Influences of hydrogen-rich saline on acute kidney injury in severely burned rats and mechanism

Abstract:

Objective: To explore the influences of hydrogen-rich saline on acute kidney injury in severely burned rats and to analyze the related mechanism.

Methods: Fifty-six Sprague Dawley rats were divided into sham injury group (n=8), burn group (n=24), and hydrogen-rich saline group (n=24) according to the random number table. Rats in sham injury group were treated by 20 ℃ water bath on the back for 15 s to simulate injury, and rats in burn group and hydrogen-rich saline group were inflicted with 30% total body surface area (TBSA) full-thickness scald (hereinafter referred to as burns) by 100 ℃ water bath on the back for 15 s. Immediately after injury, hydrogen-rich saline at the dose of 10 mL/kg were intraperitoneally injected to the rats in hydrogen-rich saline group at one time, while normal saline with the same dose were intraperitoneally injected to the rats in sham injury group and burn group. At post injury hour (PIH) 6, rats in the 3 groups were intraperitoneally injected with 4 mL·kg(-1)·%TBSA(-1) lactated Ringer's solution for resuscitation. Eight rats from sham injury group at PIH 72 and eight rats from burn group and hydrogen-rich saline group at PIH 6, 24, and 72 were sacrificed respectively after their blood samples from abdominal aorta were collected. Then their kidney tissue was harvested for histopathological observation and renal tubular injury scoring by hematoxylin and eosin staining, serum creatinine and blood urea nitrogen were detected by the clinical blood biochemical analyzer, expression distribution and mRNA expressions of tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β), and IL-6 in renal tissue were evaluated by immunohistochemical staining and real time fluorescent quantitive reverse transcription polymerase chain reaction respectively, and protein expression of high mobility group protein 1 (HMGB1) was detected by Western blotting. Data were processed with Kruskal-Wallis H test, Dunn test, one-way analysis of variance, Bonferroni test.

Results: (1) The renal tubular structure of rats in sham injury group at PIH 72 was complete with no inflammatory cell infiltration and no cellular degeneration or necrosis. Since PIH 6, the changes such as vacuolation and shape change of cells and aggregation of broken protein in renal tubules were observed in rats of burn group, and all these changes deteriorated with time. The renal injury of rats in hydrogen-rich saline group at different post injury time points were relieved compared with those of rats in burn group at the corresponding time points. The renal tubular injury scores of rats in burn group and hydrogen-rich saline group at PIH 6, 24, and 72 were significantly higher than the score in sham injury group at PIH 72 (P<0.05). The renal tubular injury scores of rats in hydrogen-rich saline group were significantly lower than those in burn group at PIH 6, 24, and 72 (P0.05), the levels of serum creatinine of rats in burn group at all the time points and hydrogen-rich saline group at the other time points were significantly higher than the level of serum creatinine of rats in sham injury group at PIH 72 (P<0.01). The levels of blood urea nitrogen of rats in burn group and hydrogen-rich saline group at PIH 6, 24, and 72 were significantly higher than the level of blood urea nitrogen of rats in sham injury group at PIH 72 (P<0.01). The levels of serum creatinine and blood urea nitrogen of rats in hydrogen-rich saline group at PIH 6, 24, and 72 were significantly lower than those in burn group at the corresponding time points (P<0.05). (3) There were certain degree of positive expressions of TNF-α, IL-1β, and IL-6 in renal tissue of rats in sham injury group at PIH 72, which were mainly observed in the cytoplasm of renal tubular epithelium cell. The expressions of above-mentioned inflammatory cytokines in renal tissue of rats in burn group at PIH 6, 24, and 72 were higher than those in sham injury group. The expressions of above-mentioned inflammatory cytokines in renal tissue of rats in hydrogen-rich saline group at all the time points were less than those in burn group at the corresponding time points. (4) Compared with those in sham injury group at PIH 72, the mRNA expression levels of TNF-α, IL-1β, and IL-6 of rats in burn group at PIH 6, 24, and 72 were significantly increased (P<0.01). The mRNA expression levels of TNF-α were significantly increased in hydrogen-rich saline group at PIH 6 and 24 (P<0.05 or P<0.01), and the mRNA expression level of IL-6 was significantly increased in hydrogen-rich saline group at PIH 6 (P0.05), and the mRNA expression levels of TNF-α, IL-1β, and IL-6 at the other time points in hydrogen-rich saline group were significantly decreased (P<0.05). (5) Compared with 0.39±0.03 in sham injury group at PIH 72, the protein expression of HMGB1 of rats in burn group at PIH 6, 24, and 72 (1.19±0.07, 1.00±0.06, 0.80±0.05) were significantly increased (P0.05). Compared with those in burn group, the protein expressions of HMGB1 of rats in hydrogen-rich saline group at PIH 6, 24, and 72 were significantly decreased (P<0.05). Conclusions: Hydrogen-rich saline can alleviate the acute kidney injury in severely burned rats through regulating the release of inflammatory cytokines in renal tissue.