Nell-1 Is a Key Functional Modulator in Osteochondrogenesis and Beyond

Abstract

Neural EGFL-like 1 (Nell-1) is a well-studied osteogenic factor that has comparable osteogenic potency with the Food and Drug Administration-approved bone morphogenic protein 2 (BMP-2). In this review, which aims to summarize the advanced Nell-1 research in the past 10 y, we start with the correlation of structural and functional relevance of the Nell-1 protein with the identification of a specific receptor of Nell-1, contactin-associated protein-like 4 (Cntnap4), for osteogenesis. The indispensable role of Nell-1 in normal craniofacial and appendicular skeletal development and growth was also defined by using the newly developed tissue-specific Nell-1 knockout mouse lines in addition to the existing transgenic mouse models. With the achievements on Nell-1's osteogenic therapeutic evaluations from multiple preclinical animal models for local and systemic bone regeneration, the synergistic effect of Nell-1 with BMP-2 on osteogenesis, as well as the advantages of Nell-1 as an osteogenic protein with antiadipogenic, anti-inflammatory, and provascularized characteristics over BMP-2 in bone tissue engineering, is highlighted, which lays the groundwork for the clinical trial approval of Nell-1. At the molecular level, besides the mitogen-activated protein kinase (MAPK) signaling pathway, we emphasize the significant involvement of the Wnt/β-catenin pathway as well as the key regulatory molecules Runt-related transcription factor 2 (Runx2) in Nell-1-induced osteogenesis. In addition, the involvement of Nell-1 in chondrogenesis and its relevant pathologies have been revealed with the participation of the nuclear factor of activated T cells 1 (Nfatc1), Runx3, and Indian hedgehog (Ihh) signaling pathways, although the mechanistic insights of Nell-1's osteochondrogenic property will be continuously evolving. With this perspective, we elucidate some emerging and novel functional properties of Nell-1 in oral-dental and neural tissues that will be the frontiers of future Nell-1 studies beyond the context of bone and cartilage. As such, the therapeutic potential of Nell-1 continues to evolve and grow with continuous pursuit.

Keywords: bone regeneration; bone remodeling; cartilage; chondrocytes; osteogenesis; osteoporosis.

Figure 1.

The domains of Nell-1 and Nell-1’s isoforms as well as the known functions of Nell-1. (A) Nell-1 has a length of 810 amino acids and is composed by a secretion signal peptide, an NH2-terminal von Willebrand factor C (vWC) domain, 4 vWC domains, and 6 epidermal growth factor–like (EGF) domains. Nell-1 short isoform, also known as Nell-1 570, is an N-terminal-truncated Nell-1 isoform. Nell-1-ΔE is lacking 1 calcium-binding EGF-like domain. The molecular weight of the full-length Nell-1 and its isoforms before and after posttranslational modifications (PTMs) based on previous publications are stated in the figure. The LamG domain of Nell-1 can bind to apoptosis-related protein 3 (APR3), heparin, and contactin-associated protein-like 4 (Cntnap4). The CC region is in charge of the formation of homo-oligomers. The EGF domains can bind to protein kinase C βI (PKCβI), while the second and third EGF domains can also bind to roundabout 2 (Robo2). The last 2 vWC domains can bind to integrin α3β1. (B) The current known functions of Nell-1.

Figure 2.

Homopentamer might be the functional format of Nell-1. The predicted 3-dimensional structure of Nell-1 as a homodimer (A), homotrimer (B), and homopentamer (C) and that of the Nell-1 pentamer and Cntnap4 complex (D) were predicted by using the PATCHDOCK server (http://bioinfo3d.cs.tau.ac.il/PatchDock/php.php) and I-TASSER server (https://zhanglab.ccmb.med.umich.edu/I-TASSER/).

Figure 3.

The function of Nell-1 in osteogenesis, chondrogenesis, and adipogenesis. During the osteogenic differentiation of the mesenchymal stem cells, Nell-1 promotes the differentiation of osteoprogenitor cells to preosteoblasts and osteoblasts. Nell-1 and Runx2 regulate each other, and Nell-1 mainly actives canonical Wnt and MAPK signaling. During the chondrogenic differentiation of the mesenchymal stem cells, Nell-1 promotes the chondrocytes’ formation and maturation through the Runx2-Nell-1-Nfatc1-Runx3-Ihh signaling cascades. Nell-1 inhibits the adipogenesis by inhibition of the major adipogenic transcription factor peroxisome proliferator-activated receptor γ (PPARγ), which is a Hedgehog-dependent effect in our study.

Figure 4.

The mechanism of Nell-1 in osteogenesis. Nell-1 could bind to Cntnap4 and integrin α3β1 on the surface of osteogenic linear cell, while Cntnap4 serves as the specific functional receptor-like protein of Nell-1. Nell-1 could activate the canonical Wnt, ERK1/2, and JNK MAPK signaling, as well as HH signaling pathways during the osteogenesis. Cntnap4 is known to be participate in Nell-1’s activation via the Wnt and MAPK signaling pathways. Nell-1 could promote the phosphorylation of Runx2, while Runx2 regulates the expression of Nell-1 directly by binding to the OSE2 sites of Nell-1 promoter.