Screening And Identification Of Nerve Growth-promoting Genes In Neurofibromatosis Type1

Neurofibromatosis l (NFl), known as von Recklinghausen disease, is amain type of Neurofibromatosis which is first reported by Daniel Friedrich vonRecklinghausen in1882. NF1is a common autosome dominant genetic disease,characterized by multiple pigmentations and neurofibroma, accompanied by irishamartoma, osteodysplasty, and dysgnosia. It’s mainly initiated from theabnormal differentiation of neural crest cells, resulting in multi-system diseases.NF1gene is localized in human chromosome17q11.2, with350,000bases,spanning about350genomic DNA, and containing60exons and more than60introns. Known as a tumor suppressor, NF1encodes a protein productneurofibromin, which is a highly-conserved protein, expressed in multipletissues and organs, and functions as a negative regulator of RAS signalingtransduction by stimulating Ras-GTPase through its main functional domainwhich is homologous to GAP and encoded by exon21-27. Meanwhile, it is alsoreported to be an intracellular positive regulator of cAMP. Defective expressionof NF1leads to the inactivation of neurofibromin and abnormal expressions ofdown-stream factors which regulated by Ras/ERK and cAMP/PKA signalingpathways, and play an important role in the regulation of cell proliferation,differentiation and tumorigenesis. The haploid missing of NF1in mouse inducedtumorigenesis. Mutations happened in other genes like p27-kip1, p53and p16contribute to the tumor development as well, but the exact mechanism is still not clear.This study is conducted to explore nerve growth stimulating factors in NF1,provides a new basis for the pathogenesis of NF1and finding new therapeutictargets. We obtained abnormal bulky terminal nerves from three diagnosed NF1patients, analyzed genes’ expression profiles between NF1patients and normalpersons through microarray analysis, and identified nerve growth stimulatingrelative factors from those genes which expression changed more than5foldscompared with controls. Meanwhile we used NGF to induce the differentiationof PC12into neurons-like cells, and identified the transcriptional andtranslational levels of genes which expression changed>10folds by RT-PCRand Western blot. After analyzing the expression of30,000genes, we found132,144and158genes have statistically significant change (P value<0.001) in threeNF1patients with macrodactylia fingers or toes along in the end of the limbsrespectively.93genes expressed in all NF1patients, and48%of them take partin DNA-protein interaction,28%involve in molecular signaling transduction,and18%participate in enzyme catalyzing. In the common expressed93genes,61genes showed a similar trend, and among them,28genes changedremarkably more than5folds compared with controls. More than75%genes inabove28genes showed down-regulation, such as metastasis associated genes,lymphocytic function associated genes (LFA1、CXCR4), intercellular adhesionrelated genes ICAM, inflammatory factors (IL1β、IL6、IL8), cell cycling relatedgenes (CDKN2B, CDK4and PLK1) and apoptosis genes (LMNB1, BIRC3andFOSL1) were all at a low expression, and that may explain the fact that NF1israrely developed to malignancy. In cell experiments, the results indicated that the expression of HOXC8is positively related to the NGF induction at bothtranscriptional and translational levels，and it may be related to nerve growth.This study identified nerve growth stimulating factors in NF1, which laid afoundation for the research of the mechanism of nerve overgrowth, and it’sessential to help us to explore new treatment and targets in the reparation andreconstruction of nerve injury.