Core 4 - David Dorman

Core 4 - David Dorman

Core 4 – David Dorman: Project cancelled.

Manganese Health Research Program: Phase 1, Core 4

Research Core Project Number:

W81XWH-05-1-0239) (Core 4) VUMC31528-R

Research Core Project: Cellular mechanisms involved in the uptake and delivery of inhaled manganese via the olfactory nerve
Core Principal Investigator (CPI): David C. Dorman, DVM, PhD
CIIT Centers for Health Research
dorman@ciit.org
Key Collaborators:

Rusty Thomas, PhD
CIIT Centers for Health Research
rthomas@ciit.org

Brian Wong, PhD
CIIT Centers for Health Research
 wong@ciit.org

Project Objectives:

  • Objective 1: Develop adenoviral and lentiviral shRNA vectors for DMT-1.  Completion of this study will require us to alter DMT-1 expression in the rat nose.
  • Objective 2: Deliver viral particles containing the shRNAs for DMT-1 to rat olfactory epithelium and demonstrate a reduction of DMT-1 mRNA and/or protein. 
  • Objective 3: Determine whether reduced DMT-1 expression alters the delivery of inhaled manganese to the rat brain.

Project Description:

Neurotoxicity is a significant public health concern associated with manganese inhalation.  Inhaled manganese is deposited in the olfactory epithelium and can be transported directly to the mammalian brain via the olfactory nerve.  We hypothesize that the divalent metal transporter (DMT-1) plays a role in the initial uptake of inhaled manganese by the rat olfactory epithelium.  Viral vectors transfected with short hairpin ribonucleic acid (shRNA) sequences that target degradation of the DMT-1 gene messenger RNA (mRNA) will be developed.  Once developed, the modified virons will be administered to the right side of the rat’s nasal cavity while media will be instilled into the left nostril, thus each animal can serve as it’s own control.  Rats treated with shRNAs and having altered olfactory epithelial DMT-1 will then be exposed to manganese chloride (54MnCl2).  Rats will be exposed to 54MnCl2 for 90 minutes and killed 1 to 8 days after the end of the inhalation exposure.  Brain and nasal tissues will be collected and evaluated for 54Mn levels,  If our hypothesis is correct, then we expect to show that brain levels of manganese (as 54Mn) will be decreased on the side of the head with defective DMT-1 transport.

Project Status:

Project started:                         May 1, 2005  
Completed:     January 2007  

Key research accomplishments:

  • Confirmation of DMT1 mRNA expression in the rat nasal epithelium.
  • Development of lentiviral vectors with green fluorescent protein (GFP) and beta galactosidase (LACZ) expression markers.
  • Development of rodent anesthesia methods and procedures to facilitate slow infusion of virus particles into the rat nasal cavity.
  • Development of nasal explant systems for in vitro virus transfection studies.  These methods allow method development efforts to rely on the use of fewer animals and takes advantage of tissue samples from naïve rats that would otherwise be discarded.

Last updated: July 2011


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