Differential Cellular Expression of Neurotrophins in Cortical Tubers of the Tuberous Sclerosis Complex

Abstract

Neurotrophins and their receptors modulate cerebral cortical development. Tubers in the tuberous sclerosis complex (TSC) are characterized histologically by disorganized cortical cytoarchitecture and thus, we hypothesized that expression of neurotrophin mRNAs and proteins might be altered in tubers. Using in situ transcription and mRNA amplification to probe cDNA arrays, we found that neurotrophin-3 (NT3) and trkB mRNA expression were reduced whereas neurotrophin-4 (NT4) and trkC mRNA expression were increased in whole tuber sections. Alterations in mRNA abundance were defined in single microdissected dysplastic neurons (DNs) and giant cells (GCs). NT3 mRNA expression was reduced in GCs and trkB mRNA expression was reduced in DNs. NT4 mRNA expression was increased in DNs and trkC mRNA expression was increased in both DNs and GCs. In three patients, TSC2 locus mutations were confirmed and the mean tuberin mRNA expression levels was reduced across all nine cases. Consistent with these observations, NT3 mRNA expression was reduced but trkC mRNA expression was increased in vitro in human NTera2 neurons ( Tubers in the tuberous sclerosis complex (TSC) are developmental abnormalities of cerebral cortical cytoarchitecture that are associated clinically with epilepsy. [1][2][3] Electrocorticography has shown that tubers are epileptogenic and seizures in TSC patients are often medically intractable despite anticonvulsant polytherapy. 4 -6 TSC is an autosomal-dominant disorder resulting from mutations in one of two genes, TSC1 or TSC2 7,8 although the mechanism by which mutations in either TSC gene leads to tuber formation is unknown. Disorganized cortical lamination and aberrant cellular morphology are important histological features of tubers. Large dysplastic neurons (DNs) and giant cells (GCs) are prominent cell types in tubers. 9 DNs and GCs share select morphological features including cytomegaly, the extension of aberrant processes often of unclear identity, ie, axons versus dendrites, and the expression of neural protein markers such as neurofilament and ␣-internexin. 1,9,10 The TSC2 knockout mouse 11 and the Eker rat strain 12 do not fully model human brain pathology in TSC, and thus, analysis of human tuber specimens provides the only direct avenue to study the mechanisms of tuber formation. One strategy to investigate the molecular pathogenesis of cytoarchitectural disorganization in tubers is to evaluate the expression of candidate genes and proteins in human tuber specimens that are relevant to cortical development. 10 Neurotrophins and their cognate receptors comprise a family of proteins that mediate proliferation, differentiation, migration, and process outgrowth during cortical development, 13,14 and thus, are ideal candidate molecules to investigate in tubers. Brainderived neurotrophic factor (BDNF), nerve growth factor (NGF), neurotrophin 3 (NT3), and neurotrophin 4 (NT4) exert their effects on neurons by binding selectively to a family of neuronal cell membrane receptors, trks A to C. 13,14 NGF signals...

Enhanced Epidermal Growth Factor, Hepatocyte Growth Factor, and Vascular Endothelial Growth Factor Expression in Tuberous Sclerosis Complex

“…15 The entire cortex from both hemispheres was removed from the subcortical structures using a microscalpel at the level of the midhippocampus. Poly(A) mRNA served as a template for in vitro cDNA synthesis with avian myeloblastosis virus reverse transcriptase, and then double-stranded template cDNA was synthesized with T4 DNA polymerase I (BoehringerMannheim) from extracted cDNA.…”

“…15 The ODR is calculated by determining the level of pixel staining density in labeled cells versus the pixel density of the noncellular background (the cell densities are digitally subtracted from the image). An ODR greater than 3 was used as a threshold to define immunopositivity for a given antibody.…”

“…Importantly, these alterations closely predicted enhanced expression of these proteins in tuber and subependymal giant cell astrocytoma (SEGA) specimens in TSC. Expression of EGF, EGFR, HGF, c-Met, and VEGF protein, as well as hypoxia inducible factor-1␣, a transcription factor that regulates VEGF levels and is also modulated by mTOR cascade activity, was enhanced in SEGAs (n ‫؍‬ 6) and tubers (n ‫؍‬ 10) from 15 Tuberous sclerosis complex (TSC) is an autosomal dominant disorder that results from mutations in the TSC1 or TSC2 genes, which encode TSC1 and TSC2 proteins, respectively. 1,2 Many individuals with TSC exhibit cognitive disability and autism, 3 and more than 75% of TSC patients develop seizures.…”

“…Previous studies have suggested that altered growth factor expression may be associated with abnormal cellular architecture in the brains of TSC patients. For example, differential expression of neurotrophins and their receptors has been observed in cortical tubers, 15 and TSC1-TSC2 mediated control of mTOR is modulated by insulin-like growth factor-1 (IGF-1). 16 -18 Recent evidence also suggests that mTOR signaling is regulated by several growth factors, such as epidermal growth factor (EGF) and hepatocyte growth factor (HGF), and that TSC1-TSC2 may regulate downstream expression of select angiogenic factors, such as vascular endothelial growth factor (VEGF), via hypoxia inducible factor-1␣ (HIF-1␣).…”

Expression of the Neural Stem Cell Markers NG2 and L1 in Human Angiomyolipoma: Are Angiomyolipomas Neoplasms of Stem Cells?

“…TS is notable for the development of benign neoplasms of the kidney, lungs, brain, and skin, as well as cortical tubers (2,6). Cortical tubers are giant cells that express neuronal and glial markers (7,8). While seizures remain the leading cause of morbidity and mortality in children, improvements in neurological care are leading to an increased population of adults with TS.…”