Eosinophilic inclusions in rat Clara cells and the effect of an inhaled corticosteroid.

Large eosinophilic cytoplasmic inclusions (ECIs) are occasionally seen in untreated rat Clara cells. Following inhalation exposure to a corticosteroid, the number of ECIs was increased. This is the first histopathological description of rat ECIs and attempted characterization by immunohistochemistry, in situ hybridization, and electron microscopy. ECIs were strongly positive for surfactant protein D (SP-D) and weakly positive for Clara cell specific protein (CCSP). Clara cell cytoplasm was positive for CCSP mRNA regardless of ECIs, but not within ECIs. Corticosteroid treatment and ECI presence did not affect the immunohistochemistry and in situ hybridization staining intensities. Electron microscopy revealed large intracytoplasmic granules with an irregular limiting membrane. The ECI number was microscopically quantified in rats from three-, six-, and twenty-four-month studies. The mean ECI counts in treated rats increased from three- to fifty-four-fold with a positive dose-related trend, when compared with vehicle controls. Although the mechanism is unclear, SP-D and to a lesser extent CCSP accumulate in the ECIs. As human bronchial epithelium does not appear to contain structures analogous to the ECI, it is suggested that the observation of an increased number of ECIs in the treated rats is not likely to be relevant for human clinical risk assessment.

Immunolocalization of CC10 in Clara cells in mouse and human lung.

Two antisera, denoted R41 and R42, were raised against a synthetic peptide from the murine Clara cell-specific protein CC10, and one antiserum, denoted R40, was raised against human recombinant uteroglobin, the human homolog of murine CC10. Purified antigen-specific antisera, denoted R40AP, R41AP, and R42AP were prepared using peptide columns. The purified antisera were characterized by dot blots, immunohistochemistry, and immunoblots. Immunohistochemistry of mouse lung showed specific labeling of Clara cells in distal bronchioles by all three antisera. In human lung, the antiuteroglobin antiserum specifically labeled Clara cells, while the anti-mouse peptide antisera had weak crossreactivity and higher background staining. Electron microscopy revealed immunogold labeling of CC10 granules in Clara cells of mouse lung with all antisera. All antisera also labeled a 5-kDa protein on immunoblots of mouse lung homogenates. The surface epithelium of the alveolar air spaces around the distal bronchioles were CC10 positive suggesting a functional activity for CC10 in the lung parenchyma distal to Clara cells. R40AP immunohistochemical staining of sections of normal human lungs and lungs from patients with surfactant protein B deficiency, bronchopneumonia, and idiopathic alveolar proteinosis illustrate the utility of the anti-human CC10 antibody for diagnostic pathology.