Abstract
Histones are considered potential risk factors that contribute to the development of septic acute kidney injury (SAKI) by inducing apoptosis and inflammation. This study aimed to explore the protective effects of heparin on septic acute kidney injury through the neutralization of extracellular histones (EH) and to uncover the underlying mechanism. C57BL mice (16 each) were randomly divided into the sham group, the sepsis group (established by cecal ligation and puncture operation, CLP), and the heparin intervention group. Mice in the heparin intervention group received a subcutaneous injection of unfractionated heparin (0.03 IU/g) 4 h after CLP. At 6 h after the operation, nine mice from each group were sacrificed by the removal of the eyeballs to harvest blood samples; the upper half of the right kidney was used as the study sample. Mice renal tubular epithelial cells cultivated in six-well plates were equally divided into five groups. We cultured cells treated with either histone (40 U), histone (40 U) + heparin (25 IU/ml), histone(40U) + lipopolysaccharides (LPS; 10 μg/ml), or histone (40 U) + LPS (10 μg/ml) + heparin (25 IU/ml) for 6 h. For the histone + heparin group and the histone + LPS + heparin group, histone (and LPS) were treated with heparin simultaneously. Mice in the heparin intervention group showed decreased levels of EH4, neutrophil gelatinase-associated lipocalin (NAGL), kidney injury molecule-1 (KIM-1), tumor necrosis factor-α (TNF-α), and interleukin (IL)-6 in the blood serum, longer average 72-h survival rate, significantly decreased kidney tissue edema, and a clearer glomerular structure coupled with decreased protein and mRNA expression levels of kidney apoptosis-related proteins (cleaved Caspase-3/Caspase-3 and Bax/Bcl-2) compared with those in the sepsis group at 6 h after CLP (P < 0.05). Meanwhile, cells in the heparin intervention group exhibited lower expression levels of serum EH4 and inflammatory cytokines, a lower apoptosis rate, and decreased expression of apoptosis-related proteins, both at protein and mRNA levels, than those in the histone-stimulated group at 6 h after stimulation (P < 0.05). Heparin may alleviate apoptosis and inflammation through the neutralization of histones, thus playing a protective role against septic acute kidney injury.
Highlights
Extracellular histones (EH) have been shown to exhibit cytotoxicity that can induce apoptosis in HK-2 cells in vitro [1]
Mice were divided into four groups: sham group, sepsis group, heparin intervention group, and histone group
Our results showed that mice in the sepsis group exhibited higher expression levels of H4 in serum (Figure 4), indicating that the levels of extracellular histones (EH) increased during the development of septic acute kidney injury (SAKI)
Summary
Extracellular histones (EH) have been shown to exhibit cytotoxicity that can induce apoptosis in HK-2 cells in vitro [1]. Zarjou et al have reported that EH can lead to a series of acute kidney injury (AKI) manifestations in vivo, such as endothelial cell activation, increased vascular permeability, and leukocyte recruitment [3]. Using a murine model of septic AKI (SAKI), animals receiving anti-histone antibody exhibited much less injury than those without treatment [4], indicating that histones may be potential targets for SAKI, the underlying mechanisms involved in SAKI are not entirely clear. It has been assumed that the increased production of reactive oxygen species (ROS) may activate cell apoptosis [5]
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