Regulation of cell morphology and viability using anodic aluminum oxide with custom-tailored structural parameters

doi:10.1007/s11706-022-0622-8

Front. Mater. Sci.

2022, Vol. 16

Issue (4) : 220622 https://doi.org/10.1007/s11706-022-0622-8

RESEARCH ARTICLE

Regulation of cell morphology and viability using anodic aluminum oxide with custom-tailored structural parameters

Zhiying ZHANG¹, Ting LIU¹, Juan LI¹(

), Yiyan GUO¹, Ruiqing LIANG¹, Jiangbo LU¹, Runguang SUN¹(

), Jun DONG²(

)

¹. School of Physics and Information Technology, Shaanxi Normal University, Xi’an 710119, China
². Department of Orthopaedics, Second Affiliated Hospital of Xi’an Jiaotong University, Xi’an 710004, China

Download: PDF(7873 KB) HTML
Export: BibTeX | EndNote | Reference Manager | ProCite | RefWorks

Abstract

Anodic aluminum oxide (AAO) with independently controlled period, porosity, and height is used as the model surface to study the single structural parameter effect on breast cancer cell behaviors, including cell polarity and cell viability. It is found that the quantity of multipolar cells and cell viability increases as the nanodent period increases from 100 to 300 nm, while the number of bipolar cells has almost no change until there is a dramatic decrease as the period increases to 300 nm. After anodizing nanodents into nanopores, the numbers of both bipolar cells and the cell viability increase significantly with the porosity increase. However, as the porosity further increases and the nanopore changes into a nanocone pillar, most of the cells become nonpolar spheres and the cell viability decreases. Increasing the height of the nanocone pillar has little effect on the cell polarity; the cell viability increases slightly with the increase of the nanocone pillar height. These results reveal the influence of individual nanostructure parameters on the cell behavior, especially the cell polarity and the cell viability, which can help to design the surface to make the cell grow as desired.

Keywords nanostructure parameter breast cancer cell cell morphology cell polarity cell viability

Corresponding Author(s): Juan LI,Runguang SUN,Jun DONG

Issue Date: 25 November 2022

Cite this article:

Zhiying ZHANG,Ting LIU,Juan LI, et al. Regulation of cell morphology and viability using anodic aluminum oxide with custom-tailored structural parameters[J]. Front. Mater. Sci., 2022, 16(4): 220622.

URL:

https://academic.hep.com.cn/foms/EN/10.1007/s11706-022-0622-8
https://academic.hep.com.cn/foms/EN/Y2022/V16/I4/220622

Fig.1 Schematic diagram of the electrochemical experiment reactor.

Tab.1 Detailed experimental conditions for fabricating the self-ordered nanostructures with different structure parameters

Tab.2 Independently adjustable structure parameters of as-fabricated self-ordered nanostructures

Fig.2 Top row: SEM images of aluminum films with different period nanodent of (a) 0 nm (plane), (b) 100 nm (D₁₀₀), (c) 200 nm (D₂₀₀), and (d) 300 nm (D₃₀₀). Middle row: SEM images of MDA-MB-231 cells culture for 48 h on the corresponding aluminum film. Bottom row: details of the pseudopodia of the corresponding cell.

Fig.3 Fluorescent images of MDA-MB-231 cells cultured for 48 h on (a) the plane aluminum film, (b) the aluminum film with 100-nm period nanodent, (c) the aluminum film with 200-nm period nanodent, and (d) the aluminum film with 300-nm period nanodent, where cell cytoskeletons are stained by FITC-phalloidin (left column), cell nuclei are stained by DAPI (middle column), and merged images of the above two (right column). (e) The ratio of multipolar cells (left slash), bipolar cells (right slash), and nonpolar cells (blank) cultured for 48 h on aluminum films with 100-nm (D₁₀₀), 200-nm (D₂₀₀), and 300-nm (D₃₀₀) period nanodent, respectively, and the values on the plane aluminum film is also shown for comparison. (f) The cell viability on nanodent structured aluminum films with different periods, which are cultured for 24, 48, 72, and 96 h. The viability of cells grown on the plane surface is used as comparison, and the viability of cells cultured for 24 h on the plane surface was denoted as 100%. The statistical signification was marked as *p < 0.05, **p < 0.01.

Fig.4 Top row: SEM top views of AAO with the same pore diameter of 80 nm and the same pore depth of 370 nm, while their periods are (a) 100 nm, (b) 200 nm, and (c) 300 nm, which are denoted by P_D100, P_D200, and P_D300 on the top right, respectively (the insert images are corresponding side views of the AAO). Bottom row: SEM images of MDA-MB-231 cell cultures for 48 h on P_D100, P_D200, and P_D300 (the insert images show details of the pseudopodia of corresponding cells).

Fig.5 Fluorescent images of MDA-MB-231 cells cultured for 48 h on (a) plane aluminum film, (b) P_D100, (c) P_D200, and (d) P_D300, where cell cytoskeletons are stained by FITC-phalloidin (left column), cell nucleus are stained by DAPI (middle column), and merged images of the above two (right column). (e) The ratio of multipolar cells (left slash), bipolar cells (right slash), and nonpolar cells (blank) cultured for 48 h on the AAO with the same pore diameter and the same pore depth, but with different nanopore periods of 100 nm (P_D100), 200 nm (P_D200), and 300 nm (P_D300), respectively, and the value on the plane aluminum film is also shown for comparison. (f) The cell viability on AAO with different nanopore periods, cultured for 24, 48, 72, and 96 h. The viability of cells grown on the plane surface is used as comparison, and the viability of cells cultured for 24 h on the plane surface was denoted as 100%. The statistical signification was marked as **p < 0.01.

Fig.6 Top row: SEM top views of nanopore structured AAO with period of 300 nm whose pore widening time are (a) 0 min, (b) 90 min, (c) 150 min, and (d) 160 min, which are denoted as P_D300, PW1, PW2, and PW3, respectively, and the pore diameters are 80, 150, and 250 nm for P_D300, PW1, and PW2, respectively. For PW3, as the pore widening time increases, the nanopore structure disappears and only the nanocone pillar is left. The insert images are corresponding side views of AAOs, clearly showing that these AAOs also have the same depth of 370 nm. Bottom row: SEM images of MDA-MB-231 cells cultured on P_D300, PW1, PW2, and PW3 for 48 h.

Fig.7 Fluorescent images of MDA-MB-231 cells cultured for 48 h on (a) P_D300, (b) PW1, (c) PW2, and (d) PW3, where cell cytoskeletons are stained by FITC-phalloidin (left column), cell nucleus are stained by DAPI (middle column), and merged images of the above two (right column). (e) The ratio of multipolar cells (left slash), bipolar cells (right slash), and the nonpolar cells (blank) cultured for 48 h on P_D300, PW1, PW2, and PW3, respectively. (f) The cell viability on P_D300, PW1, PW2, and PW3, which are cultured for 24, 48, 72, and 96 h, respectively. The viability of cells grown on plane surface is used as comparison, and the viability of cells cultured for 24 h on the plane surface is denoted as 100%. The statistical signification was marked as **p < 0.01.

Fig.8 Top row: side-view SEM images of nanocone pillar structured AAOs with different heights of (a) 370 nm (h₃₇₀), (b) 550 nm (h₅₅₀), and (c) 650 nm (h₆₅₀). Bottom row: SEM images of MDA-MB-231 cells cultured for 48 h on the corresponding AAO (the inserts are the representative cells).

Fig.9 Fluorescent images of MDA-MB-231 cells cultured for 48 h on (a) 370 nm-height, (b) 550 nm-height, and (c) 650 nm-height nanocone pillar structured AAOs, where cell cytoskeletons are stained by FITC-phalloidin (left column), cell nuclei are stained by DAPI (middle column), and merged images of above two (right column). (d) The ratio of multipolar cells (left slash), bipolar cells (right slash), and nonpolar cells (blank), which are cultured for 48 h on the nanocone pillar with heights of 370 nm (h₃₇₀), 550 nm (h₅₅₀), and 650 nm (h₆₅₀). (e) The cell viability on 370 nm-, 550 nm-, and 650 nm-height nanocone pillars cultured for 24, 48, 72, and 96 h. The viability of cells grown on plane surface is used as comparison, and the viability of cells cultured for 24 h on the plane surface is denoted as 100%. The statistical signification is marked as *p < 0.05, **p < 0.01.

Tab.3 Effect of nanostructure with independently controlled parameters on cell behaviors

1	P, Netti M Ventre . Cell Instructive Materials to Control and Guide Cell Function: Programmable Bioactive Interfaces. Woodhead Publishing, 2021
2	S, Cai C, Wu W, Yang et al.. Recent advance in surface modification for regulating cell adhesion and behaviors.Nanotechnology Reviews, 2020, 9(1): 971–989 https://doi.org/10.1515/ntrev-2020-0076
3	C J, Bettinger R, Langer J T Borenstein . Engineering substrate topography at the micro- and nanoscale to control cell function.Angewandte Chemie International Edition, 2009, 48(30): 5406–5415 https://doi.org/10.1002/anie.200805179 pmid: 19492373
4	J, Zhou X, Zhang J, Sun et al.. The effects of surface topography of nanostructure arrays on cell adhesion.Physical Chemistry Chemical Physics, 2018, 20(35): 22946–22951 https://doi.org/10.1039/C8CP03538E pmid: 30155544
5	A T, Nguyen S R, Sathe E K F Yim . From nano to micro: topographical scale and its impact on cell adhesion, morphology and contact guidance.Journal of Physics: Condensed Matter, 2016, 28(18): 183001 https://doi.org/10.1088/0953-8984/28/18/183001 pmid: 27066850
6	C, Leclech C Villard . Cellular and subcellular contact guidance on microfabricated substrates.Frontiers in Bioengineering and Biotechnology, 2020, 8: 551505 https://doi.org/10.3389/fbioe.2020.551505 pmid: 33195116
7	B A, Dalton X F, Walboomers M, Dziegielewski et al.. Modulation of epithelial tissue and cell migration by microgrooves.Journal of Biomedical Materials Research, 2001, 56(2): 195–207 https://doi.org/10.1002/1097-4636(200108)56:2<195::AID-JBM1084>3.0.CO;2-7 pmid: 11340589
8	Y, Zhu X, Liu J, Wu et al.. Micro- and nanohemispherical 3D imprints modulate the osteogenic differentiation and mineralization tendency of bone cells.ACS Applied Materials & Interfaces, 2019, 11(39): 35513–35524 https://doi.org/10.1021/acsami.9b05521 pmid: 31507175
9	Y, Song Y, Ju G, Song et al.. In vitro proliferation and osteogenic differentiation of mesenchymal stem cells on nanoporous alumina.International Journal of Nanomedicine, 2013, 8: 2745–2756 pmid: 23935364
10	J, Agudo-Canalejo D E Discher . Biomembrane adhesion to substrate topographically pattered with nanopits.Biophysical Journal, 2018, 115(7): 1292–1306 https://doi.org/10.1016/j.bpj.2018.08.006
11	X, Wu L, Li L, Wang et al.. Cell spreading behaviors on hybrid nanopillar and nanohole arrays.Nanotechnology, 2022, 33(4): 045101 https://doi.org/10.1088/1361-6528/ac084a pmid: 30485249
12	Y, Yang K, Wang X, Gu et al.. Biophysical regulation of cell behavior — cross talk between substrate stiffness and nanotechnology.Engineering, 2017, 3(1): 36–54 https://doi.org/10.1016/J.ENG.2017.01.014 pmid: 29071164
13	Y, Li Y, Xiao C Liu . The horizon of materiobiology: a perspective on material-guided cell behaviors and tissue engineering.Chemical Reviews, 2017, 117(5): 4376–4421 https://doi.org/10.1021/acs.chemrev.6b00654 pmid: 28221776
14	C, Gao S, Ito A, Obata et al.. Fabrication and in vitro characterization of electrospun poly (γ-glutamic acid)–silica hybrid scaffolds for bone regeneration.Polymer, 2016, 91: 106–117 https://doi.org/10.1016/j.polymer.2016.03.056
15	V S Saji . Supramolecular organic nanotubes for drug delivery.Materials Today Advances, 2022, 14: 100239 https://doi.org/10.1016/j.mtadv.2022.100239
16	V S, Saji H C, Choe K W K Yeung . Nanotechnology in biomedical applications: a review.International Journal of Nano and Biomaterials, 2010, 3(2): 119–139 https://doi.org/10.1504/IJNBM.2010.037801
17	V S, Saji T, Kumeria K, Gulati et al.. Localized drug delivery of selenium (Se) using nanoporous anodic aluminium oxide for bone implants.Journal of Materials Chemistry B: Materials for Biology and Medicine, 2015, 3(35): 7090–7098 https://doi.org/10.1039/C5TB00125K pmid: 32262711
18	G E Thompson . Porous anodic alumina: fabrication, characterization and applications.Thin Solid Films, 1997, 297(1–2): 192–201 https://doi.org/10.1016/S0040-6090(96)09440-0
19	S, Rahman R, Ormsby A, Santos et al.. Nanoengineered drug-releasing aluminium wire implants: comparative investigation of nanopore geometry, drug release and osteoblast cell adhesion.RSC Advances, 2015, 5(92): 75004–75014 https://doi.org/10.1039/C5RA10418A
20	G, Rajeev Simon B, Prieto L F, Marsal et al.. Advances in nanoporous anodic alumina-based biosensors to detect biomarkers of clinical significance: a review.Advanced Healthcare Materials, 2018, 7(5): 1700904 https://doi.org/10.1002/adhm.201700904 pmid: 29205934
21	A M M, Jani D, Losic N H Voelcker . Nanoporous anodic aluminium oxide: advances in surface engineering and emerging applications.Progress in Materials Science, 2013, 58(5): 636–704 https://doi.org/10.1016/j.pmatsci.2013.01.002
22	A, Ruiz-Clavijo O, Caballero-Calero M Martín-González . Revisiting anodic alumina templates: from fabrication to applications.Nanoscale, 2021, 13(4): 2227–2265 https://doi.org/10.1039/D0NR07582E pmid: 33480949
23	L, Zajączkowska M Norek . Peculiarities of aluminum anodization in AHAs-based electrolytes: case study of the anodization in glycolic acid solution.Materials, 2021, 14(18): 5362 https://doi.org/10.3390/ma14185362 pmid: 34576586
24	A M M, Jani A S, Habiballah M Z B A, Halim et al.. Nanoporous anodic aluminum oxide (NAAO) for catalytic, biosensing and template synthesis applications (a review).Current Nanoscience, 2019, 15(1): 49–63 https://doi.org/10.2174/1573413714666180308145336
25	Z, Tang D, Zhang W, Cui et al.. Fabrications, applications and challenges of solid-state nanopores: a mini review.Nanomaterials and Nanotechnology, 2016, 6: 35 https://doi.org/10.5772/64015
26	S, Liu J, Tian W Zhang . Fabrication and application of nanoporous anodic aluminum oxide: a review.Nanotechnology, 2021, 32(22): 222001 https://doi.org/10.1088/1361-6528/abe25f pmid: 33530076
27	S Ono . Nanostructure analysis of anodic films formed on aluminum-focusing on the effects of electric field strength and electrolyte anions.Molecules, 2021, 26(23): 7270 https://doi.org/10.3390/molecules26237270 pmid: 34885861
28	D Brüggemann . Nanoporous aluminium oxide membranes as cell interfaces.Journal of Nanomaterials, 2013, 2013: 460870 https://doi.org/10.1155/2013/460870
29	G O, Kim H, Lee E, Ma et al.. Viability studies of cells on nanostructured surfaces with various feature sizes.Bulletin of the Korean Chemical Society, 2017, 38(12): 1447–1454 https://doi.org/10.1002/bkcs.11325
30	J S, Park D, Moon J S, Kim et al.. Cell adhesion and growth on the anodized aluminum oxide membrane.Journal of Biomedical Nanotechnology, 2016, 12(3): 575–580 https://doi.org/10.1166/jbn.2016.2192 pmid: 27280255
31	G E J, Poinern X T, Le M, O’Dea et al.. Chemical synthesis, characterisation, and biocompatibility of nanometre scale porous anodic aluminium oxide membranes for use as a cell culture substrate for the Vero cell line: a preliminary study.BioMed Research International, 2014, 2014: 238762 https://doi.org/10.1155/2014/238762 pmid: 24579077
32	F, Mussano T, Genova F G, Serra et al.. Nano-pore size of alumina affects osteoblastic response.International Journal of Molecular Sciences, 2018, 19(2): 528 https://doi.org/10.3390/ijms19020528 pmid: 29425177
33	S, Nasrollahi S, Banerjee B, Qayum et al.. Nanoscale matrix topography influences microscale cell motility through adhesions, actin organization, and cell shape.ACS Biomaterials Science & Engineering, 2017, 3(11): 2980–2986 https://doi.org/10.1021/acsbiomaterials.6b00554 pmid: 33418718
34	X, Che J, Boldrey X, Zhong et al.. On-chip studies of magnetic stimulation effect on single neural cell viability and proliferation on glass and nanoporous surfaces.ACS Applied Materials & Interfaces, 2018, 10(34): 28269–28278 https://doi.org/10.1021/acsami.8b05715 pmid: 30080968
35	Y, Ma Y, Wen J, Li et al.. Fabrication of self-ordered alumina films with large interpore distance by Janus anodization in citric acid.Scientific Reports, 2016, 6(1): 39165 https://doi.org/10.1038/srep39165 pmid: 27958365
36	J, Li C, Li C, Chen et al.. Facile method for modulating the profiles and periods of self-ordered three-dimensional alumina taper-nanopores.ACS Applied Materials & Interfaces, 2012, 4(10): 5678–5683 https://doi.org/10.1021/am301603e pmid: 23020550
37	C, Feng Z, Zhang J, Li et al.. A bioinspired, highly transparent surface with dry-style antifogging, antifrosting, antifouling, and moisture self-cleaning properties.Macromolecular Rapid Communications, 2019, 40(6): 1800708 https://doi.org/10.1002/marc.201800708 pmid: 30468541
38	J, Lim A, Choi H W, Kim et al.. Constrained adherable area of nanotopographic surfaces promotes cell migration through the regulation of focal adhesion via focal adhesive kinase/Racl activation.ACS Applied Materials & Interfaces, 2018, 10(17): 14331–14341 https://doi.org/10.1021/acsami.7b18954 pmid: 29649358
39	Y, Song Y, Ju Y, Morita et al.. Effect of the nanostructure of porous alumina on growth behavior of MG63 osteoblast-like cells.Journal of Bioscience and Bioengineering, 2013, 116(4): 509–515 https://doi.org/10.1016/j.jbiosc.2013.04.007 pmid: 23643619
40	Maat J, ter R, Regeling C J, Ingham et al.. Organic modification and subsequent biofunctionalization of porous anodic alumina using terminal alkynes.Langmuir, 2011, 27(22): 13606–13617 https://doi.org/10.1021/la203738h pmid: 21962228
41	E, Kolos A J Ruys . Biomimetic coating on porous alumina for tissue engineering: characterisation by cell culture and confocal microscopy.Materials, 2015, 8(6): 3584–3606 https://doi.org/10.3390/ma8063584

Viewed

Full text

Abstract

Cited

Shared

Discussed