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The emerging role of NG2 in pediatric diffuse intrinsic pontine glioma.

Yadavilli S, Scafidi J, Becher OJ, Saratsis AM, Hiner RL, Kambhampati M, Mariarita S, MacDonald TJ, Codispoti KE, Magge SN, Jaiswal JK, Packer RJ, Nazarian J - Oncotarget (2015)

Bottom Line: We further demonstrate that microRNA 129-2 (miR129-2) is downregulated and hypermethylated in human DIPGs, resulting in the increased expression of NG2.NG2 knockdown retards cellular migration in vitro, while NG2 expressing neurospheres are highly tumorigenic in vivo, resulting in rapid growth of pontine tumors.This data implicates NG2 as a molecule of interest in DIPGs especially those with H3.3 mutation.

View Article: PubMed Central - PubMed

Affiliation: Research Center for Genetic Medicine, Children's National Health System, Washington, DC, USA.

ABSTRACT
Diffuse intrinsic pontine gliomas (DIPGs) have a dismal prognosis and are poorly understood brain cancers. Receptor tyrosine kinases stabilized by neuron-glial antigen 2 (NG2) protein are known to induce gliomagenesis. Here, we investigated NG2 expression in a cohort of DIPG specimens (n= 50). We demonstrate NG2 expression in the majority of DIPG specimens tested and determine that tumors harboring histone 3.3 mutation express the highest NG2 levels. We further demonstrate that microRNA 129-2 (miR129-2) is downregulated and hypermethylated in human DIPGs, resulting in the increased expression of NG2. Treatment with 5-Azacytidine, a methyltransferase inhibitor, results in NG2 downregulation in DIPG primary tumor cells in vitro. NG2 expression is altered (symmetric segregation) in mitotic human DIPG and mouse tumor cells. These mitotic cells co-express oligodendrocyte (Olig2) and astrocyte (glial fibrillary acidic protein, GFAP) markers, indicating lack of terminal differentiation. NG2 knockdown retards cellular migration in vitro, while NG2 expressing neurospheres are highly tumorigenic in vivo, resulting in rapid growth of pontine tumors. NG2 expression is targetable in vivo using miR129-2 indicating a potential avenue for therapeutic interventions. This data implicates NG2 as a molecule of interest in DIPGs especially those with H3.3 mutation.

No MeSH data available.


Related in: MedlinePlus

NG2 overexpression in PDGFB and PDGFB/H3.3 K27M/p53−/− mouse models(a) In mouse, expression of NG2 in PDGFB mouse was limited to brainstem tumor (dotted area). (b) Sporadic expression of NG2 across brainstem and cerebellum of normal mouse. (c) Injection of NSG SCID mouse with mouse DIPG cells that are H3.3.K27 mutant and have PDGFB overexpression and p53 deletion resulted in overexpression of NG2 (dotted area). (d) In the adjacent normal brain tissue of NSG SCID mouse injected with PDGFB/H3.3.K27M/p53−/− cells, NG2 overexpression was not detected. Insets are 40 × magnifications of the corresponding panel. Scale bar: 200 μM.
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Figure 2: NG2 overexpression in PDGFB and PDGFB/H3.3 K27M/p53−/− mouse models(a) In mouse, expression of NG2 in PDGFB mouse was limited to brainstem tumor (dotted area). (b) Sporadic expression of NG2 across brainstem and cerebellum of normal mouse. (c) Injection of NSG SCID mouse with mouse DIPG cells that are H3.3.K27 mutant and have PDGFB overexpression and p53 deletion resulted in overexpression of NG2 (dotted area). (d) In the adjacent normal brain tissue of NSG SCID mouse injected with PDGFB/H3.3.K27M/p53−/− cells, NG2 overexpression was not detected. Insets are 40 × magnifications of the corresponding panel. Scale bar: 200 μM.

Mentions: To test NG2 expression in preclinical DIPG murine models, we used IHC to assess NG2 expression in the PDGFB-Ink4a-ARF−/− mouse (PDGFB model) [16] as well as mouse injected with glioma cells derived from PDGFB/p53−/−/H3.3K27M model [17]. We show significant upregulation of NG2 in both PDGFB (Figure 2a) and PDGFB/p53−/−/H3.3K27M (Figure 2c) DIPG mouse models. Low NG2 expression was detected in normal adjacent brainstem regions as well as the cerebellum (Figure 2b and 2d). High level (148 fold, p < 0.05) of NG2 was also detected by Western blot assays using tumor (n = 3) and normal (n = 3) brainstem specimens obtained from PDGFB mouse model or healthy controls (Supplementary Figure 2a). Furthermore, we tested NG2 expression in three primary human DIPG lines (SF8628, SUDIPGIV and SUDIPGVI) (Supplementary Figure 2b). All three primary cells showed NG2 expression as assessed by Western blotting assays.


The emerging role of NG2 in pediatric diffuse intrinsic pontine glioma.

Yadavilli S, Scafidi J, Becher OJ, Saratsis AM, Hiner RL, Kambhampati M, Mariarita S, MacDonald TJ, Codispoti KE, Magge SN, Jaiswal JK, Packer RJ, Nazarian J - Oncotarget (2015)

NG2 overexpression in PDGFB and PDGFB/H3.3 K27M/p53−/− mouse models(a) In mouse, expression of NG2 in PDGFB mouse was limited to brainstem tumor (dotted area). (b) Sporadic expression of NG2 across brainstem and cerebellum of normal mouse. (c) Injection of NSG SCID mouse with mouse DIPG cells that are H3.3.K27 mutant and have PDGFB overexpression and p53 deletion resulted in overexpression of NG2 (dotted area). (d) In the adjacent normal brain tissue of NSG SCID mouse injected with PDGFB/H3.3.K27M/p53−/− cells, NG2 overexpression was not detected. Insets are 40 × magnifications of the corresponding panel. Scale bar: 200 μM.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4494928&req=5

Figure 2: NG2 overexpression in PDGFB and PDGFB/H3.3 K27M/p53−/− mouse models(a) In mouse, expression of NG2 in PDGFB mouse was limited to brainstem tumor (dotted area). (b) Sporadic expression of NG2 across brainstem and cerebellum of normal mouse. (c) Injection of NSG SCID mouse with mouse DIPG cells that are H3.3.K27 mutant and have PDGFB overexpression and p53 deletion resulted in overexpression of NG2 (dotted area). (d) In the adjacent normal brain tissue of NSG SCID mouse injected with PDGFB/H3.3.K27M/p53−/− cells, NG2 overexpression was not detected. Insets are 40 × magnifications of the corresponding panel. Scale bar: 200 μM.
Mentions: To test NG2 expression in preclinical DIPG murine models, we used IHC to assess NG2 expression in the PDGFB-Ink4a-ARF−/− mouse (PDGFB model) [16] as well as mouse injected with glioma cells derived from PDGFB/p53−/−/H3.3K27M model [17]. We show significant upregulation of NG2 in both PDGFB (Figure 2a) and PDGFB/p53−/−/H3.3K27M (Figure 2c) DIPG mouse models. Low NG2 expression was detected in normal adjacent brainstem regions as well as the cerebellum (Figure 2b and 2d). High level (148 fold, p < 0.05) of NG2 was also detected by Western blot assays using tumor (n = 3) and normal (n = 3) brainstem specimens obtained from PDGFB mouse model or healthy controls (Supplementary Figure 2a). Furthermore, we tested NG2 expression in three primary human DIPG lines (SF8628, SUDIPGIV and SUDIPGVI) (Supplementary Figure 2b). All three primary cells showed NG2 expression as assessed by Western blotting assays.

Bottom Line: We further demonstrate that microRNA 129-2 (miR129-2) is downregulated and hypermethylated in human DIPGs, resulting in the increased expression of NG2.NG2 knockdown retards cellular migration in vitro, while NG2 expressing neurospheres are highly tumorigenic in vivo, resulting in rapid growth of pontine tumors.This data implicates NG2 as a molecule of interest in DIPGs especially those with H3.3 mutation.

View Article: PubMed Central - PubMed

Affiliation: Research Center for Genetic Medicine, Children's National Health System, Washington, DC, USA.

ABSTRACT
Diffuse intrinsic pontine gliomas (DIPGs) have a dismal prognosis and are poorly understood brain cancers. Receptor tyrosine kinases stabilized by neuron-glial antigen 2 (NG2) protein are known to induce gliomagenesis. Here, we investigated NG2 expression in a cohort of DIPG specimens (n= 50). We demonstrate NG2 expression in the majority of DIPG specimens tested and determine that tumors harboring histone 3.3 mutation express the highest NG2 levels. We further demonstrate that microRNA 129-2 (miR129-2) is downregulated and hypermethylated in human DIPGs, resulting in the increased expression of NG2. Treatment with 5-Azacytidine, a methyltransferase inhibitor, results in NG2 downregulation in DIPG primary tumor cells in vitro. NG2 expression is altered (symmetric segregation) in mitotic human DIPG and mouse tumor cells. These mitotic cells co-express oligodendrocyte (Olig2) and astrocyte (glial fibrillary acidic protein, GFAP) markers, indicating lack of terminal differentiation. NG2 knockdown retards cellular migration in vitro, while NG2 expressing neurospheres are highly tumorigenic in vivo, resulting in rapid growth of pontine tumors. NG2 expression is targetable in vivo using miR129-2 indicating a potential avenue for therapeutic interventions. This data implicates NG2 as a molecule of interest in DIPGs especially those with H3.3 mutation.

No MeSH data available.


Related in: MedlinePlus