The Study Of Jieduquyuziyin Prescription Alleviating The Illness In Lupus Mice By Inhibiting P2X7R/NLRP3 Signaling Pathway

Objective To explore the mechanism that Jieduquyuziyin Prescription alleviates the illness in lupus mice by inhibiting P2X7R/NLRP3 signaling pathway.Methods ①The following groups were administered intragastrically for 10 weeks:18 ml/kg normal saline for model group.P2X7R inhibitor group and control group:12.5 mg/kg prednisone for western medicine group;18 ml/kg Jieduquyuziyin Prescription for Chinese medicine group and Chinese medicine combined with P2X7R inhibitor group.A-438079 80 mg/kg were given to P2X7R inhibitor group and Chinese medicine combined with P2X7R inhibitor group at the 16 week of age.All the mice were sacrificed at the 17 week of age,when the weights of the whole body,spleen and kidneys were measured,the serum and urine were collected,and the BMDM from the femur and tibia was obtained.The levels of serum anti-ds-DNA antibody,the expressions of P2X7R,NLRP3,IL-1β and IL-18,the cell surface makers of F4/80,CD86 and CD 163,and the levels of TNF-α and IL-10 in BMDM cell supernate were detected.②The levels of urine protein,serum creatinine and urea nitrogen were measured.The kidney macrophage infiltration of each group was evaluated.The kidney macrophage polarization(marked by F4/80,CD86 and CD 163)of each group was tested.The levels of TNF-α and IL-10,the expression levels of P2X7R.NLRP3.ASC and Caspase-1 p20 and the relative mRNA levels of P2X7R,NLRP3,IL-1β and IL-18 in kidneys were tested.③RAW264.7 cells were divided into 5 groups,including blank group,Chinese medicine group,LPS+ATP group,LPS+ATP+P2X7R inhibitor group and LPS+ATP+Chinese medicine group Then,the cell viability,the levels of TNF-α and IL-10 in the cell supernatant,the expression levels of CD86,CD163,P2X7R,NLRP3,ASC,Caspase-1 p20 and NEK7,the relative mRNA levels of IL-1β and IL-18 and the levels of ROS were measured.The cell surface markers including F4/80,CD86 and CD163 were testedResults ① Compared with model group,western medicine group,Chinese medicine group.P2X7R inhibitor group and Chinese medicine combined with P2X7R inhibitor group were improved at aspects of hair loss,lymph nodes enlargement,slow movement,spleen index and kidney index;levels of serum anti-ds-DNA antibody were significantly lower;M2 type BMDM marked by F4/80 and CD 163 was increasing and M1 type BMDM marked by F4/80 and CD86 was decreasing;the relative mRNA levels of P2X7R,NLRP3.IL-1β and IL-18 were significantly decreasing.②Compared with model group,western medicine group,Chinese medicine group,P2X7R inhibitor group and Chinese medicine combined with P2X7R inhibitor group had lower levels of urine protein,serum creatinine and urea nitrogen;the infiltration of macrophages in the kidney were less,with less layers of cell proliferation;the distribution of macrophages in the kidney were less,with the domination of M2 type;the levels of TNF-α and IL-10 in the kidney were decreasing;the expression levels of P2X7R.NLRP3,ASC and Caspase-1 p20 in the kidney were decreasing;the relative mRNA levels of P2X7R,NLRP3,IL-1β and IL-18 were significantly decreasing.③After collecting the RAW 264.7 cells,the cell viability ratio of Chinese medicine group were significantly increasing compared with blank group;the numbers of M1 type macrophage marked by F4/80 and CD86 were decreasing;the expression of CD 163 was increasing;the expression of CD86,P2X7R,NLRP3,ASC,Caspase-1 p20 and NEK7 were decreasing;the levels of TNF-α in the cell supernatant were decreasing and the levels of IL-10 increasing;the expression levels of IL-1βand IL-18 mRNA were decreasing;the level of ROS was significantly decreasing.Compared with LPS+ATP group,LPS+ATP+P2X7R inhibitor group and LPS+ATP+Chinese medicine group have increased cell viability,lower number of M1 type macrophages marked by F4/80 and CD86,up-regulated CD 163,down-regulated expressions of CD86,P2X7R,NLRP3,ASC.Caspase-1 p20 and NEK7,lower expression of TNF-α in the cell supernatant,an increased level of IL-10 and a decreased level of ROSConclusion①Jieduquyuziyin Prescription could reverse the imbalance of BMDM polarization via P2X7R/NLRP3 pathway to relieve the systemic condition in MRL/Ipr mice.② Jieduquyuziyin Prescription could reverse the imbalance of MRL/lpr mice macrophage polarization via P2X7R/NLRP3 pathway to relieve the kidney damage.③ Jieduquyuziyin Prescription could reverse the imbalance of RAW264.7 cells macrophage polarization via P2X7R/NLRP3 pathway to relieve inflammation.