Isoniazid Induced Bombyx Mori Fat Body Injury And Its Transcriptome Analysis

Tuberculosis is one of the three diseases harmful to human health, due to its high incidence and high mortality, had been a global public health issues. Isoniazid(INH) remains first-line agent for the treatment of tuberculosis. However, INH has been associated with drug-induced liver injury(DILI) under long-term treatment. In this paper, Bombyx mori(silkworm) as a model animal, used to detect the damage in Bombyx mori fat boby induced by INH after orally exposed to INH. We comparative analyze genes’ expression between two silkworm fat body transcriptome libraries with the use of RNASeq technology. Discuss the potential effects on Bombyx mori fat boby metabolism and function of INH.1 INH induced silkworm fat body damage that similar to mammalian liver damageThe 72 h old of the 5th instar of Bombyx mori Dazao were orally exposed to INH during 15 min. High performance liquid chromatography(HPLC) detected a certain amount of original INH in midgut、blood(plasma) and fat body of Silkworm. ROS agent detected that content of active oxygen content was observed in the INH group of silkworm fat body and hemolymph more than CK group; The degree of fat body damage induced by INH discovered by HE and DAPI staining found that there is more broken cells and nuclei in the INH treatment group compared with CK treatment group. Meanwhile the detection of enzyme activities of alanine aminotransferase(ALT) and aspartate aminotransferase(AST) which were the Liver Damage Index in silkworm hemolymph showed that their activities had a significantly increased along with INH dose and time of treatment. It indicted that INH induced Bombyx fat body injury were similar to INHinduced acute liver injury in a mammal.2 INH induced gene transcription level change in silkworm fat bodyThe 72 h old of the 5th instar of Bombyx mori Dazao were orally administered 100 μL1000 mg/kg(the volume of solution is100 μL) INH after 8h of oral administration to extract fat body. Building two INH/CK transcriptome libraries based on RNA-Seq. All Clean reads blast to NCBI can mapped 10113 functional genes, in which 2211 differential genes including1908 genes expressed only in one library and 303 significantly differentially expressed genes between two libraries. There were 652 genes only expressed in INH group and 1264 genes silence; beside of that, 85 genes up-regulated expression and 218 genes down- regulated expression.3 Differentially expressed genes induced by INH in silkworm fat body of in GO functions and KEGG pathway analysisCompared of two DGE libraries, specifically expressed genes and significantly differentially expressed genes GO function showed: Biological Process had most number of genes, including biological regulation, cellular processes, positioning, metabolism, pigmentation and other GO terms; binding and catalytic activity contained about 90% genes in molecular function; Meanwhile, each GO terms assicated with cell and biological membrane had more differentially expressed genes in cellular component.According to the characteristics of drug metabolism, a KEGG cluster ananlysis was performed on the 10 pathways, the pathways with the greatest representation in the differentially expressed genes were nucleotide damage pathways(46members), cellular processes(52 members), extracellular signal transduction pathways(31 members) apoptosis pathway(29 members), protein metabolism(27 members). Screened INH major drug metabolism and INH-induced silkworm fat body injury KEGG pathway based on the distribution of differentially expressed genes in KEGG pathway, CYP4E3 and GST expression level had changed a lot in drug metabolism pathway during INH metabolism and decomposition process, and GST activity in fat body of female silkworm and male silkworm significantly reduced. Genes related to protein structure 、 transport and carbohydrate metabolism had significantly differentially expressed in protein processing in endoplasmic reticulum. AMPK(ATP synthesis important enzyme) and cytochrome complex(mitochondrial respiratory chain important complexes) in the process of apoptosis played an important role. The paper also detected the mitochondrial respiratory chain enzyme activity varies INH and measurement time, different enzymatic activity changes appeared different trends, but the activity of cytochrome existed typical timeand dose- response to INH. Atg6 and Atg8 expression had nothing changed.