| ObjectivesSepsis and septic shock have become the main diseases to cause the death of critically ill patients, liver is one of the major organs easily being damaged and involved by sepsis. Sepsis can lead to the dysfunction of hepatocellular mitochondria, which may result in edema, apoptosis and necrosis of liver cells. AQP8 is a kind of important aquapor in located in mitochondrial membrane, critical to the regulation of water permeability inside and outside of mitochondria, playing a very important role in maintaining the normal structure and function of mitochondria. This paper further discusses the influence of the expression of AQP8 which is located hepatocyte mitochondrial membrane of rat with sepsis on its hepatocellular function and mitochondrial function.MethodsSelect 24 SD rats and divided them into two groups randomly:the control group (C group) and the sepsis group (S group), with 12 rats at each group. Adopt cecum ligation perforation (CLP) method to establish rat models with sepsis. Kill rats 18 hours after establishing models, take out part of their livers, and analyze with colorimetry and fluorescence analysis method to detect hepatocellular ATP contents; and extract hepatocellular mitochondria by differential centrifugation, test the potential of mitochondrial membrane and the swelling degree of mitochondria with laser confocal microscope and flow cytometry; test the activities of Na+-K+-ATP enzyme, Mg2+-ATPenzyme, Ca2+-ATP enzyme and Ca2+-Mg2+-ATP enzyme on mitochondrial membrane by the means of spectrophotometry; break down hepatocellular mitochondria to isolate mitochondrial membrane proteins, use Western Blot method to detect the contents of AQP-8 protein, and real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) method to detect the expression of AQP-8 mRNA in liver tissues; take blood from abdominal aorta, adopt enzyme coupling rate method to detect the activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and aspartate aminotransferase mitochondria isoenzyme (m-AST).ResultsThe concentrations of serum ALT, AST and m-AST are (29.81±13.16) U/L, (99.78±41.67) U/L and (46.23±24.24)U/L in normal control group, while (233.02± 10.16) U/L, (742.56±441.41) U/L and (412.78±252.56) U/L in sepsis group, compared with normal control group, the transaminase concentration of sepsis group increases significantly (P<0.01). The ATP contents are (94.65±17.79) nmol/mg and (19.27±8.76) nmol/mg in the hepatocyte of normal control group and that of sepsis group respectively, the ATP contents in control group decrease significantly(P<0.01). The activities of Na+-K+-ATP enzyme, Mg2+-ATP enzyme, Ca2+-ATP enzyme and Ca2+-Mg2+-ATP enzyme are (4.74±0.84) U/mgprot, (4.49±0.73)U/mgprot, (3.48±0.43)U/mgprot and (4.68±0.81)U/mgprotin the mitochondria of normal control group respectively, while (2.81±0.81)U/mgprot, (2.59±1.03)U/mgprot, (1.63±1.26)U/mgprot and (2.54±0.97)U/mgprot in sepsis group separately. So compared with control group, the activities of ATP enzymes decrease apparently (P<0.01). The hepatocellular mitochondrial membrane potential and the swelling degree of mitochondria are (7.65±3.54) and (0.59±0.16) respectively in normal control group, and (1.73±1.06) and (1.07±0.43) in sepsis group, showing that membrane potential of sepsis grouped decreases noticeably, and its swelling degree increases significantly compared with control group (P<0.01), with apparent statistical differences. The expression quantity of AQP-8 located in mitochondrial membrane and AQP-8 mRNA transcription level are (1.24±0.05) and 1.00 in normal control group, while (0.68±0.03) and (0.34±0.19) in sepsis group, down by 32.3% and 65.7% than control group respectively (P<0.01), showing significant statistical differences.Conclusions(1) The mRNA transcription quantity of AQP-8 mRNA in sepsis group decreases significantly, resulting in the apparent decrease of expression quantity of AQP-8 in hepatocellular mitochondrial membrane.(2) ATP contents in the hepatocytes of sepsis group decreases significantly.(3) The membrane potential of sepsis group decreases noticeably, and its swelling degree increases significantly.(4) The activities of all ATP enzymes on the hepatocellular mitochondrial membrane of sepsis group decrease apparently.(5) The expression quantity of AQP8 and ATP contents in the hepatocytes of sepsis group are in positive correlation with the activities of Na+-K+-ATP enzyme, Ca2+-ATP enzyme Mg2+-ATP enzyme and Ca2+-Mg2+-ATP enzyme.(6) The mechanism of mitochondrial dysfunction in liver cells caused by sepsis is as follows:the decreased expression of AQP8 and mRNA in hepatocellular mitochondria can lead to the reduced synthesis of AQP-8, causing the swelling of mitochondria, and lowering the potential of mitochondrial membrane, as well as the activities of Na+-K+-ATP enzyme, Mg2+-ATP enzyme, Ca2+-ATP enzyme and Ca2+-Mg2+-ATP enzyme on mitochondrial membrane, thus damaging mitochondrial function, which can result in the decreased synthesis of hepatocellular mitochondrial ATP, liver cell dysfunction, and even the damage to liver cells. |
PDF Full Text Request