimental, which have been directed towards a better understanding of the significance and potential pathogenic role of immunologic disturbances in CAG and P A. Two recent reviews are available [110,160] which cover more fully the clinical importance and sequelae of CAG and the pathogenic role of nonimmunologic factors.
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II. Gastric Autoimmunity - Historical Aspects
Gastritis
15
III. Gastric Autoantibodies
It is now established that autoantibodies against two separate components of the gastric parietal cell may be identified in patients with CAG or PA. It is of interest that despite involvement of chief cells in the atrophic damage, antibodies to constituents of the pepsin-producing cells have not been demonstrable so far.
A. Intrinsic Factor Antibodies (IF A)
Fig.2. Diagrammatic illustration of antibodies to IF. Type I or 'blocking' antibody reacts with the vitamin B12 binding site. Type II or 'binding' antibody reacts with a site distant from the B12 binding site and may combine with IF alone or IF-vitamin B12 complex.
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IF, a glycoprotein with a molecular weight of approximately 60,000 [70] is secreted by the gastric parietal cell in man [78). Its function in promoting absorption of dietary vitamin B12 relates to its capacity to bind the vitamin [63] and form a stable complex for transport from the stomach to receptors on the distal ileal epithelial cell microvilli where active absorption of vitamin B12 takes place [31,41,75]. Normally IF is secreted in amounts vastly in excess of that required to maintain adequate vitamin B12 absorption [7). Thus vitamin B12 absorption may often be preserved even in the presence of advanced atrophic gastritis and total achlorhydria. Two distinct antibodies to IF (IF A) have been identified (fig. 2). Type I or blocking antibody [1,6,144,145] is directed against the binding site of vitamin B12 on the IF molecule and blocks subsequent combination of IF with free
16
vitamin. Type II or binding antibody [97,142,143,167] is directed against an antigenic site distant from that related to vitamin B12 binding and thus may react with IF alone or the preformed IF - vitamin B12 complex. IF which has reacted with type-II antibody is still capable of combining with type-I antibody, thus confirming the existence of two distinct antigenically reactive sites on the IF molecule [62,145]. Several techniques have been described for detecting and quantifying IF A. Type-I IF A is detected by demonstrating that radioactively labelled vitamin B12 is inhibited from binding to IF of normal human gastric juice by the serum being tested. Quantitation of type-I IFA is dependent upon separating free-labelled vitamin B12 from the larger IF-vitamin B12 complex by such methods as dialysis [1], charcoal [6, 67], zinc sulphate-barium sulphate precipitation [139], or zirconyl phosphate gel [72]. Increased sensitivity in detecting type-I IFA is reported [71] utilizing pretreatment of serum with unlabelled vitamin B12 (saturating B12-binding sites of serum proteins) and subsequent adsorption of unbound excess to albumin-coated charcoal. This maneuver also eliminates false-positive results which can arise from recent vitamin B12 injections. The charcoal-binding test is usually preferred for routine testing, and a reversed charcoal test using sera containing high titer type-I antibody can be used for the assay of IF in gastric juice [6,67,98]. Detecting and quantifying type-II IFA requires separation of IF-vitamin B12 complex from antibody-bound IF-vitamin B12 complex and electrophoresis [97,167] immune co precipitation [32,95,140,145] and radio immunodiffusion [142] have been used for this purpose. The salt precipitation method is the one preferred for routine testing. Both antibodies may also be tested for in one system utilizing uptake of radioactive vitamin B12 by guinea pig ileal mucosal homogenate and altering the order of reactant addition to detect either type-lor type-II IFA [12]. A recent report describes a radioimmunoassay for simultaneous detection of both antibodies and evidence is presented for its greater sensitivity over earlier techniques [141]. Immunofluorescence is generally held to detect antibody to a component of the gastric parietal cell which is distinct and separate from intrinsic factor (vide infra). However, using elaborate immunoabsorptive techniques, FISHER and TAYLOR [54] have demonstrated in testing sera known to contain IFA that a part of the fluorescence seen, using human fundic mucosa as substrate, can be attributable to IF antibody. It is also believed that IF antibodies do not fix complement; however, some evidence to the contrary has recently been reported [94].
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STRICKLAND
Gastritis
17
Fig. 3. Immunofluorescent staining of parietal cells in human gastric corpus mucosa by serum from a patient with PA. Note the diffuse cytoplasmic pattern of staining.
It is probable that the complement-fixing antibody to gastric mucosal homogenate [89,116] and that detected by immunofluorescence and seen to react diffusely with the cytoplasm of parietal cells (fig.3) [167] are directed against the same antigen. The immunofluorescent technique, however, is the more sensitive of the two. The antigen is organ- and cell-specific, showing no cross-reaction with other parts of the gastrointestinal tract or the thyroid. It is not species specific [127], being present even in the 'Hauptdriisen-Zellen' of amphibian stomach [163]. Ultracentrifugation of gastric mucosal homogenate reveals the antigen to be confined to the microsomal fraction. It is destroyed by papain and trypsin and inactivated by surface active detergents but not by treatment with phospholipases, ribonuclease, or dilute alkali [19]. Recent work [183] has strengthened the belief that the parietal cell antigen is a lipoprotein and is associated with smooth cytoplasmic membranes. The application of IgG fractions of PCA containing serum, coupled to horseradish peroxidase, onto sections of
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B. Parietal Cell Antibody (PCA)
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human gastric mucosa indicated a localization of bound antibody to the microvillus membrane of the canalicular system of the parietal cell [79]. The routine detection of PCA is best accomplished by the immunofluorescent technique using unfixed cryostat sections of gastric fundic mucosa. Where possible, human fundic mucosa from patients with duodenal ulcer should be used. Rat mucosa should no longer be used due to the presence in some human sera of a heteroantibody to rat parietal cells [126]. This heteroantibody is not indicative of gastric mucosal damage, whereas the parietal cell autoantibody is such an indicator [158]. The complement fixation technique is utilized where it is desirable to estimate antibody titer [87].
1. 'Classical' (Adult Onset) PA There is a striking incidence of gastric autoantibodies in the serum of patients with Addisonian PA. PCA is detected in up to 90% of all patients [90, 135,143,167,176]. The incidence is somewhat lower in patients over 60 years of age [135,167,181], but this does not correlate with the known duration of disease [99, 143, 167]; however, the onset of atrophic gastritis cannot be defined in individual patients and probably exists for many years prior to diagnosis. Repeated tests for PCA in individual patients have not revealed disappearance or decrease in titer of PC A over periods of up to 9 years [184]. Nevertheless itis possible that the absence of PCA in a minority of patients with classical PAis due to total atrophy of the corpus mucosa and loss ofantigenic stimulus [52, 99]. The prevalence of overt PAis considered to be lower among some racial groups; however, studies in both Chinese [91] and Indians [40] indicate that gastric autoimmunity as judged by the frequency of PCA approximates that seen in Caucasian populations. Environmental factors such as high-dietary vitamin B12 intake may protect some racial groups against the onset of overt PA. PCA can also be detected in gastric juice [50,99,155] and mononuclear cells of the gastric mucosa [17] but with a lower frequency than in serum [155]. The frequency in gastric juice was 75% and in mucosal plasma cells was 60% [155]. It is present in serum predominately as IgG [38, 155] and in gastric juice as both IgG and IgA [50,155]. It is not known whether the IgA antibody in gastric juice is of secretory type; however, a lack of correlation observed between immunoglobulin classes of PC A in serum and gastric juice suggests that they are derived wholly or in part from different sources [155].
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C. Autoantibodies in Pernicious Anemia
19
Gastritis
Table I. Frequency of intrinsic factor antibody (IFA) in serum and gastric juice before and after acid dissociation in 40 patients with pernicious anaemia Author
Number IFA serum
gastric juice before after disdissociation sociation
ROSE and CHANARIN [136] GOLDBERG and BLUESTONE [65]
28 12
11 7
10 6
16 10
total (both sites)
19
IJ1
Type-I or blocking IFA is found in the serum of up 74% of patients with PA [141,143, 176] and type-II or binding IFA is detected in up to 48% of PA sera [62, 141, 143]. Type-II IFA appears to occur only when type-I IFA is also present [62, 143]. This difference in prevalence of the two types ofIF A may reflect a greater immunogenicity of the antigenic site for blocking antibody compared to that of binding antibody [144]. Both types ofIFA have also been detected in gastric juice [51,66,141,146,155]. In addition, it has been shown that type-I IFA may be entirely complexed to IF in gastric juice and will be detected only after acid dissociation of the complexes [65,136]. Using this technique, it has become evident that type-I IF A may be more common in PA gastric juice than in serum [65,136] (table I). Type-II IFA has been observed in gastric mucosal mononuclear cells but with a lower frequency (30%) than in serum or gastric juice [155]. IFA is predominately ofIgG class in serum and has been characterized as IgG or IgA in gastric juice [65,155]. Light chain heterogeneity of IF A has been demonstrated in serum of patients with PA [21], indicating that gastric autoantibodies do not result from a primary derangement of a single clone of antibody-forming cells. In two instances IF A in gastric juice has been identified as secretory IgA [65,66]. However, the immunoglobulin class of type-II IFA-containing mononuclear cells in the gastric mucosa was shown to be solely of IgG type by a double-labelling technique [18]. These studies of the frequency and immunoglobulin class of peA or IF A in various sites thus indicate that while the local (gastric) secretory IgA system may contribute to gastric autoantibody formation, these phenomena are not based solely on this system. This contrasts significantly with the predominant role of IgA-secreting cells in response to ingested exogenous antigens.
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lOne patient without IFA in serum or gastric juice had agammaglobulinemia.
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2. Juvenile Vitamin B12 Deficiency Three distinct forms of juvenile vitamin B12 deficiency exist [69,76,85, 119,123]. In one the gastric mucosa is normal and acid secretion is preserved, but there is a congenital lack of IF secretion leading to an early onset (usually before 2 years of age) of vitamin B12 deficiency. Gastric autoantibodies are absent in these patients, and gastric mucosal damage has not been observed later in life [123]. However, there are a few reports [111,123,124] of classical adult onset PA occurring in family members of patients with congenital IF deficiency, and it has been suggested that a partial defect in IF secretion might increase susceptibility to autoimmune gastric damage in later life [134]. In the Griisbeck-Imerslund syndrome the gastric mucosa is normal and secretes both acid and IF, but a selective defect in absorption of vitamin B12 across the ileal mucosa leads to vitamin B12 deficiency. Gastric autoantibodies have in general not been observed [114], although in one report PCA was observed in 5 of 19 patients [134]. The third form usually has an onset of vitamin B12 deficiency in later childhood, and here the gastric lesion is similar to that observed in classical adult onset P A. Both IFA and PCA are found, and there is a very strong association with polyendocrine disorder and moniliasis [125].
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Fig. 4. Macroscopic appearance of gastric mucosa in PA showing preserved antral folds (right) in contrast to thin atrophic corpus mucosa lacking rugae (left).
Gastritis
21
D. Autoantibodies in Atrophic Gastritis Previous reports of the frequency of PCA in atrophic gastritis being as high as 60% [28,52] may not reflect its true prevalence in this disease. A recent biopsy study carried out in a random sample of an adult Finnish rural population revealed CAG in 28% of the sample [153]. The incidence of PCA in this sample was 8%, and IFA was not detected in any subject [93]. Chronic gastritis was present in 90% of subjects with PCA. Within the group with CAG the ratio of those with PCA to those without PCA was 1 : 4. PCA was predominately found in females. The incidence of CAG and PCA rose with increasing age. Other studies are a\-ailable which confirm that PCA is rarely if ever found in the presence of a healthy gastric mucosa [2,49,188] and that the incidence of both PCA [33,96,185] and CAG [15,82,102, 153] rise with advancing age. A complete survey of an Australian rural population over 20 years of age revealed the prevalence of PC A in serum to be 4.8%; it rose from 2.5% in the 21-30 age group to 9.6% in persons over 80 years of age [80]. Thus PCA appears to be less frequent in CAG than in PA, and serum IFA is rarely observed. When the latter is discovered, further investigation will often reveal a profound decrease in IF secretion and vitamin B12 malabsorption which then becomes by definition PA.
Earlier studies in patients with CAG with or without gastric autoantibodies could not clearly separate the two groups on the basis of morphologic changes in the gastric corpus mucosa or in its function [28,49,169]. However, recent attention to the structure and function of the antral mucosa provides clear evidence for a differing distribution of gastritis among patients with or without gastric autoantibodies. The availability of a sensitive radioimmunoassay for the antral hormone gastrin led to its measurement in the serum of patients with PA [118]. Fasting gastrin levels were found to be greatly elevated above normal and were often in the range seen in the ZollingerEllison syndrome [118,60]. Reference to earlier studies of the gastric mucosa in PA provided the basis for an understanding of the mechanism of hypergastrinemia in this disease. The gastric atrophy in PAis confined to the corpus mucosa, and the antral mucosa is spared [35, 115, 122] (fig. 4,5). The lack of acid inhibition of gastrin release from an intact antral mucosa thus seemed to be an adequate
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E. Gastric Morphology and Function in Atrophic Gastritis
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a
Fig. 5. Corpus mucosa (a) from a patient with PA. PCA and IFA positive. Serum gastrin elevated. Note gastric atrophy with intestinal metaplasia. HE. x 250. Antral mucosa (b) from a patient with PA showing essentially normal structure. HE. x 250.
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b
Gastritis
23
a
b
explanation for the hypergastrinemia of PA [118]. It was subsequently found that hyper gas trinemi a is also present in patients without PA but with CAG and circulating gastric autoantibodies [60, 106]. Further, in patients with CAG without circulating autoantibodies serum gastrin levels were not elevated despite the presence of achlorhydria [106]. These patients were shown to have a severe antritis, whereas in the former group the antral mucosa was essentially normal [61,156] (fig. 5-7). Utilizing the serum gastrin response to a standard protein meal as an indirect measure of the gastrin (G) cell mass, it was shown that patients with
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Fig. 6. Corpus (a) and pyloric antral (b) mucosa from a patient with indigestion and achlorhydria who developed gastric carcinoma 21 years after \::iopsy diagnosis of atrophic gastritis. Gastric antibodies negative. Fasting gastrin normal. Note multifocal atrophy and intestinal metaplasia in corpus and severe atrophy and metaplasia in antrum. HE. x 50 (a); x 150 (b).
24
STRICKLAND
1,200
N
1
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,
900
•
,.
2
3
••
0
600
I
.§ OJ
a.
,,-
:s
300
Gastritis R.G.
STRICKLAND
Division of Gastroenterology, Department of Medicine, School of Medicine, University of New Mexico, Albuquerque, N. M.
Contents I. Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . II. Gastric Autoimmunity - Historical Aspects. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III. Gastric Autoantibodies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . A. Intrinsic Factor Antibodies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . B. Parietal Cell Antibody. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . C. Autoantibodies in Pernicious Anemia. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. Classical (adult onset) Pernicious Anemia. . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Juvenile Vitamin B12 Deficiency ................................... D. Autoantibodies in Atrophic Gastritis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E. Gastric Morphology and Function in Atrophic Gastritis ................ F. Significance of Gastric Autoantibodies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV. Cell-Mediated Immunity. .. . . .. . . .. . . . .. . . . . . . .. . . .. . . ... . . .. . . . . . . . . . . V. Immune Deficiency ...................................... , . . . . . . . .. . . . VI. Genetic Influences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII. Associated Diseases. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII. Effects of Corticosteroids. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IX. Experimental Autoimmune Gastritis ................. . . . . . . . . . . . . . . . . . . . X. Conclusions ......................................................... Acknowledgments. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
12 14 15 15 17 18 18 20 21 21 26 29 30 31 32 33 34 36 38 38
Several problems are encountered in attempting to classify human gastritis. Accurate diagnosis rests on a histological examination of the mucosa. While single specimens obtained by peroral biopsy are representative in the more severe forms of chronic gastritis, this may not be so when the inflam-
Downloaded by: Université René Descartes Paris 5 193.51.85.197 - 12/24/2019 8:55:20 AM
I. Introduction
STRICKLAND
13
mation or atrophy has a focal distribution. Most previous studies of gastritis have focused on morphological and functional disturbances in the gastric corpus, and little attention has been given to simultaneous investigation of the pyloric antral mucosa which, on the basis of histology and function, must be considered a separate structure (fig. I). Short gastric pits and straight glands lined by parietal or acid-producing cells and chief or pepsinogen-producing cells distinguish the corpus mucosa. The antral mucosa takes on a more villous appearance; the glands are tortuous and lined for the most part by mucussecreting cells. Interspersed between these cells are several types of enterochromaffin cells; the best characterized being the gastrin secreting (G) cells. Many potential pathogenic factors have been identified in gastritis, but few of them are sufficiently well-documented to justify attempts to classify this disease on etiological grounds. Finally it is clear that gastritis is often clinically silent, and its prevalence in the general population is uncertain. This has raised doubts as to its significance once discovered. A generally accepted classification is that based on morphology: (A)
Downloaded by: Université René Descartes Paris 5 193.51.85.197 - 12/24/2019 8:55:20 AM
Fig.I. Normal histological appearance of gastric pyloric antral mucosa (left) and corpus mucosa (right). HE. x 150.
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14
acute; (B) chronic, (1) superficial, (2) atrophic, (a) multifocal, (b) diffuse, (c) gastric atrophy; (C) indeterminate, (1) hypertrophic, (2) eosinophilic. This terminology is generally used to describe changes in the corpus mucosa, but a similar range of histological findings may occur in the antral mucosa. The only form of human gastritis in which substantial evidence favors the pathogenic role of immunologic factors is chronic atrophic gastritis (CAG). This includes the gastric lesion of pernicious anemia (PA) where the essential defect is failure of production of intrinsic factor (IF) to such a degree that absorption of vitamin B12 is severely impaired.
Investigation into the phenomenon of acquired resistance to oral hog IF in patients with PA resulted in the discovery by SCHWARTZ [149] in 1958 of a serum factor which interfered with the action of hog IF in promoting vitamin B12 absorption. In the same year TAYLOR and MORTON [166] reported the induction of antibody to IF in rabbits immunized with human gastric juice and the ability of this antibody to block IF mediated vitamin B12 absorption. TAYLOR [164] and SCHWARTZ [150] then demonstrated independently a similar inhibitory factor in the y-globulin fraction of serum from patients with PA who had never received oral treatment with hog IF. The hypothesis that the inhibitor was an autoantibody to IF was subsequently confirmed in 1962 by the studies of JEFFRIES et al. [97] who showed that the y-globulin fraction ofPA serum could combine with IF-vitamin B12 complex and retard its electrophoretic mobility in starch gel. In the same year complement-fixing antibody to a saline extract of human gastric mucosa was demonstrated in PA sera by IRVINE et al. [89] and MARKSON and MOORE [116], and TAYLOR et al. [167] showed by immunofluorescence that this antibody reacted specifically with gastric parietal cells. These studies were the first to identify a direct link between diffuse chronic inflammatory atrophy of the gastric mucosa and organspecific immunologic abnormalities. They marked the beginning of a new era of investigation into the pathogenesis of this disease. This chapter presents a review of subsequent studies, both human and experimental, which have been directed towards a better understanding of the significance and potential pathogenic role of immunologic disturbances in CAG and P A. Two recent reviews are available [110,160] which cover more fully the clinical importance and sequelae of CAG and the pathogenic role of nonimmunologic factors.
Downloaded by: Université René Descartes Paris 5 193.51.85.197 - 12/24/2019 8:55:20 AM
II. Gastric Autoimmunity - Historical Aspects
Gastritis
15
III. Gastric Autoantibodies
It is now established that autoantibodies against two separate components of the gastric parietal cell may be identified in patients with CAG or PA. It is of interest that despite involvement of chief cells in the atrophic damage, antibodies to constituents of the pepsin-producing cells have not been demonstrable so far.
A. Intrinsic Factor Antibodies (IF A)
Fig.2. Diagrammatic illustration of antibodies to IF. Type I or 'blocking' antibody reacts with the vitamin B12 binding site. Type II or 'binding' antibody reacts with a site distant from the B12 binding site and may combine with IF alone or IF-vitamin B12 complex.
Downloaded by: Université René Descartes Paris 5 193.51.85.197 - 12/24/2019 8:55:20 AM
IF, a glycoprotein with a molecular weight of approximately 60,000 [70] is secreted by the gastric parietal cell in man [78). Its function in promoting absorption of dietary vitamin B12 relates to its capacity to bind the vitamin [63] and form a stable complex for transport from the stomach to receptors on the distal ileal epithelial cell microvilli where active absorption of vitamin B12 takes place [31,41,75]. Normally IF is secreted in amounts vastly in excess of that required to maintain adequate vitamin B12 absorption [7). Thus vitamin B12 absorption may often be preserved even in the presence of advanced atrophic gastritis and total achlorhydria. Two distinct antibodies to IF (IF A) have been identified (fig. 2). Type I or blocking antibody [1,6,144,145] is directed against the binding site of vitamin B12 on the IF molecule and blocks subsequent combination of IF with free
16
vitamin. Type II or binding antibody [97,142,143,167] is directed against an antigenic site distant from that related to vitamin B12 binding and thus may react with IF alone or the preformed IF - vitamin B12 complex. IF which has reacted with type-II antibody is still capable of combining with type-I antibody, thus confirming the existence of two distinct antigenically reactive sites on the IF molecule [62,145]. Several techniques have been described for detecting and quantifying IF A. Type-I IF A is detected by demonstrating that radioactively labelled vitamin B12 is inhibited from binding to IF of normal human gastric juice by the serum being tested. Quantitation of type-I IFA is dependent upon separating free-labelled vitamin B12 from the larger IF-vitamin B12 complex by such methods as dialysis [1], charcoal [6, 67], zinc sulphate-barium sulphate precipitation [139], or zirconyl phosphate gel [72]. Increased sensitivity in detecting type-I IFA is reported [71] utilizing pretreatment of serum with unlabelled vitamin B12 (saturating B12-binding sites of serum proteins) and subsequent adsorption of unbound excess to albumin-coated charcoal. This maneuver also eliminates false-positive results which can arise from recent vitamin B12 injections. The charcoal-binding test is usually preferred for routine testing, and a reversed charcoal test using sera containing high titer type-I antibody can be used for the assay of IF in gastric juice [6,67,98]. Detecting and quantifying type-II IFA requires separation of IF-vitamin B12 complex from antibody-bound IF-vitamin B12 complex and electrophoresis [97,167] immune co precipitation [32,95,140,145] and radio immunodiffusion [142] have been used for this purpose. The salt precipitation method is the one preferred for routine testing. Both antibodies may also be tested for in one system utilizing uptake of radioactive vitamin B12 by guinea pig ileal mucosal homogenate and altering the order of reactant addition to detect either type-lor type-II IFA [12]. A recent report describes a radioimmunoassay for simultaneous detection of both antibodies and evidence is presented for its greater sensitivity over earlier techniques [141]. Immunofluorescence is generally held to detect antibody to a component of the gastric parietal cell which is distinct and separate from intrinsic factor (vide infra). However, using elaborate immunoabsorptive techniques, FISHER and TAYLOR [54] have demonstrated in testing sera known to contain IFA that a part of the fluorescence seen, using human fundic mucosa as substrate, can be attributable to IF antibody. It is also believed that IF antibodies do not fix complement; however, some evidence to the contrary has recently been reported [94].
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Fig. 3. Immunofluorescent staining of parietal cells in human gastric corpus mucosa by serum from a patient with PA. Note the diffuse cytoplasmic pattern of staining.
It is probable that the complement-fixing antibody to gastric mucosal homogenate [89,116] and that detected by immunofluorescence and seen to react diffusely with the cytoplasm of parietal cells (fig.3) [167] are directed against the same antigen. The immunofluorescent technique, however, is the more sensitive of the two. The antigen is organ- and cell-specific, showing no cross-reaction with other parts of the gastrointestinal tract or the thyroid. It is not species specific [127], being present even in the 'Hauptdriisen-Zellen' of amphibian stomach [163]. Ultracentrifugation of gastric mucosal homogenate reveals the antigen to be confined to the microsomal fraction. It is destroyed by papain and trypsin and inactivated by surface active detergents but not by treatment with phospholipases, ribonuclease, or dilute alkali [19]. Recent work [183] has strengthened the belief that the parietal cell antigen is a lipoprotein and is associated with smooth cytoplasmic membranes. The application of IgG fractions of PCA containing serum, coupled to horseradish peroxidase, onto sections of
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B. Parietal Cell Antibody (PCA)
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human gastric mucosa indicated a localization of bound antibody to the microvillus membrane of the canalicular system of the parietal cell [79]. The routine detection of PCA is best accomplished by the immunofluorescent technique using unfixed cryostat sections of gastric fundic mucosa. Where possible, human fundic mucosa from patients with duodenal ulcer should be used. Rat mucosa should no longer be used due to the presence in some human sera of a heteroantibody to rat parietal cells [126]. This heteroantibody is not indicative of gastric mucosal damage, whereas the parietal cell autoantibody is such an indicator [158]. The complement fixation technique is utilized where it is desirable to estimate antibody titer [87].
1. 'Classical' (Adult Onset) PA There is a striking incidence of gastric autoantibodies in the serum of patients with Addisonian PA. PCA is detected in up to 90% of all patients [90, 135,143,167,176]. The incidence is somewhat lower in patients over 60 years of age [135,167,181], but this does not correlate with the known duration of disease [99, 143, 167]; however, the onset of atrophic gastritis cannot be defined in individual patients and probably exists for many years prior to diagnosis. Repeated tests for PCA in individual patients have not revealed disappearance or decrease in titer of PC A over periods of up to 9 years [184]. Nevertheless itis possible that the absence of PCA in a minority of patients with classical PAis due to total atrophy of the corpus mucosa and loss ofantigenic stimulus [52, 99]. The prevalence of overt PAis considered to be lower among some racial groups; however, studies in both Chinese [91] and Indians [40] indicate that gastric autoimmunity as judged by the frequency of PCA approximates that seen in Caucasian populations. Environmental factors such as high-dietary vitamin B12 intake may protect some racial groups against the onset of overt PA. PCA can also be detected in gastric juice [50,99,155] and mononuclear cells of the gastric mucosa [17] but with a lower frequency than in serum [155]. The frequency in gastric juice was 75% and in mucosal plasma cells was 60% [155]. It is present in serum predominately as IgG [38, 155] and in gastric juice as both IgG and IgA [50,155]. It is not known whether the IgA antibody in gastric juice is of secretory type; however, a lack of correlation observed between immunoglobulin classes of PC A in serum and gastric juice suggests that they are derived wholly or in part from different sources [155].
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C. Autoantibodies in Pernicious Anemia
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Table I. Frequency of intrinsic factor antibody (IFA) in serum and gastric juice before and after acid dissociation in 40 patients with pernicious anaemia Author
Number IFA serum
gastric juice before after disdissociation sociation
ROSE and CHANARIN [136] GOLDBERG and BLUESTONE [65]
28 12
11 7
10 6
16 10
total (both sites)
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IJ1
Type-I or blocking IFA is found in the serum of up 74% of patients with PA [141,143, 176] and type-II or binding IFA is detected in up to 48% of PA sera [62, 141, 143]. Type-II IFA appears to occur only when type-I IFA is also present [62, 143]. This difference in prevalence of the two types ofIF A may reflect a greater immunogenicity of the antigenic site for blocking antibody compared to that of binding antibody [144]. Both types ofIFA have also been detected in gastric juice [51,66,141,146,155]. In addition, it has been shown that type-I IFA may be entirely complexed to IF in gastric juice and will be detected only after acid dissociation of the complexes [65,136]. Using this technique, it has become evident that type-I IF A may be more common in PA gastric juice than in serum [65,136] (table I). Type-II IFA has been observed in gastric mucosal mononuclear cells but with a lower frequency (30%) than in serum or gastric juice [155]. IFA is predominately ofIgG class in serum and has been characterized as IgG or IgA in gastric juice [65,155]. Light chain heterogeneity of IF A has been demonstrated in serum of patients with PA [21], indicating that gastric autoantibodies do not result from a primary derangement of a single clone of antibody-forming cells. In two instances IF A in gastric juice has been identified as secretory IgA [65,66]. However, the immunoglobulin class of type-II IFA-containing mononuclear cells in the gastric mucosa was shown to be solely of IgG type by a double-labelling technique [18]. These studies of the frequency and immunoglobulin class of peA or IF A in various sites thus indicate that while the local (gastric) secretory IgA system may contribute to gastric autoantibody formation, these phenomena are not based solely on this system. This contrasts significantly with the predominant role of IgA-secreting cells in response to ingested exogenous antigens.
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lOne patient without IFA in serum or gastric juice had agammaglobulinemia.
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2. Juvenile Vitamin B12 Deficiency Three distinct forms of juvenile vitamin B12 deficiency exist [69,76,85, 119,123]. In one the gastric mucosa is normal and acid secretion is preserved, but there is a congenital lack of IF secretion leading to an early onset (usually before 2 years of age) of vitamin B12 deficiency. Gastric autoantibodies are absent in these patients, and gastric mucosal damage has not been observed later in life [123]. However, there are a few reports [111,123,124] of classical adult onset PA occurring in family members of patients with congenital IF deficiency, and it has been suggested that a partial defect in IF secretion might increase susceptibility to autoimmune gastric damage in later life [134]. In the Griisbeck-Imerslund syndrome the gastric mucosa is normal and secretes both acid and IF, but a selective defect in absorption of vitamin B12 across the ileal mucosa leads to vitamin B12 deficiency. Gastric autoantibodies have in general not been observed [114], although in one report PCA was observed in 5 of 19 patients [134]. The third form usually has an onset of vitamin B12 deficiency in later childhood, and here the gastric lesion is similar to that observed in classical adult onset P A. Both IFA and PCA are found, and there is a very strong association with polyendocrine disorder and moniliasis [125].
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Fig. 4. Macroscopic appearance of gastric mucosa in PA showing preserved antral folds (right) in contrast to thin atrophic corpus mucosa lacking rugae (left).
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D. Autoantibodies in Atrophic Gastritis Previous reports of the frequency of PCA in atrophic gastritis being as high as 60% [28,52] may not reflect its true prevalence in this disease. A recent biopsy study carried out in a random sample of an adult Finnish rural population revealed CAG in 28% of the sample [153]. The incidence of PCA in this sample was 8%, and IFA was not detected in any subject [93]. Chronic gastritis was present in 90% of subjects with PCA. Within the group with CAG the ratio of those with PCA to those without PCA was 1 : 4. PCA was predominately found in females. The incidence of CAG and PCA rose with increasing age. Other studies are a\-ailable which confirm that PCA is rarely if ever found in the presence of a healthy gastric mucosa [2,49,188] and that the incidence of both PCA [33,96,185] and CAG [15,82,102, 153] rise with advancing age. A complete survey of an Australian rural population over 20 years of age revealed the prevalence of PC A in serum to be 4.8%; it rose from 2.5% in the 21-30 age group to 9.6% in persons over 80 years of age [80]. Thus PCA appears to be less frequent in CAG than in PA, and serum IFA is rarely observed. When the latter is discovered, further investigation will often reveal a profound decrease in IF secretion and vitamin B12 malabsorption which then becomes by definition PA.
Earlier studies in patients with CAG with or without gastric autoantibodies could not clearly separate the two groups on the basis of morphologic changes in the gastric corpus mucosa or in its function [28,49,169]. However, recent attention to the structure and function of the antral mucosa provides clear evidence for a differing distribution of gastritis among patients with or without gastric autoantibodies. The availability of a sensitive radioimmunoassay for the antral hormone gastrin led to its measurement in the serum of patients with PA [118]. Fasting gastrin levels were found to be greatly elevated above normal and were often in the range seen in the ZollingerEllison syndrome [118,60]. Reference to earlier studies of the gastric mucosa in PA provided the basis for an understanding of the mechanism of hypergastrinemia in this disease. The gastric atrophy in PAis confined to the corpus mucosa, and the antral mucosa is spared [35, 115, 122] (fig. 4,5). The lack of acid inhibition of gastrin release from an intact antral mucosa thus seemed to be an adequate
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E. Gastric Morphology and Function in Atrophic Gastritis
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a
Fig. 5. Corpus mucosa (a) from a patient with PA. PCA and IFA positive. Serum gastrin elevated. Note gastric atrophy with intestinal metaplasia. HE. x 250. Antral mucosa (b) from a patient with PA showing essentially normal structure. HE. x 250.
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b
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a
b
explanation for the hypergastrinemia of PA [118]. It was subsequently found that hyper gas trinemi a is also present in patients without PA but with CAG and circulating gastric autoantibodies [60, 106]. Further, in patients with CAG without circulating autoantibodies serum gastrin levels were not elevated despite the presence of achlorhydria [106]. These patients were shown to have a severe antritis, whereas in the former group the antral mucosa was essentially normal [61,156] (fig. 5-7). Utilizing the serum gastrin response to a standard protein meal as an indirect measure of the gastrin (G) cell mass, it was shown that patients with
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Fig. 6. Corpus (a) and pyloric antral (b) mucosa from a patient with indigestion and achlorhydria who developed gastric carcinoma 21 years after \::iopsy diagnosis of atrophic gastritis. Gastric antibodies negative. Fasting gastrin normal. Note multifocal atrophy and intestinal metaplasia in corpus and severe atrophy and metaplasia in antrum. HE. x 50 (a); x 150 (b).
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