Sinusoidal wavy surfaces for curvature-guided migration of T lymphocytes
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 . 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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2019, Biophysical JournalCitation Excerpt :More recent findings showed that cell migration is also directed by purely mechanical cues, such as stiffness gradients (durotaxis) (15), topographic pattern gradients (topotaxis) (16), and substrate anisotropy (ratchetaxis) (17–19). Lastly, microscopic curved topographies have gained greater interest for their resemblance with smooth biological tissues (20–23). A recurrent factor in these directed migration mechanisms is the predominant role of the nucleus (24,25).
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These authors contributed equally to the work.