siRNA-based screening was performed using 20 nM final siRNA complexed to NeuroMag transfection reagent
This paper shows the high efficiency of NeuroMag transfection reagent from OZ Biosciences to transfect primary Retinal Ganglion Cells with siRNA.Moreover this article illustrates the usefulness of Magnetofection for high content screening.article reference: Proc Natl Acad Sci U S A. 2013 Feb 19.
Functional genomic screening identifies dual leucine zipper kinase as a key mediator of retinal ganglion cell death.Welsbie DS, Yang Z, Ge Y, Mitchell KL, Zhou X, Martin SE, Berlinicke CA, Hackler L Jr, Fuller J, Fu J, Cao LH, Han B, Auld D, Xue T, Hirai SI, Germain L, Simard-Bisson C, Blouin R, Nguyen JV, Davis CH, Enke RA, Boye SL, Merbs SL, Marsh-Armstrong N, Hauswirth WW, Diantonio A, Nickells RW, Inglese J, Hanes J, Yau KW, Quigley HA, Zack DJ.Abstract
Glaucoma, a major
cause of blindness worldwide, is a neurodegenerative optic neuropathy in
which vision loss is caused by loss of retinal ganglion cells (RGCs).
To better define the pathways mediating RGC death and identify targets
for the development of neuroprotective drugs, we developed a
high-throughput RNA interference screen with primary RGCs and used it to
screen the full mouse kinome. The screen identified dual leucine zipper
kinase (DLK) as a key neuroprotective target in RGCs. In cultured RGCs,
DLK signaling is both necessary and sufficient for cell death. DLK
undergoes robust posttranscriptional up-regulation in response to axonal
injury in vitro and in vivo. Using a conditional knockout approach, we
confirmed that DLK is required for RGC JNK activation and cell death in a
rodent model of optic neuropathy. In addition, tozasertib, a small
molecule protein kinase inhibitor with activity against DLK, protects
RGCs from cell death in rodent glaucoma and traumatic optic neuropathy
models. Together, our results establish a previously undescribed
drug/drug target combination in glaucoma, identify an early marker of
RGC injury, and provide a starting point for the development of more
specific neuroprotective DLK inhibitors for the treatment of glaucoma,
nonglaucomatous forms of optic neuropathy, and perhaps other CNS
neurodegenerations.
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