A study to investigate the effect of inhaled and intranasal pathogens on pro-infammatory markers using (existing) techniques in order to acquire experience with these techniques.

This is a phase 0 study to investigate the effects of inhaled and intranasal lipopolysaccharide (LPS) on pro-inflammatory markers sampled by hypertonic saline-induced sputum (SI)
and nasal lavage (NAL) in healthy subjects. The main objectives are to set up the SI and LPS challenge techniques, to evaluate the sampling success, to investigate the pro-inflammatory effects of inhaled and intranasal LPS, and to evaluate the repeatability of the LPS inflammatory response. In part 1 of the study LPS will be substituted by normal saline.



This study contributes to further development of medicine of obstructive airway diseases.



Subjects will be recruited in the Netherlands.

To acquire experience with sputum induction and LPS challenge techniques.

After spututum induction challenges, after LPS challenges and during the course of the study (safety endpoints).

Primary objectives Part 1:

1. To set up the SI and LPS (in part 1 substituted by Normal Saline) challenge techniques;

2. To evaluate the sampling success (i.e., the percentage of analysable sputum and NAL samples), pre- and post-LPS challenge; in terms of non-squamous cell counts (total cell numbers and differentials);

3. To identify “LPS-responders”, defined as those subjects with an absolute increase (pre- vs. post challenge) in sputum neutrophil count of >=10%.



Primary objectives part 2:

1. To investigate the pro-inflammatory effects of inhaled LPS (at a previously applied dose of 60 μg) within the lower airways of healthy subjects, by hypertonic saline-induced (NaCl 4.5%) sputum;

2. To investigate the pro-inflammatory effects of low-dose intranasal LPS (at a previously applied dose using the split nose method) within the upper airways of healthy subjects, using NAL;

3. To evaluate the repeatability of the LPS inflammatory response, all procedures need to be repeated with the same subjects in a second identical study period;

4. Additional subjects need to be tested, to validate the techniques used.



Intervention Part 1:

On day 1, after baseline spirometry, sputum will be induced by inhalation of hypertonic saline (NaCl 4.5% aerosol generated by ultrasonic nebulization), during 3 times 5 min (at approximately 15 min intervals). Sputum samples will be collected in a clean plastic container on melting ice and processed within 2 hrs. This will serve as the baseline sputum sample. On day 2, an inhaled LPS* challenge will be performed (approx. 24 hrs after baseline SI) followed by SI 6 hrs later (approx. 30 hrs after baseline SI); to assess the acute LPS effects in the lower airways. On day 3, an intranasal LPS challenge will be performed, using the split-nose method, followed by a NAL 6 hrs post challenge.



Intervention Part 2:

Part One study participants (those actually exposed to SI and LPS challenge, will also be invited to participate in a second study period (washout >=3 weeks), identical to the first study period (to test SI and LPS challenge repeatability). Only those will be invited again, from whom an analysable sputum sample could be obtained during Part One. To validate the techniques used, additional subjects will be included, each of them will participate in two identical study periods as well. For individual subjects, all study related measurements will be performed in similar within-day time frames (±3 hrs).