Cell lines (T17, T48, and T62) to screen for TrkA and/or TrkB agonists.
Neurotrophins play an essential role in the development and maintenance of the nervous system. One such neutrophin, nerve growth factor (NGF) binds mainly tyrosine kinase A (TrkA). Docking of TrkA by NGF initiates receptor dimerization, phosphorylation of cytoplasmic tyrosine residues on the receptor, and a cascade of cell-signaling events. These signals lead to the prevention of apoptotic cell death, promotion of cellular differentiation and axon elongation, and up-regulation of choline acetyl transferase. Several neuronal cell types implicated in various diseases express TrkA and therefore respond to NGF. In neurodegenerative processes such as mild cognitive impairment, loss of TrkA density correlates with neuronal atrophy and precedes neuronal death and severe cognitive impairment in Alzheimer's disease (AD). In mild cognitive impairment and AD progression, loss of TrkA correlates with cognitive decline. For example, in basal forebrain neurons of the aged rat, the expression of NGF receptors is decreased, but can be reversed by NGF administration.
Another neutrophin, brain-derived neurotrophic factor (BDNF) acts on two transmembrane receptors: the p75 neurotrophin receptor (p75NTR) and the tyrosine kinase B (TrkB) receptor tyrosine kinase. BDNF binding to TrkB triggers its dimerization through conformational changes and autophosphorylation of tyrosine residues in its intracellular domain, resulting in activation of the three major signaling pathways. BDNF protects hippocampal neurons from glutamate toxicity, rescues cerebellar neurons from programmed cell death, reduces ischemic injury, and improves functional recovery and postinjury regeneration. It has been suggested that NGF or BDNF therapy may delay the onset of AD and ameliorate peripheral diabetic neuropathies. Despite the therapeutic potential of NGF and BDNF, clinical trials have been disappointing, which is due to inherent drawbacks associated with the use of polypeptides applied as drugs, in vivo instability, and pleiotropic effects.
Dr. Ye and his colleagues developed the T17, T48, and T62 cell line to screen for NGF-like compounds that would act as TrkA agonists. The cells were stably transfected with rat TrkA or TrkB and cultured in DMEM medium containing 1 mM pyruvate, 10% FBS, and 300 Âµg/ml G418. TrkA or TrkB plasmid was transfected into this SN56 cell and selected by 300 mg/ml G418. TrkA expressing T17 cells were generated and cultured in DMEM medium containing 1 mM pyruvate and 10% FBS and antibiotics.
Publication: Jang et.al. Proc Natl Acad Sci USA. 104:16329-16334. (2007)