The Effects of Nell-1 Deficiency in Cranial Neural Crest Cells
Objectives: Nell-1 is a craniosynostosis associated molecule and an osteoinductive protein. However, the exact role of Nell-1 in development and osteogenic differentiation of cranial neural crest (CNC) cells has not been fully understood due to lacking proper animal model. This study aims to create CNC cell Nell-1 deficient model to further evaluate Nell-1’s role in the pathogenesis of craniofacial anomalies. Methods: To investigate the specific effects of Nell-1 deficiency in CNC cells in attributing to craniofacial anomalies, the Nell-1flox/flox mice were mated with CMV-Cre mice or Wnt1-Cre mice to obtain Nell-1flox/flox;CMV-Cre or Nell-1flox/flox;Wnt1-Cre mice for comprehensive analyses. Genotyping and gene expression profile were performed by qPCR. The MicroCT, histology and immunohistochemisty (IHC) were used for phenotypic evaluation. Primary calvarial cells representing CNC cells from Nell-1flox/flox mice with or without Cre expression were used for cell adhesion, proliferation and osteogenic differentiation assay. Results: The majority of the Nell-1flox/flox;CMV-Cre mice were alive at birth with craniofacial abnormalities manifesting from severe dysplasia of mandible and defects of eye sockets to reduction of skull dimension with calvaria defects. The neonatal lethality happened only with severe cases, which differs from homozygous ENU-induced Nell-1 deficient mice. Significantly, Nell-1flox/flox;Wnt1-Cre mice also exhibited craniofacial anomaly including defect of eye sockets, widen suture/fontanelle and microcephaly when compared to Cre-negative littermates. At cellular level, the Nell-1-knockout calvarial cells have decreased cell adhesion and osteogenic differentiation capacity, but increased cell proliferation activity. With transduction of adenoviral CMV-Cre for Nell-1 knockout at different stages, the calvarial cells showed that the expression of Tnxb, Ocn and Col1 was significantly reduced in all differentiation stage, while expression of Runx2 remained the same compared to normal calvarial cells in early differentiation stage. Conclusions: Collectively, Nell-1 plays an important role in CNC development, and lacking Nell-1 in CNC cells is responsible to craniofacial anomaly.
Division: AADR/CADR Annual Meeting
Meeting:2016 AADR/CADR Annual Meeting (Los Angeles, California) Location: Los Angeles, California
Year: 2016 Final Presentation ID:1299 Abstract Category|Abstract Category(s):Craniofacial Biology
Authors
Yu, Mengliu
( School of Dentistry, University of California, Los Angeles
, Los Angeles
, California
, United States
; Zhejiang University
, Hangzhou
, Zhejiang
, China
)
Tanjaya, Justine
( School of Dentistry, University of California, Los Angeles
, Los Angeles
, California
, United States
)
Li, Chenshuang
( School of Dentistry, University of California, Los Angeles
, Los Angeles
, California
, United States
)
Pan, Hsin Chuan
( School of Dentistry, University of California, Los Angeles
, Los Angeles
, California
, United States
)
Wang, Huiming
( Zhejiang University
, Hangzhou
, Zhejiang
, China
)
Ting, Kang
( School of Dentistry, University of California, Los Angeles
, Los Angeles
, California
, United States
)
Soo, Chia
( University of California - Los Angeles
, Los Angeles
, California
, United States
)
Zhang, Xinli
( School of Dentistry, University of California, Los Angeles
, Los Angeles
, California
, United States
)
Support Funding Agency/Grant Number: This work was supported by the CIRM Early Translational II Research Award TR2-01821, NIH/NIDCR (grants R21 DE0177711 and RO1 DE01607), UC Discovery Grant 07-10677, Eli & Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA Innovatio
Financial Interest Disclosure: Drs. X.Z., K.T., and C.S. are inventors of Nell-1 related patents and K.T. and C.S.
are inventors of perivascular stem cell-related patents filed from UCLA. Drs. X.Z., K.T., and C.S. are founders and/or
board members of Bone Biologics Inc. which sublicens