Effect Of DHFZT On Activation Of Kupffer Cells Of Liver In Rats With Severe Acute Pancreatitis

Background: Severe acute pancreatitis(SAP)is a common clinical emergency,with a dangerous condition and high mortality.SAP is often accompanied with the damage of intestinal mucosal barrier function,and with the translocation of a large number of intestinal bacteria and endotoxin,which enters the blood circulation through the portal vein and liver.Various harmful substances are released throughout the body causing systemic inflammatory response syndrome(SIRS),which eventually leads to multiple organ dysfunction,failure or even death.The liver is the indispensable place for intestinal bacteria and endotoxin to enter the blood.Under physiological conditions,the liver Kupffer cells act as the largest tissue-fixed macrophage population in the body,and can recognize the danger signal in microenvironment through the pattern recognition receptor(PRR).Liver Kupffer cells have two active forms of M1 and M2 depending on the activation pathway of different helper T cells.M1 Kupffer cells primarily mediate inflammatory responses,while M2 Kupffer cells primarily mediate anti-inflammatory and tissue repair.F4/80 was expressed on the mature activated Kupffer cell membrane.The proteins specifically expressed on the M1 and M2 Kupffer cell membranes are co-stimulatory factors CD86 and CD163,respectively.Activated Kupffer cells are able to clear pathogens to reduce body infections,but they may aggravate the inflammatory response,and even more often lead to multiple organ failure under certain factors.Previous studies have found that a large number of intestinal bacteria and endotoxins in SAP still undergo damage or even functional failure of other organs after liver treatment,the reason is not clear.We hypothesized that activation of liver Kupffer cells may play an important role in SAP leading to SIRS and multiple organ function damage.In recent years,the team research found that Dahuang Fuzi Decoction(DHFZT)has a significant effect in alleviating SAP and its complications.DHFZT may realize the therapeutic effect on SAP and its complications by regulating the activation of liver Kupffer cells.Objective: To investigate the changes of liver Kupffer cell activation in severe acute pancreatitis and the effect of DHFZT on the activation of liver Kupffer and the regulation mechanism of inflammation.Methods: The 24 male SD rats provided by the Experimental Animal Center of Dalian Medical University were randomly divided into sham operation group(SH,n=6),sham operation DHFZT group(SH-DHFZT,n=6),severe acute pancreatitis group(SAP n=6)and DHFZT group(DHFZT,n=6).After anesthetizing the rats,the sham operation group and sham operation DHFZT group had opened the abdomen and gently turned the pancreas several times before closing the abdomen.SAP model was induced by retrograde injection of 4% sodium taurocholate(1ml/kg)into the pancreatic duct.After the modeling was completed,the sham group and the SAP group had retention enema with Saline of 2 ml every 12 hours after surgery.The SH-DHFZT and DHFZT group was treated with 2 ml DHFZT retention enema at the same point.Rats were sacrificed48 hours after surgery,and abdominal aorta blood was collected for plasma endotoxin,blood amylase and inflammatory factors.Rat pancreas,lung,liver and ileal tissue samples were collected,and tissue sections were harvested after paraffin dehydration.HE staining was used to analyze the histomorphological changes of DHFZT before and after treatment.The expression of protein F4/80,CD86,CD163 in liver Kupffer cells was analyzed by immunohistochemistry and Western-blotting.Results:1.The levels of serum endotoxin,amylase,blood calcium: the level of serum endotoxin and amylase in the model group and treatment group were higher than those in the sham operation group(P<0.05).Compared with the model group,the content of the serum endotoxin and amylase in the treatment group was significantly lower(P<0.05).The serum calcium level in the model group was lower than that in the sham operation group(P<0.05).Compared with the model group,the serum calcium in the treatment group was higher(P<0.05).2.There were no obvious abnormalities in the structure of pancreas,lung,ileum and liver in SH group and SH-DHFZT group.The expressions of F4/80,CD86 and CD163 in SH-DHFZT group were not significantly different from those in SH group.The expression of F4/80,CD86 and CD163 increased in the tissues of SAP group at 48 h after operation(P<0.05),compared with SAP group,the expression of F4/80 and CD86 in DHFZT group was decreased,expression of CD163 was increased(P<0.05),and the damage of pancreas,lung and ileum tissue structure was alleviated.3.The levels of serum IL-1?,i NOS,TNF-? and IL-10 in the SH-DHFZT group did not change significantly compared with the SH group.The levels of serum IL-1?,i NOS,TNF-? and IL-10 in the SAP group and DHFZT group were significantly higher than those in the SH group(P<0.05).The levels of IL-1?,i NOS and TNF-? in the DHFZT group were lower than those in the SAP group(P<0.05).while IL-10 increased,the difference was statistically significant(P<0.05).