Background: Gliotoxin is a cytotoxic metabolite of Aspergillus fumigatus. However, little is known about its role in the cellular pathogenesis of invasive pulmonary aspergillosis. We therefore studied the effects of gliotoxin in mediating apoptosis and necrosis in alveolar epithelial type II pneumocytes (TIIP). Methods: TIIP cells were exposed to gliotoxin and studied via 3 assays. Terminal deoxynucleotidyl transferase assay (TUNEL) was used to assess apoptosis. DNA fragmentation by TUNEL was assessed by in situ through-end labeling of single and double strand DNA fragments using terminal deoxyribonucleotidyl transferase (TdT). The uptake of trypan blue (TB) into TIIP was used to detect necrosis associated with cell membrane disruption. The XTT assay quantifies cell viability by measuring cell metabolism after exposure of TIIP cells to gliotoxin. Results: TUNEL assay demonstrated dose dependent DNA fragmentation following exposure to gliotoxin. Exposure to 0.01, 1.0 and 10 ÂµM for 2hrs resulted in 12.5Â±4.42, 22.03Â±8.81 and 75.4Â±10.01 % DNA fragmentation respectively: exposure for 6hrs, at the same concentrations, resulted in 23.60Â±13.87, 34.8Â±12.66, and 80.1Â±1.5 % DNA fragmentation. The TB assay detected significant concentration and timepoint-dependent necrosis at concentrations ≥ 0.1ÂµM (p 8804; 0.004) in a reciprocal relation to the degree of apoptosis. The XTT assay detected significant concentration-dependent cytotoxicity at 24 hours (p 8804; 0.03) Conclusions: Gliotoxin mediates cytotoxicity of TIIP via apoptosis and necrosis. These data provide further insights into the possible role of gliotoxin in the cellular pathogenesis of invasive pulmonary aspergillosis.
Full conference title:
46th Interscience Conference on Antimicrobial Agents and Chemotherapy
- ICAAC 46th