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  • Effects of Short-term Electronic(e)-Cigarette Aerosol Exposure in the Mouse Larynx. The Laryngoscope Easwaran, M., Maria, C. S., Martinez, J. D., Hung, B., Yu, X., Soo, J., Kimura, A., Gross, E. R., Erickson-DiRenzo, E. 2023

    Abstract

    The effects of electronic cigarettes (e-cigarettes) on the larynx are relatively unknown. This study examined the short-term effects of e-cigarette inhalation on cellular and inflammatory responses within the mouse laryngeal glottic and subglottic regions after exposure to pod-based devices (JUUL).Male C57BL6/J mice (8-9 weeks) were assigned to control (n = 9), JUUL flavors Mint (JMi; n = 10) or Mango (JMa; n = 10). JUUL mice were exposed to 2 h/day for 1, 5, and 10 days using the inExpose inhalation system. Control mice were in room air. Vocal fold (VF) epithelial thickness, cell proliferation, subglandular area and composition, inflammatory cell infiltration, and surface topography were evaluated in the harvested larynges. Mouse body weight and urinary nicotine biomarkers were also measured. Chemical analysis of JUUL aerosols was conducted using selective ion flow tube mass spectrometry.JUUL-exposed mice had reduced body weight after day 5. Urinary nicotine biomarker levels indicated successful JUUL exposure and metabolism. Quantitative analysis of JUUL aerosol indicated that chemical constituents differ between JMi and JMa flavors. VF epithelial thickness, cellular proliferation, glandular area, and surface topography remained unchanged after JUUL exposures. Acidic mucus content increased after 1 day of JMi exposure. VF macrophage and T-cell levels slightly increased after 10 days of JMi exposures.Short-term e-cigarette exposures cause minimal flavor- and region-specific cellular and inflammatory changes in the mouse larynx. This work provides a foundation for long-term studies to determine if these responses are altered with multiple e-cigarette components and concentrations.N/A Laryngoscope, 2023.

    View details for DOI 10.1002/lary.31043

    View details for PubMedID 37698394

  • Method for Collecting Single Epithelial Cells from the Mouse Larynx. The Laryngoscope Khan, M. I., Easwaran, M., Martinez, J. D., Kimura, A., Erickson-DiRenzo, E. 2023

    Abstract

    The larynx is lined by specialized epithelial cell populations. Studying molecular changes occurring in individual epithelial cell types requires a reliable method for removing these cells from the larynx. Our objective was to develop a method to harvest individual epithelial cells from the mouse larynx while minimizing contamination from non-laryngeal sites and non-epithelial laryngeal cells.Mice were euthanized, and the larynx was carefully exposed and separated from non-laryngeal sites. A small dental brush was inserted into the laryngeal inlet and rotated to obtain epithelial cells. Cells were transferred to collection media, counted, and cytospin preparations stained for laryngeal epithelial (i.e., Pan-Keratin, EpCAM, NGFR, p63, K5, β-tubulin, MUC5AC) and non-epithelial (i.e., vimentin) cell markers. Histopathology was completed on brushed laryngeal tissue sections to evaluate the depth of cell collection. Preliminary Single-cell RNA sequencing (scRNA-seq) was performed to confirm this method can capture diverse laryngeal cell types.We collected 6000-8000 cells from a single larynx and 35000-40000 cells from combining brushings from three tissues. Histopathology demonstrated brushing removed the epithelial layer of the larynx and some underlying tissue. Immunofluorescence staining demonstrated the phenotype of harvested cells was primarily epithelial. Preliminary scRNA-seq was successfully conducted and displayed nine unique cell clusters.We developed a reliable method of harvesting individual epithelial cells from the mouse larynx. This method will be useful for collection of laryngeal cells for a variety of downstream cellular and molecular assays, including scRNA-seq, protein analyses, and cell-culture-based experiments, following laryngeal injury.N/A Laryngoscope, 2023.

    View details for DOI 10.1002/lary.30970

    View details for PubMedID 37602769