Patterning Fluid Shear Stress Landscapes with Multiphoton Inner Laser Lithography (MILL) for Live Cell Adhesion and Translocation

Lim, Y. J.; Zhang, J.; Lin, H.; Xu, T.; Li, Y.; Zhang, Z.; Hicks, S. M.; Chudinov, I.; Nechipurenko, D. Y.; Gardiner, E. E.; Lee, S.

doi:10.1101/2022.06.17.496569

bioRxiv · 10.1101/2022.06.17.496569

Patterning Fluid Shear Stress Landscapes with Multiphoton Inner Laser Lithography (MILL) for Live Cell Adhesion and Translocation

Lim, Y. J.·Zhang, J.·Lin, H.·Xu, T.·Li, Y.·Zhang, Z.·Hicks, S. M.·Chudinov, I.·Nechipurenko, D. Y.·Gardiner, E. E.·Lee, S.

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Abstract

Heterogenous fluid shear stress is known to provide mechanical cues for cell adhesion and translocation. To assemble 3D microstructures using current fabrication methods in a single channel and recapitulate in vivo heterogenous fluid flow would require hours of fabrication and specialized equipment. Inspired by the traditional art form of inside painting, we developed a technique for 3D fabrication of micro-patterned flow channels and mixed in vivo fluid flow in a matter of minutes. We termed this technique Multiphoton Inner Laser Lithography (MILL). We further showed that when combined with adaptive optics, MILL is compatible with both flat and curved channel shapes. MILL recapitulated in vivo tissue topology and 3D fluid flow within tissue stroma (low fluid shear, 0 - 3.5 dynes/cm2) and blood vessel (high fluid shear, 0 - 80 dynes/cm2). We demonstrate fibroblast cell and platelets adhere and translocate differently between laminar flow patterns that are homogenous versus heterogeneous in real time. Parallel strips of MILL channels were assembled for simultaneous platelet function test to quantify the efficacy of an antithrombotic GPVI Fab (~2000 microthrombi per test). The MILL technique can be readily reproduced in vivo fluid flow in minutes and benefit preclinical screening of drug pharmacokinetics. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=98 SRC="FIGDIR/small/496569v2_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@1efbe9eorg.highwire.dtl.DTLVardef@cbab5aorg.highwire.dtl.DTLVardef@15ed255org.highwire.dtl.DTLVardef@42d14e_HPS_FORMAT_FIGEXP M_FIG C_FIG Significant pointsO_LIMILL channels are made from commercially available off the shelf components using a standard multiphoton imaging system C_LIO_LIVarying degrees of in vivo heterogenous laminar flow is shown to directly influence cells translocating on thinly coated collagen surfaces C_LIO_LIParallel strips of MILL channels were assembled for simultaneous platelet function tests (~2000 microthrombi per test). C_LI

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Lim, Y. J., Zhang, J., Lin, H., Xu, T., Li, Y., Zhang, Z., Hicks, S. M., Chudinov, I., Nechipurenko, D. Y., Gardiner, E. E., Lee, S.. 2022-06-19. Patterning Fluid Shear Stress Landscapes with Multiphoton Inner Laser Lithography (MILL) for Live Cell Adhesion and Translocation. https://doi.org/10.1101/2022.06.17.496569

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