Preliminary Characterization of the Interaction Mechanisms of Engineered Nanomaterials with DNA

Featuring Bryant Nelson
National Institute of Standards and Technology, Gaithersburg, MD

Location:  2430 Engineering Hall
Free and open to the public

Abstract:

Exposure to engineered nanomaterials through normal manufacturing processes (via inhalation or skin absorption) or through intentional therapeutic or diagnostic procedures (via oral ingestion or subcutaneous injection) is an immediate and significant concern for human health and safety. This concern derives from the observation that certain nanomaterials have the potential to generate reactive oxygen species (ROS) in biological environments. ROS in the form of free radicals are naturally present in the body, however, an overabundance of free radicals can overwhelm the body’s natural antioxidant defense system and lead to oxidative stress. Oxidative damage to the body’s genomic DNA is one of the consequences of oxidative stress. It is not known at a fundamental molecular level if engineered nanomaterials promote or hinder the formation of free radicals in the body or how these nanomaterials might interact with specific molecular targets such as DNA. By using simple solutions of calf thymus DNA and various types of gold nanoparticles (Au-NPs) as preliminary model systems, the present work aims to elucidate the fundamental interaction mechanisms of nanomaterials with DNA that will enable continued and deeper research into the potential mechanisms of nanomaterial interactions (if any) with the DNA in mammalian cells.

Isotope-dilution liquid chromatography/mass spectrometry (LC/MS) and isotope-dilution liquid chromatography/tandem mass spectrometry (LC/MS/MS) methods were developed and utilized to monitor the occurrence of DNA damage in solutions of ct-DNA + AuNPs. DNA damage was quantified by detecting and measuring the formation of oxidatively-induced DNA lesions such as 8-hydroxydeoxyguanosine (8-OH-dG), 8-hydroxydeoxyadenosine (8-OH-dA), (5'S)-8,5'-cyclo-2'-deoxyadenosine (S-cdA) and (5'R)-8,5'-cyclo-2'-deoxyadenosine (R-cdA) in the bicomponent solutions. AuNPs with diameters from 2 to 60 nm were tested. Samples of the ct-DNA + AuNPs solutions, with NP concentrations of 1, 10 and 90 nmol/L, were incubated at 37 °C for 1 h and the DNA was precipitated, washed and enzymatically digested into its component bases. Sample digests and control digests were analyzed for NP induced DNA damage and surprisingly, test samples demonstrated a size-dependent interaction of the Au-NPs with DNA. The 10 nm Au-NPs produced a significant and consistent reduction, versus control samples, in the measured level of 8-OH-dG that was not observable with either the 30 or 60 nm Au-NPs.