Chest
Volume 115, Issue 3, March 1999, Pages 829-835
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Laboratory and Animal Investigations
Bacterial Endotoxin Is an Active Component of Cigarette Smoke

https://doi.org/10.1378/chest.115.3.829Get rights and content

Background

Chronic bronchitis in cigarette smokers shares many clinical and histologic features with environmental lung diseases attributed to bacterial endotoxin (lipopolysaccharide [LPS]) inhalation. Experimental LPS inhalation mimics many of the acute effects of cigarette smoke in the lower airway. Therefore, we reasoned that LPS may be a biologically active component of cigarette smoke.

Design

The Limulus amebocyte lysate (LAL) assay was used to measure LPS in the tobacco and filter tip components of unsmoked 1R4F experimental cigarettes and commercially available “light” cigarettes, as well as in mainstream (MS) and sidestream (SS) smoke particles generated with an automated smoking machine and collected on ventilator mainflow filters.

Setting and participants

Blood LPS activity and plasma cytokine concentrations were measured in groups of healthy smokers and nonsmokers who reported to the walk-in clinic at the Baltimore VA Medical Center for unrelated complaints.

Measurements

Blood LPS levels were measured by LAL assay and plasma levels of tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), soluble TNF receptors I and II (sTNFR I and sTNFR II) were measured by enzyme-linked immunosorbent assay.

Results

Bioactive LPS was detected in both the tobacco portion (1R4F, 17.8 ± 1.0μ g/cigarette; light, 26.8 ± 7.3 μg/cigarette [mean ± SE]) and filter tips (1R4F, 0.67 ± 0.55 μg/cigarette; light, 0.70 ± 0.39 μg/cigarette) of cigarettes. Bioactive LPS was also detected in both MS (1R4F, 120 ± 64 ng/cigarette; light: 45.3 ± 16 ng/cigarette) and SS smoke (1R4F, 18 ± 1.5 ng/cigarette; light: 75 ± 49 ng/cigarette). Although systemic absorption of inhaled LPS may occur, we failed to detect any differences between nonsmokers and smokers in median blood LPS levels (median values, 66.75 and 72.1 pg/mL, respectively; p = 0.55) or plasma concentrations of TNF-α (0 vs 0 pg/mL, respectively; p = 0.71), sTNFR I (1,469 vs 1,576 pg/mL, respectively), sTNFR II (2,011 vs 3,110 pg/mL, respectively), or IL-6 (8.8 vs 0 pg/mL, respectively; p = 0.20).

Conclusions

Smoking one pack of cigarettes per day delivers a dose of respirable LPS that is comparable to the levels of LPS associated with adverse health effects in cotton textile workers. Thus, we suggest that the bioactive LPS in cigarette smoke may contribute to the pathogenesis of chronic bronchitis that develops in susceptible cigarette smokers.

Section snippets

Reagents

Pyrogen-free water, chromogenic Limulus amebocyte lysate (LAL) assay reagents using the diazo-coupling method, LPS standard from Escherichia coli 0113:H10 (specific activity, 9.73 endotoxin units [EU])/ng), and pyrogen-free polystyrene microtiter plates were obtained from Associates of Cape Cod (Falmouth, MA). E coli 0111B4 LPS prepared by trichloroacetic acid precipitation was obtained from Sigma Chemical Co (St. Louis, MO); its specific activity was determined to be 1 EU/ng in our laboratory.

LPS Content in Unsmoked Cigarettes

We measured the levels of extractable LPS bioactivity in unsmoked 1R4F research cigarettes and commercially available light cigarettes. The tobacco and filter tip portions of the cigarettes were analyzed separately (Fig 1). Tobacco from 1R4F and light cigarettes contained LPS bioactivity equivalent to 17.8 ± 1.0 and 26.8 ± 7.3 μg LPS/cigarette (mean ± SE), respectively. The unsmoked filter tips from these cigarettes contained LPS bioactivity equivalent to 0.67 ± 0.55 and 0.70 ± 0.39 μg

Discussion

Based on the reported LPS content in other agricultural products, we reasoned that cigarette tobacco would contain biologically significant quantities of LPS, one of the most potent inflammatory stimuli known. We extracted 6 to 9 μg of biologically active LPS from each gram of tobacco in either research or commercially available cigarettes. This LPS level is similar to levels reported in other noncombusted agricultural products.2 Temperatures at the burning cigarette tip (880°F or 471°C)19

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    Presented in part at the Thomas L. Petty Aspen Lung Conference, Aspen, CO, June 7–10, 1995.

    Supported by Public Health Service grant CA52741 and HL40945. Drs. Dubin and Costa were supported by Pulmonary Research Training Fellowships from the American Lung Association of Maryland.

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