Expression And Function Of Circular RNA Hsa_circ₀011940 In Peripheral Blood Of Patients With Anti-tuberculosis Drug-induced Liver Injury

Objective To screen and validate the differentially expressed circular RNA?circRNA?in the plasma of patients with anti-tuberculosis drug-induced liver injury?ADLI?,the mechanism action of circRNA as a miRNA sponge in ADLI was further explored from cytological experiments.Methods Research was conducted from both crowds and cells.1 Crowd study:Blood samples and clinical data of inpatients diagnosed as pulmonary tuberculosis in the Fourth Hospital of Tangshan City were collected.According to strict exclusion criteria,16 patients in the liver injury group and non-liver injury group were selected,Screenning differential expression of circRNA in two groups by chip technology and predicting the target miRNA of the circRNA;Then 150 patients in the liver injury group and non-liver injury group were selected to detect the differential expression of circRNA and miRNA by qRT-PCR.The expression levels of inflammatory factors?NF-?B,TNF-?,IL-6?were detected by ELISA;t-test or ?2 test was used to analyze the basic data of patients,has circ 0011940 and miR-21 expression,ALT,AST,NF-?B,TNF-? and IL-6 protein levels Whether the change was statistically significant,Pearson correlation analysis judged the correlation between has circ 0011940 and liver function indicators?ALT,AST?,miR-21,inflammatory factors?NF-?B,TNF-?,IL-6?.2 Cytological experiments:The liver injury cell model was constructed with the optimal concentration of three drugs.The recombinant over-expression plasmid was constructed by whole-gene synthesis method,and the cells were divided into six groups:blank group?drug group?circRNA overexpression group?overexpression negative control?circRNA knockdown group?knockdown negative group.HE staining was used to observe the morphological changes of the cells in each group.The ALT and AST contents were detected by the Lai method.The mRNA expression of has circ 0011940 and NF-?B,TNF-? and IL-6 were detected by qRT-PCR.The expression levels of NF-?B protein in each group were detected by Western Blot.The expression levels of TNF-? and IL-6 protein in each group were detected by ELISA.In order to verify whether circRNA acts as a miRNA sponge to regulate the inflammatory response in ADLI,cells were further divided into blank group,drug group,miR-21 overexpression group,overexpression negative control group,miR-21 knockdown group,and knockdown negative control group.Six groups were selected to detect changes in various indicators of each group of cells.Results 1 Population study:A total of 6661 differentially expressed circRNAs were screened by peripheral blood in ADLI patients using circRNA microarray.Among them,6389 were down-regulated and 272 were up-regulated.The selected target circular has circ 0011940 was expressed in plasma of ADLI patients.Down-regulation?P<0.05?,its ability to bind to miR-21 as an RNA sponge resulted in increased expression of miR-21 in peripheral blood of ADLI patients,consistent with the predicted results;and inflammatory factor NF-?B in peripheral blood of ADLI patients and The expression levels of inflammatory factors TNF-a and IL-6 were also increased?all P<0.05?;has circ 0011940 and liver function indicators?ALT,AST?,miR-21,inflammatory factors?NF-?B,IL-6,TNF-??was negatively correlated?r =-0.674,-0.545,-0.309,-0.779,-0.684,-0.492,all P<0.05?.2 Cell study:Compared with the blank control group,the number of cell morphology changes in the drug group decreased,the content of ALT and AST increased,the expression of has circ 0011940 decreased,and the mRNA and protein levels of inflammatory factors NF-?B,TNF-?,IL-6 increased.P<0.05);overexpression of has_circ₀011940 reduced the mRNA and protein levels of inflammatory factors NF-?B,TNF-?,IL-6?all P<0.05?,while knocking down has_circ₀011940 aggravated the process of hepatocyte injury induced by anti-tuberculosis drugs.Inflammatory response?P<0.05?.In the reverse validation experiments,the expression of miR-21 in the drug group was higher than that in the blank group;knocking down miR-21 reduced the inflammatory response in the process of hepatocyte injury induced by anti-tuberculosis drugs;while overexpression of miR-21 aggravated anti-tuberculosis The inflammatory response in the process of hepatocyte injury induced by drugs?all P<0.05?.Conclusion 1 The expression level of has_circ₀011940 is decreased in the process of anti-tuberculosis drug-induced liver injury,and negatively correlated with liver function indexes?ALT,AST?,miR-21,inflammatory factors?NF-?B,IL-6,TNF-??.2 Has_circ₀011940 can reduce the inflammatory response in the process of anti-tuberculosis drug-induced liver injury by inhibiting the NF-?B signaling pathway as a miR-21 sponge.Figure 15;Table 15;reference 161.