Biochemical, immunohistochemical, and molecular biomarkers in rats following inhalation of cigarette mainstream smoke (MS) or a mixture of cigarette sidestream smoke (SS) and MS
Presented at Frontiers in Aerosol Dosimetry Research
Objective: Identify new biomarkers responsive to cigarette smoke in a rat inhalation model for reduced exposure research. Rats were exposed to either fresh air, MS, or SS/MS-mix (89% SS / 11% MS) from the reference cigarette 2R1 for up to 4 weeks (5d/week, 2x3 h/day). Total particulate matter (TPM) concentrations were 100 and 250 microg/l for MS and 100 microg/l for SS/MS-mix. At equal TPM doses, neutrophils in bronchoalveolar lavage fluid (BALF), as a marker for inflammation, were increased mainly with MS. Several biomarkers were seen predominantly with SS/MS-mix: 10-fold increase in 4-ABP adducts, decreased frequency of immature erythrocytes, and 15% body weight reduction. For both SS/MS-mix and MS there was a 2-fold increase in 8-OHdG adducts (lung immunohistochemistry), as well as an increase in cytochrome P450(CYP)1A1 activity (lung homogenates) and a decrease in CYP2B1 (confirmed by gene expression analysis). There was no effect on lung epithelial permeability (albumin in BALF), bone marrow micronuclei, or CYP2E1 activity. With MS250 only, a 20% decrease of apolipoprotein-a1 in serum and 3-fold increase of LDH in BALF were seen. Gene expression analysis, done for MS100 only, showed differential regulation of metabolism- and stress-related genes. 4-ABP adducts and 8-OHdG adducts were confirmed for SS/MS-mix and shown to respond to MS. Lung inflammation was identified as mainly an MS effect. The ratio of CYP1A1 to CYP2B1 activity and differential gene regulation were identified as promising biomarkers. This multi-endpoint approach, which can discriminate SS/MS-mix and MS at similar TPM concentrations, may be useful for evaluating potentially reduced exposure prototypes.