A Minimally Invasive Model Of Cardiopulmonary Bypass In Rats And PPAR Agonist Ameliorates Renal Injury Induced By Cardiopulmonary Bypass

An increasing number of patients are undergoing cardiac surgery and in most cases a cardiopulmonary bypass(CPB) is required.This technique is aggressive and the patients are subjected to a high risk of organ injury.To better understand its pathophysiology and design protective strategies,an appropriate animal model may prove useful.We developed a minimally invasive model of CPB in rats,which was proved to be economical,easy to operate and reproducible.In addition,we investigated renal injury in a certain condition in this model and explored the possible mechanisms of the injury.Peroxisome proliferator-activated receptor(PPAR) belongs to nuclear hormone superfamily of ligand-dependent transcription factors expressed in many tissues of the body.PPAR activity is thought to be related not only to lipid metabolism and glucose homeostasis but also to inflammatory responses,cell proliferation and differentiation, as well as apoptosis.Through PPAR activation,PPAR agonists exert some beneficial effects on the kidney against to the injury induced by ischemia-reperfusion as well as nephrotoxicity.PPAR agonists also showed protective effects in cardiomyocytes from apoptosis and maintained ventricle function in an animal CPB model.However, researches concerning PPAR agonists in renal protection in CPB have not been reported.Therefore,in the present study,we aimed to determine the biochemical and morphological renal effects of pioglitazone,a PPAR-γ,agonist,in the rat model of CPB,and to explore the potential mechanisms of this agent.PartⅠ:A minimally invasive model of eardiopulmonary bypass in rats and its renal injuryObjectiveThis study was to develop a minimally invasive model of cardiopulmonary bypass in rats.In addition,we hypothesized that CPB would insult renal function and tried to elucidate the probable mechanisms.Materials and methodsEighty male Sprague-Dawley rats were randomized into 8 groups(n=10):sham group,CPB when ended,2hours after CPB,6 hours after CPB,12 hours after CPB, 24 hours after CPB,48 hours after CPB and 98 hours after CPB.The left femoral artery was cannulated for blood pressure monitoring.The tail artery was cannulated for arterial inflow and the right external jugular vein was cannulated for vein drainage.The rats were exposed to either 60 minutes of CPB at a flow rate of 100 mL/kg·min,or to cannulation only(sham group).Mean arterial pressure,heart rate, rectal temperature,blood gas analysis were monitored peri-CPB.Rats were sacrificed at the time according to the grouping after re-anesthesia.Blood sample was collected via abdominal aorta,while urine sample via bladder.The bilateral kidneys were harvested.One was kept in formalin for microscopic examination, another one was immediately frozen in liquid nitrogen for protein analysis.Serum creatinine,urinary NGAL were measured as indicators for renal function or renal injury.Levels of serum TNF-α,tissue TNF-α,NF-κB,MPO,MDA,CAT,SOD and GSH were also measured.Statistical analyses were performed by using SPSS 13.0.A value of p＜0.05 was considered statistically significant.ResultsAll rats survived until sacrificed.Rats underwent CPB showed significant difference with sham group in mean arterial pressure,heart rate,rectal temperature, blood gas analysis during the operation and the first 2 hours after CPB.Serum creatinine was elevated at 12 hours after CPB and showed statistic significance compared with the sham group.Serum creatinine peaked at 48 hours after CPB. Urinary NGAL increased at 2 hours after CPB and peaked at 12 hours,which was much earlier than serum creatinine.The typical histological features of the rats subjected to CPB included vacuolation,loss of brush border in proximal tubular cells, and sloughing of tubular cells into the lumen,leading to cast obstruction.Interstitial oedema with leucocyte infiltration can produce widely spaced tubules.TNF-α,NF-κB,MPO,and MDA in kidney tissue and serum TNF-αcontinuously increased after CPB.Most of them peaked 6-24 hours after CPB and decreased in the following time.However,the values were still higher than those in the sham group at 96 hours.On the contrary,the activities of kidney antioxidative enzymes and the concentration of reductive GSH decreased remarkably.ConclusionsA minimally invasive rat cardiopulmonary bypass model with excellent survival is developed.CPB induces renal injury in the model.Inflammatory response and oxidative stress are activated by CPB,which might contribute to the injury.PartⅡ:PPAR agonist ameliorates renal injury in a rat cardiopulmonary bypass modelObjectiveInflammatory response and oxidative stress contribute to the injury induced by CPB in rat model.We hypothesized that pretreatment with PPAR agonist, pioglitazone,would have beneficial effects on renal function after CPB.Materials and methodsThirty male Sprague-Dawley rats were randomized into 3 groups(n=10):sham group(2ml/day of saline for 7 days),control group(2ml/day of saline for 7 days) and PPAR group(pioglitazone,at a dose of 20mg/kg·day for 7 days).The rats were exposed to either 60 minutes of CPB at a flow rate of 100 mL/kg·min(sham and control group),or to cannulation only(sham group).Mean arterial pressure,heart rate, rectal temperature,blood gas analysis were monitored peri-CPB.Blood samples were collected at the time of pre-CPB(T1),CPB when ended(T2),2 hours post-CPB(T3) and 12 hours post-CPB(T4).Kidneys and urinary samples were collected when the rats were sacrificed 12 hours after CPB.Serum creatinine,TNF-αand urinary NGAL were measured.The levels of oxidative stress including MDA, GSH and the activities of SOD and CAT were measured in the kidney.Renal TNF-α and the activation of NF-κB were also determined.Statistical analyses were performed by using SPSS 13.0.A value of p＜0.05 was considered statistically significant.ResultsThere was no statistical significance observed between control and PPAR group in mean arterial pressure,heart rate,rectal temperature and blood gas analysis. Urinary NGAL was significantly decreased at 12 hours after CPB in PPAR group compared with control group,while,serum creatinine showed no statistical significance between the two groups at 12 hours.Vacuolation,loss of brush border in proximal tubular cells,and sloughing of tubular cells into the lumen,oedema with leucocyte infiltration were found in epithelial cells in histomorphologic studies of the CPB rats,which was significantly reversed in pioglitazone pretreated rats.Compared with sham group,GSH content and the activity of SOD and CAT significantly decreased in CPB groups.PPAR group had higher levels of these antioxidants than the untreated group.Renal MDA,TNF-αand NF-κB were notably increased in all CPB groups relative to the sham group,and pioglitazone attenuated these changes significantly.ConclusionsPretreatment with pioglitazone preserves renal function after cardiopulmonary bypass.Furthermore,inflammatory response and oxidative stress were significantly reduced in the treated animals.