Elsevier

Biomaterials

Volume 51, May 2015, Pages 151-160
Biomaterials

Sinusoidal wavy surfaces for curvature-guided migration of T lymphocytes

https://doi.org/10.1016/j.biomaterials.2015.01.071Get rights and content

Abstract

Micro/nanofabricated surfaces have been widely used for the study of topography-guided migration of cells. While the current studies mostly utilized micro/nanostructures containing sharp edges, internal tissues guiding migration of cells such as blood and lymphatic vessels, bone cavities, perivascular tracks have smooth microscale topographical structures. To overcome these limitations, we fabricated sinusoidal wavy surfaces with various wavelengths by deep X-ray lithography enabling precise and simultaneous control of amplitudes and wavelengths. Using these surfaces, we systematically studied curvature-guided migration of T cells. The majority of T cells migrated along the concave surfaces of sinusoidal wavy structures and as wavelength increased (or curvature decreased), preference to concave surfaces decreased. Integrin-mediated adhesion augmented the tendency of T cells crawling along grooves of highly curved wavy surfaces. To understand mechanisms of curvature-guided migration of T cells, T cells were treated with small molecule drugs such as blebbistatin and CK636, inhibiting myosin II activity and lamellipodia formation, respectively. While lamellipodia-inhibited T cells frequently crossed ridges, myosin II-inhibited T cells were mostly confined within concave surfaces. These results suggest that lamellipodia regulate local actin polymerization in response to surface curvature to maintain T cells within concave surfaces while myosin II-mediated contractile forces push T cells out of concave surfaces to make T cells less sensitive to surface curvature.

Introduction

Migration of cells guided by various micro/nanoscale topographical structures in vivo has been observed by intravital microscopy during immune surveillances/responses [1], [2] and cancer invasion/metastasis [3], [4], but the mechanisms by which such structures guide migration of cells have not been completely understood [5], [6]. Micro/nanofabricated surfaces containing well-defined topographical structures have been useful for the study of topography-guided migration of cells [7], [8], [9], [10]. Most of these microfabricated structures had sharp edges [7], [8], [9], but tissue structures known to guide migration of cells such as blood and lymphatic vessels, bone cavities and perivascular tracks have smooth microscale topography [11], [12], [13].

Various methods such as laser interference lithography [14], [15] and spontaneous buckling [16], [17] or wrinkling of elastomers [18], [19] have been applied to fabricate sinusoidal wavy structures, but precise and simultaneous control of wavelength λ and of amplitude A of wavy structures has been challenging. To overcome these limitations, we used deep X-ray lithography (DXRL) based on synchrotron radiation to fabricate sinusoidal wavy structures with λ = 20, 40, 80, and 160 μm and A = 10 μm. DXRL allows fabrication of features with height up to several millimeters with excellent sidewall quality [20], [21], so sinusoidal wavy structures with height ∼1 mm were fabricated. By orienting vertically-fabricated sinusoidal wavy structures horizontally and replicating them using polymers, we generated sinusoidal wavy topographical surfaces with precisely-controlled λ and fixed A. Using these surfaces, we systematically studied curvature-guided migration of T cells. T cells are immune cells that migrate virtually everywhere in the body to orchestrate cell-mediated immune responses; thus understanding motility of T cells under complex microenvironments is essential for designing therapeutic strategies against various immune-related diseases [22], [23], [24].

Section snippets

Fabrication of sinusoidal wavy surfaces

To fabricate substrates containing sinusoidal waves, deep X-ray LIGA (German acronym for lithographie, galvanoformung, abformung, or lithography, electroforming, and molding) process was used as previously described [25], [26]. Briefly, a 1.1-mm-thick poly(methyl methacrylate) (PMMA) template with step-gradient periods of 20, 40, 80, and 160 μm with an amplitude of 10 μm was prepared by deep X-ray lithography (Fig. 2A(i)). An X-ray mask was fabricated by standard UV-photolithography followed by

Design and fabrication of sinusoidal wavy surfaces with various wavelengths

To fabricate sinusoidal wavy structures with various λ, X-ray lithography, electroforming and molding (German acronym LIGA) processes were used [25], [26]. An X-ray mask containing sinusoidal wavy patterns with λ = 20, 40, 80 and 160 μm and A = 10 μm (Fig. 1A) was prepared by standard photolithography and gold electroplating. The curvature of a sinusoidal wave A cos2π/λx, is (2πλ)2Acos2πλx/(1+((2πλ)Asin2πλ)2)3/2. Since curvature is a function of both λ and A, varying λ over a large range of

Discussion

Migration of cells guided by topographical structures has been previously studied using micro/nanofabricated surfaces [7], [8], but these studies have used structures with sharp edges due to difficulties in fabrication of smooth structures with well defined geometries; structures with sharp edges may not be physiologically relevant. To address this limitation, we devised a new method to fabricate sinusoidal wavy surfaces with precisely controlled λ and A First, sinusoidal wavy structures with

Conclusions

In summary, sinusoidal wavy surfaces with precisely controlled amplitude and wavelength were successfully fabricated by deep X-ray LIGA followed by UV-assisted CFL. Using these surfaces with well-controlled curvatures, effects of surface curvature on the motility of T cells were systematically investigated. The functions of lamellipodia and myosin II on curvature-guided migration of T cells were also investigated. The method developed in this study will be useful to study how surface curvature

Acknowledgments

This work was supported by the National Research Foundation of Korea (NRF) grants funded by the Korea government (MSIP) (Grant No. 2012-004146 to JD and Grant No. 2014R1A2A1A01006527 and No.2011-0030075 to DSK and Brain Korea 21 Plus Program to JD and DSK).

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    These authors contributed equally to the work.

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