Prednisolone has a positive effect on the kidney but not on the liver of brain dead rats: a potencial role in complement activation

Abstract

Background: Contradictory evidence has been published on the effects of steroid treatments on the outcomes of kidney and liver transplantation from brain dead (BD) donors. Our study aimed to evaluate this disparity by investigating the effect of prednisolone administration on BD rats.

Methods: BD induction was performed in ventilated rats by inflating a Fogarty catheter placed in the epidural space. Prednisolone (22.5 mg/kg) was administered 30 min prior to BD induction. After four hours of determination of BD: serum, kidney and liver tissues samples were collected and stored. RT-qPCR, routine biochemistry and immunohistochemistry were performed.

Results: Prednisolone treatment reduced circulating IL-6 and creatinine plasma levels but not serum AST, ALT or LDH. Polymorphonuclear influx assessed by histology, and inflammatory gene expression were reduced in the kidney and liver. However, complement component 3 (C3) expression was decreased in kidney but not in liver. Gene expression of HSP-70, a cytoprotective protein, was down-regulated in the liver after treatment.

Conclusions: This study shows that prednisolone decreases inflammation and improves renal function, whilst not reducing liver injury. The persistence of complement activation and the negative effect on protective cellular mechanisms in the liver may explain the disparity between the effects of prednisolone on the kidney and liver of BD rats. The difference in the molecular and cellular responses to prednisolone administration may explain the contradictory evidence of the effects of prednisolone on different organ types from brain dead organ donors.

Figure 1

Blood pressure registry. The record started with the brain death induction, we considered the time “0” as the end of brain death induction and the starting of BD period. As a black-continuous line the mean in each minute of blood pressure for the prednisolone brain death group. As a grey-continuous line the mean in each minute for the saline brain death group.

Figure 2

Plasmatic levels of kidney function markers and liver injury markers. Creatinine (Sham operated + Saline: 36 ± 9.24 μM; Sham operated + prednisolone treatment: 36.88 ± 4.29 μM; BD + Saline 86.63 ± 18.37 μM; BD + prednisolone treatment: 50.38 ± 6.91 μM), AST (Sham operated + Saline: 71.50 ± 14.19 U/l; Sham operated + prednisolone treatment: 72.88 ± 11.66 U/l; BD + Saline: 138.0 ± 72.76 U/l; BD + prednisolone treatment: 146.3 ± 35.07 U/l), ALT (Sham operated + Saline: 54.38 ± 13.97 U/l; Sham operated + prednisolone treatment: 45.88 ± 15.82 U/l; BD + Saline: 106.3 ± 58.38 U/l; BD + prednisolone treatment: 87.50 ± 18.33 U/l), LDH (Sham + Saline: 157.1 ± 41.54 U/l; Sham + prednisolone treatment: 172.4 ± 104.2 U/l; BD + Saline: 342.8 ± 228.0 U/l; BD + prednisolone treatment: 270 ± 152.9 U/l) and IL-6 (Sham operated + Saline: 15.63 ± 29.69 pg/ml; Sham operated + prednisolone: undetectable; BD + Saline: 2.07 × 10 ⁵± 9.59 × 10 ⁴ pg/ml; BD + prednisolone treatment: 1.18 × 10 ³± 2.69 × 10 ³ pg/ml).

Figure 3

PMN infiltration quantification and stanning. Kidneys from A) Sham operated + Saline, B) Sham operated + prednisolone treatment, C) BD + Saline, D) BD + prednisolone treatment rats. PMN infiltration in livers from E) Sham operated + Saline, F) Sham operated + prednisolone treatment, G) BD + Saline, H) BD + prednisolone treatment rats, 40x magnification. I) Quantification of PMN infiltration in kidney and J) liver.

Figure 4

Relative expression of inflammatory genes.A) Kidney samples. IL-6 (Sham operated + Saline: 0.08 ± 0.09; Sham operated + prednisolone treatment: 0.05 ± 0.05; BD + Saline: 6.04 ± 3.79; BD + prednisolone treatment: 0.34 ± 0.59), IL-1 β (Sham operated + Saline: 0.89 ± 0.22; Sham operated + prednisolone treatment: 0.27 ± 0.11; BD + Saline: 1.95 ± 0.69; BD + prednisolone treatment: 0.40 ± 0.30), TNF- α (Sham operated + Saline: 0.69 ± 0.34; Sham operated + prednisolone treatment: 0.20 ± 0.10; BD + Saline: 0.99 ± 0.49; BD + prednisolone treatment: 0.20 ± 0.16), MCP-1 (Sham operated + Saline: 0.46 ± 0.13; Sham operated + prednisolone treatment: 0.13 ± 0.07; BD + Saline 4.46 ± 1.02; BD + prednisolone treatment: 0.39 ± 0.44). B) Liver samples. IL-6 (Sham operated + Saline: 0.11 ± 0.05; Sham operated + prednisolone treatment: 0.04 ± 0.03; BD + Saline: 2.98 ± 0.84; BD + prednisolone treatment: 0.60 ± 0.45), IL-1 β (Sham operated + Saline: 1.09 ± 0.42; Sham operated + prednisolone treatment: 0.50 ± 0.20; BD + Saline: 1.91 ± 0.27; BD + prednisolone treatment: 0.61 ± 0.26), TNF- α (Sham operated + Saline: 7.91 ± 2.13; Sham operated + prednisolone treatment: 4.51 ± 1.57; BD + Saline: 4.04 ± 0.66; BD + prednisolone treatment: 1.3 ± 0.71), MCP-1 (Sham operated + Saline: 0.71 ± 0.17; Sham operated + prednisolone treatment: 0.32 ± 0.09; BD + Saline: 1.06 ± 0.23; BD + prednisolone treatment: 0.36 ± 0.24).

Figure 5

Relative expression of C3, HO-1 and HSP-70. Ratio of Bcl2/BAX expression. A) Kidney samples. C3 (Sham operated + Saline: 0.18 ± 0.04; Sham operated + prednisolone treatment: 0.16 ± 0.02; BD + Saline: 17.27 ± 10.74; BD + prednisolone treatment: 1.47 ± 0.26), HO-1 (Sham operated + Saline: 0.03 ± 0.01; Sham operated + prednisolone treatment: 0.04 ± 0.01; BD + Saline: 3.05 ± 2.06; BD + prednisolone treatment: 0.42 ± 0.23), HSP-70 (Sham operated + Saline: 0.06 ± 0.09; Sham operated + prednisolone treatment: 0.03 ± 0.07; BD + Saline: 0.03 ± 0.02; BD + prednisolone treatment: 0.03 ± 0.01), Ratio Bcl2/BAX (Sham operated + Saline: 0.37 ± 0.15; Sham operated + prednisolone treatment: 0.38 ± 0.15; BD + Saline: 0.69 ± 0.19; BD + prednisolone treatment: 0.69 ± 0.28). B) Liver samples. C3 (Sham operated + Saline: 546.9 ± 109.4; Sham operated + prednisolone treatment: 534.2 ± 92.18; BD + Saline: 687.4 ± 96.13; BD + prednisolone treatment: 952.2 ± 125.3), HO-1 (Sham operated + Saline: 0.47 ± 0.11; Sham operated + prednisolone treatment: 0.45 ± 0.13; BD + Saline: 5.78 ± 2.65; BD + prednisolone treatment: 12.47 ± 4.96), HSP-70 (Sham operated + Saline: 0.05 ± 0.09; Sham operated + prednisolone treatment: 0.05 ± 0.11; BD + Saline: 0.03 ± 0.04; BD + prednisolone treatment: 0.003 ± 0.005), Ratio Bcl2/BAX (Sham operated + Saline: 0.18 ± 0.03; Sham operated + prednisolone treatment: 0.18 ± 0.01; BD + Saline: 0.05 ± 0.01; BD + prednisolone treatment: 0.05 ± 0.01).