Neutrophil Gene expression in Acute-on-chronic liver failure (ACLF), Chronic liver disease (CLD) and Healthy Controls_AIIMSND_Aug2020

To address the molecular basis of immune-dysfunction in Acute-on-chronic liver failure (ACLF), we carried out gene expression profiling of blood derived neutrophil from ACLF, belonging to both sterile inflammatory and sepsis conditions. Peripheral whole blood was subjected to PMN enrichment by double gradient centrifugation, and RNA isolation was done by TRIZOL method, followed by microarray experiment using Agilent one-color platform. We compared the gene expression of these neutrophils with that of Chronic liver disease (CLD) patients and healthy controls (HC) for baseline relative quantification, and found unique set of upregulated and downregulated genes in ACLF. We validated the expression of the most differentially expressed genes by quantitative RT-PCR and also stratified the patients into survivors and non-survivors, sepsis and sterile-inflammation. We found an upregulated 3-gene signature of ELANE-MPO-CD177 to be associated with 28-day mortality, irrespective of presence or absence of sepsis. Total RNA from blood enriched neutrophils was subjected to one-color microarray experiment for the following study groups: ACLF sepsis (n=6), ACLF sterile inflammation (n=6), CLD (n=6), Healthy controls HC (n=6). Differential gene expression analysis was done for CLD vs HC, ACLF overall (n=12) vs CLD, and ACLF sepsis vs sterile inflammation.