Zhi-Cheng Xiao

F3/Contactin Acts as a Functional Ligand for Notch during Oligodendrocyte Maturation

Axon-derived molecules are temporally and spatially required as positive or negative signals to coordinate oligodendrocyte differentiation. Increasing evidence suggests that, in addition to the inhibitory Jagged1/Notch1 signaling cascade, other pathways act via Notch to mediate oligodendrocyte differentiation. The GPI-linked neural cell recognition molecule F3/contactin is clustered during development at the paranodal region, a vital site for axoglial interaction. Here, we show that F3/contactin acts as a functional ligand of Notch. This trans-extracellular interaction triggers gamma-secretase-dependent nuclear translocation of the Notch intracellular domain. F3/Notch signaling promotes oligodendrocyte precursor cell differentiation and upregulates the myelin-related protein MAG in OLN-93 cells. This can be blocked by dominant negative Notch1, Notch2, and two Deltex1 mutants lacking the RING-H2 finger motif, but not by dominant-negative RBP-J or Hes1 antisense oligonucleotides. Expression of constitutively active Notch1 or Notch2 does not upregulate MAG. Thus, F3/contactin specifically initiates a Notch/Deltex1 signaling pathway that promotes oligodendrocyte maturation and myelination.

A TAG1-APP signalling pathway through Fe65 negatively modulates neurogenesis

The release of amyloid precursor protein (APP) intracellular domain (AICD) may be triggered by extracellular cues through γ-secretase-dependent cleavage. AICD binds to Fe65, which may have a role in AICD-dependent signalling; however, the functional ligand has not been characterized. In this study, we have identified TAG1 as a functional ligand of APP. We found that, through an extracellular interaction with APP, TAG1 increased AICD release and triggered Fe65-dependent activity in a γ-secretase-dependent manner. TAG1, APP and Fe65 colocalized in the neural stem cell niche of the fetal ventricular zone. Neural precursor cells from TAG1−/−, APP−/− and TAG1−/−;APP−/− mice had aberrantly enhanced neurogenesis, which was significantly reversed in TAG1−/− mice by TAG1 or AICD but not by AICD mutated at the Fe65 binding site. Notably, TAG1 reduced normal neurogenesis in Fe65+/+ mice. Abnormally enhanced neurogenesis also occurred in Fe65−/− mice but could not be reversed by TAG1. These results describe a TAG1–APP signalling pathway that negatively modulates neurogenesis through Fe65.

Fetal Microchimerism in the Maternal Mouse Brain: A Novel Population of Fetal Progenitor or Stem Cells Able to Cross the Blood–Brain Barrier?

We investigated whether fetal cells can enter the maternal brain during pregnancy. Female wild‐type C57BL/6 mice were crossed with transgenic Green Mice ubiquitously expressing enhanced green fluorescent protein (EGFP). Green Mouse fetal cells were found in the maternal brain. Quantitative real‐time polymerase chain reaction (PCR) of genomic DNA for the EGFP gene showed that more fetal cells were present in the maternal brain 4 weeks postpartum than on the day of parturition. After an excitotoxic lesion to the brain, more fetal cells were detected in the injured region. The presence of fetal cells in the maternal brain was also confirmed by quantitative real‐time PCR for the sex‐determining region of the Y chromosome. Four weeks postpartum, EGFP‐positive Green Mouse fetal cells in the maternal brain were found to adopt locations, morphologies, and expression of immunocytochemical markers indicative of perivascular macrophage‐, neuron‐, astrocyte‐, and oligodendrocyte‐like cell types. Expression of morphological and immunocytochemical characteristics of neuron‐ and astrocyte‐like cell types was confirmed on identification of fetal cells in maternal brain by Y chromosome fluorescence in situ hybridization. Although further studies are required to determine whether such engraftment of the maternal brain has any physiological or pathophysiological functional significance, fetomaternal microchimerism provides a novel model for the experimental investigation of the properties of fetal progenitor or stem cells in the brain without prior in vitro manipulation. Characterization of the properties of these cells that allow them to cross both the placental and blood–brain barriers and to target injured brain may improve selection procedures for isolation of progenitor or stem cells for brain repair by intravenous infusion.

Nogo‐A at CNS paranodes is a ligand of Caspr: possible regulation of K+ channel localization

We report Nogo‐A as an oligodendroglial component congregating and interacting with the Caspr–F3 complex at paranodes. However, its receptor Nogo‐66 receptor (NgR) does not segregate to specific axonal domains. CHO cells cotransfected with Caspr and F3, but not with F3 alone, bound specifically to substrates coated with Nogo‐66 peptide and GST–Nogo‐66. Binding persisted even after phosphatidylinositol‐ specific phospholipase C (PI‐PLC) removal of GPI‐linked F3 from the cell surface, suggesting a direct interaction between Nogo‐66 and Caspr. Both Nogo‐A and Caspr co‐immunoprecipitated with Kv1.1 and Kv1.2, and the developmental expression pattern of both paralleled compared with Kv1.1, implicating a transient interaction between Nogo‐A–Caspr and K+ channels at early stages of myelination. In pathological models that display paranodal junctional defects (EAE rats, and Shiverer and CGT−/− mice), distances between the paired labeling of K+ channels were shortened significantly and their localization shifted toward paranodes, while paranodal Nogo‐A congregation was markedly reduced. Our results demonstrate that Nogo‐A interacts in trans with axonal Caspr at CNS paranodes, an interaction that may have a role in modulating axon–glial junction architecture and possibly K+‐channel localization during development.

Optic nerve regeneration in polyglycolic acid-chitosan conduits coated with recombinant L1-Fc.

Autografts have been extensively studied to facilitate optic nerve (ON) regeneration in animal experiments, but the clinical application of this approach to aid autoregeneration has not yet been attempted. This study aims to explore the guided regeneration by an artificial polyglycolic acid–chitosan conduit coated with recombinant L1-Fc. Consistent with previous studies; in vitro assay showed that both chitosan, a natural biomaterial, and the neural cell adhesion molecule L1-Fc enhanced neurite outgrowth. Rat optic nerve transection was used as an in vivo model. The implanted PGA-chitosan conduit was progressively degraded and absorbed, accompanied by significant axonal regeneration as revealed by immunohistochemistry, anterograde and retrograde tracing. The polyglycolic acid–chitosan conduit coated with L1-Fc showed more effective to promote axonal regeneration and remyelination. Taken together, our observations demonstrated that the L1-Fc coated PGA–chitosan conduits provided a compatible and supportive canal to guild the injured nerve regeneration and remyelination.

Recombinant DNA vaccine encoding multiple domains related to inhibition of neurite outgrowth: a potential strategy for axonal regeneration

Myelin‐derived proteins, such as tenascin‐R (TN‐R), myelin associate glycoprotein (MAG), and Nogo‐A, inhibit the CNS regeneration. By targeting specifically the inhibitory epitopes, we have investigated whether vaccination with a recombinant DNA molecule encoding multiple domains of myelin inhibitors may be useful in CNS repair. We show here that the recombinant DNA vaccine is able to activate the immune system but does not induce experimental autoimmune encephalomyelitis (EAE) in Lewis rats. Importantly, it promotes axonal regeneration in a spinal cord injury model. Thus, the application of DNA vaccine, encoding multiple specific domains of major inhibitory proteins and/or their receptors, provides another promising approach to overcome the inhibitory barriers during CNS regeneration.

Protein-tyrosine Phosphatase α Acts as an Upstream Regulator of Fyn Signaling to Promote Oligodendrocyte Differentiation and Myelination

The tyrosine kinase Fyn plays a key role in oligodendrocyte differentiation and myelination in the central nervous system, but the molecules responsible for regulating Fyn activation in these processes remain poorly defined. Here we show that receptor-like protein-tyrosine phosphatase α (PTPα) is an important positive regulator of Fyn activation and signaling that is required for the differentiation of oligodendrocyte progenitor cells (OPCs). PTPα is expressed in OPCs and is up-regulated during differentiation. We used two model systems to investigate the role of PTPα in OPC differentiation: the rat CG4 cell line where PTPα expression was silenced by small interfering RNA, and oligosphere-derived primary OPCs isolated from wild-type and PTPα-null mouse embryos. In both cell systems, the ablation of PTPα inhibited differentiation and morphological changes that accompany this process. Although Fyn was activated upon induction of differentiation, the level of activation was severely reduced in cells lacking PTPα, as was the activation of Fyn effector molecules focal adhesion kinase, Rac1, and Cdc42, and inactivation of Rho. Interestingly, another downstream effector of Fyn, p190RhoGAP, which is responsible for Rho inactivation during differentiation, was not affected by PTPα ablation. In vivo studies revealed defective myelination in the PTPα−/− mouse brain. Together, our findings demonstrate that PTPα is a critical regulator of Fyn activation and of specific Fyn signaling events during differentiation, and is essential for promoting OPC differentiation and central nervous system myelination.

Nogo‐66 and myelin‐associated glycoprotein (MAG) inhibit the adhesion and migration of Nogo‐66 receptor expressing human glioma cells

Malignant gliomas are common and aggressive brain tumours associated with significant morbidity and mortality. We showed in this report that substratum adherence and migration by human U87MG glioma cells in culture were significantly attenuated by the extracellular domains of Nogo‐A (Nogo‐66) and the myelin‐associated glycoprotein (MAG). U87MG cells contained significant amounts of endogenous Nogo‐66 receptor (NgR), and treatment of the cells with phosphatidylinositol‐specific phospholipase C (PI‐PLC) or NgR antibodies resulted in an increase in their ability to adhere to, or migrate through, Nogo‐66‐ and MAG‐coated substrates. Nogo‐66 and MAG may therefore modulate glioma growth and migration by acting through the NgR, a phenomenon that has potential therapeutic implications.

Oligodendrocytes regulate formation of nodes of Ranvier via the recognition molecule OMgp

The molecular mechanisms underlying the involvement of oligodendrocytes in formation of the nodes of Ranvier (NORs) remain poorly understood. Here we show that oligodendrocyte-myelin glycoprotein (OMgp) aggregates specifically at NORs. Nodal location of OMgp does not occur along demyelinated axons of either Shiverer or proteolipid protein (PLP) transgenic mice. Over-expression of OMgp in OLN-93 cells facilitates process outgrowth. In transgenic mice in which expression of OMgp is down-regulated, myelin thickness declines, and lateral oligodendrocyte loops at the node-paranode junction are less compacted and even join together with the opposite loops, which leads to shortened nodal gaps. Notably, each of these structural abnormalities plus modest down-regulation of expression of Na(+) channel alpha subunit result in reduced conduction velocity in the spinal cords of the mutant mice. Thus, OMgp that is derived from glia has distinct roles in regulating nodal formation and function during CNS myelination.

Cell Surface Sialylation and Fucosylation Are Regulated by L1 via Phospholipase Cγ and Cooperate to Modulate Neurite Outgrowth, Cell Survival and Migration

Background Cell surface glycosylation patterns are markers of cell type and status. However, the mechanisms regulating surface glycosylation patterns remain unknown. Methodology/Principal Findings Using a panel of carbohydrate surface markers, we have shown that cell surface sialylation and fucosylation were downregulated in L1−/y neurons versus L1+/y neurons. Consistently, mRNA levels of sialyltransferase ST6Gal1, and fucosyltransferase FUT9 were significantly reduced in L1−/y neurons. Moreover, treatment of L1+/y neurons with L1 antibodies, triggering signal transduction downstream of L1, led to an increase in cell surface sialylation and fucosylation compared to rat IgG-treated cells. ShRNAs for both ST6Gal1 and FUT9 blocked L1 antibody-mediated enhancement of neurite outgrowth, cell survival and migration. A phospholipase Cγ (PLCγ) inhibitor and shRNA, as well as an Erk inhibitor, reduced ST6Gal1 and FUT9 mRNA levels and inhibited effects of L1 on neurite outgrowth and cell survival. Conclusions Neuronal surface sialylation and fucosylation are regulated via PLCγ by L1, modulating neurite outgrowth, cell survival and migration.