Nontypeable Haemophilus influenzae activates the NLRP3 inflammasome and leads to Caspase-1 dependent secretion of IL-1beta – a possible pathway of exacerbations in COPD

The inflammasome is a cytosolic protein complex important for early inflammatory responses of the innate immunity. Inflammasome activation has been described for several types of infection, and our data indicate that also nontypeable Haemophilus influenzae (NTHi) induces the assembly of the inflammasome components. The inflammasome consists of NOD-like receptors such as NLRP3, the adaptor protein ASC and Caspase-1 which cleaves pro-IL-1β into its active form.

Murine macrophages (RAW 264.7) and human lung tissue were stimulated with NTHi 106cfu/ml for 24–48h. Supernatants and cells were collected to measure the inflammatory response using cytokine ELISA, PCR and Western Blot. To assess the relevance of the inflammasome for the inflammatory response, a Caspase-1 inhibitor (CI) was added 8h after in-vitro infection.

Western Blot analysis of cells and lung tissue showed the activation of Caspase-1 after NTHi infection and a significant induction of IL-1β expression (RAW: control 24h and 48h at the lower detection limit vs. NTHi 24h 408±64pg/ml and NTHi 48h 717±72pg/ml, both n=6, p<0.01). Moreover, the NTHi-dependent expression of NLRP3 in respiratory cells was demonstrated (fold change to β-actin: control 24h 147±28 vs. NTHi 24h 197±28, n=9, p<0.05), whereas ASC expression was significantly reduced by NTHi (fold change to β-actin: control 24h 89±7 vs. NTHi 24h 44±2, n=4, p<0.05). Inhibition of Caspase-1 in human lung tissue led to a significant reduction of IL-1β levels (IL-1β: NTHi 24h 408±64pg/ml vs. NTHi+CI 24h 174±12pg/ml, n=6, p<0.01; NTHi 48h 717±72pg/ml vs. NTHi+CI 48h 432±49pg/ml, n=6, p<0.01).

Our data clearly demonstrate the participation of the NRLP3-inflammasome in NTHi-induced inflammation in pulmonary cells and tissues. Our findings concerning Caspase-1 mediated IL-1β-upregulation emphasize the role of the inflammasome in respiratory tract infections. Furthermore, these results provide new insights into the pathogenesis of infectious exacerbations in COPD.