Endogenous Expression of Matriptase in Neural Progenitor Cells Promotes Cell Migration and Neuron Differentiation

Sheau-Ling Lee,Hsiu-Hui Tung,Jung-Da Fang,Pao-Yi Huang,Hsiao-Chin Chou

doi:10.1074/jbc.m110.153866

Sheau-Ling Lee, Hsiu-Hui Tung + Show 3 more

Open Access

https://doi.org/10.1074/jbc.m110.153866

Copy DOI

Abstract

Recent studies show that type II transmembrane serine proteases play important roles in diverse cellular activities and pathological processes. Their expression and functions in the central nervous system, however, are largely unexplored. In this study, we show that the expression of one such member, matriptase (MTP), was cell type-restricted and primarily expressed in neural progenitor (NP) cells and neurons. Blocking MTP expression or MTP activity prevented NP cell traverse of reconstituted basement membrane, whereas overexpression of MTP promoted it. The NP cell mobilization induced by either vascular endothelial growth factor or hepatocyte growth factor was also impaired by knocking down MTP expression. MTP acts upstream of matrix metalloproteinase 2 in promoting NP cell mobility. In embryonic stem cell differentiation to neural cells, MTP knockdown had no effect on entry of embryonic stem cells into the neural lineage. High MTP expression or activity, however, shifts the population dynamics from NP cells toward neurons to favor neuronal differentiation. This is the first report to demonstrate the direct involvement of type II transmembrane serine protease in NP cell function.

Highlights

Progenitor (NP) cells in both locations is enhanced after brain injuries, and streams of neuroblasts are attracted by and move toward the injured sites [3,4,5,6,7]
It has been shown that MMP2 and MMP9 expressed by endothelial cells promote neural progenitor (NP) cell migration [18]
MTP Is Expressed in NP Cells, Neurons, and during Neural Lineage Differentiation by Embryonic Stem (ES) Cells—NP cells, both isolated from the SVZ of the adult mouse brain LV (Fig. 1A, aNPC) and differentiated from embryonic stem cells (Fig. 1A, ESC-NPC), expressed MTP mRNA

Summary

EXPERIMENTAL PROCEDURES

Cell Culture Reagents—Glasgow modification of Eagle’s medium, Dulbecco’s modified Eagle’s medium (DMEM)/ F-12 (50/50), neural basal medium, fetal bovine serum (FBS), knock-out serum replacement, glutamine, sodium pyruvate, N2 supplement, B27 supplement, 2-mercaptoethanol, bovine serum albumin fraction V (BSA-V), and Hank’s buffered saline solution were purchased from Invitrogen. Astrocyte Differentiation—NP cells were plated on the polyD-lysine/laminin-coated surface and cultured in N2B27 medium supplemented with 2% FBS and 50 ng/ml of ciliary neurotrophic factor for 10 days. After shaking overnight at 250 rpm in a 37 °C incubator, medium was removed, and the attached primary astrocytes were kept in culture with medium containing 100 ␮M cytosine ␤-D-arabinofuranoside (Sigma) for 3 days. For MTP staining, tissue sections were heated 20 min at 95 °C in 10 mM citrate (pH 6.0) for antigen retrieval, incubated with sheep primary antibody, with rabbit anti-sheep IgG before incubation with the peroxidase-labeled polymer-conjugated anti-rabbit antibody. The collected cell spheroids were either cultured directly with freshly prepared neuronal differentiation medium or transfected with siRNA or DNA plasmid using Lipofectamine 2000 (see section). Western Blot—Protein extracts were prepared from brain tissues and cultured cells as described previously [21, 22].

RESULTS

NP cell culturea

DISCUSSION