Effects of agmatine on excessive inflammatory reaction and proliferation of splenic cells in mice with trauma / 中华危重病急救医学

ABSTRACT

ObjectiveTo observe protective effects of agmatine (AGM) on inflammatory response and spleen immune function in mice with trauma.Methods Forty-eight adult male C57BL/6 mice were randomly divided into three groups (n= 16 each), including control group, model group (bilateral femoral fracture and removal of 35% of the total blood volume), and AGM group (trauma/hemorrhage & AGM 200 mg/kg). Eight mice in each group were sacrificed at 3 hours and 24 hours, respectively, after modeling, and blood samples and tissue homogenate of spleen and liver were collected. The contents of tumor necrosis factor-α (TNF-α), interleukins (IL-6, IL-1β) in serum and liver tissue were determined with enzyme linked immunosorbent assay (ELISA). Serum aspartate transaminase (AST), alanine aminotransferase (ALT) and lactic dehydrogenase (LDH) were determined with automatic biochemistry analyzer. Spleen proliferation response stimulated with concanavalin A (ConA) was evaluated with methyl thiazolyl tetrazolium colourimetry (MTT).γ-interferon (IFN-γ) and IL-2 releases were determined with ELISA.Results Compared with control group, 3 hours after trauma/hemorrhage, the levels of serum TNF-α, IL-6, and IL-1β in model group were significantly elevated [TNF-α (ng/L) 145.38±31.50 vs. 23.06±11.14, IL-6 (ng/L) 496.94±50.76 vs. 47.13±17.47, IL-1β (ng/L) 321.31±43.02 vs. 29.25±16.24,allP< 0.01]. It was found that AGM treatment could alleviate the increase in serum pro-inflammatory mediators induced by trauma/hemorrhage, such as TNF-α (ng/L111.56±25.47 vs. 145.38±31.50), IL-6 (ng/L 412.56±44.33 vs. 496.94±50.76), IL-1β (ng/L 273.38±45.25 vs. 321.31±43.02,P< 0.05 orP< 0.01). Twenty-four hours after trauma/hemorrhage, serum pro-inflammatory mediators were recovered to the levels in control group. There was no significant difference in TNF-α and IL-6 levels at 3 hours after trauma/hemorrhage among groups. Compared with control group, the expressions of liver TNF-α and IL-6 in model group were increased at 24 hours following trauma [TNF-α (ng/mg) 32.93±4.90 vs. 26.58±2.33, IL-6 (ng/mg) 11.20±1.66 vs. 8.38±0.89,bothP< 0.01]. However, AGM inhibited the level of TNF-α (ng/mg28.92±3.16 vs. 32.93±4.90) and IL-6 (ng/mg 9.03±1.28 vs. 11.20±1.66) in the liver as induced by trauma/hemorrhage (P< 0.05 andP< 0.01). At 24 hours after modeling, model group and AGM group had distinctly higher serum AST, ALT, LDH levels than those of control group [AST (U/L) 405.9±31.2, 245.7±22.1 vs. 128.2±15.9; ALT (U/L) 92.1±6.3, 51.6±5.0 vs. 30.1±3.2; LDH (U/L) 606.7±36.3, 478.7±25.3 vs. 384.0±16.6, allP< 0.01]. Nevertheless,the increase in serum AST, ALT and LDH was alleviated in AGM group (allP< 0.01). Meantime, trauma/hemorrhage produced a noticeable depression of proliferation of splenic cells and IFN-γ and IL-2 release stimulated with ConA compared with control group [proliferation rate (40.97±4.13)% vs. (89.99±7.76)%, IFN-γ(ng/L) 91.6±12.3 vs. 353.2±21.5,IL-2 (ng/L) 53.4±6.4 vs. 91.0±12.2,allP< 0.01]. In contrast, AGM notably restored the capacity of proliferation response of splenic cells [proliferation rate (74.86±5.75)% vs. (40.97±4.13)%, P< 0.01],enhanced the release of IFN-γ and IL-2 stimulated with ConA [IFN-γ (ng/L) 327.8±23.6 vs. 91.6±12.3, IL-2 (ng/L) 74.8±10.4 vs. 53.4±6.4, bothP< 0.01].Conclusion AGM can dramatically alleviate spleen immunosuppression, excessive inflammation and organ damage induced by trauma/hemorrhage.