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Complementary effects of two growth factors in multifunctionalized silk nanofibers for nerve reconstruction.

Dinis TM, Vidal G, Jose RR, Vigneron P, Bresson D, Fitzpatrick V, Marin F, Kaplan DL, Egles C - PLoS ONE (2014)

Bottom Line: PC12 cells grown on the fibers confirmed the bioavailability and bioactivity of the NGF, which was not significantly released from the fibers.Organotypic culture of rat fetal DRGs confirmed the complementary effect of both growth factors in multifunctionalized nanofibers, which allowed glial cell migration, alignment and parallel axonal growth in structures resembling the 'bands of Bungner' found in situ.Our bioactive silk tubes thus represent new options for a biological and biocompatible nerve guidance conduit.

View Article: PubMed Central - PubMed

Affiliation: Université de Technologie de Compiègne, CNRS UMR 7338: BioMécanique et BioIngénierie, Centre de Recherches, BP 20529, Compiègne, France; Department of Biomedical Engineering, Tufts University, Medford, Massachusetts, United States of America.

ABSTRACT
With the aim of forming bioactive guides for peripheral nerve regeneration, silk fibroin was electrospun to obtain aligned nanofibers. These fibers were functionalized by incorporating Nerve Growth Factor (NGF) and Ciliary NeuroTrophic Factor (CNTF) during electrospinning. PC12 cells grown on the fibers confirmed the bioavailability and bioactivity of the NGF, which was not significantly released from the fibers. Primary neurons from rat dorsal root ganglia (DRGs) were grown on the nanofibers and anchored to the fibers and grew in a directional fashion based on the fiber orientation, and as confirmed by growth cone morphology. These biofunctionalized nanofibers led to a 3-fold increase in neurite length at their contact, which was likely due to the NGF. Glial cell growth, alignment and migration were stimulated by the CNTF in the functionalized nanofibers. Organotypic culture of rat fetal DRGs confirmed the complementary effect of both growth factors in multifunctionalized nanofibers, which allowed glial cell migration, alignment and parallel axonal growth in structures resembling the 'bands of Bungner' found in situ. Graftable multi-channel conduits based on biofunctionalized aligned silk nanofibers were developed as an organized 3D scaffold. Our bioactive silk tubes thus represent new options for a biological and biocompatible nerve guidance conduit.

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Growth factors release from electrospun functionalized silk and stability.A) No NGF was detected from the silk electrospun after 5 days in PBS. B) As NGF, no CNTF was detected from the electrospun. Degradation of growth factors was observed in PBS with 0.1% BSA. After 5 days, more than 50% of NGF could be detected unlike 10% for CNTF.
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pone-0109770-g002: Growth factors release from electrospun functionalized silk and stability.A) No NGF was detected from the silk electrospun after 5 days in PBS. B) As NGF, no CNTF was detected from the electrospun. Degradation of growth factors was observed in PBS with 0.1% BSA. After 5 days, more than 50% of NGF could be detected unlike 10% for CNTF.

Mentions: NGF (1 µg.mL−1) and CNTF (0.1 µg.mL−1) were added to the spinning solution to functionalize nanofibres. The release of NGF and CNTF from fibroin nanofibers after electrospinning was monitored by ELISA at different time points after sample incubation in PBS. For each batch of neurotrophic factors, one group contained soluble NGF (80 pg. mL−1) or CNTF (400 pg. mL−1) as a positive control. The detected amount from these controls decreased rapidly for NGF (37% after 24h and 50.1% after 48h) and even faster for CNTF (74% after 24h and 80.5% after 48h), suggesting rapid degradation of these neurotrophic factors. NGF and CNTF release was not detected from the nanofibers over 4 days, suggesting that both growth factors were trapped, strongly adsorbed to the nanofibers or released below the detection level of the ELISA kit (Figure 2 A and B).


Complementary effects of two growth factors in multifunctionalized silk nanofibers for nerve reconstruction.

Dinis TM, Vidal G, Jose RR, Vigneron P, Bresson D, Fitzpatrick V, Marin F, Kaplan DL, Egles C - PLoS ONE (2014)

Growth factors release from electrospun functionalized silk and stability.A) No NGF was detected from the silk electrospun after 5 days in PBS. B) As NGF, no CNTF was detected from the electrospun. Degradation of growth factors was observed in PBS with 0.1% BSA. After 5 days, more than 50% of NGF could be detected unlike 10% for CNTF.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4196919&req=5

pone-0109770-g002: Growth factors release from electrospun functionalized silk and stability.A) No NGF was detected from the silk electrospun after 5 days in PBS. B) As NGF, no CNTF was detected from the electrospun. Degradation of growth factors was observed in PBS with 0.1% BSA. After 5 days, more than 50% of NGF could be detected unlike 10% for CNTF.
Mentions: NGF (1 µg.mL−1) and CNTF (0.1 µg.mL−1) were added to the spinning solution to functionalize nanofibres. The release of NGF and CNTF from fibroin nanofibers after electrospinning was monitored by ELISA at different time points after sample incubation in PBS. For each batch of neurotrophic factors, one group contained soluble NGF (80 pg. mL−1) or CNTF (400 pg. mL−1) as a positive control. The detected amount from these controls decreased rapidly for NGF (37% after 24h and 50.1% after 48h) and even faster for CNTF (74% after 24h and 80.5% after 48h), suggesting rapid degradation of these neurotrophic factors. NGF and CNTF release was not detected from the nanofibers over 4 days, suggesting that both growth factors were trapped, strongly adsorbed to the nanofibers or released below the detection level of the ELISA kit (Figure 2 A and B).

Bottom Line: PC12 cells grown on the fibers confirmed the bioavailability and bioactivity of the NGF, which was not significantly released from the fibers.Organotypic culture of rat fetal DRGs confirmed the complementary effect of both growth factors in multifunctionalized nanofibers, which allowed glial cell migration, alignment and parallel axonal growth in structures resembling the 'bands of Bungner' found in situ.Our bioactive silk tubes thus represent new options for a biological and biocompatible nerve guidance conduit.

View Article: PubMed Central - PubMed

Affiliation: Université de Technologie de Compiègne, CNRS UMR 7338: BioMécanique et BioIngénierie, Centre de Recherches, BP 20529, Compiègne, France; Department of Biomedical Engineering, Tufts University, Medford, Massachusetts, United States of America.

ABSTRACT
With the aim of forming bioactive guides for peripheral nerve regeneration, silk fibroin was electrospun to obtain aligned nanofibers. These fibers were functionalized by incorporating Nerve Growth Factor (NGF) and Ciliary NeuroTrophic Factor (CNTF) during electrospinning. PC12 cells grown on the fibers confirmed the bioavailability and bioactivity of the NGF, which was not significantly released from the fibers. Primary neurons from rat dorsal root ganglia (DRGs) were grown on the nanofibers and anchored to the fibers and grew in a directional fashion based on the fiber orientation, and as confirmed by growth cone morphology. These biofunctionalized nanofibers led to a 3-fold increase in neurite length at their contact, which was likely due to the NGF. Glial cell growth, alignment and migration were stimulated by the CNTF in the functionalized nanofibers. Organotypic culture of rat fetal DRGs confirmed the complementary effect of both growth factors in multifunctionalized nanofibers, which allowed glial cell migration, alignment and parallel axonal growth in structures resembling the 'bands of Bungner' found in situ. Graftable multi-channel conduits based on biofunctionalized aligned silk nanofibers were developed as an organized 3D scaffold. Our bioactive silk tubes thus represent new options for a biological and biocompatible nerve guidance conduit.

Show MeSH