Effects of Lonicera japonica Flower Bud Extract on Citrobacter rodentium-Induced Digestive Tract Infection

Abstract

Background: Although antibiotic therapy is currently a gold standard for bacterial infections, it is not used for severe diseases like enterohemorrhagic Escherichia coli, in which the Shiga toxin is overproduced by antibiotic action. The Lonicera japonica flower bud (LJF) is an herbal component used against purulent diseases in traditional Japanese and Chinese medicine. We investigated the effects of LJF extract (LJFE) on Citrobacter rodentium-induced digestive tract infection in a mouse model. Methods:Citrobacter rodentium and LJFE were orally administered to C57BL/6 mice. The survival rate and bacterial colonization in the large intestine, mesenteric lymph node, and blood of mice were evaluated. Cytokines secreted from intraperitoneal macrophages of LJFE-treated mice were measured using ELISA. Moreover, the phagocytic activity of intraperitoneal macrophages against Citrobacter rodentium was compared between LJFE- or chlorogenic acid (CGA)-treated mice. Results: LJFE significantly increased the survival rate and decreased Citrobacter rodentium colonization in mice. Moreover, the values of tumor necrosis factor-α, interleukin-1β, and interferon-γ secreted from macrophages were increased following LJFE treatment. While macrophages of LJFE-treated mice showed a significant phagocytic activity, macrophages of CGA-treated mice only showed a phagocytic tendency. Conclusions: LJF may be useful for treating Citrobacter rodentium-induced digestive tract infection.

Keywords: Citrobacter rodentium; Lonicera japonica; bacterial digestive tract infection; chlorogenic acid; intraperitoneal macrophages.

Figure 1

Three-dimensional high-performance liquid chromatogram of Lonicera japonica flower bud extract (LJFE).

Figure 2

Effects of Lonicera japonica flower bud extract (LJFE) on the survival rate of Citrobacter-infected mice. Eight mice were orally infected with Citrobacter, and their survival rate was plotted. Dotted line, control group without LJFE treatment; dashed line: LJFE-treated group (1 g/kg); solid line, LJFE-treated group (2 g/kg). * p < 0.05, ** p < 0.01 compared with the control group (Kaplan–Meier log rank test).

Figure 3

Effects of Lonicera japonica flower bud extract (LJFE) on the large intestine histology of Citrobacter-infected mice. Hematoxylin and eosin staining were performed. The yellow arrow points to the bacterial mass. (a) Normal mouse, (b) Citrobacter-infected control group without LJFE-treatment, (c) Citrobacter-infected group treated with LJFE (2 g/kg).

Figure 4

Effects of Lonicera japonica flower bud extract (LJFE) on bacterial colonization in Citrobacter-infected mice. Bacteria colonies in the (a) large intestine, (b) mesenteric lymph nodes, and (c) blood were represented. Data are presented as mean ± S.D. (n = 6). ** p < 0.01, N.S. not significant, Turkey’s multiple comparison test.

Figure 5

Effects of Lonicera japonica flower bud extract (LJFE) on mice serum cytokine levels. Mice were orally administered LJFE (2 g/kg) for seven days, and the serum levels interleukin (a) (IL)-1β, (b) interferon (IFN)-γ, (c) tumor necrosis factor (TNF)-α, and (d) IL-6 were measured by ELISA. Data are presented as mean ± S.D. (n = 6). * p < 0.05, ** p < 0.01, t-test.

Figure 6

Ex Vivo effects of Lonicera japonica flower bud extract (LJFE) on the phagocytic activity of intraperitoneal macrophages against Citrobacter in mice. Mice were orally administered LJFE (2 g/kg) for seven days, and their peritoneal macrophages were cultured with Citrobacter. Bacterial colonies at 1 h after the start of phagocytosis were demonstrated. Data are presented as mean ± S.D. (n = 6). ** p < 0.01, t-test.

Figure 7

Ex Vivo effect of Lonicera japonica flower bud extract (LJFE) on the ³H-thymidine uptake of mice intraperitoneal macrophages. Mice were orally administered LJFE (2 g/kg) for seven days, and the ³H-thymidine uptake of peritoneal macrophages was measured. Data are presented as mean ± S.D. (n = 6). ** p < 0.01, t-test.

Figure 8

Ex Vivo effect of Lonicera japonica flower bud extract (LJFE) on the levels of cytokines secreted from mice intraperitoneal macrophages. Mice were orally administered LJFE (2 g/kg) for seven days, and the levels of interleukin (a) (IL)-1β, (b) interferon (IFN)-γ, and (c) tumor necrosis factor (TNF)-α in the culture supernatant of intraperitoneal cells collected from mice were measured by ELISA. Data are presented as mean ± S.D. (n = 6). * p < 0.05, ** p < 0.01, t-test.

Figure 9

Ex Vivo effects of chrologenic acid (CGA) on the phagocytic activity of intraperitoneal macrophages against Citrobacter in mice. Bacterial colonies at 1 h after the start of phagocytosis were demonstrated. Data are presented as mean ± S.D. (n = 6). t-test.