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Silibinin suppresses TGFβ2-induced lens epithelial cell migration and epithelial–mesenchymal transition

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Abstract

Growth factor-induced migration of lens epithelial cell (LEC) toward the posterior of lens capsule bag and their epithelial–mesenchymal transition (EMT) is the key process involved in the pathogenesis of posterior capsular opacification (PCO). Silibinin, a natural flavonolignan, confers therapeutic effects to different cells by regulation of signalling pathways; however, its role in the prevention of migration and EMT of LECs is yet to be analysed. In this study, the inhibitory capabilities of silibinin on migration and EMT were analysed in response to TGFβ2 stimulation in HLE B-3 cells. The anti-migratory effect of silibinin was analysed using wound healing assay. Transcriptional and translational expression of genes related to LEC migration, EMT, and transcription factors related to EMT were studied by quantitative real-time PCR and Western blotting. Immunofluorescence analysis was utilized to study the localization of fibronectin. Silibinin reduced the viability of LECs in a concentration-dependent manner and inhibited the wound healing capacity of LECs induced by TGFβ2. Silibinin also suppressed alteration in the EMT-related markers such as cytoskeletal proteins, cell adhesion markers, extracellular matrix molecules, and transcription factors. Analysis of downstream signalling revealed that treatment with silibinin decreased phosphorylated Akt (Ser473, Thr308), PDK1 (Ser241), PTEN (Ser380), c-Raf (Ser259), and GSK3β (Ser9) in TGFβ-stimulated cells. The effect of silibinin treatment on phosphorylated Akt resembled that of the PI3K inhibitor LY294002. Our results suggest that silibinin can suppress LEC migration and EMT, which involves the inactivation of the PI3K-Akt signalling pathway. Silibinin might be a good candidate for PCO prevention; however, functional evaluation of silibinin using in vivo models is a pre-requisite.

Graphical abstract

Inhibitory effect of silibinin on TGFβ2-induced migration and EMT of LECs.

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Abbreviations

Akt:

protein kinase B

BSA:

bovine serum albumin

COX1/2:

cyclooxygenase 1/2

CTGF:

connective tissue growth factor

DABCO:

1,4-diazabicyclo(2.2.2)octane

DAPI:

4′,6-diamidino-2-phenylindone

ECL:

enhanced chemiluminescence

EGF:

epidermal growth factor

EMT:

epithelial–mesenchymal transition

Erk:

extracellular signal-regulated kinase

GAPDH:

glyceraldehyde 3-phosphate dehydrogenase

GSK3β:

glycogen synthase kinase 3 beta

HGF:

hepatocyte growth factor

HRP:

horseradish peroxidase

IC50 :

half maximal inhibitory concentration

IOL:

intraocular lens

IOP:

intraocular pressure

JNK:

C-jun N-terminal kinase

LEC:

lens epithelial cell

MAPK:

mitogen-activated protein kinase

MMP9:

matrix metalloproteinase 9

MTT:

(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

NdYAG:

neodymium-doped yttrium aluminum garnet

pAkt:

phosphor Akt

PBS:

phosphate-buffered saline

PCO:

posterior capsular opacification

PDK1:

3-phosphoinositide-dependent protein kinase 1

PI3K:

phosphatidylinositol 3-kinase

PTEN:

phosphatase and tensin homolog

qRT PCR:

quantitative real-time polymerase chain reaction

SDS:

sodium dodecyl sulfate

TBS:

tris-buffered saline

TBST:

tris-buffered saline plus tween 20

TGFβ2:

transforming growth factor-beta 2

VEGF:

vascular endothelial growth factor

ZEB:

zinc finger E-box binding homeobox

ZO1/TJP1:

zona occludens/tight junction protein 1

αSMA:

alpha smooth muscle actin

References

  • Awasthi N, Guo S and Wagner BJ 2009 Posterior capsular opacification: a problem reduced but not yet eradicated. Arch. Ophthalmol. 127 555–562

    Article  Google Scholar 

  • Chandler HL, Webb TR, Barden CA, et al. 2010 The effect of phosphorylated Akt inhibition on posterior capsule opacification in an ex vivo canine model. Mol. Vis. 16 2202–2214

    CAS  PubMed  PubMed Central  Google Scholar 

  • Cufí S, Bonavia R, Vazquez-Martin A, et al. 2013 Silibinin meglumine, a water-soluble form of milk thistle silymarin, is an orally active anti-cancer agent that impedes the epithelial-to-mesenchymal transition (EMT) in EGFR-mutant non-small-cell lung carcinoma cells. Food Chem. Toxicol. 60 360–368

    Article  Google Scholar 

  • Dai X, Liu J, Zheng H, et al. 2017 Nano-formulated curcumin accelerates acute wound healing through Dkk-1-mediated fibroblast mobilization and MCP-1-mediated anti-inflammation. NPG Asia Mater. 9 368

    Article  Google Scholar 

  • Deep G, Gangar SC, Agarwal C and Agarwal R 2011 Role of E-cadherin in antimigratory and antiinvasive efficacy of silibinin in prostate cancer cells. Cancer Prev. Res. 4 1222–1232

    Article  CAS  Google Scholar 

  • Eo HJ, Park GH and Jeong JB 2016 Inhibition of wnt signaling by silymarin in human colorectal cancer cells. Biomol. Ther. 24 380–386

    Article  CAS  Google Scholar 

  • Ezhilarasan D, Evraerts J, Brice S, et al. 2016 Silibinin inhibits proliferation and migration of human hepatic stellate LX-2 cells. J. Clin. Exp. Hepatol. 6 167–174

    Article  Google Scholar 

  • Fan Y, Hou T, Dan W, et al. 2020 Silibinin inhibits epithelial–mesenchymal transition of renal cell carcinoma through autophagy–dependent Wnt/β–catenin signaling. Int. J. Mol. Med. 45 1341–1350

    CAS  PubMed  PubMed Central  Google Scholar 

  • Ganatra DA, Vasavada AR, Vidya NG, Gajjar DU and Rajkumar S 2018 Trichostatin a restores expression of adherens and tight junction proteins during transforming growth factor β-mediated epithelial-to-mesenchymal transition. J. Ophthal. Vis. Res. 13 274–283

    Article  Google Scholar 

  • Guo R, Meng Q, Guo H, et al. 2016 TGF-β2 induces epithelial-mesenchymal transition in cultured human lens epithelial cells through activation of the PI3K/Akt/mTOR signaling pathway. Mol. Med. Rep. 13 1105–1110

    Article  CAS  Google Scholar 

  • Hales AM, Schulz MW, Chamberlain CG and Mcavoy JW 1994 TGF-β1 induces Lens cells to accumulate αsmooth muscle actin, a marker for subcapsular cataracts. Curr. Eye Res. 13 885–890

    Article  CAS  Google Scholar 

  • Hogan FS, Krishnegowda NK, Mikhailova M and Kahlenberg MS 2007 Flavonoid, silibinin, inhibits proliferation and promotes cell-cycle arrest of human colon cancer. J. Surg. Res. 143 58–65

    Article  CAS  Google Scholar 

  • Huang RYJ, Guilford P and Thiery JP 2012 Early events in cell adhesion and polarity during epithelial-mesenchymal transition. J. Cell Sci. 125 4417–4422

    Article  CAS  Google Scholar 

  • Jiang J, Shihan MH, Wang Y and Duncan MK 2018 Lens epithelial cells initiate an inflammatory response following cataract surgery. Investig. Ophthalmol. Vis. Sci. 59 4986–4997

    Article  CAS  Google Scholar 

  • Karahan E, Er D and Kaynak S 2014 An overview of Nd:YAG laser capsulotomy. Med. Hypothesis, Discov. Innov. Ophthalmol. J. 3 45–50

    Google Scholar 

  • Kayastha F, Madhu H, Vasavada A and Johar K 2014 Andrographolide reduces proliferation and migration of lens epithelial cells by modulating PI3K/Akt pathway. Exp. Eye Res. 128 23–26

    Article  CAS  Google Scholar 

  • Kim WK, Kwon Y, Jang M, et al. 2019 β-catenin activation down-regulates cell-cell junction-related genes and induces epithelial-to-mesenchymal transition in colorectal cancers. Sci. Rep. 9 18440

    Article  CAS  Google Scholar 

  • Kubo E, Shibata T, Singh DP and Sasaki H 2018 Roles of TGF β and FGF signals in the lens: Tropomyosin regulation for posterior capsule opacity. Int. J. Mol. Sci. 19 3093

    Article  Google Scholar 

  • Li F, Sun Y, Jia J, et al. 2018 Silibinin attenuates TGF-β1-induced migration and invasion via EMT suppression and is associated with COX-2 downregulation in bladder transitional cell carcinoma. Oncol. Rep. 40 3543–3550

    CAS  PubMed  Google Scholar 

  • Li J, Ye W, Xu W, et al. 2020 Activation of autophagy inhibits epithelial to mesenchymal transition process of human lens epithelial cells induced by high glucose conditions. Cell. Signal. 75 109768

    Article  CAS  Google Scholar 

  • Liu H, Mao Y, Xia B, et al. 2020a Curcumin inhibits proliferation and epithelial-mesenchymal transition in lens epithelial cells through multiple pathways. Biomed Res. Int. 2020 6061894

    PubMed  PubMed Central  Google Scholar 

  • Liu R, Wang Q, Ding Z, et al. 2020b Silibinin augments the antifibrotic effect of valsartan through inactivation of TGF-β1 signaling in kidney. Drug Des. Devel. Ther. 14 603–611

    Article  CAS  Google Scholar 

  • Loh CY, Chai JY, Tang TF, et al. 2019 The E-Cadherin and N-Cadherin Switch in epithelial-to-mesenchymal transition: signaling, therapeutic implications, and challenges. Cells 8 1118

    Article  CAS  Google Scholar 

  • Mamuya FA, Wang Y, Roop VH, et al. 2014 The roles of αV integrins in lens EMT and posterior capsular opacification. J. Cell. Mol. Med. 18 656–670

    Article  CAS  Google Scholar 

  • Medici D, Hay ED and Olsen BR 2008 Snail and slug promote epithelial-mesenchymal transition through β-catenin-T-cell factor-4-dependent expression of transforming growth factor-β3. Mol. Biol. Cell 19 4875–4887

    Article  CAS  Google Scholar 

  • Menko AS, Bleaken BM, Libowitz AA, et al. 2014 A central role for vimentin in regulating repair function during healing of the lens epithelium. Mol. Biol. Cell 25 776–790

    Article  CAS  Google Scholar 

  • Raj SM, Vasavada AR, Johar SRK, Vasavada VA and Vasavada VA 2007 Post-operative capsular opacification: a review. Int. J. Biomed. Sci. 3 237–250

    PubMed  PubMed Central  Google Scholar 

  • Sameri S, Saidijam M, Bahreini F and Najafi R 2020 Cancer chemopreventive activities of silibinin on colorectal cancer through regulation of E-cadherin/β-catenin pathway. Nutr. Cancer. https://doi.org/10.1080/01635581.2020.1800764

    Article  PubMed  Google Scholar 

  • Shirai K, Kitano-Izutani A, Miyamoto T, Tanaka SI and Saika S 2014 Wound healing and epithelial-mesenchymal transition in the lens epithelium: Roles of growth factors and extracellular matrix; in Lens epithelium and posterior capsular opacification (Springer, Japan) pp. 159–174

  • Shukal D, Bhadresha K, Shastri B, Mehta D, Vasavada A, and Johar K 2020 Dichloroacetate prevents TGFβ-induced epithelial-mesenchymal transition of retinal pigment epithelial cells. Exp. Eye Res. 197

  • Singh RP, Gu M and Agarwal R 2008a Silibinin inhibits colorectal cancer growth by inhibiting tumor cell proliferation and angiogenesis. Cancer Res. 68 2043–2050

    Article  CAS  Google Scholar 

  • Singh RP, Raina K, Sharma G and Agarwal R 2008b Silibinin inhibits established prostate tumor growth, progression, invasion, and metastasis and suppresses tumor angiogenesis and epithelial-mesenchymal transition in transgenic adenocarcinoma of the mouse prostate model mice. Clin. Cancer Res. 14 7773–7780

    Article  CAS  Google Scholar 

  • Valenta T, Hausmann G and Basler K 2012 The many faces and functions of Î 2-catenin. EMBO J. 31 2714–2736

    Article  CAS  Google Scholar 

  • Wernecke L, Keckeis S, Reichhart N, Strauß O and Salchow DJ 2018 Epithelial-mesenchymal transdifferentiation in pediatric lens epithelial cells. Investig. Ophthalmol. Vis. Sci. 59 5785–5794

    Article  CAS  Google Scholar 

  • Wertheimer CM, Shajari M, Kohnen T, et al. 2018 Comparison of fibrotic response in the human lens capsular bag after femtosecond laser–assisted cataract surgery and conventional phacoemulsification. J. Cataract Refract. Surg. 44 750–755

    Article  Google Scholar 

  • Wormstone IM and Eldred JA 2016 Experimental models for posterior capsule opacification research. Exp. Eye Res. 142 2–12

    Article  CAS  Google Scholar 

  • Wu K, Zeng J, Li L, et al. 2010 Silibinin reverses epithelial-to-mesenchymal transition in metastatic prostate cancer cells by targeting transcription factors. Oncol. Rep. 23 1545–1552

    Article  CAS  Google Scholar 

  • Wu K, Ning Z, Zeng J, et al. 2013 Silibinin inhibits β-catenin/ZEB1 signaling and suppresses bladder cancer metastasis via dual-blocking epithelial-mesenchymal transition and stemness. Cell. Signal. 25 2625–2633

    Article  CAS  Google Scholar 

  • Xu S, Zhang H, Wang A, et al. 2020 Silibinin suppresses epithelial-mesenchymal transition in human non-small cell lung cancer cells by restraining RHBDD1. Cell. Mol. Biol. Lett. 25 36

    Article  CAS  Google Scholar 

  • Yao K, Ye PP, Tan J, Tang XJ and Shen Tu XC 2008 Involvement of PI3K/Akt pathway in TGF-β2-mediated epithelial mesenchymal transition in human lens epithelial cells. Ophthal. Res. 40 69–76

    Article  CAS  Google Scholar 

  • Zhang YE 2017 Non-Smad signaling pathways of the TGF-β family. Cold Spring Harb. Perspect. Biol. 9 a022129

    Article  Google Scholar 

  • Zhang J, Shemezis JR, McQuinn ER, et al. 2013 AKT activation by N-cadherin regulates beta-catenin signaling and neuronal differentiation during cortical development. Neural Dev. 8 7

    Article  CAS  Google Scholar 

  • Zhu XX, Ding YH, Wu Y, et al. 2016 Silibinin: a potential old drug for cancer therapy. Expert Rev. Clin. Pharmacol. 9 1323–1330

    Article  CAS  Google Scholar 

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Acknowledgements

This work was a part of PBM’s PhD programme, Iladevi Cataract and IOL Research Centre, Ahmedabad, India, and Manipal Academy of Higher Education, Manipal, India, and the authors acknowledge Indian Council of Medical Research, India, for providing Senior Research Fellowship (BMS/Nan/2020_6123) to PBM.

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Authors and Affiliations

  1. Department of Cell and Molecular Biology, Iladevi Cataract and IOL Research Centre, Ahmedabad, India

    Pooja B Malaviya, Dhaval K Shukal, Tusha Sharma, Abhay R Vasavada & Kaid Johar SR

  2. Manipal Academy of Higher Education, Manipal, India

    Pooja B Malaviya & Dhaval K Shukal

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  1. Pooja B Malaviya
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  2. Dhaval K Shukal
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  3. Tusha Sharma
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  4. Abhay R Vasavada
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  5. Kaid Johar SR
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Correspondence to Kaid Johar SR.

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Communicated by Dipankar Nandi.

Corresponding editor: Dipankar Nandi

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Malaviya, P.B., Shukal, D.K., Sharma, T. et al. Silibinin suppresses TGFβ2-induced lens epithelial cell migration and epithelial–mesenchymal transition. J Biosci 47, 51 (2022). https://doi.org/10.1007/s12038-022-00286-3

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