Jungner Åsa, Vallius Suvi, Bruschettini Matteo, Romantsik Olga, Gram Magnus, Ley David
Department of Clinical Sciences Lund, Pediatrics, Skane University Hospital, Lund University, Lund, Sweden.
Department of Clinical Sciences Lund, Pediatric Surgery and Neonatal Care, Skane University Hospital, Lund University, Lund, Sweden.
Intensive Care Med Exp. 2017 Oct 4;5(1):45. doi: 10.1186/s40635-017-0153-2.
Infants with congenital heart defects (CHD) are at risk for white matter brain injury. This novel rat pup model characterizes the systemic effects of intravasal cell-free hemoglobin and hyperoxia, hypothesizing that immature endogenous scavenging systems relate to increased vulnerability to conditions present during cardiopulmonary bypass (CPB).
Plasma pharmacokinetics of cell-free human hemoglobin (Hb) was determined after intraperitoneal (i.p.) administration in postnatal day 6 (P6) rat pups. Cell-free hemoglobin degradation, scavenger- and oxidative stress responses in altered oxygen environments were evaluated in P6 rat pups exposed to i.p. cell-free Hb or vehicle and subjected to hyperoxia or normoxia for 24 h. Plasma and liver were analyzed for free heme, haptoglobin, hemopexin, heme-oxygenase 1, and 8-OHdG at 3-120 h post-injection. Baseline scavenging properties were evaluated in P0-P12 rat pups.
Cell-free Hb displayed peak plasma concentrations of 3.6 ± 0.5 mg/mL (mean ± SD) at 3 h post-administration. Animals exposed to cell-free Hb demonstrated a 30-fold increase in plasma haptoglobin and a decrease in plasma hemopexin to 1/6 of concentrations observed in pups exposed to vehicle. Exposure to cell-free Hb and hyperoxia mediated increased plasma concentrations of free heme (72.7 ± 19.5 μM, mean ± SD) compared to exposure to cell-free Hb and normoxia (49.3 ± 13.1 μM) at 3 h, and an elevated hepatic mRNA expression of heme-oxygenase 1. mRNA expression of haptoglobin and hemopexin was increased in animals exposed to hemoglobin with a mitigated response in pups exposed to hemoglobin and hyperoxia. Animals exposed to hyperoxia displayed an increase in hepatic transcription of scavenger proteins at 24 h. Combined exposure to cell-free Hb and hyperoxia mediated an increased DNA-oxidation at 6 h, whereas all insults conveyed a decrease in DNA-oxidation at 120 h.
In this study, we present a novel rat pup model with scavenging characteristics and brain maturation similar to newborns with CHD. We have confirmed a distinct scavenger response after exposure to systemic cell-free hemoglobin. We have indications of an accelerated metabolism of cell-free Hb and of an altered transcription of scavenger proteins in a hyperoxic environment. We believe that this model will prove valuable in future delineation of inflammatory and oxidative end-organ damage following CPB.
患有先天性心脏病（CHD）的婴儿存在脑白质损伤风险。这种新型幼鼠模型表征了血管内无细胞血红蛋白和高氧的全身效应，推测未成熟的内源性清除系统与体外循环（CPB）期间存在的状况导致的易感性增加有关。
在出生后第6天（P6）的幼鼠腹腔内（i.p.）给药后，测定无细胞人血红蛋白（Hb）的血浆药代动力学。在暴露于腹腔内无细胞Hb或赋形剂并在高氧或常氧环境下处理24小时的P6幼鼠中，评估无细胞血红蛋白降解、清除剂和氧化应激反应。在注射后3 - 120小时分析血浆和肝脏中的游离血红素、触珠蛋白、血红素结合蛋白、血红素加氧酶1和8-羟基脱氧鸟苷。评估P0 - P12幼鼠的基线清除特性。
给药后3小时，无细胞Hb的血浆峰值浓度为3.6±0.5 mg/mL（平均值±标准差）。暴露于无细胞Hb的动物血浆触珠蛋白增加30倍，血浆血红素结合蛋白降至暴露于赋形剂的幼鼠中观察到浓度的1/6。与暴露于无细胞Hb和常氧（49.3±13.1 μM）相比，暴露于无细胞Hb和高氧在3小时时介导血浆游离血红素浓度增加（72.7±19.5 μM，平均值±标准差），并且肝脏血红素加氧酶1的mRNA表达升高。暴露于血红蛋白的动物中触珠蛋白和血红素结合蛋白的mRNA表达增加，而暴露于血红蛋白和高氧的幼鼠中反应减轻。暴露于高氧的动物在24小时时肝脏清除蛋白的转录增加。联合暴露于无细胞Hb和高氧在6小时时介导DNA氧化增加，而所有损伤在120小时时均导致DNA氧化减少。
在本研究中，我们提出了一种具有与患有CHD的新生儿相似的清除特征和脑成熟度的新型幼鼠模型。我们证实了暴露于全身无细胞血红蛋白后有明显的清除反应。我们有迹象表明无细胞Hb的代谢加速以及在高氧环境中清除蛋白的转录改变。我们相信该模型在未来描绘CPB后的炎症和氧化终末器官损伤方面将被证明是有价值的。
