Identification of Aeromonas hydrophila cytotoxic enterotoxin-induced genes in macrophages using microarrays.
A cytotoxic enterotoxin (Act) of Aeromonas hydrophila possesses several biological activities, and it induces an inflammatory response in the host. In this study, we used microarrays to gain a global and molecular view of the cellular transcriptional responses to Act and to identify important genes up-regulated by this toxin. Total RNA was isolated at 0, 2, and 12 h from Act-treated macrophages and applied to Affymetrix MGU74 arrays, and the data were processed using a multi-analysis approach to identify genes that might be critical in the inflammatory process evoked by Act. Seventy-six genes were significantly and consistently up-regulated. Many of these genes were immune-related, and several were transcription factors, adhesion molecules, and cytokines. Additionally, we identified several apoptosis-associated genes that were significantly up-regulated in Act-treated macrophages. Act-induced apoptosis of macrophages was confirmed by annexin V staining and DNA laddering. Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay were used to verify increased expression of some inflammatory and apoptosis-associated genes identified by the microarray analysis. To further confirm Act-induced increases in gene expression, real-time RT-PCR was also used for selected genes. Taken together, the array data provided for the first time a global view of Act-mediated signal transduction and clearly demonstrated an inflammatory response and apoptosis mediated by this toxin in host cells at the molecular level.
Galindo
Cristi L
CL
Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas 77555-1070, USA.
Sha
Jian
J
Ribardo
Deborah A
DA
Fadl
Amin A
AA
Pillai
Lakshmi
L
Chopra
Ashok K
AK
D016980:Aeromonas hydrophila / Q000472:pathogenicity; D000818:Animals; D001426:Bacterial Proteins; D001427:Bacterial Toxins / Q000633:toxicity*; D001483:Base Sequence; D015815:Cell Adhesion Molecules / Q000235:genetics; D002460:Cell Line; D018925:Chemokines / Q000235:genetics; D016207:Cytokines / Q000235:genetics; D004247:DNA / Q000235:genetics; D004768:Enterotoxins / Q000633:toxicity*; D005246:Feedback; D020869:Gene Expression Profiling; D007249:Inflammation / Q000209:etiology / Q000235:genetics; D008264:Macrophages / Q000187:drug effects* / Q000378:metabolism*; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D014157:Transcription Factors / Q000235:genetics; D015854:Up-Regulation / Q000187:drug effects
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D015815:Cell Adhesion Molecules; D018925:Chemokines; D016207:Cytokines; D004768:Enterotoxins; D014157:Transcription Factors; C082174:cytolytic enterotoxin protein, Aeromonas; D004247:DNA