Eradication of Helicobacter pylori may be beneficial in the management of Alzheimer’s disease
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- Kountouras, J., Boziki, M., Gavalas, E. et al. J Neurol (2009) 256: 758. doi:10.1007/s00415-009-5011-z
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Infectious agents have been proposed as potential causes of Alzheimer’s disease (AD). Recently, we documented a high prevalence of Helicobacter pylori (Hp) infection in patients with AD. We aim to access the effect of Hp eradication on the AD cognitive (MMSE: Mini Mental State Examination and CAMCOG: Cambridge Cognitive Examination for the Elderly) and functional (FRSSD: Functional Rating Scale for Symptoms of Dementia) status parameters. In the first part of the study, a total of 50 consecutive patients with AD and 30 age-matched anaemic controls underwent an upper gastrointestinal endoscopy, and gastric mucosal biopsies were obtained to detect the presence of Hp infection by histologic analysis and rapid urease test. Serum anti-Hp-specific IgG level was analysed by enzyme-linked immunosorbent assay. In the second part, Hp-positive AD patients received a triple eradication regimen (omeprazole, clarithromycin and amoxicillin), and all patients were followed up for 2 years, while under the same treatment with cholinesterase inhibitors. Hp was detected in 88% of AD patients and in 46.7% of controls (P < 0.001). Hp eradication was successful in 84.8% of treated patients. At the 2-year clinical endpoint, cognitive and functional status parameters improved in the subgroup of patients where Hp eradication was successful (P < 0.001 and P = 0.049 for MMSE and CAMCOG, respectively; P < 0.001 for FRSSD), but not in the other patients. Hp eradication may positively influence AD manifestations, suggesting a possible common link between Hp and AD.
KeywordsAlzheimer’s diseaseHelicobacter pyloriHistologic analysisRapid urease test
Alzheimer’s disease (AD) is by far the most common cause of dementia of ageing ; it is a horribly debilitating disease that will increase in prevalence as the populations of the USA and Europe continue to age. The disease is characterised by progressive impairment in memory, visuospatial skills, complex cognition, language and personality in its earlier stages.
In general, cognitive performance in ageing individuals is frequently adequate until the individual suffers a challenge, including severe infection; systemic infections are vulnerable to cognitive decline, and pharmacologic strategies to decrease neuroinflammation associated with infection might be important for reducing neurobehavioural deficits in the elderly [3, 10]. Specifically, although the early events underlying AD remain uncertain, the consideration that microorganisms can cause AD has recently been addressed [15, 19, 46]; infiltration of the brain by pathogens acts as a trigger or co-factor for AD, with Herpes simplex virus type 1 (HSV1) and Chlamydophila (Chlamydia) pneumoniae being most frequently implicated [15, 46]. These pathogens may cause the neurological damage that results in AD by eliciting inflammation. In this regard, an infection-based animal model demonstrates that following intranasal inoculation of BALB/c mice with Chlamydia pneumoniae, amyloid plaques/deposits consistent with those observed in the AD brain develop, thereby implicating this infection in the aetiology of AD .
Helicobacter pylori (Hp) is a gram-negative, spiral, flagellated bacterium, comprising more than 1,400 genes, that colonises the gastric mucosa of most humans worldwide, mainly affecting older adults in the developed world, including Greece . It is associated with various upper gastrointestinal (GI) diseases [6, 43] and has also been implicated in a variety of extradigestive vascular conditions, including ischaemic heart disease , ischaemic cerebrovascular  and functional vascular disorders caused by vascular dysregulation, frequently detected in AD.
The association of Hp infection (Hp-I) and AD has also only recently been addressed by three studies [24, 26, 31]. A higher seropositivity for anti-Hp IgG antibodies was reported in patients with AD than in age-matched controls, but this serological test has limitations because it does not discriminate between current and old infections . Such a distinction is essential because current Hp-I induces humoral and cellular immune responses that, owing to the sharing of homologous epitopes (molecular mimicry), cross-react with components of nerves [25, 26], thereby affecting or perpetuating neural tissue damage. Moreover, eradication of Hp-I might delay AD progression, particularly at early disease stages, including mild cognitive impairment (MCI). Based on the histological analysis of gastric mucosa biopsy for the documentation of Hp-I, Kountouras and associates  reported a higher prevalence of Hp-I in patients with AD than in age-matched controls, accompanied by increased homocysteine (Hcy) concentration, an independent risk factor for dementia and AD, thereby suggesting an association between these two diseases. The authors also reported similar data in patients with MCI compared with age-matched controls . Demonstrating the association of Hp and AD and proving the benefit of eradicating Hp-I in the clinical course of the disease may have a major impact on its treatment. However, before antibiotic therapy for Hp-I becomes an established step in the management of AD, sufficient evidence must be provided that AD parameters are positively influenced by the eradication of Hp-I.
The objective of this series was to evaluate the effect of Hp eradication on cognitive and functional status parameters of patients with AD. We have therefore designed methods to confirm and quantify our hypothesis that Hp eradication therapy has a beneficial effect on these AD parameters in Hp-positive patients with AD.
Materials and methods
This was a two-part series. Part 1 was designed to evaluate the prevalence of Hp-I infection in AD. Fifty consecutive patients with documented AD and 30 age-matched anaemic controls were included in this part of the study.
All participants had been referred to the Memory and Dementia Outpatient Clinic by their caregivers, mainly relatives, who certified a cognitive deterioration and/or other cognitive functional disturbances in the participants for at least a period of 6 months. Patients were diagnosed with probable Alzheimer’s dementia according to the NINCS-ADRDA and DSM-IV criteria . Screening procedure for their evaluation was conducted at their first visit to the Memory and Dementia Outpatient Clinic. Patients and controls underwent neuropsychological assessment that included measurement for cognitive deterioration (MMSE: Mini Mental State Examination and CAMCOG: Cambridge Cognitive Examination for the Elderly), functional disorders (FRSSD: Functional Rating Scale for Symptoms of Dementia), neuropsychological disorders (NPI: Neuropsychiatric Inventory) and depression (GDS: Geriatric Depression Scale, HDRS: Hamilton’s Depression Rate Scale). The aforementioned scale battery required 90 min on average. MMSE was used as a screening test for cognitive deterioration, assessing orientation in time and place, naming, repetition, immediate and late recall, ideational apraxia and constructional praxis (total score 30). CAMCOG was used for a more thorough investigation of the above-mentioned cognitive functions (total score 107). FRSSD assessed patients’ ability to carry out routine tasks such as eating, dressing and toileting (total score 42). FRSSD total score over seven revealed functional difficulties in everyday living, while total score 5–7 was regarded as borderline. The twelve-item NPI was the only scale provided to the caregiver assessing patient’s neuropsychiatric symptoms, such as hallucinations, apathy-indifference, aggressiveness, sleep and eating disorders and depression. Both frequency (scale 1–4) and severity (scale 1–3) of every disorder was measured (total score 120). Depression being the major cause of dementia was regarded as an exclusion criterion. It was estimated through scales of GDS (total score 15, cutoff 5) and HDRS (total score 55, cutoff 16), both provided by the patients themselves. Neuropsychological assessment was conducted by the same neuropsychologist for all patients. Apart from the above-mentioned assessment, MRI tomography was conducted as diagnostic neuroimaging technique in order to confirm temporal lobe and hippocampal formation atrophy. It was also used in order to exclude other causes of dementia (stroke, tumour, fronto-temporal dementia, etc.). Patients with vascular, Lewy-body, fronto-temporal and other types of dementia were excluded from the study. We also excluded patients with known or subclinical thyroid disorders as well as patients with depression. None of the patients had previously been treated with cholinesterase inhibitors (ChEIs), memantine or any other pharmacological treatment for dementia.
Inclusion of cognitively normal controls required MMSE score >24, CAMCOG score >85 and FRSSD score <5. Moreover, in addition to scores above cutoff in cognitive tests, subjective memory complaints should be absent for a control participant in order to be regarded as cognitively normal.
All patients and controls underwent diagnostic upper GI endoscopy after informed consent. Apart from upper GI endoscopy, the control subjects underwent lower endoscopy to investigate mild iron-deficiency anaemia; the GI mucosa appeared to be without obvious macroscopic abnormalities. Participants were excluded if they had taken H2-receptor antagonists, proton pump inhibitors, antibiotics, bismuth compounds or nonsteroidal anti-inflammatory drugs in the preceding 4 weeks. Participants were also excluded if they had undergone previous gastric surgery; received anticoagulant therapy; were alcohol abusers; had allergy to penicillin or macrolides; had gastric cancer or other neoplasms; or had severe cardiac, pulmonary, kidney or liver disease.
All participants and/or their relatives signed a consent form prior to enrollment, and the study protocol was approved by the local ethics committee. All patients received the same ChEI during the 2-year follow-up period of the study. None of the participants in this study received oral drugs that could influence cognitive state, other than the medication prescribed by the researchers.
According to standard clinical practice in Europe, the Hp eradication regimen comprised a 1-week course consisting of omeprazole (20 mg bid), clarithromycin (500 mg bid), and amoxycillin (1 g bid), followed by omeprazole 20 mg once daily for 1 month .
Hp detection methods were described previously . Biopsy urease test and histopathology process were also described previously . Success of the Hp eradication regimen was evaluated by control endoscopy at least 8 weeks after cessation of therapy, and patients were considered as Hp negative if both histology and the rapid urease test proved negative. The neuropsychologist assessing cognitive and functional state in this study was masked to the Hp status of the patients.
The follow-up study population was classified into three AD groups: patients for whom Hp eradication treatment was successful (group A); those for whom eradication of Hp had failed, and they refused and/or were noncompliant with their eradication therapy (group B); and those who were Hp negative at baseline (group C, Fig. 1).
Mann–Whitney U-test, Fisher’s exact test, chi-square, odds ratios, 95% CI, two-tailed t-test and one-way analysis of variance were used. Significance was set at P < 0.05. The analysis was performed by using the statistical software package SPSS (Statistical Package for Social Sciences, version 13.0; SPSS Inc., Chicago, IL).
Helicobacter pylori positivity in patients with Alzheimer’s disease and anaemic controls
Patients (n = 50)
Controls (n = 30)
Odds ratio (95% CI)
Mean ± SD age (range), years
65.0 ± 6.9 (53–80)
62.2 ± 8.6 (44–70)
Positive urease test (gastric mucosa)
Mean ± SD serum anti H. pylori IgG (U/ml)
34.0 ± 40.1
17.0 ± 18.1
Histologically confirmed presence of H. pylori
Outcome of Hp eradication therapy
One Hp-negative and four Hp-positive patients finally refused to participate and were excluded from the second part of the study. Of the remaining 40 Hp-positive AD patients, 33 received and 7 refused to receive eradication therapy. Hp eradication was successful in 28 of these 33 (84.8%) patients (group A, Fig. 1). Treatment was unsuccessful in the remaining five patients; apart from the 7 Hp-positive AD patients who refused to receive eradication therapy, 16 additional Hp-positive patients were recruited in the second part of the study who also refused to receive eradication treatment, thereby remaining Hp-positive throughout the follow-up period (group B, Fig. 1). All group A patients were compliant with their eradication therapy as determined by the number of tablets and capsules remaining after therapy. Adverse effects were generally mild, including mild abdominal pain, occasional nausea or vomiting, diarrhoea and stomatitis. None of the patients discontinued their therapy because of these mild adverse effects.
When compared with baseline values (49.3 ± 30.4 U/ml), the mean serum IgG anti-Hp level was significantly reduced in group A patients at 3-month follow-up (24.5 ± 12.9 U/ml) (P < 0.001). In group B patients, this parameter had increased at 3-month follow-up (32.6 ± 14.7 U/ml at 3 months vs. 28.2 ± 11.2 U/ml baseline; P = 0.03). In group C patients, who did not receive Hp eradication therapy, both mean serum IgG anti-Hp values at baseline and at 3 months were within normal limit levels (6.7 ± 2.4 U/ml at 3 months vs. 7.1 ± 2.1 U/ml baseline; P = 0.04).
Outcome of AD parameters
Baseline Mean Mini Mental State Examination (MMSE) (F = 0.986, dfwg = 60, dfbg = 2, P = 0.379), Cambridge Cognitive Test (CAMCOG) (F = 1.238, dfwg = 60, dfbg = 2, P = 0.302) and Functional Rating Scale for Symptoms of Dementia (FRSSD) (F = 1.646, dfwg = 60, dfbg = 2, P = 0.202) scores did not differ significantly among the three groups.
Comparison of mean Mini Mental State Examination (MMSE), Cambridge Cognitive Examination for the Elderly (CAMCOG) and Functional Rating Scale for Symptoms of Dementia (FRSSD) parameters for all patients with Alzheimer’s disease at baseline and after 1 and 2 years of follow-up
Mean ± SD measurement value
17.46 ± 6.09
19.6 ± 6.08
19.92 ± 5.94
17.07 ± 6.15
14.39 ± 7.1
11.25 ± 5.87
21.2 ± 5.63
17.8 ± 9.67
16.4 ± 11.01
52.07 ± 24.3
57.71 ± 25.49
62.86 ± 22.42
55 ± 17.58
46.4 ± 19.98
38.3 ± 15.55
65.5 ± 14.84
56 ± 35.35
53.33 ± 25.38
12.53 ± 6.81
10.38 ± 5.98
9.15 ± 6.52
15.32 ± 8.15
18.6 ± 7.59
21.53 ± 8.21
10.2 ± 2.48
10 ± 4.06
17.2 ± 8.98
MDM (95% CI)
Change from baseline at 1 year
2.14 (2.82 to 1.46)
−2.67 (−1.55 to −3.8)
−3.4 (2.05 to −8.85)
5.64 (8.7 to −2.58)
−8.6 (−4.38 to −12.81)
−9.5 (174.74 to −193.73)
−2.15 (−1.05 to −3.24)
3.28 (5.01 to 1.54)
−0.2 (4.47 to −4.87)
Change from baseline at 2 years
2.46 (3.28 to 1.64)
−5.82 (−4.68 to −6.95)
−4.8 (2.35 to −11.95)
5.13 (10.23 to −0.028)
−18.3 (−16.27 to −20.32)
−17.66 (17.84 to −53.17)
3.38 (−2.1 to −4.66)
6.15 (8.92 to 3.38)
7 (19.19 to −5.19)
In the first part of this series, by documenting a higher prevalence of Hp-I in an AD cohort compared with an age- and sex-matched control group, we established for the first time a significant relationship between Hp-I infection and AD . Hp-I was determined by histologic detection of organisms in mucosal biopsy specimens, considered as the actual gold standard for the diagnosis of this infection. A higher seropositivity for anti-Hp IgG antibodies was also reported in 30 patients with AD than in age-matched controls .
It is important to consider whether the rate of Hp-I in the control group has been negatively influenced by the coexistence of anaemia. Existing data show that anaemia does not protect against Hp development. Anaemic controls have been used before , and the frequency of Hp-I in the anaemic control group matches that of the general population in Greece and that reported for other ethnic populations . Moreover, it is unlikely that individuals with iron-deficiency anaemia are protected against Hp-I because it is thought that the infection is actually associated with iron- and/or vitamin B12-deficiency anaemia . In addition, eradication of Hp-I may be associated with reversal of iron and/or vitamin B12 deficiency and improvement in anaemia .
In the second part of the study we obtained an acceptable eradication rate of 84.8%. Similar eradication rates have been achieved by others . Moreover, Hp eradication was beneficial for the cognitive and functional state in patients in whom Hp was successfully eradicated (group A), thereby possibly altering the progressive nature of AD and increasing survival of AD patients. This might not be the case in groups B and C despite the fact that all groups were maintained on the same ChEI they received at baseline. In this regard, parameters significantly associated with reduced survival at diagnosis include mainly increased severity of cognitive impairment and decreased functional level strongly related with shortened survival [4, 28]. Longer term follow-up, however, is required to evaluate the possible beneficial effect of Hp eradication therapy.
The results of this study suggest that eradication therapy may somehow improve the degenerative process in AD. It should be noted, however, that the number of patients in the groups, particularly in group, C was small, and it may not, therefore, be possible to draw definitive conclusions. Future studies are needed to focus on the influence of Hp on the degenerative process in the brain. It is conceivable that Hp induces relative mechanisms and/or agents such as the synthesis of various mediators (e.g., cytokines), which may be detrimental to the degeneration of the demented brain. Specifically, there is evidence for abnormal cellular immune and apoptotic mechanisms playing an important role in the Hp-associated GI pathologies and potentially affecting the neurodegenerative process in AD. Interestingly, molecular mimicry of host structures by the saccharide portion of lipopolysaccharides of the GI pathogens Campylobacter jejuni (C. jejuni) and Hp is thought to be connected with the development of autoimmune sequelae observed in neuropathies. Campylobacter jejuni, a principal cause of gastroenteritis, is the most common antecedent infection in Guillain-Barré syndrome (GBS), an inflammatory autoimmune neuropathy sharing comparable pathogenic mechanisms with AD . Chemical analyses of the core oligosaccharides of neuropathy-associated C. jejuni strains have revealed structural homology with human gangliosides. Serum antibodies against gangliosides are found in one-third of patients with GBS, but are generally absent in enteritis cases. Collective data suggest that the antibodies are induced by antecedent infection with C. jejuni, and subsequently react with nerve tissue, causing damage , possibly by apoptosis. In addition, 46% of patients with GBS have specific IgG antibodies to VacA of Hp in the cerebrospinal fluid, which are probably associated with some components of the peripheral nerve myelin, thereby showing considerable demyelination of the peripheral nerves; the sequence homology found between VacA and human [Na(+)/K(+)] ATPase A subunit suggests that antibodies to VacA involve ion channels in abaxonal Schwann cell plasmalemma, resulting in demyelination in these patients [5, 27]. In this regard, it is relevant to speculate that such anti-Hp-mediated apoptotic mechanisms might also lead to degeneration of ganglion cells, thereby contributing to AD neuropathy or other degenerative neuropathies, such as glaucoma, defined as ocular AD. Support for this theory is provided by our observations indicating that the titre of anti-Hp IgG antibodies in the aqueous humour of patients with glaucoma may reflect the severity of glaucomatous damage .
The question raised is how exactly Hp-I influences the pathophysiology of AD. This bacterium may be involved in the pathophysiology of AD by one of the following mechanisms: (1) Promoting platelet and platelet–leukocyte aggregation [24, 26]. Platelet activation and aggregation have also been proposed to play possible pathophysiologic roles in the development of AD . (2) Inducing chronic atrophic gastritis with a concomitant decrease in vitamin B12 and folate concentrations, thereby increasing the concentration of Hcy, an independent risk factor for AD and a potent contributor to vascular disorders implicated in endothelial damage and neurodegeneration via oxidative injury. Specifically, an increased serum Hcy concentration has been shown in our AD patients with Hp-I . Chronic gastritis, as a result of Hp-I, can lead to malabsorption of vitamins B12 and folate, which results in failure of methylation by 5-methyl-tetrahydrofolic acid and hence accumulation of Hcy . The elevated Hcy, in turn, could trigger endothelial damage and neurodegeneration via oxidative injury and result in atherothrombotic disorders and AD. (3) Releasing large amounts of proinflammatory and vasoactive substances, such as cytokines [interleukin (IL)-1, IL-6, IL-8, IL-10, IL-12, tumour necrosis factor (TNF)-α, interferon-γ], eicosanoids (leucotrienes, prostaglandins catalysed by cyclo-oxygenase enzymes) and acute phase proteins (fibrinogen, C-reactive protein) [24, 25], involved in a number of vascular disorders possibly including AD and other AD-related neuropathies such as glaucoma . (4) Stimulating mononuclear cells to produce a tissue factor-like procoagulant that converts fibrinogen into fibrin . (5) Causing the development of cross mimicry between endothelial and Hp antigens. (6) Producing reactive oxygen metabolites (ROMs) and circulating lipid peroxides [24, 25], which have been implicated in the pathophysiology of AD [31, 45]; accumulating evidence suggests that ROMs are potent deleterious agents causing cell death or other forms of irreversible cell damage, and oxidative stress participates in the neuronal loss in AD [25, 48]. ROMs accumulation impairs endothelial barrier function and promotes leucocyte adhesion, induces alterations in normal vascular function and might result in the development of AD , events that are also triggered in Hp-induced GI injury . (7) Influencing the apoptotic process that plays a potential role in the pathogenesis of many neurodegenerative diseases including AD, glaucomatous neuropathy [24, 25] or Down syndrome; the latter predisposes to the early onset of the neurodegeneration of AD . In particular, increased endothelin-1 (a potent constrictor of arterioles and venules), nitric oxide (NO) and inducible nitric oxide synthase (iNOS) levels are associated with Hp-I . Relevant data in AD indicate that endothelin-1-like immunoreactivity in the AD brains is significantly increased in the frontal and occipital cortex compared to those in control brains, thereby explaining the decreased cerebral blood flow in AD patients . Besides, recent evidence in humans indicates that the expression of the nitrergic system, the synthesis of NO, the peroxynitrite reactive production and protein tyrosine nitration are activated over the entire course of AD, and that the presence of amyloid-beta peptide (Abeta) increases the presence of neuronal nitric oxide synthase (nNOS) and iNOS isoforms over the course of AD in pyramidal-like neurones ; the overproduction of NO, the increase in both peroxynitrite and superoxide production, the mitochondrial membrane depolarisation and the caspase activation contribute to neuronal death [7, 17], mainly via apoptosis. NO is a rapidly diffusing gas and a potent neurotoxin that may facilitate the apoptotic death of retinal ganglion cells in glaucomatous optic neuropathy [11, 51], and probably in AD neuropathy [7, 32]. Further studies, however, are needed to clarify the aforementioned points.
Finally, a few shortcomings need to be addressed in our series. In contrast to data presented for other intracellular infectants [Chlamydia pneumoniae, Herpes simplex virus (HSV)-1, Borrelia species] found in brain regions demonstrating considerable AD pathology [2, 14, 30, 37, 38, 42], most of the aforementioned mechanisms of how Hp infection could influence the pathophysiology of AD are not necessarily specific to the regional pathology observed in the AD brain, nor is there a specific temporal sequence indicated by which Hp infection would directly injure the brain, at the present time. However, Hp, an extracellular bacterium, could affect the brain and other target organs, such as the heart, indirectly, through the release of numerous cytokines, including TNF-α acting at a distance. TNF-α and IL-6 (TNF-α is the main trigger for the production of IL-6 by a variety of cells) play important roles in the regulation of the synthesis of other acute phase proteins, which are established risk factors for atherosclerosis, such as fibrinogen and factor VIII. These cytokines also have profound effects on lipid metabolism directly at the site of the atherosclerotic lesion, but could influence the atheroma process through blood circulating levels, distant production of cytokines or through stimulating circulating white blood cells to produce them, thereby contributing to the pathogenesis of brain diseases including AD [11, 36, 49].
In addition, a Hp eradication regimen might have influenced other infections in the AD, particularly those of Chlamydia peneumoniae and Borrelia species, and could explain, at least partly, the positive improvement observed in our AD patients at the post-treatment period. However, the prevalence of Chlamydia peneumoniae and possibly Borrelia species infections are very low in both child and adult Greek populations [44, 52, 53], and, moreover, our participants did not report or present any characteristic clinical features suggesting the aforementioned infectious diseases.
Besides, we did not investigate the APOE gene (NCBI Entrez gene 348), the strongest genetic risk factor for later onset AD . In this respect, there is evidence suggesting that some relationship exists between the APOE4 gene product and the pathobiology of the intracellular Chlamydophila (Chlamydia) pneumoniae involved in AD pathogenesis [8, 9]. Moreover, the risk of developing AD is much greater in patients HSV-1-positive in brain who also possess an APOE4 allele than in those with only one of these factors . Therefore, future relative studies might clarify the potential relationship between this gene and the pathobiology of the extracellular Hp species.