| ObjectiveTo explore the mechanism of NLRP3 inflammatory corpuscle signaling pathway in recurrent pelvic inflammatory disease(RPID)patients with spleen deficiency and liver stagnation syndrome.RT-PCR,ELISA laboratory tests were used to detect the Nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3)mRNA expression,cysteine aspartate protease-1(Caspase-1),interleukin-1β(IL-1β)and interleukin-18(IL-18)in serum of healthy subjects(HS)and RPID patients.The correlation between symptom,physical signs,disease course,disease level,pathogen infection with NLRP3 inflammatory corpuscle signaling pathway in RPID patients were also analyzed.MethodsThe gynaecological patients ware from gynecology clinic of the Affiliated Hospital of Chengdu University of Traditional Chinese Medicine from September 2016 to November 2018,outpatient diagnosed as RPID,TCM syndrome differentiation was spleen deficiency and liver stagnation syndrome,and 22 patients who met the inclusion criteria were used as observation group.18 healthy women were used as control group.Blood samples were collected on the day after informed consent was obtained,and 3 ml of elbow venous blood was taken,1 ml of which was stored in EDTA anticoagulant blood collection tube,and Reverse Transcription-Polymerase Chain Reaction(RT-PCR)was used to detect the expression level of the target gene NLRP3 mRNA in peripheral blood mononuclear cells;another 2 ml was stored in serum tubes to prepare serum samples,and serum-associated immunosorbent assay(ELISA)was used to detect Caspase-1,IL-1β,IL-18 content,while the observation group filled in the symptoms and signs score table,and according to the patient’s willingness to culture mycoplasma,PCR method to detect chlamydia and gonococcus.Results1.Laboratory indicator test results1.1 RT-PCR was used to detect the expression of NLRP3 mRNA in peripheral blood mononuclear cells: the expression level of NLRP3 mRNA in the observation group was 0.187±0.171,and in the control group was 0.096±0.109.The expression level of NLRP3 mRNA in the observation group was higher than that in the control group,the difference was not statistically significant(P > 0.05).1.2 ELISA detection of serum Caspase-1,IL-1β,IL-18 content1.2.1 Peripheral serum Caspase-1 expression level: The expression level of Caspase-1 in the observation group was 12.355±7.515 ng/ml,and the expression level of Caspase-1 in the control group was 15.148±9.594 ng/ml.The expression level of Caspase-1 in the observation group was slightly lower than that in the control group,the difference was not statistically significant(P>0.05).1.2.2 Peripheral serum IL-1β expression level: The expression level of IL-1β in the observation group was 7.053±2.217 pg/ml,and the expression level of IL-1β in the control group was 7.958±3.752 pg/ml.The expression level of IL-1β in the observation group was slightly lower than that in the control group,but the difference was not statistically significant(P > 0.05).1.2.3 Peripheral serum IL-18 expression level: The expression level of IL-18 in the observation group was 19.617±3.704 pg/ml,and the expression level of IL-18 in the control group was 39.544±9.870 pg/ml.The expression level of IL-18 in the observation group was significantly lower than that in the control group,and the difference was statistically significant(P<0.01).2.Correlation analysis2.1 Correlation analysis of related indicators of NLRP3 inflammatory corpuscle signaling pathway2.1.1 Correlation analysis between the indicators in the observation group: In the observation group,NLRP3 mRNA was positively correlated with Caspase-1,the correlation coefficient was 0.141,and the expression of NLRP3 mRNA was negatively correlated with the expression of IL-1β and IL-18,the correlation coefficients were-0.213,-0.44,the correlation was not statistically significant(P>0.05).2.1.2 Correlation analysis between the indicators in the control group: In the control group,NLRP3 mRNA was positively correlated with the expression of Caspase-1,IL-1β and IL-18.NLRP3 mRNA was positively correlated with IL-1β(r=0.505),and the correlation was statistically significant(P <0.05).NLRP3 mRNA was positively correlated with Caspase-1 and IL-18(r=0.205,0.139),the correlation was statistically significant(P <0.05);Caspase-1 was positively correlated with IL-1β(r=0.575),and the correlation was statistically significant(P <0.05).2.2 Correlation between NLRP3 inflammatory corpuscle signaling pathway related factors and TCM syndrome score,signs,and pathogenesisThe expression level of NLRP3 mRNA in the observation group was positively correlated with TCM syndrome score,physical score and disease course(r=0.098,0.219,0.010);Caspase-1 was negatively correlated with TCM syndrome score and disease course(r=-0.153,-0.194),positively correlated with physical score(r=0.112);IL-1β was negatively correlated with TCM syndrome score(r=-0.209),positively correlated with physical score and duration(r=0.042,0.296);IL-18 was negatively correlated with TCM syndrome score and disease course(r=-0.104,-0.238),and positively correlated with physical score(r=0.015),but the correlation was not statistically significant(P>0.05).2.3 Analysis of related indicators of NLRP3 inflammatory corpuscle signaling pathway related factors and disease grade and mycoplasma infectionThe expression level of NLRP3 mRNA in the observation group was positively correlated with the patient’s disease grade and mycoplasma infection(r=0.113,0.430);Caspase-1 was negatively correlated with the disease grade(r=-0.252),and positively correlated with mycoplasma infection(r=0.233);IL-1β was negatively correlated with disease grade and mycoplasma infection(r=-0.051,-0.465);IL-18 had almost no correlation with disease grade(r=-0.003),and was positively correlated with mycoplasma infection(r=0.072).None of the above was statistically significant(P>0.05).Conclusions1.NLRP3 mRNA,Caspase-1,and IL-1β were not statistically significant between observation group and control group.The NLRP3 inflammatory corpuscle signaling pathway related factors were not significantly associated with the patient’s TCM syndrome score,physical score,course of disease,disease grade and mycoplasma infection.The role of NLRP3 inflammatory corpuscle signaling pathway in the RPID in spleen deficiency and liver stagnation syndrome is not clear.2.The expression level of IL-18 in the observation group was significantly lower than that in the control group,suggesting that RPID may be related to the low expression of IL-18.3.In the control group,NLRP3 mRNA transcription level was significantly positively correlated with IL-1β,and Caspase-1 with IL-1β,but there was no significant correlation among the indicators in the observation group.To some extent,it is suggested that RPID are related to the imbalance of immune function in patients. |
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