Comparative Analysis of 4NQO- and Ethanol-Induced Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma in Male and Female Mice: Association With Peripheral Blood Inflammatory Markers.

Background: Considering that peripheral blood biomarkers are prognostic predictors for several human tumors, this study aimed to comparatively analyze the association of hematological alterations with the incidence of epithelial dysplasia (ED) and oral squamous cell carcinoma (OSCC) in male and female mice treated with 4-nitroquinoline-N-oxide (4NQO) and ethanol (EtOH).

Methods: 120 C57Bl/6J mice (60 males and 60 females) were allocated to four groups (n = 15). They were treated firstly either with 5 mg/mL propylene glycol (PPG) or 100 μg/mL 4NQO in the drinking water for 10 weeks, followed by sterilized water (H₂O) or 8% EtOH (v/v) for 15 weeks, as follows: PPG/H₂O, PPG/EtOH, 4NQO/H₂O, and 4NQO/EtOH (CEUA-UFU, #020/21). After killing, tongues were collected for histopathological analysis of ED and OSCC. Blood samples were processed for complete blood count and calculation of neutrophil-to-lymphocyte ratio (NLR) and systemic immune-inflammation index (SII).

Results: The incidence of OSCC in females from the 4NQO/EtOH group (60%) was lower when compared to males (93%). Neutrophils, NLR, and SII increased from control animals (PPG/H₂O and PPG/EtOH) to ED and OSCC-bearing male and female mice (4NQO/H₂O and 4NQO/EtOH), while lymphocytes decreased. Females from the 4NQO/H₂O group with ED also had higher neutrophils, NRL, and SII values than females with normal tongues.

Conclusion: The low incidence of 4NQO- and ethanol-induced OSCC in females indicates that the sex bias in OSCC may not be associated with extrinsic risk factors alone. Neutrophil and lymphocyte counts, NRL, and SII were significantly altered during multistep carcinogenesis and thus could be explored as biomarkers for ED and OSCC development.