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Endotoxin (endotoxin) is the presence of the cell wall lipopolysaccharide (lipopolysaccharide, LPS) component of the outer membrane of Gram-negative bacteria. Endotoxemia (endotoxemia, ETM) to participate in the clinical variety of diseases, development, such as sepsis, burns, acute respiratory distress syndrome (ARDS), acute and chronic liver disease, severe pancreatitis, severe cases can lead to infection the shock (septic shock), disseminated intravascular coagulation (DIC) and multiple organ dysfunction (MODS). That the monocyte / macrophage system is the main target cells of endotoxin, but also produce a major effector cells of endotoxin tolerance. Toll-like receptor 4 (TLR4) in endotoxin signal transduction plays an important role. IL-1 receptor-associated kinase-M (IRAK-M) is an important negative regulator of the signal transduction pathway factor. The Xuebijing injection safflower, red peony, Chuanxiong Angelica, Salvia, traditional Chinese medicine extracted from. Domestic study reported that the drug directly endotoxin to reduce endogenous inflammatory mediators uncontrolled release, improve microcirculation, protection and restoration of stress damage to the organ, reduce sepsis (sepsis) and the MODS of mortality due to other reasons. The purpose of this research project is to be an objective evaluation of the in vitro test the blood will net \The first part of blood will be the net directly in and the role of endotoxin in vitro research purposes: understanding the role of Xuebijing vitro without direct neutralizing or antagonistic toxins. Methods: Different concentrations of blood Xuebijing and standard endotoxin after mixing a water bath, and then using the Limulus test (the chromogenic substrate assay) the endotoxin content in the mixture. Results: With increasing concentrations of Xuebijing, compared with the positive control group, the endotoxin content of anti-rise, but the difference was not statistically significant (p> 0.05). With the extension of the water bath time, each experimental group average toxin water show a downward trend, but no significant difference (p> 0.05). Conclusion: The blood will net vitro directly in and endotoxin role. Affect the purpose the second part Xuebijing endotoxin-stimulated THP-1 cells: understanding blood will net whether blocking endotoxin-stimulated THP-1 cells secretion of TNF-α; further explore the possible mechanism of action, whether blocking endotoxin signal transduction induced by endotoxin tolerance. Methods: advance with different concentrations of the blood will net (10,25,50 mg / ml concentration gradient) treatment of THP-1 cells, the control group did not add Xuebijing different time (respectively 4, 12, 24 hours ), and then 10ng/ml of endotoxin to stimulate THP-1 cells four hours. Levels of TNF-α in the supernatant was measured by ELISA; groups total cellular RNA was extracted using real-time quantitative PCR to determine and compare the cells in each group TLR4 and IRAK-M mRNA expression differences. Results: (1) under the the different concentrations Xuebijing and different time role in each experimental group cells TNF-α secretion levels had no significant change (p> 0.05). (2) only for 24 hours, TLR4 expression only with the Xuebijing increasing concentrations of a slight increase, but still no statistically significant difference compared with the control group; 50mg/ml concentration, 24 hours of TLR4 expression is 1.547 times of 4 hours, and the difference was statistically significant (p <0.05). (3) the role of 4 hours and 12 hours, the concentration of blood will net group IRAK-M expression of the control group had no significant difference; extended to 24 hours, the high concentration (50mg/ml) blood will net group IRAK-M upregulation of 1.349 times the control group (p <0.05), while the low was no significant difference between the concentration group and the control group; and, in the expression of IRAK-M The high concentration Xuebijing group, along with the prolonged duration of action also raised, but no significant difference (p> 0.05). THP-1 cells conclusion: (1) The blood shall be net did not block endotoxin stimulate the secretion of TNF-α. (2) high concentrations of blood will net long role may at the same time raised the THP-1 cells TLR4 and IRAK-M mRNA expression, but the exact meaning remains unclear.
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