Berberine induces neuronal differentiation through inhibition of cancer stemness and epithelial-mesenchymal transition in neuroblastoma cells

C. R. Naveen, Sagar Gaikwad, Reena Agrawal-Rajput

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Background Berberine, a plant alkaloid, has been used since many years for treatment of gastrointestinal disorders. It also shows promising medicinal use against metabolic disorders, neurodegenerative disorders and cancer; however its efficacy in neuroblastoma (NB) is poorly explored. Hypothesis EMT is important in cancer stemness and metastasis resulting in failure to differentiate; thus targeting EMT and related pathways can have clinical benefits. Study design Potential of berberine was investigated for (i) neuronal differentiation and cancer stemness inhibition, (ii) underlying molecular mechanisms regulating cancer-stemness and (iii) EMT reversal. Methods Using neuro2a (N2a) neuroblastoma cells (NB); we investigated effect of berberine on neuronal differentiation, cancer-stemness, EMT and underlying signalling by immunofluorescence, RT-PCR, Western blot. High glucose-induced TGF-β mediated EMT model was used to test EMT reversal potential by Western blot and RT-PCR. STRING analysis was done to determine and validate functional protein-interaction networks. Results We demonstrate berberine induces neuronal differentiation accompanying increased neuronal differentiation markers like MAP2, β-III tubulin and NCAM; generated neurons were viable. Berberine attenuated cancer stemness markers CD133, β-catenin, n-myc, sox2, notch2 and nestin. Berberine potentiated G0/G1 cell cycle arrest by inhibiting proliferation, cyclin dependent kinases and cyclins resulting in apoptosis through increased bax/bcl-2 ratio. Restoration of tumor suppressor proteins, p27 and p53, indicate promising anti-cancer property. The induction of NCAM and reduction in its polysialylation indicates anti-migratory potential which is supported by down regulation of MMP-2/9. It increased epithelial marker laminin and smad and increased Hsp70 levels also suggest its protective role. Molecular insights revealed that berberine regulates EMT via downregulation of PI3/Akt and Ras-Raf-ERK signalling and subsequent upregulation of p38-MAPK. TGF-β secretion from N2a cells was potentiated by high glucose and negatively regulated by berberine through modulation of TGF-β receptors II and III. Berberine reverted mesenchymal markers, vimentin and fibronectin, with restoration of epithelial marker E-cadherin, highlighting the role of berberine in reversal of EMT. Conclusion Collectively, the study demonstrates prospective use of berberine against neuroblastoma as elucidated through inhibition of fundamental characteristics of cancer stem cells: tumorigenicity and failure to differentiation and instigates reversal in the EMT.

Original languageEnglish (US)
Pages (from-to)736-744
Number of pages9
JournalPhytomedicine
Volume23
Issue number7
DOIs
StatePublished - Jun 15 2016
Externally publishedYes

Fingerprint

Berberine
Epithelial-Mesenchymal Transition
Neuroblastoma
Neoplasms
Neural Cell Adhesion Molecules
Down-Regulation
Western Blotting
G1 Phase Cell Cycle Checkpoints
Protein Interaction Maps
Tumor Suppressor Protein p53
Glucose
Catenins
Nestin
Polymerase Chain Reaction
Cyclins
Neoplastic Stem Cells
Cyclin-Dependent Kinases
Differentiation Antigens
Laminin
p38 Mitogen-Activated Protein Kinases

Keywords

  • Berberine
  • Cancer stemness
  • Cell cycle
  • Differentiation
  • EMT
  • Neuroblastoma

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology
  • Pharmaceutical Science
  • Drug Discovery
  • Complementary and alternative medicine

Cite this

Berberine induces neuronal differentiation through inhibition of cancer stemness and epithelial-mesenchymal transition in neuroblastoma cells. / Naveen, C. R.; Gaikwad, Sagar; Agrawal-Rajput, Reena.

In: Phytomedicine, Vol. 23, No. 7, 15.06.2016, p. 736-744.

Research output: Contribution to journalArticle

@article{153f8a9228a845d6bba164ec84666f01,
title = "Berberine induces neuronal differentiation through inhibition of cancer stemness and epithelial-mesenchymal transition in neuroblastoma cells",
abstract = "Background Berberine, a plant alkaloid, has been used since many years for treatment of gastrointestinal disorders. It also shows promising medicinal use against metabolic disorders, neurodegenerative disorders and cancer; however its efficacy in neuroblastoma (NB) is poorly explored. Hypothesis EMT is important in cancer stemness and metastasis resulting in failure to differentiate; thus targeting EMT and related pathways can have clinical benefits. Study design Potential of berberine was investigated for (i) neuronal differentiation and cancer stemness inhibition, (ii) underlying molecular mechanisms regulating cancer-stemness and (iii) EMT reversal. Methods Using neuro2a (N2a) neuroblastoma cells (NB); we investigated effect of berberine on neuronal differentiation, cancer-stemness, EMT and underlying signalling by immunofluorescence, RT-PCR, Western blot. High glucose-induced TGF-β mediated EMT model was used to test EMT reversal potential by Western blot and RT-PCR. STRING analysis was done to determine and validate functional protein-interaction networks. Results We demonstrate berberine induces neuronal differentiation accompanying increased neuronal differentiation markers like MAP2, β-III tubulin and NCAM; generated neurons were viable. Berberine attenuated cancer stemness markers CD133, β-catenin, n-myc, sox2, notch2 and nestin. Berberine potentiated G0/G1 cell cycle arrest by inhibiting proliferation, cyclin dependent kinases and cyclins resulting in apoptosis through increased bax/bcl-2 ratio. Restoration of tumor suppressor proteins, p27 and p53, indicate promising anti-cancer property. The induction of NCAM and reduction in its polysialylation indicates anti-migratory potential which is supported by down regulation of MMP-2/9. It increased epithelial marker laminin and smad and increased Hsp70 levels also suggest its protective role. Molecular insights revealed that berberine regulates EMT via downregulation of PI3/Akt and Ras-Raf-ERK signalling and subsequent upregulation of p38-MAPK. TGF-β secretion from N2a cells was potentiated by high glucose and negatively regulated by berberine through modulation of TGF-β receptors II and III. Berberine reverted mesenchymal markers, vimentin and fibronectin, with restoration of epithelial marker E-cadherin, highlighting the role of berberine in reversal of EMT. Conclusion Collectively, the study demonstrates prospective use of berberine against neuroblastoma as elucidated through inhibition of fundamental characteristics of cancer stem cells: tumorigenicity and failure to differentiation and instigates reversal in the EMT.",
keywords = "Berberine, Cancer stemness, Cell cycle, Differentiation, EMT, Neuroblastoma",
author = "Naveen, {C. R.} and Sagar Gaikwad and Reena Agrawal-Rajput",
year = "2016",
month = "6",
day = "15",
doi = "10.1016/j.phymed.2016.03.013",
language = "English (US)",
volume = "23",
pages = "736--744",
journal = "Phytomedicine",
issn = "0944-7113",
publisher = "Urban und Fischer Verlag Jena",
number = "7",

}

TY - JOUR

T1 - Berberine induces neuronal differentiation through inhibition of cancer stemness and epithelial-mesenchymal transition in neuroblastoma cells

AU - Naveen, C. R.

AU - Gaikwad, Sagar

AU - Agrawal-Rajput, Reena

PY - 2016/6/15

Y1 - 2016/6/15

N2 - Background Berberine, a plant alkaloid, has been used since many years for treatment of gastrointestinal disorders. It also shows promising medicinal use against metabolic disorders, neurodegenerative disorders and cancer; however its efficacy in neuroblastoma (NB) is poorly explored. Hypothesis EMT is important in cancer stemness and metastasis resulting in failure to differentiate; thus targeting EMT and related pathways can have clinical benefits. Study design Potential of berberine was investigated for (i) neuronal differentiation and cancer stemness inhibition, (ii) underlying molecular mechanisms regulating cancer-stemness and (iii) EMT reversal. Methods Using neuro2a (N2a) neuroblastoma cells (NB); we investigated effect of berberine on neuronal differentiation, cancer-stemness, EMT and underlying signalling by immunofluorescence, RT-PCR, Western blot. High glucose-induced TGF-β mediated EMT model was used to test EMT reversal potential by Western blot and RT-PCR. STRING analysis was done to determine and validate functional protein-interaction networks. Results We demonstrate berberine induces neuronal differentiation accompanying increased neuronal differentiation markers like MAP2, β-III tubulin and NCAM; generated neurons were viable. Berberine attenuated cancer stemness markers CD133, β-catenin, n-myc, sox2, notch2 and nestin. Berberine potentiated G0/G1 cell cycle arrest by inhibiting proliferation, cyclin dependent kinases and cyclins resulting in apoptosis through increased bax/bcl-2 ratio. Restoration of tumor suppressor proteins, p27 and p53, indicate promising anti-cancer property. The induction of NCAM and reduction in its polysialylation indicates anti-migratory potential which is supported by down regulation of MMP-2/9. It increased epithelial marker laminin and smad and increased Hsp70 levels also suggest its protective role. Molecular insights revealed that berberine regulates EMT via downregulation of PI3/Akt and Ras-Raf-ERK signalling and subsequent upregulation of p38-MAPK. TGF-β secretion from N2a cells was potentiated by high glucose and negatively regulated by berberine through modulation of TGF-β receptors II and III. Berberine reverted mesenchymal markers, vimentin and fibronectin, with restoration of epithelial marker E-cadherin, highlighting the role of berberine in reversal of EMT. Conclusion Collectively, the study demonstrates prospective use of berberine against neuroblastoma as elucidated through inhibition of fundamental characteristics of cancer stem cells: tumorigenicity and failure to differentiation and instigates reversal in the EMT.

AB - Background Berberine, a plant alkaloid, has been used since many years for treatment of gastrointestinal disorders. It also shows promising medicinal use against metabolic disorders, neurodegenerative disorders and cancer; however its efficacy in neuroblastoma (NB) is poorly explored. Hypothesis EMT is important in cancer stemness and metastasis resulting in failure to differentiate; thus targeting EMT and related pathways can have clinical benefits. Study design Potential of berberine was investigated for (i) neuronal differentiation and cancer stemness inhibition, (ii) underlying molecular mechanisms regulating cancer-stemness and (iii) EMT reversal. Methods Using neuro2a (N2a) neuroblastoma cells (NB); we investigated effect of berberine on neuronal differentiation, cancer-stemness, EMT and underlying signalling by immunofluorescence, RT-PCR, Western blot. High glucose-induced TGF-β mediated EMT model was used to test EMT reversal potential by Western blot and RT-PCR. STRING analysis was done to determine and validate functional protein-interaction networks. Results We demonstrate berberine induces neuronal differentiation accompanying increased neuronal differentiation markers like MAP2, β-III tubulin and NCAM; generated neurons were viable. Berberine attenuated cancer stemness markers CD133, β-catenin, n-myc, sox2, notch2 and nestin. Berberine potentiated G0/G1 cell cycle arrest by inhibiting proliferation, cyclin dependent kinases and cyclins resulting in apoptosis through increased bax/bcl-2 ratio. Restoration of tumor suppressor proteins, p27 and p53, indicate promising anti-cancer property. The induction of NCAM and reduction in its polysialylation indicates anti-migratory potential which is supported by down regulation of MMP-2/9. It increased epithelial marker laminin and smad and increased Hsp70 levels also suggest its protective role. Molecular insights revealed that berberine regulates EMT via downregulation of PI3/Akt and Ras-Raf-ERK signalling and subsequent upregulation of p38-MAPK. TGF-β secretion from N2a cells was potentiated by high glucose and negatively regulated by berberine through modulation of TGF-β receptors II and III. Berberine reverted mesenchymal markers, vimentin and fibronectin, with restoration of epithelial marker E-cadherin, highlighting the role of berberine in reversal of EMT. Conclusion Collectively, the study demonstrates prospective use of berberine against neuroblastoma as elucidated through inhibition of fundamental characteristics of cancer stem cells: tumorigenicity and failure to differentiation and instigates reversal in the EMT.

KW - Berberine

KW - Cancer stemness

KW - Cell cycle

KW - Differentiation

KW - EMT

KW - Neuroblastoma

UR - http://www.scopus.com/inward/record.url?scp=84966372765&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84966372765&partnerID=8YFLogxK

U2 - 10.1016/j.phymed.2016.03.013

DO - 10.1016/j.phymed.2016.03.013

M3 - Article

VL - 23

SP - 736

EP - 744

JO - Phytomedicine

JF - Phytomedicine

SN - 0944-7113

IS - 7

ER -