Leukotriene B4, administered via intracerebroventricular injection, attenuates the antigen-induced asthmatic response in sensitized guinea pigs

Abstract

Background: Despite intensive studies focused on the pathophysiology of asthmatic inflammation, little is known about how cross-talk between neuroendocrine and immune systems regulates the inflammatory response during an asthmatic attack. We recently showed corresponding changes of cytokines and leukotriene B4 (LTB4) in brain and lung tissues of antigen-challenged asthmatic rats. Here, we investigated how LTB4 interacts with the neuroendocrine-immune system in regulating antigen-induced asthmatic responses in sensitized guinea pigs.

Methods: Ovalbumin-sensitized guinea pigs were challenged by inhalation of antigen. Vehicle, LTB4 or U75302 (a selective LTB4 BLT1 receptor inhibitor) was given via intracerebroventricular injection (i.c.v.) 30 min before challenge. Airway contraction response was evaluated using Penh values before and after antigen challenge. The inflammatory response in lung tissue was evaluated 24 h after challenge. The LTB4 content of lung and brain homogenate preparations was detected by reversed phase high-performance liquid chromatography (RP-HPLC). Plasma levels of adrenocorticotropic hormone (ACTH) and corticosterone (CORT) were measured using ELISA kits.

Results: Antigen challenge impaired pulmonary function and increased inflammatory cell infiltration in lung tissue. These responses could be significantly suppressed by LTB4, 30 ng i.c.v., in ovalbumin-sensitized guinea pigs. LTB4 content of lung and brain homogenates from antigen-challenged guinea pigs was significantly increased. In addition, administration of LTB4 via i.c.v. markedly increased CORT and ACTH level in plasma before antigen challenge, and there were further increases in CORT and ACTH levels in plasma after antigen challenge. U75302, 100 ng i.c.v., completely blocked the effects of LTB4. In addition, U75302, 100 ng via i.c.v. injection, markedly decreased LTB4 content in lung homogenates, but not in brain homogenates.

Conclusions: Increased LTB4 levels in brain during asthmatic attacks down-regulates airway contraction response and inflammation through the BLT1 receptor. Stimulation of the hypothalamic-pituitary-adrenal axis by LTB4 may result in an increase in systemic glucocorticoids which, in turn, would feed back to suppress the asthmatic response.

Figure 1

LTB₄ attenuates the antigen-induced asthmatic response in guinea pigs, and U75302 blocks the inhibitory effect of LTB₄. Animals were challenged for 30 s with aerosolized 1% ovalbumin and Penh values were measured before and 1, 2, 3, 4, 5, 10 and 15 min after the antigen challenge on day 21 after ovalbumin sensitization. Vehicle, LTB₄ or U75302 was administered via i.c.v. before antigen challenge. Penh values are expressed as the percent change from baseline. A: LTB₄ or U75302 i.c.v. did not increase the Penh value in saline-sensitized control guinea pigs. B: The antigen challenge-induced increase in Penh value was dose-dependently suppressed by LTB₄ i.c.v. in ovalbumin-sensitized guinea pigs. C: U75302 100 ng i.c.v. pretreatment completely blocked the inhibitory effects of LTB₄ on antigen-induced increases in airway resistance. Data are expressed as mean ± S.E.M. (n = 8 per group). ^#P < 0.05, ^##P   0.01 vs. NS-vehicle group; *P < 0.05, **P < 0.01 vs. OVA-vehicle group; †P < 0.05, ††P < 0.01 vs. OVA-LTB₄ 30 ng group. OVA, ovalbumin; NS, saline.

Figure 2

Effects of LTB₄, 30 ng via i.c.v. injection, on inflammatory cells in bronchoalveolar lavage fluid (BALF). Guinea pigs were treated as described in Methods, and BALF was harvested 24 h after OVA challenge. Total inflammatory cells in BALF were counted, and cell classification was performed on a minimum of 200 cells to classify lymphocytes, eosinophils, macrophages and neutrophils. Data are expressed as mean ± S.E.M. (n = 8 per group). ^#P < 0.05, ^##P < 0.01 vs. the NS-vehicle group;*P < 0.05, **P < 0.01 vs. the OVA-vehicle group; ^†P < 0.05, ^††P < 0.01 vs. the OVA-LTB₄ 30 ng group. OVA, ovalbumin; NS, saline.

Figure 3

LTB₄ via i.c.v. suppresses antigen-induced eosinophil infiltration in lung tissue. A-D are representative histopathological pictures of lung. Compared with NS-vehicle (A), there is a marked infiltration of eosinophil cells in the peribronchiolar space and perivascular space in the OVA-vehicle group (B). LTB₄, 30 ng via i.c.v. injection, significantly inhibited OVA-induced eosinophil cell infiltration (C). U75302 (100 ng pretreatment via i.c.v.) blocked the inhibitory effect of LTB₄on eosinophil cell infiltration (D). Eosinophil cell infiltration was scored based on the severity of inflammation (E). A non-parametric test, the Mann--Whitney U-test, was used to compare differences in eosinophil cell infiltration in lung tissues. Data are expressed as mean ± S.E.M. (n = 8 per group). ^##P < 0.01 vs. the NS-vehicle group;**P < 0.01 vs. the OVA-vehicle group; ^††P < 0.01 vs. the OVA-LTB₄ 30 ng group. OVA, ovalbumin; NS, saline.

Figure 4

LTB₄ content in lung and cerebral cortex homogenates from antigen-challenged asthmatic guinea pigs. LTB₄ content was detected by reversed phase high-performance liquid chromatography (RP-HPLC). Data are expressed as the mean ± S.E.M. (n = 8 per group). ^#P < 0.05, ^##P < 0.01 vs. the NS-vehicle group;*P < 0.05, **P < 0.01 vs. the OVA-vehicle group; ^†P < 0.05, ^††P < 0.01 vs. the OVA-LTB₄ 30 ng group. OVA, ovalbumin; NS, saline.

Figure 5

LTB₄ via i.c.v. injection increases plasma CORT and ACTH levels in guinea pigs. Plasma CORT (A) and ACTH (B) levels were measured using commercial ELISA kits 30 min after either vehicle, LTB₄ or U75302 were administered via i.c.v., both before antigen challenge and 180 min after antigen challenge. Data are expressed as mean ± S.E.M (n = 8 in each group). ANOVA and Student--Newman--Keuls multiple comparisons test were used to compare differences between groups. **P < 0.01 value is significantly different from after antigen challenge; ^#P < 0.05 vs. the OVA-vehicle group after antigen challenge; P < 0.05 vs. the OVA-vehicle group after injection (i.c.v.); ^†P < 0.05 vs. the OVA-LTB₄ group after antigen challenge. OVA, ovalbumin; NS, saline.