Persistence of Innate Immune Pathways in Late Stage Human Bacterial and Fungal Keratitis: Results from a Comparative Transcriptome Analysis

Abstract

Microbial keratitis (MK) is a major cause of blindness worldwide. Despite adequate antimicrobial treatment, tissue damage can ensue. We compared the human corneal transcriptional profile in late stage MK to normal corneal tissue to identify pathways involved in pathogenesis. Total RNA from MK tissue and normal cadaver corneas was used to determine transcriptome profiles with Illumina HumanHT-12 v4 beadchips. We performed differential expression and network analysis of genes in bacterial keratitis (BK) and fungal keratitis (FK) compared with control (C) samples. Results were validated by RTqPCR for 45 genes in an independent series of 183 MK patients. For the microarray transcriptome analysis, 27 samples were used: 12 controls, 7 BK culture positive for Streptococcus pneumoniae (n = 6), Pseudomonas aeruginosa (n = 1), and 8 FK, culture positive for Fusarium sp. (n = 5), Aspergillus sp. (n = 2), or Lasiodiplodia sp. (n = 1). There were 185 unique differentially expressed genes in BK, 50 in FK, and 339 common to both [i.e., genes with fold-change (FC) < −4 or ≥4 and false discovery rate (FDR) adjusted P < 0.05]. MMP9 had the highest FC in BK (91 FC, adj p = 3.64 E-12) and FK (FC 64, adj. p = 6.10 E-11), along with other MMPs (MMP1, MMP7, MMP10, MMP12), pro-inflammatory cytokines (IL1B, TNF), and PRRs (TLR2, TLR4). HIF1A and its induced genes were upregulated uniquely in BK. Immune/defense response and extracellular matrix terms were the most enriched Gene Ontology terms in both BK and FK. In the network analysis, chemokines were prominent for FK, and actin filament reorganization for BK. Microarray and RTqPCR results were highly correlated for the same samples tested with both assays, and with the larger RTqPCR series. In conclusion, we found a great deal of overlap in the gene expression profile of late stage BK and FK, however genes unique to fungal infection highlighted a corneal epithelial wound healing response and for bacterial infection the prominence of HIF1A-induced genes. These sets of genes may provide new targets for future research into therapeutic agents.

Bacteria induce autophagy in a human ocular surface cell line

“…Microbial keratitis or inflammation resulting from infection of the cornea can lead to tissue damage, and vision loss (Lakhundi et al, 2017). Even when following a strict antibiotic regimen, corneal ulceration can readily occur resulting in perforation in 30% of cases and loss of the eye in 25% of patients (Burton et al, 2011; Chidambaram et al, 2017; Poole et al, 2002). The primary bacterial pathogens responsible for causing keratitis include Stapylococcus aureus, Pseudomonas aeruginosa, Serratia marcescens, Moraxella sp., Haemophilus influenzae, coagulase-negative Staphylococci, and Stretococcous sp (Kowalski et al, 2013).…”

Whole Exome Sequencing Identifies New Host Genomic Susceptibility Factors in Empyema Caused by Streptococcus pneumoniae in Children: A Pilot Study

“…The gene MEIS1 was found to be down-regulated in both corneal tissue from humans (GSE58291 [ 54 ]; Figure 4 A) and lung tissue from mice (GSE45644 [ 51 ]; Figure 4 B) suffering pneumonia caused by S. pneumoniae . In mice, the p -value is in the multiple-test correction limit, which could reflect the low sample size available for cases and controls.…”

“…We found a significant result of association for the transcript of APOBR gene in four studies (GSE42464 [ 49 ], GSE49533 [ 50 ], GSE49755 [ 52 ], and GSE58291 [ 54 ]). The APOBR encodes the receptor of the APOB protein.…”

“…The direct molecular link existing between APOB and APOBR is very suggestive of an association of pneumonia caused by S. pneumoniae and the APOB pathway. Furthermore, we have observed a tissue-dependent regulation in the data from different studies, three of them showing over-expression of APOBR in lung [ 49 , 50 ] and cornea [ 54 ], (two tissues that are directly exposed to the air and therefore a possible direct contact with the pathogen), while another study indicates a down-regulation of APOBR in plasma [ 52 ]. This result seems most promising for the understanding of the mechanisms of infection of S. pneumoniae , but it needs further validation in larger cohorts.…”

“…We filtered the results from these queries by selecting only those studies on humans and mice. We detected only six studies containing suitable data to validate our gene candidates, including three studies in mice (GEO accession numbers: GSE42464 [ 49 ], GSE49533 [ 50 ], and GSE45644 [ 51 ], all of them carried out in lung tissue), and three in humans (GSE49755 [ 52 ] in plasma, GSE6269 [ 53 ], in blood, and GSE58291 [ 54 ], in corneal tissue). Table S1 contains the microarray sample IDs of the datasets analyzed that passed all the quality filters and were publicly available (February 10, 2018).…”