Toll‑like receptor 4 contributes to acute kidney injury after cardiopulmonary resuscitation in mice

Abstract

Toll-like receptor 4 (TLR4) activation mediates renal injury in regional ischemia and reperfusion (I/R) models generated by clamping renal pedicles. However, it remains unclear whether TLR4 is causal in the kidney injury following global I/R induced by cardiac arrest (CA) and cardiopulmonary resuscitation (CPR). The present study used wild‑type (C3H/HeN) and TLR4‑mutant (C3H/HeJ) mice to produce the CA/CPR model. CA was induced by injection of cold KCl and left untreated for different time periods. After resuscitation (72 h), the level of blood urea nitrogen (BUN) and serum creatinine (Scr), as well as histological changes in renal tissue were assessed to evaluate the severity of acute kidney injury (AKI). The expression of TLR4, intercellular adhesion molecule‑1 (ICAM‑1), myeloperoxidase (MPO) and growth‑regulated oncogene‑β (GRO‑β) in kidney tissues was detected. The results demonstrated that the levels of Scr and BUN increased significantly in C3H/HeN and C3H/HeJ mice after CPR. CPR also resulted in increased expression of TLR4, ICAM‑1, GRO‑β and MPO in a CA‑duration dependent manner. However, there was decreased expression of ICAM‑1, GRO‑β and MPO in C3H/HeJ mice compared with that in C3H/HeN mice. C3H/HeJ mice were resistant to AKI as demonstrated by the minor changes in renal histology and function following CPR. In conclusion, mice suffered from AKI after successful CPR and severe AKI occurred in mice with prolonged CA duration. TLR4 and its downstream signaling events that promote neutrophil infiltration via ICAM‑1 and GRO‑β may be important in mediating inflammatory responses to renal injury after CPR.

Figure 1

Plasma concentrations of Scr and BUN after CPR. The plasma concentrations of (A) Scr and (B) BUN were significantly increased after CPR with the extension of cardiac arrest duration. (C) Scr and (D) BUN levels in model-ej group were significantly lower than those in the model-en group ^*P<0.05, ^**P<0.01 and ^***P<0.001. Scr, serum creatinine; BUN, blood urea nitrogen; CPR, cardiopulmonary resuscitation.

Figure 2

Expression of TLR4 mRNA and protein following CPR. CPR increased the (A) mRNA and (B) protein expression of TLR4 in the kidney tissues. As the duration of cardiac arrest was prolonged, TLR4 expression increased gradually. ^*P<0.05, ^**P<0.01 and ^***P<0.001. TLR4, toll-like receptor 4; CPR, cardiopulmonary resuscitation; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

Figure 3

Expression of ICAM-1 and GRO-β after CPR. Expression of (A) ICAM-1 and (B) GRO-β increased in the kidney tissue in the 3, 5 and 8 min CA groups compared with the sham animals. In experiment 2, CPR induced the elevated (C) ICAM-1 and (D) GRO-β expression. However, the expression of ICAM-1 and GRO-β in C3H/HeJ mice was lower than that in the C3H/HeN mice. ^*P<0.05, ^**P<0.01 and ^***P<0.001. ICAM-1, intercellular adhesion molecule-1; GRO-β, growth-regulated oncogene-β; CPR, cardiopulmonary resuscitation; CA, cardiac arrest.

Figure 4

Activity of MPO after CPR. (A) With the extension of cardiac arrest duration MPO activity was increased significantly. (B) There was a less intense upregulation of MPO activity in the model-ej group compared with that in model-en CPR group. ^*P<0.05, ^**P<0.01 and ^***P<0.001. MPO, myeloperoxidase; CPR, cardiopulmonary resuscitation.

Figure 5

Light microscopy results. (A) The epithelium of the tubule showed signs of denaturation, swelling and vacuolization after CPR (magnification, ×400). (B) Damage scores increased in kidney tissues as the duration of cardiac arrest was prolonged. (C) The histological abnormalities were severe after CPR in model-ej and model-en groups (magnification, ×400). (D) The renal damage score was lower in the model-ej group compared with that in the model-en group ^**P<0.01, ^***P<0.001.

Figure 6

Electron microscopy results. (A) The loss of microvilli and increased cell vacuolization in microvilli occurred in the model-ej and model-en groups. The abnormalities were more severe in the model-en group than that in model-ej group. (B) The glomerular epithelium presented more evident swollen endoplasmic reticulum and mitochondria with broken or absent cristae, some of which appeared to be vacuolized, in the model-en group than in the model-ej group.