Interaction Between Cigarette Smoke and Mycoplasma Infection: A Murine Model

Figures & data

Figure 1 Four groups of mice were studied over an eight week time interval. Group 1 mice were exposed to cigarette smoke for 6 hours per day, 5 days per week. Group 2 mice had the same cigarette exposure as group 1 and additionally had Mycoplasma pneumoniae (Mp) respiratory infection induced at weeks 2 and 6. Group 3 mice only had Mp inoculations at weeks 2 and 6 while group 4 mice had saline inoculation at these time points. *All mice were sacrificed at week 8 for study analyses.

Table 1 Bronchoalveolar lavage cell counts

Groups	Total Cell Counts (×10^4/ml)	% Macs	% Black Macs	% Neuts	% Lymphs	% Eos
1 Smoking alone	17.1 ± 6.5	99.0 ± 0.4	67.0 ± 2.2**	0.8 ± 0.3	0.2 ± 0.1	0.03 ± 0.03
2 Smoking plus Mp	20.5 ± 6.1	88.1 ± 3.3^+	60.8 ± 5.4**	0.6 ± 0.6	11.3 ± 3.3^dag	0.0 ± 0.0
3 Mp alone	16.7 ± 2.4	77.3 ± 3.3^+	0 ± 0	8.0 ± 1.2^+ +	12.0 ± 3.0^dag	0.0 ± 0.1
4 Saline control	6.0 ± 0.8*	95.7 ± 0.9	0 ± 0	3.1 ± 0.8^+ +	1.5 ± 0.5	0.2 ± 0.1

Mp = Mycoplasma pneumoniae, Macs = macrophages, Black macs = macrophages that engulf particulate cigarette smoke, Neuts = neutrophils, Lymphs = lymphocytes, Eos = eosinophils. Mean ± SEM.

* p = 0.0003 group 4 vs. 1–3. ⁺ p = 0.0002 groups 2 or 3 vs. 1 or 4. ** p = 0.001 groups 1 or 2 vs. groups 3 or 4. ⁺⁺ p = 0.001 groups 3 or 4 vs. 1 or 2 and p = 0.005 groups 3 vs. 4. ^dag p = 0.0009 groups 2 or 3 vs. groups 1 or 4.

Figure 2 There is greater neutrophil degranulation (immunoperoxidase stain, arrows) in the outer wall of a small airway (diameter ≤ 200 μm) for a smoking plus Mp mouse (group 2) compared to a smoking without Mp mouse (group 1).

Table 2 Measurement of fixed and specific lung volume

	1	2	3	4
Groups	Smoking alone	Smoking plus Mp	Mp alone	Saline control
Fixed lung volume (ml)	0.67 ± 0.03	0.66 ± 0.04	0.66 ± 0.02	0.60 ± 0.017
Specific lung volume (lung volume/body weight, ml/g)	0.035 ± 0.002	0.035 ± 0.002	0.030 ± 0.001**	0.027 ± 0.001**

Mp = Mycoplasma pneumoniae.

Data expressed as Mean ± SEM.

* p = 0.001 group 4 vs. 1–3.

** p = 0.001 groups 1 or 2 vs. groups 3 or 4.

Figure 3 Lung tissue relative expression levels of MMP-9 mRNA by real-time RT-PCR. Group 1, smoking only is used as the baseline standard. Group 2, smoking plus Mp has a 10-fold increase in MMP-9 compared to group 1. Group 3 = Mp only and group 4 = saline control. Data expressed are medians with interquartile ranges. Group 2 is significantly increased compared to the other groups. Sample sizes are reduced due to RNA quality variability, only good quality accepted based on housekeeping gene 18sRNA.