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Rat Nasal Respiratory Mucosa-Derived Ectomesenchymal Stem Cells Differentiate into Schwann-Like Cells Promoting the Differentiation of PC12 Cells and Forming Myelin In Vitro.

Zhang J, Gao X, Zou H, Liu J, Zhang Z - Stem Cells Int (2015)

Bottom Line: When cocultured with dREMSCs for 5 days, PC12 cells differentiated into mature neuron-like cells with long neurites.More importantly, dREMSCs could form myelin structures with the neurites of PC12 cells at 21 days in vitro.Our data indicated that REMSCs, a kind of EMSCs, could differentiate into SC-like cells and have the ability to promote the differentiation of PC12 cells and form myelin in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopedics, The Third Affiliated Hospital of Suzhou University, Changzhou 213003, China.

ABSTRACT
Schwann cell (SC) transplantation as a cell-based therapy can enhance peripheral and central nerve repair experimentally, but it is limited by the donor site morbidity for clinical application. We investigated weather respiratory mucosa stem cells (REMSCs), a kind of ectomesenchymal stem cells (EMSCs), isolated from rat nasal septum can differentiate into functional Schwann-like cells (SC-like cells). REMSCs proliferated quickly in vitro and expressed the neural crest markers (nestin, vimentin, SOX10, and CD44). Treated with a mixture of glial growth factors for 7 days, REMSCs differentiated into SC-like cells. The differentiated REMSCs (dREMSCs) exhibited a spindle-like morphology similar to SC cells. Immunocytochemical staining and Western blotting indicated that SC-like cells expressed the glial markers (GFAP, S100β, Galc, and P75) and CNPase. When cocultured with dREMSCs for 5 days, PC12 cells differentiated into mature neuron-like cells with long neurites. More importantly, dREMSCs could form myelin structures with the neurites of PC12 cells at 21 days in vitro. Our data indicated that REMSCs, a kind of EMSCs, could differentiate into SC-like cells and have the ability to promote the differentiation of PC12 cells and form myelin in vitro.

No MeSH data available.


Related in: MedlinePlus

Morphology of REMSCs, SC-like cells, and PC12 cells cocultured with REMSCs or SC-like cells was observed under SEM. REMSCs cultured in DF12 medium (a) and SC-like cells cultured in SCDM (b), PC12 cells (marked with red) cocultured with REMSCs in DF12 medium (c1, c2), and PC12 cells (marked with red) cocultured with SC-like cells in SCDM (d1, d2). The imaging showed that the neurites of PC12 cells in (d1, d2) were longer than that in (c1, c2), and the neurites grew along with SC-like cells. Bar: 20 μm for all pictures.
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fig8: Morphology of REMSCs, SC-like cells, and PC12 cells cocultured with REMSCs or SC-like cells was observed under SEM. REMSCs cultured in DF12 medium (a) and SC-like cells cultured in SCDM (b), PC12 cells (marked with red) cocultured with REMSCs in DF12 medium (c1, c2), and PC12 cells (marked with red) cocultured with SC-like cells in SCDM (d1, d2). The imaging showed that the neurites of PC12 cells in (d1, d2) were longer than that in (c1, c2), and the neurites grew along with SC-like cells. Bar: 20 μm for all pictures.

Mentions: To detect the ability of SC-like cells to induce the differentiation of PC12 cells and form myelin structures with the neurites of PC12 cells, we cocultured SC-like cells with PC12 cells which are neuron-like cells. PC12 cells were labeled with CM-Dil (Red); SC-like cells or REMSCs were infected with GFP virus (Green). PC12 cells in group A (PC12 cells cultured alone) and group B (PC12 cells cultured in SCDM) were round red and have few neurites (Figures 5(a) and 5(b)). In group C (PC12 cells and REMSCs were cocultured in DF12 medium), short neurites could be detected (Figure 5(c)), while, in group D (PC12 cells and SC-like cells were cocultured in SCDM), PC12 cells grew with long neurites (Figure 5(d)). Compared with group A (3.2 ± 0.4%), the percentage of positive neurite-bearing cells was significantly increased to 38.0 ± 2.3% (P < 0.01) and 57.9 ± 2.6% (P < 0.01), respectively, in group C and group D, but group B (3 ± 0.3%) (P > 0.05) had no significant difference (Figure 5(e)). Also, the percentage of positive neurite-bearing cells in group D significantly increased compared with group C (P < 0.01) (Figure 5(e)). Likewise, compared with group A (10.1 ± 0.5 μm), the length of the longest neurite significantly increased to 72 ± 3.7 μm (P < 0.01) and 223 ± 7.5 μm (P < 0.01), respectively, in group C and group D (Figure 5(f)). There was no significant difference between group A and group B (9.7 ± 0.4 μm) (Figure 5(f)). To further investigate the differentiation of PC12 cells, Western blotting was used to examine the expression levels of NF-H, Synapsin II, GAP-43, and PSD-95 in PC 12 cells. As shown in Figure 6(a), compared with group A, the level of GAP-43, NF-H, Synapsin II, and PSD-95 significantly (P < 0.01) increased in group C and group D. There was significant difference between group C and group D in NF-H, Synapsin II, PSD-95, and GAP-43 (P < 0.05). Immunofluorescent staining showed that differentiated PC12 cells in group D expressed NF-H (35.3 ± 0.42%) and PSD-95 (56.7 ± 0.47%) (Figures 6(b) and 6(c)). The level of NGF was examined as well. SC-like cells were strongly positive for NGF expression as compared with REMSCs (Figure 7). To assess the myelination capacity of SC-like cells, PC12 cells were cultured with SC-like cells or REMSCs. Consisting with the morphology observed under phase contrast microscopy, observation under scanning electron microscopy (SEM) also shows that SC-like cells were bipolar and spindle-like shaped (Figure 8(b)). When PC12 cells were cocultured with REMSCs the neurites were shorter (Figures 8(c1) and 8(c2)) and, in contrast, when cocultured with SC-like cells for 7 days, the neurites of PC12 cells were longer and grew along with SC-like cells (Figures 8(d1) and 8(d2)). After 21 days of coculture, transmission electron microscopy (TEM) showed that SC-like cells could form myelin sheath with neurites (Figure 9(c)). On the contrary, REMSCs could not form myelin structures with PC12 neurites (Figure 9(b)), and REMSCs could not form myelin structures without neurites (Figure 9(a)).


Rat Nasal Respiratory Mucosa-Derived Ectomesenchymal Stem Cells Differentiate into Schwann-Like Cells Promoting the Differentiation of PC12 Cells and Forming Myelin In Vitro.

Zhang J, Gao X, Zou H, Liu J, Zhang Z - Stem Cells Int (2015)

Morphology of REMSCs, SC-like cells, and PC12 cells cocultured with REMSCs or SC-like cells was observed under SEM. REMSCs cultured in DF12 medium (a) and SC-like cells cultured in SCDM (b), PC12 cells (marked with red) cocultured with REMSCs in DF12 medium (c1, c2), and PC12 cells (marked with red) cocultured with SC-like cells in SCDM (d1, d2). The imaging showed that the neurites of PC12 cells in (d1, d2) were longer than that in (c1, c2), and the neurites grew along with SC-like cells. Bar: 20 μm for all pictures.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig8: Morphology of REMSCs, SC-like cells, and PC12 cells cocultured with REMSCs or SC-like cells was observed under SEM. REMSCs cultured in DF12 medium (a) and SC-like cells cultured in SCDM (b), PC12 cells (marked with red) cocultured with REMSCs in DF12 medium (c1, c2), and PC12 cells (marked with red) cocultured with SC-like cells in SCDM (d1, d2). The imaging showed that the neurites of PC12 cells in (d1, d2) were longer than that in (c1, c2), and the neurites grew along with SC-like cells. Bar: 20 μm for all pictures.
Mentions: To detect the ability of SC-like cells to induce the differentiation of PC12 cells and form myelin structures with the neurites of PC12 cells, we cocultured SC-like cells with PC12 cells which are neuron-like cells. PC12 cells were labeled with CM-Dil (Red); SC-like cells or REMSCs were infected with GFP virus (Green). PC12 cells in group A (PC12 cells cultured alone) and group B (PC12 cells cultured in SCDM) were round red and have few neurites (Figures 5(a) and 5(b)). In group C (PC12 cells and REMSCs were cocultured in DF12 medium), short neurites could be detected (Figure 5(c)), while, in group D (PC12 cells and SC-like cells were cocultured in SCDM), PC12 cells grew with long neurites (Figure 5(d)). Compared with group A (3.2 ± 0.4%), the percentage of positive neurite-bearing cells was significantly increased to 38.0 ± 2.3% (P < 0.01) and 57.9 ± 2.6% (P < 0.01), respectively, in group C and group D, but group B (3 ± 0.3%) (P > 0.05) had no significant difference (Figure 5(e)). Also, the percentage of positive neurite-bearing cells in group D significantly increased compared with group C (P < 0.01) (Figure 5(e)). Likewise, compared with group A (10.1 ± 0.5 μm), the length of the longest neurite significantly increased to 72 ± 3.7 μm (P < 0.01) and 223 ± 7.5 μm (P < 0.01), respectively, in group C and group D (Figure 5(f)). There was no significant difference between group A and group B (9.7 ± 0.4 μm) (Figure 5(f)). To further investigate the differentiation of PC12 cells, Western blotting was used to examine the expression levels of NF-H, Synapsin II, GAP-43, and PSD-95 in PC 12 cells. As shown in Figure 6(a), compared with group A, the level of GAP-43, NF-H, Synapsin II, and PSD-95 significantly (P < 0.01) increased in group C and group D. There was significant difference between group C and group D in NF-H, Synapsin II, PSD-95, and GAP-43 (P < 0.05). Immunofluorescent staining showed that differentiated PC12 cells in group D expressed NF-H (35.3 ± 0.42%) and PSD-95 (56.7 ± 0.47%) (Figures 6(b) and 6(c)). The level of NGF was examined as well. SC-like cells were strongly positive for NGF expression as compared with REMSCs (Figure 7). To assess the myelination capacity of SC-like cells, PC12 cells were cultured with SC-like cells or REMSCs. Consisting with the morphology observed under phase contrast microscopy, observation under scanning electron microscopy (SEM) also shows that SC-like cells were bipolar and spindle-like shaped (Figure 8(b)). When PC12 cells were cocultured with REMSCs the neurites were shorter (Figures 8(c1) and 8(c2)) and, in contrast, when cocultured with SC-like cells for 7 days, the neurites of PC12 cells were longer and grew along with SC-like cells (Figures 8(d1) and 8(d2)). After 21 days of coculture, transmission electron microscopy (TEM) showed that SC-like cells could form myelin sheath with neurites (Figure 9(c)). On the contrary, REMSCs could not form myelin structures with PC12 neurites (Figure 9(b)), and REMSCs could not form myelin structures without neurites (Figure 9(a)).

Bottom Line: When cocultured with dREMSCs for 5 days, PC12 cells differentiated into mature neuron-like cells with long neurites.More importantly, dREMSCs could form myelin structures with the neurites of PC12 cells at 21 days in vitro.Our data indicated that REMSCs, a kind of EMSCs, could differentiate into SC-like cells and have the ability to promote the differentiation of PC12 cells and form myelin in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopedics, The Third Affiliated Hospital of Suzhou University, Changzhou 213003, China.

ABSTRACT
Schwann cell (SC) transplantation as a cell-based therapy can enhance peripheral and central nerve repair experimentally, but it is limited by the donor site morbidity for clinical application. We investigated weather respiratory mucosa stem cells (REMSCs), a kind of ectomesenchymal stem cells (EMSCs), isolated from rat nasal septum can differentiate into functional Schwann-like cells (SC-like cells). REMSCs proliferated quickly in vitro and expressed the neural crest markers (nestin, vimentin, SOX10, and CD44). Treated with a mixture of glial growth factors for 7 days, REMSCs differentiated into SC-like cells. The differentiated REMSCs (dREMSCs) exhibited a spindle-like morphology similar to SC cells. Immunocytochemical staining and Western blotting indicated that SC-like cells expressed the glial markers (GFAP, S100β, Galc, and P75) and CNPase. When cocultured with dREMSCs for 5 days, PC12 cells differentiated into mature neuron-like cells with long neurites. More importantly, dREMSCs could form myelin structures with the neurites of PC12 cells at 21 days in vitro. Our data indicated that REMSCs, a kind of EMSCs, could differentiate into SC-like cells and have the ability to promote the differentiation of PC12 cells and form myelin in vitro.

No MeSH data available.


Related in: MedlinePlus