Overview
The goal of this observational study is to learn whether two inflammatory proteins, interleukin-8 (IL-8) measured in saliva and interleukin-6 (IL-6) measured in blood serum, can help distinguish adults with oral potentially malignant disorders (OPMDs), such as oral lichen planus and leukoplakia, from adults with healthy oral mucosa, and whether smoking changes these protein levels.
The main questions it aims to answer are:
- Do salivary IL-8 and serum IL-6 levels differ between people with OPMDs and healthy controls, and does this difference depend on smoking status?
- How well do IL-8 and IL-6, alone or combined, discriminate people with OPMDs from healthy controls?
Researchers compared healthy never-smokers, healthy current smokers, never-smokers with OPMDs, and current smokers with OPMDs. An additional exploratory group of current smokers with oral squamous cell carcinoma (OSCC) was compared with current smokers with OPMDs.
Participants provided an unstimulated whole saliva sample and a blood sample at a single visit, and completed a clinical, periodontal, and tobacco-use assessment. No treatment was assigned as part of the study.
Description
Design: Cross-sectional observational diagnostic study, reported with reference to the STROBE guidelines.
Population: 250 adults (18 years or older). The primary cohort comprised 200 participants in four equal groups (n = 50 each): healthy never-smokers, healthy current smokers, never-smokers with histopathologically confirmed OPMDs, and current smokers with histopathologically confirmed OPMDs. OPMDs included oral lichen planus and oral leukoplakia; lesion subtype, anatomical site, and presence and grade of epithelial dysplasia were recorded. An exploratory cohort of 50 current smokers with histopathologically confirmed oral squamous cell carcinoma (OSCC) was analyzed separately and was not part of the primary comparisons.
Assessments: Smoking status (never vs. current) and tobacco exposure (daily cigarette consumption, smoking duration, shisha use, electronic-cigarette use, dual-tobacco use); demographic data; plaque index, percentage of sites with bleeding on probing, and periodontal status (healthy, gingivitis, periodontitis); histopathological classification (dysplasia for OPMDs, tumor grade for OSCC).
Biomarkers: Unstimulated whole saliva was collected for IL-8 and serum for IL-6. Concentrations were measured by immunoassay and expressed in pg/mL.
Statistical analysis: Group comparisons used t-tests or Mann-Whitney U tests and chi-square or Fisher's exact tests. Linear regression models of log-transformed IL-8 and IL-6 included disease status, smoking status, and their interaction, with HC3 robust standard errors. Logistic regression evaluated each biomarker individually and jointly, with adjustment for age, sex, and smoking, and an exploratory model adding periodontal variables. ROC analysis estimated AUCs with 95% bootstrap confidence intervals; sensitivity and specificity were calculated at Youden-index thresholds. Exploratory prediction models (logistic regression) were evaluated with repeated stratified five-fold cross-validation (20 repetitions), comparing four predictor sets: IL-8; IL-8 + IL-6; both biomarkers + smoking status; and these plus demographic and periodontal variables.
Sample size: With 100 OPMD patients and 100 controls, two-sided alpha = 0.05 and 80% power, the minimum detectable difference was approximately 0.40 SD, and the minimum detectable AUC against 0.50 was approximately 0.61.
Eligibility
Inclusion Criteria:
All participants:
- Age 18 years or older
- Written informed consent provided before enrollment
- Smoking status classifiable as never-smoker or current smoker
Healthy controls:
- Clinically healthy oral mucosa
- No clinical evidence or previous history of oral potentially malignant disorders (OPMDs) or oral cancer
OPMD group:
- Clinically identified oral lichen planus or oral leukoplakia, confirmed histopathologically
- No definitive treatment of the lesion before biological-sample collection
OSCC group (exploratory):
- Histopathologically confirmed oral squamous cell carcinoma
- Current smoker
Exclusion Criteria:
- Active oral infection unrelated to the study lesion
- Recent major dental surgery or invasive dental procedure
- Active or recent systemic infection
- Uncontrolled diabetes mellitus
- Major systemic inflammatory or autoimmune disease
- Clinically significant salivary gland dysfunction
- Current systemic treatment likely to substantially affect inflammatory cytokine concentrations (e.g., systemic corticosteroids or immunomodulatory agents)
- Essential clinical data or smoking-status classification unavailable
- Biological samples insufficient, unsuitable for analysis, or failing predefined laboratory quality-control criteria
