Microtopographical cues promote peripheral nerve regeneration via transient mTORC2 activation

Thomson, Sydney; Charalambous, Chloe; Smith, Carol-Anne; Tsimbouri, Penelope; Déjardin, Théophile; Kingham, Paul J.; Hart, Andrew; Riehle, Mathis O.

doi:10.1016/j.actbio.2017.07.031

Cited by 26 publications

(24 citation statements)

References 91 publications

(112 reference statements)

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“…Moreover, mTORC2 has been shown to be involved in the regulation of actin dynamics and morphology of neurons in a Rac/PAK-dependent signaling pathway that controls cofilin phosphorylation (Huang et al, 2013 ; Thomanetz et al, 2013 ). Very recently, it has been demonstrated in DRG neurons that topographical features can potentiate mTORC2 guiding neurite outgrowth (Thomson et al, 2017 ). On ns-Zr15 the mTORC2 might already be induced by the cell/nanotopography interaction and thus rapamycin treatment does not further affect neurite outgrowth.…”

Section: Resultsmentioning

confidence: 99%

Proteomic Dissection of Nanotopography-Sensitive Mechanotransductive Signaling Hubs that Foster Neuronal Differentiation in PC12 Cells

Maffioli

Schulte

Nonnis

et al. 2018

Front. Cell. Neurosci.

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Neuronal cells are competent in precisely sensing nanotopographical features of their microenvironment. The perceived microenvironmental information will be “interpreted” by mechanotransductive processes and impacts on neuronal functioning and differentiation. Attempts to influence neuronal differentiation by engineering substrates that mimic appropriate extracellular matrix (ECM) topographies are hampered by the fact that profound details of mechanosensing/-transduction complexity remain elusive. Introducing omics methods into these biomaterial approaches has the potential to provide a deeper insight into the molecular processes and signaling cascades underlying mechanosensing/-transduction but their exigence in cellular material is often opposed by technical limitations of major substrate top-down fabrication methods. Supersonic cluster beam deposition (SCBD) allows instead the bottom-up fabrication of nanostructured substrates over large areas characterized by a quantitatively controllable ECM-like nanoroughness that has been recently shown to foster neuron differentiation and maturation. Exploiting this capacity of SCBD, we challenged mechanosensing/-transduction and differentiative behavior of neuron-like PC12 cells with diverse nanotopographies and/or changes of their biomechanical status, and analyzed their phosphoproteomic profiles in these settings. Versatile proteins that can be associated to significant processes along the mechanotransductive signal sequence, i.e., cell/cell interaction, glycocalyx and ECM, membrane/f-actin linkage and integrin activation, cell/substrate interaction, integrin adhesion complex, actomyosin organization/cellular mechanics, nuclear organization, and transcriptional regulation, were affected. The phosphoproteomic data suggested furthermore an involvement of ILK, mTOR, Wnt, and calcium signaling in these nanotopography- and/or cell mechanics-related processes. Altogether, potential nanotopography-sensitive mechanotransductive signaling hubs participating in neuronal differentiation were dissected.

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Section: Resultsmentioning

confidence: 99%

Proteomic Dissection of Nanotopography-Sensitive Mechanotransductive Signaling Hubs that Foster Neuronal Differentiation in PC12 Cells

Maffioli

Schulte

Nonnis

et al. 2018

Front. Cell. Neurosci.

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“…Neonatal Sprague Dawley rats (1–3 days old) dissection and Dorsal Root Ganglia (DRG) extraction were performed according to previous protocols [ 23 ]. DRG organotypic explants after delicate microsurgical root cleaning were immediately cultured (a single one for each well) according to the following conditions: Condition 1 (control): single DRG grown on poly- l -lysine (PLL)-coated coverslip for 48 h at 37 °C, 5% CO 2 in 200 μL of L-15 media (Sigma-Aldrich), 50 μg/ml N -acetyl-cysteine (NAC, Sigma-Aldrich), 1% penicillin-streptomycin (Pen-S, GE Healthcare), 10 ng/ml NGF 2.5S (NGF, Invitrogen), and 5% human platelet lysate (hPL, Antibodies) or 10% fetal bovine serum (FBS, Sigma Aldrich).…”

Section: Methodsmentioning

confidence: 99%

“…Neonatal Sprague Dawley rats (1-3 days old) dissection and Dorsal Root Ganglia (DRG) extraction were performed according to previous protocols [23]. DRG organotypic explants after delicate microsurgical root cleaning were immediately cultured (a single one for each well) according to the following conditions:…”

Section: Functional Cocultures With Primary Neuronsmentioning

confidence: 99%

Human platelet lysate as a potential clinical-translatable supplement to support the neurotrophic properties of human adipose-derived stem cells

et al. 2020

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Background The autologous nerve graft, despite its donor site morbidity and unpredictable functional recovery, continues to be the gold standard in peripheral nerve repair. Rodent research studies have shown promising results with cell transplantation of human adipose-derived stem cells (hADSC) in a bioengineered conduit, as an alternative strategy for nerve regeneration. To achieve meaningful clinical translation, cell therapy must comply with biosafety. Cell extraction and expansion methods that use animal-derived products, including enzymatic adipose tissue dissociation and the use of fetal bovine serum (FBS) as a culture medium supplement, have the potential for transmission of zoonotic infectious and immunogenicity. Human-platelet-lysate (hPL) serum has been used in recent years in human cell expansion, showing reliability in clinical applications. Methods We investigated whether hADSC can be routinely isolated and cultured in a completely xenogeneic-free way (using hPL culture medium supplement and avoiding collagenase digestion) without altering their physiology and stem properties. Outcomes in terms of stem marker expression (CD105, CD90, CD73) and the osteocyte/adipocyte differentiation capacity were compared with classical collagenase digestion and FBS-supplemented hADSC expansion. Results We found no significant differences between the two examined extraction and culture protocols in terms of cluster differentiation (CD) marker expression and stem cell plasticity, while hADSC in hPL showed a significantly higher proliferation rate when compared with the usual FBS-added medium. Considering the important key growth factors (particularly brain-derived growth factor (BDNF)) present in hPL, we investigated a possible neurogenic commitment of hADSC when cultured with hPL. Interestingly, hADSC cultured in hPL showed a statistically higher secretion of neurotrophic factors BDNF, glial cell-derived growth factor (GDNF), and nerve-derived growth factor (NFG) than FBS-cultured cells. When cocultured in the presence of primary neurons, hADSC which had been grown under hPL supplementation, showed significantly enhanced neurotrophic properties. Conclusions The hPL-supplement medium could improve cell proliferation and neurotropism while maintaining stable cell properties, showing effectiveness in clinical translation and significant potential in peripheral nerve research.

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“…The use of microtopographical patterns have been reported to promote the regeneration of peripheral nerves (Farrukh, Zhao, & del Campo, 2018; Gomez, Lee, Nickels, & Schmidt, 2007; Thomson et al, 2017; Y. Yang, Zhang, Chai, & Gu, 2018). During axonal regeneration, different cell types in the nerve undergo dynamic changes that are controlled by external cues.…”

Section: Introductionmentioning

confidence: 99%

mentioning

confidence: 99%

Microtopographical patterns promote different responses in fibroblasts and Schwann cells: A possible feature for neural implants

Mobini

Kuliasha

Siders

et al. 2020

J Biomedical Materials Res

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The chronic reliability of bioelectronic neural interfaces has been challenged by foreign body reactions (FBRs) resulting in fibrotic encapsulation and poor integration with neural tissue. Engineered microtopographies could alleviate these challenges by manipulating cellular responses to the implanted device. Parallel microchannels have been shown to modulate neuronal cell alignment and axonal growth, and Sharklet™ microtopographies of targeted feature sizes can modulate bio-adhesion of an array of bacteria, marine organisms, and epithelial cells due to their unique geometry. We hypothesized that a Sharklet™ micropattern could be identified that inhibited fibroblasts partially responsible for FBR while promoting Schwann cell proliferation and alignment. in vitro cell assays were used to screen the effect of Sharklet™ and channel micropatterns of varying dimensions from 2 to 20 μm on fibroblast and Schwann cell metrics (e.g., morphology/alignment, nuclei count, metabolic activity), and a hierarchical analysis of variance was used to compare treatments. In general, Schwann cells were found to be more metabolically active and aligned than fibroblasts when compared between the same pattern. 20 μm wide channels spaced 2 μm apart were found to promote Schwann cell attachment and alignment while simultaneously inhibiting fibroblasts and warrant further in vivo study on neural interface devices. No statistically significant trends between cellular responses and geometrical parameters were identified because mammalian cells can change their morphology dependent on their environment in a manner dissimilar to bacteria. Our results showed although surface patterning is a strong physical tool for modulating cell behavior, responses to micropatterns are highly dependent on the cell type.

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Microtopographical cues promote peripheral nerve regeneration via transient mTORC2 activation

Abstract: Graphical abstract

Cited by 26 publications

References 91 publications

Proteomic Dissection of Nanotopography-Sensitive Mechanotransductive Signaling Hubs that Foster Neuronal Differentiation in PC12 Cells

Proteomic Dissection of Nanotopography-Sensitive Mechanotransductive Signaling Hubs that Foster Neuronal Differentiation in PC12 Cells

Human platelet lysate as a potential clinical-translatable supplement to support the neurotrophic properties of human adipose-derived stem cells

Microtopographical patterns promote different responses in fibroblasts and Schwann cells: A possible feature for neural implants

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