Alcohol exacerbates murine pulmonary tuberculosis

Abstract

Alcohol consumption has been described as a risk factor for infection with Mycobacterium tuberculosis, but its contribution to tuberculosis has been difficult to isolate from other adverse socioeconomic factors. Our objective was to evaluate the impact of alcohol consumption on pulmonary infection with M. tuberculosis in a murine model. BALB/c mice were maintained on the Lieber-DeCarli liquid ethanol diet or a liquid control diet and infected intratracheally with low-dose M. tuberculosis H37Rv. Lung organism burdens, lung and lung-associated lymph node CD4(+)- and CD8(+)- lymphocyte numbers and rates of proliferation, and CD4(+)-lymphocyte cytokine production levels were compared between the groups. The alcohol-consuming mice had significantly higher lung organism burdens than the control mice, and the CD4(+)- and CD8(+)-lymphocyte responses to pulmonary infection with M. tuberculosis were blunted in the alcohol group. Lymphocyte proliferation and production of gamma interferon were decreased in the CD4(+) lymphocytes from the alcohol-consuming mice. Additionally, lung granulomas were significantly smaller in the alcohol-consuming mice. In conclusion, murine alcohol consumption is associated with decreased control of pulmonary infection with M. tuberculosis, which is accompanied by alterations in the region-specific CD4(+)- and CD8(+)-lymphocyte responses and defective lung granuloma formation.

FIG. 1.

Weights of mice in the groups consuming the LED and the LCD over 9 weeks.

FIG. 2.

Lung burden of M. tuberculosis H37Rv in mice in the LED and LCD groups at serial time points after inoculation with M. tuberculosis. There are significantly more organisms recovered from the lungs of the LED mice than from the LCD mice at day 28 (*, P < 0.05) and a trend toward a greater burden at day 35 (+, P = 0.06). Error bars indicate standard errors of the means.

FIG. 3.

(A and B) Numbers of LALN CD4⁺ (A) and CD8⁺ (B) lymphocytes recovered at serial time points after M. tuberculosis infection in the LED and LCD groups. There are no differences between the groups at any time point for LALN CD4⁺ or CD8⁺ lymphocytes. (C and D) Numbers of lung CD4⁺ (C) and CD8⁺ (D) lymphocytes recovered at serial time points after M. tuberculosis infection in the LED and LCD groups. There are significantly fewer lung CD4⁺ lymphocytes in the LED group than in the LCD group at days 21 and 28 and fewer lung CD8⁺ lymphocytes at days 14, 21, and 28 (*, P < 0.05). (E and F) Numbers of LALN (E) and lung (F) CD4⁺ lymphocytes in mice consuming the LED and LCD diets, but not infected with M. tuberculosis, at days 14 and 28. There were no differences between the groups at either site at either time point. Error bars indicate standard errors of the means.

FIG. 4.

(A and B) Numbers of Ki67⁺ CD4⁺ (A) and CD8⁺ (B) LALN lymphocytes recovered at serial time points after M. tuberculosis infection in the LED and LCD groups. There are significantly fewer Ki67⁺ CD4⁺ lymphocytes in the LED group than in the LCD group at days 28 and 35 (*, P < 0.05; results for four mice at day 14) and a trend to fewer Ki67⁺ lymphocytes in the LED group at the day 21 time point (+, P = 0.057) (A). There are no differences in the numbers of Ki67⁺ CD8⁺ LALN lymphocytes between the groups except at the day 35 time point, when there are fewer Ki67⁺ CD8⁺ lymphocytes in the LED group than in the LCD group (*, P < 0.05; results for three mice at day 14 and four mice at day 35). (C and D) Numbers of lung Ki67⁺ CD4⁺ (C) or CD8⁺ (D) lymphocytes recovered at serial time points after M. tuberculosis infection in the LED and LCD groups. (C) There are significantly fewer Ki67⁺ CD4⁺ lung lymphocytes in the LED group than in the LCD group at day 21 (*, P < 0.05) and trends to fewer Ki67⁺ cells at days 14 and 28 (+, P < 0.08; results for four mice at day 14). (D) There are no differences in the numbers of Ki67⁺ CD8⁺ lung lymphocytes between the groups except at the day 14 time point, when there are fewer Ki67⁺ CD8⁺ lymphocytes in the LED group than in the LCD group (*, P < 0.05; results for four mice at day 14). Error bars indicate standard errors of the means.

FIG. 5.

(A and B) IFN-γ production elicited from LALN (A) and lung (B) CD4⁺ lymphocytes by 48 h of in vitro incubation with M. tuberculosis-infected PMs. For both the LALN and lung lymphocytes, there is significantly less IFN-γ produced at the day 21 time point by lymphocytes from the LED group than from the LCD group (*, P < 0.05). Error bars indicate standard errors of the means.

FIG. 6.

Representative lung sections from mice in the LCD and LED groups stained with hematoxylin and eosin. All magnifications, ×100. (A) Day 21 LCD mouse; (B) day 21 LED mouse; (C) day 28 LCD mouse; (D) day 28 LED mouse; (E) day 35 LCD mouse; (F) day 35 LED mouse.