Clinical Trial Details
— Status: Completed
Administrative data
| NCT number |
NCT02976922 |
| Other study ID # |
H-15012527 |
| Secondary ID |
|
| Status |
Completed |
| Phase |
N/A
|
| First received |
|
| Last updated |
|
| Start date |
March 14, 2016 |
| Est. completion date |
July 7, 2017 |
Study information
| Verified date |
November 2022 |
| Source |
University of Copenhagen |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Interventional
|
Clinical Trial Summary
Recurrent aphthous stomatitis (RAS) is one of the most common ulcerative diseases affecting
the oral mucosa. The aetiology remains unknown, but several local, systemic, immunologic,
genetic, allergic, nutritional, and microbial factors have been proposed as causative agents.
Clinically, RAS is characterised by recurrent bouts of one or several rounded, shallow,
painful oral ulcers at intervals of a few months or days. The aim of this study is to
characterise the salivary and faecal microbiome in 20 patients with RAS and compare the
findings with those of 20 healthy controls. The study also includes a double-blind randomized
placebo-controlled intervention with probiotics (Lactobacillus reuteri-containing lozenges 2
tablets daily for 3 months) or placebo. The salivary and faecal microbiome in RAS patients is
compared before and after treatment. This study will improve our understanding of the
pathogenesis in RAS and provide us with knowledge on potential future therapeutic approaches.
Description:
Purpose of the study The purpose of this study is to characterise the salivary and faecal
microbiome in patients with recurrent aphthous stomatitis (RAS) and to compare the findings
to those of a matched healthy control group. Another purpose is to investigate the effect of
treatment with probiotics on the microbiome in whole saliva and faeces as well as the
severity and number of RAS outbreaks. Moreover, a smear will be taken from present aphthous
ulcers before and after treatment with probiotics in order to characterise the local
microbiota.
It is assumed that the microbiome in whole saliva and faeces from patients with RAS (minor or
major type aphthous ulcers) differs from the microbiome in whole saliva and faeces from age-
and gender-matched control persons.
It is also hypothesised that probiotic treatment has a beneficial effect on the mucosal pain
and reduces the severity of RAS and the frequency of outbreaks. Furthermore, it is assumed
that treatment with probiotic has an effect on microbiome in whole saliva and faeces from
patients with RAS.
Background Recurrent aphthous stomatitis (RAS) is one of the most common ulcerative diseases
affecting the oral mucosa. The prevalence varies from 5-25%. The aetiology remains unknown,
but several local, systemic, immunologic, genetic, allergic, nutritional, and microbial
factors have been proposed as causative agents.
Clinically, RAS is characterised by recurrent bouts of one or several rounded, shallow,
painful oral ulcers at intervals of a few months or days.
RAS can be classified into three different types: minor, major and herpetiform. Minor RAS
comprises about 80% of the cases. This type is characterised by aphthous ulcers with a
diameter of about 5 mm, often localised in the buccal and labial mucosa. The ulcers heal
within 7-10 days without formation of scars. Major RAS appear in 10-15% of the patients. The
ulcers are often crateriform with a diameter of 10-30 mm, and typically present on the labial
mucosa, the soft palate, the tonsillar region and oropharynx. The healing time may vary from
2 to 6 weeks and leaving cicatrix. These ulcers are very painful and food intake may be
compromised. The prevalence of the herpetiform type is 5-10%.
This type is often localised in the floor of the mouth and the ventral part of the tongue,
and it characterised by aggregation of multiple small ulcers.
The aetiology remains unknown, but a number of local and systemic factors are assumed
increase the predisposition, including mucosal trauma, stressful events, hormonal changes,
smoking cessation, allergy to various food substances as well as vitamin- and/or mineral
deficiency. The is also a genetic predisposition as the possibility of developing RAS is 90%
if both parents have RAS and 20% if one of the parents has RAS.
Microbial factors A number of microorganisms have been suggested involved in the
aetiopathogenesis of RAS. These include oral streptococci, especially Streptococcus mitis,
but the cross-reaction between oral streptococci and oral mucosal antigens is unspecific and
considered clinically insignificant. Helicobacter pylori, which is associated with gastritis
and duodenal ulcers may also be seen in dental plaque and therefore also be suggested
involved in the pathogenesis of RAS, but the anti-H. pylori seropositivity has not been found
increased in patients with RAS. Various vira have been found in biopsies from aphthous ulcers
such as cytomegalovirus and Epstein-Barr virus. Herpes simplex, varicella zoster and
adenovirus have also been assumed to play a role in the aetiopathogenesis. However, it is has
not been possible to show a causal relationship between virus infection and RAS, and viral
DNA may be present due to a secondary infection.
Novel technologies improve our ability to make more in depth analyses of the microbiota in
patients with RAS. While working on this study protocol one study on the microbiome in
patients with RAS has been published showing an increased proportion of Bacteroides species
in the oral mucosa of patients with RAS compared to healthy controls. Thus, as previous
studies also suggest the oral microbiota appears to play a role in the development of RAS as
well as in perpetuation of RAS.
In this study we will characterise the microbiome in whole saliva, faeces and the aphthous
ulcers of patients with RAS (with active lesions) and analyse how it differs from the
microbiome of healthy subjects. This characterisation could be essential for our
understanding the pathogenesis of RAS and contribute to the development of novel strategies
for managing these patients.
Immunopathological factors Several studies indicate that the pathogenesis of RAS includes a
number of cell-mediated mechanisms, but the exact immunopathogenesis is still not clarified.
Phagocytic and cytotoxic T-lymphocytes may be a role in the destruction of the oral
epithelium and the presence of these immune cells is regulated and maintained by local
release of cytokines. Patients with outbreak of RAS have an increased presence of gamma-delta
T-cells in the blood compared to healthy control persons and patients without active RAS.
Moreover, patients with RAS have an increased presence of Tumour Necrosis Factor (TNF)-alpha
compared to those who do not have RAS, but also other proinflammatory cytokines like
interleukin-2 and -6 are likely to play a role in the pathogenesis of RAS. Finally,
aphthous-like ulcers have been found associated to a number of inflammatory bowel diseases
such as ulcerative colitis and Crohn's disease and celiac disease as well as cyclic
neutropenia, HIV-infection and immunoglobulin A (IgA)-deficiency. Accordingly, an immune
component may be involved in RAS which contribute to a change in the intestinal microbiota or
vice versa. In this study, we will characterise the microbiota in saliva and in faeces from
patients with RAS before, during and after treatment with probiotics.
The study design and methods The project comprises a cross-sectional study which investigates
the microbiome in whole saliva and faeces from 20 patients with RAS and 20 healthy control
persons. The project also includes a double-blind randomized placebo-controlled intervention
with probiotics (Lactobacillus reuteri-containing lozenges 2 tablets daily for 3 months) or
placebo. The salivary and faecal microbiome in RAS patients is compared before and after
treatment.
The patients with RAS are recruited among patients referred to the Oral Medicine Clinic,
Faculty of Health and Medical Sciences, University of Copenhagen. The 20 healthy controls are
recruited via www.forsoegsperson.dk
Methods include:
- Interview re. onset of RAS, oral symptoms (assessed by means of a visual analogue
scale), comorbidity, medication intake, smoking and alcohol habits, oral hygiene habits.
- Collection of chewing paraffin-stimulated whole saliva.
- Oral examination including evaluation of aphthous ulcers (Ulcer Severity Score),
registration of dental status and periodontal status.
- A smear from one of the aphthous ulcers.
- A faecal sample will be collected by the subjects themselves in their homes after
thorough instruction and handled according to manual from the Statens Serum Institut.
From the sample 250 mg is collected and kept in a freezer kept at -80°C until further
processing and analysis.
- A blood test including haemoglobin (HgB), C reactive protein (CRP), iron, cobalamin,
folate, transferrin, ferritin and vitamin D levels.
Extraction of bacterial DNA from saliva, faecal and smear samples (at baseline, 7-day and
90-day of intervention with probiotics or placebo, respectively) will be carried out in the
laboratory at Department of Odontology, University of Copenhagen, using previously
established procedures based on instructions from the Human Microbiome Project.
Metagenomic analyses on extracted bacterial DNA from saliva, smear and faecal samples are
made by Beijing Genomics Institute. Illumina 16S rDNA sequencing allows determination of
bacterial taxonomy and phylogenetic diversity.
