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1. Three-dimensional microscale hanging drop arrays with geometric control for drug screening and live tissue imaging

   https://doi.org/10.1126/sciadv.abc1323  

   Ganguli, A. ; Mostafa, A. ; Saavedra, C. ; Kim, Y. ; Le, P. ; Faramarzi, V. ; Feathers, R. W. ; Berger, J. ; Ramos-Cruz, K. P. ; Adeniba, O. ; et al ( April 2021 , Science Advances)

   null (Ed.)

   Existing three-dimensional (3D) culture techniques are limited by trade-offs between throughput, capacity for high-resolution imaging in living state, and geometric control. Here, we introduce a modular microscale hanging drop culture where simple design elements allow high replicates for drug screening, direct on-chip real-time or high-resolution confocal microscopy, and geometric control in 3D. Thousands of spheroids can be formed on our microchip in a single step and without any selective pressure from specific matrices. Microchip cultures from human LN229 glioblastoma and patient-derived mouse xenograft cells retained genomic alterations of originating tumors based on mate pair sequencing. We measured response to drugs over time with real-time microscopy on-chip. Last, by engineering droplets to form predetermined geometric shapes, we were able to manipulate the geometry of cultured cell masses. These outcomes can enable broad applications in advancing personalized medicine for cancer and drug discovery, tissue engineering, and stem cell research. 

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2. Deterministic Single Cell Encapsulation in Asymmetric Microenvironments to Direct Cell Polarity

   https://doi.org/10.1002/advs.202206014  

   Cho, Ik Sung ; Gupta, Prerak ; Mostafazadeh, Nima ; Wong, Sing Wan ; Saichellappa, Saiumamaheswari ; Lenzini, Stephen ; Peng, Zhangli ; Shin, Jae‐Won ( December 2022 , Advanced Science)

   Various signals in tissue microenvironments are often unevenly distributed around cells. Cellular responses to asymmetric cell‐matrix adhesion in a 3D space remain generally unclear and are to be studied at the single‐cell resolution. Here, the authors developed a droplet‐based microfluidic approach to manufacture a pure population of single cells in a microscale layer of compartmentalized 3D hydrogel matrices with a tunable spatial presentation of ligands at the subcellular level. Cells elongate with an asymmetric presentation of the integrin adhesion ligand Arg‐Gly‐Asp (RGD), while cells expand isotropically with a symmetric presentation of RGD. Membrane tension is higher on the side of single cells interacting with RGD than on the side without RGD. Finite element analysis shows that a non‐uniform isotropic cell volume expansion model is sufficient to recapitulate the experimental results. At a longer timescale, asymmetric ligand presentation commits mesenchymal stem cells to the osteogenic lineage. Cdc42 is an essential mediator of cell polarization and lineage specification in response to asymmetric cell‐matrix adhesion. This study highlights the utility of precisely controlling 3D ligand presentation around single cells to direct cell polarity for regenerative engineering and medicine.

    

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3. Probing Single-Cell Mechanical Allostasis Using Ultrasound Tweezers

   https://doi.org/10.1007/s12195-019-00578-z  

   Qian, Weiyi ; Chen, Weiqiang ( January 2019 , Cellular and Molecular Bioengineering)

   Introduction— In response to external stress, cells alter their morphology, metabolic activity, and functions to mechanically adapt to the dynamic, local environment through cell allostasis. To explore mechanotransduction in cellular allostasis, we applied an integrated micromechanical system that combines an ‘ultrasound tweezers’-based mechanical stressor and a Förster resonance energy transfer (FRET)-based molecular force biosensor, termed “actinin-sstFRET,” to monitor in situ single-cell allostasis in response to transient stimulation in real time. Methods— The ultrasound tweezers utilize 1 Hz, 10-second transient ultrasound pulses to acoustically excite a lipid-encapsulated microbubble, which is bound to the cell membrane, and apply a pico- to nano-Newton range of forces to cells through an RGD-integrin linkage. The actinin-sstFRET molecular sensor, which engages the actin stress fibers in live cells, is used to map real-time actomyosin force dynamics over time. Then, the mechanosensitive behaviors were examined by profiling the dynamics in Ca2+ influx, actomyosin cytoskeleton (CSK) activity, and GTPase RhoA signaling to define a single-cell mechanical allostasis. Results—By subjecting a 1 Hz, 10-second physical stress, single vascular smooth muscle cells (VSMCs) were observed to remodeled themselves in a biphasic mechanical allostatic manner within 30 minutes that caused them to adjust their contractility and actomyosin activities. The cellular machinery that underscores the vital role of CSK equilibrium in cellular mechanical allostasis, includes Ca2+ influx, remodeling of actomyosin CSK and contraction, and GTPase RhoA signaling. Mechanical allostasis was observed to be compromised in VSMCs from patients with type II diabetes mellitus (T2DM), which could potentiate an allostatic maladaptation. Conclusions— By integrating tools that simultaneously permit localized mechanical perturbation and map actomyosin forces, we revealed distinct cellular mechanical allostasis profiles in our micromechanical system. Our findings of cell mechanical allostasis and maladaptation provide the potential for mechanophenotyping cells to reveal their pathogenic contexts and their biophysical mediators that underlie multi-etiological diseases such as diabetes, hypertension, or aging. 

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4. Biocompatible 3D Printed MXene Microlattices for Tissue‐Integrated Antibiotic Sensing

   https://doi.org/10.1002/admt.202301517  

   Tiwari, Anand P. ; Panicker, Sreejith S. ; Huddy, Julia E. ; Rahman, Md Saifur ; Hixon, Katherine R. ; Scheideler, William J. ( December 2023 , Advanced Materials Technologies)

   3D continuous mesoscale architectures of nanomaterials possess the potential to revolutionize real‐time electrochemical biosensing through higher active site density and improved accessibility for cell proliferation. Herein, 3D microporous Ti₃C₂T_XMXene biosensors are fabricated to monitor antibiotic release in tissue engineering scaffolds. The Ti₃C₂T_X‐coated 3D electrodes are prepared by conformal MXene deposition on 3D‐printed polymer microlattices. The Ti₃C₂T_XMXene coating facilitates direct electron transfer, leading to the efficient detection of common antibiotics such as gentamicin and vancomycin. The 3D microporous architecture exposes greater electrochemically active MXene surface area, resulting in remarkable sensitivity for detecting gentamicin (10–1 mM) and vancomycin (100–1 mM), 1000 times more sensitive than control electrodes composed of 2D planar films of Ti₃C₂T_XMXene. To characterize the suitability of 3D microporous Ti₃C₂T_XMXene sensors for monitoring drug elution in bone tissue regeneration applications, osteoblast‐like (MG‐63) cells are seeded on the 3D MXene microlattices for 3, 5, and 7 days. Cell proliferation on the 3D microporous MXene is tracked over 7 days, demonstrating its promising biocompatibility and its clinical translation potential. Thus, 3D microporous Ti₃C₂T_XMXene can provide a platform for mediator‐free biosensing, enabling new applications for in vivo monitoring of drug elution.

    

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5. 3D-O-RAN: Dynamic Data Driven Open Radio Access Network Systems

   https://doi.org/10.1109/MILCOM55135.2022.10017706  

   Restuccia, Francesco ; Blasch, Erik ; Ashdown, Andrew ; Ashdown, Jonathan ; Turck, Kurt ( November 2022 , MILCOM 2022 - 2022 IEEE Military Communications Conference (MILCOM))

   This position paper introduces a Dynamic Data Driven Open Radio Access Network System (3D-O-RAN). The key objective of 3D-O-RAN is to support congested, contested and contaminated tactical settings where multimedia sensors, application constraints and operating wireless conditions may frequently change over space, time and frequency. 3D-O-RAN is compliant with the O-RAN specification for beyond 5G cellular systems to reduce costs and guarantee interoperability among vendors. Moreover, 3D-O-RAN integrates computational, sensing, and cellular networking components in a highly-dynamic, feedback-based, data-driven control loop. Specifically, 3D-O-RAN is designed to incorporate heterogeneous data into the network control loop to achieve a system-wide optimal operating point. Moreover, 3D-O-RAN steers the multimedia sensor measurement process in real time according to the required application needs and current physical and/or environmental constraints. 3D-O-RAN uses (i) a semantic slicing engine, which takes into account the semantic of the application to optimally compress the multimedia stream without losing in classification accuracy; (ii) a dynamic data driven neural network certification system that translates mission-level constraints into technical-level constraints on neural network latency/accuracy, and occupation of hardware/software resources. Realistic use-case scenarios of 3D-O-RAN in a tactical context demonstrate system performance. 

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