Gene expression network in PiZZ Alpha1 Antitrypsin Deficiency and its impact on the disease evolution

Abstract

Introduction Chronic obstructive pulmonary disease (COPD) is a complex disease with both genetic and environmental determinants, the most important of which is cigarette smoking. However, genetic variations like severe a1-antitrypsin deficiency (AATD) is known as a genetic risk factor for COPD. Severe AATD patients present an heterogeneous respiratory clinical evolution not explained by sociodemographic or clinical factors. We analyzed the gene-expression profiles of blood samples collected from AATD Pi*ZZ patients and healthy controls to identify differentialy expressed mRNA and miRNAs that may be involved in the progression of the AATD. However, this first analysis are based on the comparison between severe and mild AATD Pi*ZZ patients. Methods In this pilot high-throughput microarray experiment, RNA and miRNA microarrays hibridization was carried out using Genechip Human Gene Array Plate 2.1 and Genechip miRNA array plate 4.1 respectively. Background correction and quantile normalization were performed for the raw array data using the Oligo R package. Limma (Linear Models for Microarray Data) package was used to perform statistical testing of differential mRNA and miRNA expression between the 2 study groups defined as having a p<0.05 with at least 1.5-fold change (FC) (logFC 0.58) between the groups. Functional and enrichment analysis were also performed using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene-ontology (GO) terms. Finally, an integration of miRNA DEGs and their target mRNA DEGs was performed using ”multiMiR” R package. All the analysis was performed using R Studio (3.2.5 version). Results A total of 12 PiZZ AATD patients (n=6 in severe group and n=6 in mild disease group) were included in this pilot high-throughput microarray experiment. Both AATD groups presented similar characteristics in terms of current age, gender, exacerbations, comorbidities, replacement treatment and time from the last infusion. Severe patients were diagnosed younger than mild patients and all of them received inhaled corticosteroids (ICS). Two patients from the severe group were current smokers and therefore were not receiving replacement therapy. Limma analysis was finally performed in 4 severe and 6 mild patients according to quality control results. A total of 205 DEGs (114 upregulated and 91 downregulated) and 28 miRNA with different expression (20 upregulated and 8 downregulated) were observed in severe PiZZ AATD compared to mild patients. Of those, we found a validated interaction between hsa-miR-335-5p and 12 target upregulated mRNA. These transcripts participate dierctly in important pathways like cytokine-mediated signaling pathway, MAPK /mk2 signaling cascade, JNK cascade and angiogenesis. Conclusions Despite the limitations, the findings reported in this analysis provide valuable information about the different gene profile of severe PiZZ AATD patients. Hsa-miR-335-5p regulated important transcription factors that are involved in the activation of pathways directly related to inflammation and angiogenesis in severe PiZZ AATD patients and could play an important role in the pathogenesis of this rare condition. Nevertheless, further analysis are needed in order to validate our results.