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Adult human brain neural progenitor cells (NPCs) and fibroblast-like cells have similar properties in vitro but only NPCs differentiate into neurons.

Park TI, Monzo H, Mee EW, Bergin PS, Teoh HH, Montgomery JM, Faull RL, Curtis MA, Dragunow M - PLoS ONE (2012)

Bottom Line: This gradual change in cellular composition resulted in a progressive decline in neurogenic potential without the apparent loss of self-renewal in our cultures.These results demonstrate that while AhNPCs and FbCs behave similarly under proliferative conditions, they are two different cell populations.This information is vital for the interpretation and reproducibility of AhNPC experiments and suggests an ideal time frame for conducting AhNPC-based experiments.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Clinical Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Auckland, New Zealand.

ABSTRACT
The ability to culture neural progenitor cells from the adult human brain has provided an exciting opportunity to develop and test potential therapies on adult human brain cells. To achieve a reliable and reproducible adult human neural progenitor cell (AhNPC) culture system for this purpose, this study fully characterized the cellular composition of the AhNPC cultures, as well as the possible changes to this in vitro system over prolonged culture periods. We isolated cells from the neurogenic subventricular zone/hippocampus (SVZ/HP) of the adult human brain and found a heterogeneous culture population comprised of several types of post-mitotic brain cells (neurons, astrocytes, and microglia), and more importantly, two distinct mitotic cell populations; the AhNPCs, and the fibroblast-like cells (FbCs). These two populations can easily be mistaken for a single population of AhNPCs, as they both proliferate under AhNPC culture conditions, form spheres and express neural progenitor cell and early neuronal markers, all of which are characteristics of AhNPCs in vitro. However, despite these similarities under proliferating conditions, under neuronal differentiation conditions, only the AhNPCs differentiated into functional neurons and glia. Furthermore, AhNPCs showed limited proliferative capacity that resulted in their depletion from culture by 5-6 passages, while the FbCs, which appear to be from a neurovascular origin, displayed a greater proliferative capacity and dominated the long-term cultures. This gradual change in cellular composition resulted in a progressive decline in neurogenic potential without the apparent loss of self-renewal in our cultures. These results demonstrate that while AhNPCs and FbCs behave similarly under proliferative conditions, they are two different cell populations. This information is vital for the interpretation and reproducibility of AhNPC experiments and suggests an ideal time frame for conducting AhNPC-based experiments.

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Photomicrograph and bar graph illustrating the decrease in neurogenic capabilities observed with increased culture duration.(A, C) Illustrates the extent of neuronal (βIII-tubulin) and astrocytic (GFAP) differentiation seen after 3 weeks of differentiating AhNPCs from cultures at passage 3. (B, D) Illustrates the same parameters when AhNPC cultures from passage 6 were differentiated. These images demonstrate the dramatic decline in the neurogenic capabilities of the AhNPC cultures with increased culture duration. (E) Quantification of the percentage of differentiated neurons and astrocytes derived from the 3 independent cases using automated images analysis. Statistical analysis using one-way Anova demonstrates the significant reduction in the percentage of βIII-tubulin cells (*  =  P<0.05, **  =  P<0.01).
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pone-0037742-g003: Photomicrograph and bar graph illustrating the decrease in neurogenic capabilities observed with increased culture duration.(A, C) Illustrates the extent of neuronal (βIII-tubulin) and astrocytic (GFAP) differentiation seen after 3 weeks of differentiating AhNPCs from cultures at passage 3. (B, D) Illustrates the same parameters when AhNPC cultures from passage 6 were differentiated. These images demonstrate the dramatic decline in the neurogenic capabilities of the AhNPC cultures with increased culture duration. (E) Quantification of the percentage of differentiated neurons and astrocytes derived from the 3 independent cases using automated images analysis. Statistical analysis using one-way Anova demonstrates the significant reduction in the percentage of βIII-tubulin cells (*  =  P<0.05, **  =  P<0.01).

Mentions: Greater than 60% of our cultures continued to proliferate for longer than 8–10 months (>6 passages), formed NSs and expressed the NPC marker Nestin (Figure 1 B). However, the ability of these cultures to generate neurons and astrocytes (their neurogenic ability) deteriorated with successive passages, reaching insignificant levels by 5–6 months (∼ 6 passages; Figure 3). The morphological composition of the cultures also progressively changed, going from phase bright monolayer and sphere-dominated cultures (<5 months) to relatively homogenous bipolar monolayer cultures with limited sphere formations (>6 months). These long-term cultured cells showed strong resemblance to the FbCs reported previously by our laboratory [21], except for their bipolar morphology and the level of Nestin expression.


Adult human brain neural progenitor cells (NPCs) and fibroblast-like cells have similar properties in vitro but only NPCs differentiate into neurons.

Park TI, Monzo H, Mee EW, Bergin PS, Teoh HH, Montgomery JM, Faull RL, Curtis MA, Dragunow M - PLoS ONE (2012)

Photomicrograph and bar graph illustrating the decrease in neurogenic capabilities observed with increased culture duration.(A, C) Illustrates the extent of neuronal (βIII-tubulin) and astrocytic (GFAP) differentiation seen after 3 weeks of differentiating AhNPCs from cultures at passage 3. (B, D) Illustrates the same parameters when AhNPC cultures from passage 6 were differentiated. These images demonstrate the dramatic decline in the neurogenic capabilities of the AhNPC cultures with increased culture duration. (E) Quantification of the percentage of differentiated neurons and astrocytes derived from the 3 independent cases using automated images analysis. Statistical analysis using one-way Anova demonstrates the significant reduction in the percentage of βIII-tubulin cells (*  =  P<0.05, **  =  P<0.01).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3366988&req=5

pone-0037742-g003: Photomicrograph and bar graph illustrating the decrease in neurogenic capabilities observed with increased culture duration.(A, C) Illustrates the extent of neuronal (βIII-tubulin) and astrocytic (GFAP) differentiation seen after 3 weeks of differentiating AhNPCs from cultures at passage 3. (B, D) Illustrates the same parameters when AhNPC cultures from passage 6 were differentiated. These images demonstrate the dramatic decline in the neurogenic capabilities of the AhNPC cultures with increased culture duration. (E) Quantification of the percentage of differentiated neurons and astrocytes derived from the 3 independent cases using automated images analysis. Statistical analysis using one-way Anova demonstrates the significant reduction in the percentage of βIII-tubulin cells (*  =  P<0.05, **  =  P<0.01).
Mentions: Greater than 60% of our cultures continued to proliferate for longer than 8–10 months (>6 passages), formed NSs and expressed the NPC marker Nestin (Figure 1 B). However, the ability of these cultures to generate neurons and astrocytes (their neurogenic ability) deteriorated with successive passages, reaching insignificant levels by 5–6 months (∼ 6 passages; Figure 3). The morphological composition of the cultures also progressively changed, going from phase bright monolayer and sphere-dominated cultures (<5 months) to relatively homogenous bipolar monolayer cultures with limited sphere formations (>6 months). These long-term cultured cells showed strong resemblance to the FbCs reported previously by our laboratory [21], except for their bipolar morphology and the level of Nestin expression.

Bottom Line: This gradual change in cellular composition resulted in a progressive decline in neurogenic potential without the apparent loss of self-renewal in our cultures.These results demonstrate that while AhNPCs and FbCs behave similarly under proliferative conditions, they are two different cell populations.This information is vital for the interpretation and reproducibility of AhNPC experiments and suggests an ideal time frame for conducting AhNPC-based experiments.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Clinical Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Auckland, New Zealand.

ABSTRACT
The ability to culture neural progenitor cells from the adult human brain has provided an exciting opportunity to develop and test potential therapies on adult human brain cells. To achieve a reliable and reproducible adult human neural progenitor cell (AhNPC) culture system for this purpose, this study fully characterized the cellular composition of the AhNPC cultures, as well as the possible changes to this in vitro system over prolonged culture periods. We isolated cells from the neurogenic subventricular zone/hippocampus (SVZ/HP) of the adult human brain and found a heterogeneous culture population comprised of several types of post-mitotic brain cells (neurons, astrocytes, and microglia), and more importantly, two distinct mitotic cell populations; the AhNPCs, and the fibroblast-like cells (FbCs). These two populations can easily be mistaken for a single population of AhNPCs, as they both proliferate under AhNPC culture conditions, form spheres and express neural progenitor cell and early neuronal markers, all of which are characteristics of AhNPCs in vitro. However, despite these similarities under proliferating conditions, under neuronal differentiation conditions, only the AhNPCs differentiated into functional neurons and glia. Furthermore, AhNPCs showed limited proliferative capacity that resulted in their depletion from culture by 5-6 passages, while the FbCs, which appear to be from a neurovascular origin, displayed a greater proliferative capacity and dominated the long-term cultures. This gradual change in cellular composition resulted in a progressive decline in neurogenic potential without the apparent loss of self-renewal in our cultures. These results demonstrate that while AhNPCs and FbCs behave similarly under proliferative conditions, they are two different cell populations. This information is vital for the interpretation and reproducibility of AhNPC experiments and suggests an ideal time frame for conducting AhNPC-based experiments.

Show MeSH