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Chemically-induced RAT mesenchymal stem cells adopt molecular properties of neuronal-like cells but do not have basic neuronal functional properties.

Barnabé GF, Schwindt TT, Calcagnotto ME, Motta FL, Martinez G, de Oliveira AC, Keim LM, D'Almeida V, Mendez-Otero R, Mello LE - PLoS ONE (2009)

Bottom Line: Moreover, increased intracellular cysteine after treatment indicates an impairment of redox circuitry during chemical induction, and in vitro electrophysiological recordings (patch-clamp) of the chemically induced MSC did not indicate neuronal properties as these cells do not exhibit Na(+) or K(+) currents and do not fire action potentials.Our findings suggest that a disruption of redox circuitry plays an important role in this specific chemical induction protocol, which might result in cytoskeletal alterations and loss of functional ion-gated channels followed by cell death.Despite the neuronal-like morphology and neural protein expression, induced rat bone marrow MSC do not have basic functional neuronal properties, although it is still plausible that other methods of induction and/or sources of MSC can achieve a successful neuronal differentiation in vitro.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Fisiologia, Universidade Federal de São Paulo (UNIFESP), São Paulo, São Paulo, Brazil.

ABSTRACT
Induction of adult rat bone marrow mesenchymal stem cells (MSC) by means of chemical compounds (beta-mercaptoethanol, dimethyl sulfoxide and butylated hydroxyanizole) has been proposed to lead to neuronal transdifferentiation, and this protocol has been broadly used by several laboratories worldwide. Only a few hours of MSC chemical induction using this protocol is sufficient for the acquisition of neuronal-like morphology and neuronal protein expression. However, given that cell death is abundant, we hypothesize that, rather than true neuronal differentiation, this particular protocol leads to cellular toxic effects. We confirm that the induced cells with neuronal-like morphology positively stained for NF-200, S100, beta-tubulin III, NSE and MAP-2 proteins. However, the morphological and molecular changes after chemical induction are also associated with an increase in the apoptosis of over 50% of the plated cells after 24 h. Moreover, increased intracellular cysteine after treatment indicates an impairment of redox circuitry during chemical induction, and in vitro electrophysiological recordings (patch-clamp) of the chemically induced MSC did not indicate neuronal properties as these cells do not exhibit Na(+) or K(+) currents and do not fire action potentials. Our findings suggest that a disruption of redox circuitry plays an important role in this specific chemical induction protocol, which might result in cytoskeletal alterations and loss of functional ion-gated channels followed by cell death. Despite the neuronal-like morphology and neural protein expression, induced rat bone marrow MSC do not have basic functional neuronal properties, although it is still plausible that other methods of induction and/or sources of MSC can achieve a successful neuronal differentiation in vitro.

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Ratio between Hcy e Cys contents obtained for non-induced (control, CTR) and chemically-induced MSC.No cellular change in Hcy content was observed in the induction protocol (p = 0.23). Total Cys content increased almost four times (p = 0.00006) in chemically-induced MSC compared to non-induced ones. Data represented as mean±standard deviation.
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pone-0005222-g006: Ratio between Hcy e Cys contents obtained for non-induced (control, CTR) and chemically-induced MSC.No cellular change in Hcy content was observed in the induction protocol (p = 0.23). Total Cys content increased almost four times (p = 0.00006) in chemically-induced MSC compared to non-induced ones. Data represented as mean±standard deviation.

Mentions: In order to evaluate whether the chemical compounds used in the neuronal induction could promote alterations in the redox balance, we analyzed the Cys and Hcy cellular contents. No cellular change in Hcy content was observed in chemically-induced MSC. However, total Cys content increased almost four times in induced MSC as compared to non-induced cells (figure 6).


Chemically-induced RAT mesenchymal stem cells adopt molecular properties of neuronal-like cells but do not have basic neuronal functional properties.

Barnabé GF, Schwindt TT, Calcagnotto ME, Motta FL, Martinez G, de Oliveira AC, Keim LM, D'Almeida V, Mendez-Otero R, Mello LE - PLoS ONE (2009)

Ratio between Hcy e Cys contents obtained for non-induced (control, CTR) and chemically-induced MSC.No cellular change in Hcy content was observed in the induction protocol (p = 0.23). Total Cys content increased almost four times (p = 0.00006) in chemically-induced MSC compared to non-induced ones. Data represented as mean±standard deviation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2667250&req=5

pone-0005222-g006: Ratio between Hcy e Cys contents obtained for non-induced (control, CTR) and chemically-induced MSC.No cellular change in Hcy content was observed in the induction protocol (p = 0.23). Total Cys content increased almost four times (p = 0.00006) in chemically-induced MSC compared to non-induced ones. Data represented as mean±standard deviation.
Mentions: In order to evaluate whether the chemical compounds used in the neuronal induction could promote alterations in the redox balance, we analyzed the Cys and Hcy cellular contents. No cellular change in Hcy content was observed in chemically-induced MSC. However, total Cys content increased almost four times in induced MSC as compared to non-induced cells (figure 6).

Bottom Line: Moreover, increased intracellular cysteine after treatment indicates an impairment of redox circuitry during chemical induction, and in vitro electrophysiological recordings (patch-clamp) of the chemically induced MSC did not indicate neuronal properties as these cells do not exhibit Na(+) or K(+) currents and do not fire action potentials.Our findings suggest that a disruption of redox circuitry plays an important role in this specific chemical induction protocol, which might result in cytoskeletal alterations and loss of functional ion-gated channels followed by cell death.Despite the neuronal-like morphology and neural protein expression, induced rat bone marrow MSC do not have basic functional neuronal properties, although it is still plausible that other methods of induction and/or sources of MSC can achieve a successful neuronal differentiation in vitro.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Fisiologia, Universidade Federal de São Paulo (UNIFESP), São Paulo, São Paulo, Brazil.

ABSTRACT
Induction of adult rat bone marrow mesenchymal stem cells (MSC) by means of chemical compounds (beta-mercaptoethanol, dimethyl sulfoxide and butylated hydroxyanizole) has been proposed to lead to neuronal transdifferentiation, and this protocol has been broadly used by several laboratories worldwide. Only a few hours of MSC chemical induction using this protocol is sufficient for the acquisition of neuronal-like morphology and neuronal protein expression. However, given that cell death is abundant, we hypothesize that, rather than true neuronal differentiation, this particular protocol leads to cellular toxic effects. We confirm that the induced cells with neuronal-like morphology positively stained for NF-200, S100, beta-tubulin III, NSE and MAP-2 proteins. However, the morphological and molecular changes after chemical induction are also associated with an increase in the apoptosis of over 50% of the plated cells after 24 h. Moreover, increased intracellular cysteine after treatment indicates an impairment of redox circuitry during chemical induction, and in vitro electrophysiological recordings (patch-clamp) of the chemically induced MSC did not indicate neuronal properties as these cells do not exhibit Na(+) or K(+) currents and do not fire action potentials. Our findings suggest that a disruption of redox circuitry plays an important role in this specific chemical induction protocol, which might result in cytoskeletal alterations and loss of functional ion-gated channels followed by cell death. Despite the neuronal-like morphology and neural protein expression, induced rat bone marrow MSC do not have basic functional neuronal properties, although it is still plausible that other methods of induction and/or sources of MSC can achieve a successful neuronal differentiation in vitro.

Show MeSH
Related in: MedlinePlus