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.

View original source ↗Export metadata ↓

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

Explore related subjects

bioengineering Preprint

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

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

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Wall stiffening is a primary contributor to motility loss in Crohn's disease: an electromechanical modeling study

Fibrotic strictures are among the most disabling complications of Crohn's disease, permanently narrowing the bowel and impairing motility, yet no approved therapy reverses them. Chronic inflammation alters pacemaker-network coupling, smooth-muscle excitability, and calcium-dependent contractility, while fibrosis thickens the bowel wall, narrows the lumen, and changes tissue mechanics. The relative contributions of these coupled electrical, contractile, and structural alterations to motility loss remain unclear. To address this gap, we develop an integrated electromechanical finite-element framework for fibrostenosing Crohn's disease that couples a fibrosis-driven growth model with a FitzHugh-Nagumo electromechanical model. A full-factorial 25 design of experiments is used to quantify the relative effects of electrical diffusivity, excitation threshold, peak active stress, wall stiffness, and hypertrophic remodeling on cyclic lumen-volume deformation. Motility is quantified by the standard deviation of lumen volume over one contraction cycle. Within the parameter ranges examined, increased wall stiffness emerged as the dominant contributor to motility loss, followed by impaired smooth-muscle contractility. Changes in excitation threshold, hypertrophic remodeling, and electrical diffusivity produced substantially smaller effects. Pairwise interactions were small relative to the dominant main effects, indicating that the mechanisms contributed largely through their individual effects. Our findings suggest that limiting wall stiffening while preserving smooth-muscle contractile function may provide a therapeutic strategy for maintaining intestinal motility in fibrostenosing Crohn's disease.

bioengineering↗

bioRxiv2026

Lactate Receptor Activation Alleviates Senescence and Preserves Homeostasis of Aged Arteries

Arteries are among the first tissues to exhibit age-related dysfunction, yet the metabolic mechanisms driving vascular senescence remain poorly understood. Here, analysis of human aortic transcriptomic data identified HCAR1, encoding the lactate receptor GPR81, as one of the genes most significantly downregulated with age. We therefore investigated whether age-associated loss of GPR81 contributes to cellular senescence within the vessel wall. Senescent human endothelial cells and vascular smooth muscle cells accumulated neutral and oxidized lipids and exhibited increased labile iron and ferroptosis. Silencing GPR81 in early-passage cells recapitulated this metabolic phenotype together with multiple hallmarks of cellular senescence. Moreover, endothelial-specific deletion of GPR81 in young mice was sufficient to induce senescent cell accumulation, impaired lipid homeostasis, endothelial dysfunction, and elastin disorganization. Conversely, pharmacological activation of GPR81 with the agonist CHBA restored fatty acid metabolism, promoted glycolytic reprogramming, and attenuated ferroptotic stress and senescence-associated phenotypes. In lamin A knock-in (LAKI) progeroid mice, CHBA reduced arterial lipid accumulation and cellular senescence, shifted vascular cell composition toward a youthful state, improved endothelial integrity, and restored extracellular matrix homeostasis. Together, these findings identify age-associated loss of GPR81 as a driver of vascular metabolic dysfunction and cellular senescence and establish pharmacological GPR81 activation as a promising therapeutic strategy for preserving vascular homeostasis and mitigating age-associated cardiovascular disease.

bioengineering↗

bioRxiv2026

Targeted and bilateral blood flow monitoring in middle cerebral artery using diffuse correlation spectroscopy

Objective: To develop and validate a dual-probe Diffuse Correlation Spectroscopy (DCS) system for non-invasive and simultaneous, monitoring of cerebral blood flow (CBF) in the bilateral Middle Cerebral Artery (MCA) territories, and expanding the utility of conventional DCS limited to cortical-volume-based CBF measurements to vessel-specific cerebral perfusion monitoring. Methods: A dual-probe DCS system was designed for non-invasive monitoring of MCA-specific perfusion. Probe placement and protocol optimization study has been performed using anatomical landmarks, motor and speech activation tasks in healthy volunteers. System stability and repeatability were further evaluated in a pilot cohort of 30 healthy (age, 25{+/-}7 years) participants using optimized probe position and protocol. A bilateral MCA ischemic Lacunar Infract stroke case report also validated the feasibility of the system in clinical settings. Results: Measurements demonstrated superior sensitivity towards MCA-territory perfusion at targeted probe locations compared to off-MCA positions. In pilot cohort, significant increase of 30.34 {+/-} 21.56% and 36.48 {+/-} 21.22% in rCBF corresponding to hand squeeze and speech task respectively showed reproducible physiological responsiveness of the system (p<0.001). Measurement done on a patient with bilateral MCA ischemic Lacunar Infract stroke showed a significant change of 30% during speech for both the MCAs but no significant change is observed for hand squeeze tasks. Conclusion: The custom built dual-probe DCS system enables non-invasive, operator-independent, targeted and continuous monitoring of rCBF within bilateral MCA territories. Significance: This approach enables the potential use of DCS system for bilateral and vessel-specific monitoring of cerebral perfusion in the MCA territories.

bioengineering↗