Session Type: Abstract Submissions (ACR)
Background/Purpose: Metallothioneins bind heavy metals with high affinity and can serve as storage proteins for labile Zn2+ which in turn can regulate immune system activity through interactions with Toll-like receptor (TLR) signal transduction. Nephrogenic Systemic Fibrosis (NSF) is a generalized progressive fibrotic disorder described in some patients with renal insufficiency exposed to various gadolinium based contrast agents (GdBCA). The GdBCA Omniscan activates expression and production of several proinflammatory and profibrotic cytokines and growth factors in normal differentiated human macrophages via TLR4 and TLR7.signaling. Since some GdBCA are capable of inducing transmetallation by displacing Zn2+from proteins the effect of GdBCA on expression levels of metallothionein genes in normal human macrophages and fibroblasts was examined in this study.
Methods: Terminally differentiated macrophages generated from two normal buffy coats were exposed for 24 hour to either 50 mM Omnisan or saline. Total RNA was isolated, labeled and hybridized to Affymetrix human U133 2.0 Plus microarrays. Volcano plots were used to identify differentially expressed genes between Omniscan treated and saline treated cells employing parametric testing assuming equal variances. Differential gene expression was confirmed by real-time PCR on the same RNA samples. Validation experiments utilizing 3 additional differentiated macrophage isolates and two early passage (<6) normal human dermal fibroblasts examined the effect 1 mM Omniscan on the expression of metallothionein genes.
Results: Microarray analyses showed marked downregulation (~1.5 to 4 fold) of expression of multiple metallothionein genes (MT1E, MT1F, MT1G, MT1H, MT1M, MT1X and MT2A) in cells treated with 50 mM Omniscan compared to saline treated controls. These results were confirmed by real time-PCR analysis. Lower doses (1 mM) of Omniscan also downregulated the expression of these same genes as well as MT3 and MT4 whereas the expression of MT1A and MT1B was upregulated in normal human differentiated macrophages as well as in normal human dermal fibroblasts.
Conclusion: Global gene expression microarrays and real-time PCR analysis showed that exposure of terminally differentiated normal human macrophages to 50 mM Omniscan downregulated expression of multiple metallothionein genes in comparison to saline treated controls. Exposure of differentiated macrophages and normal human dermal fibroblasts to 1 mM Omniscan also decreased expression of these genes but increased expression of MT1A and MT1B suggesting that changes in metallothionein expression following GdBCA exposure in macrophages and dermal fibroblasts may play a role in the pathogenesis of the severe fibrotic process of NSF pathogenesis.
P. J. Wermuth,
F. Del Galdo,
S. A. Jimenez,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/downregulated-expression-of-metallothionein-genes-in-response-to-the-gadolinium-contrast-agent-omniscan-in-normal-human-differentiated-macrophages-and-dermal-fibroblasts/