skip to main content

Attention:

The NSF Public Access Repository (PAR) system and access will be unavailable from 11:00 PM ET on Thursday, February 13 until 2:00 AM ET on Friday, February 14 due to maintenance. We apologize for the inconvenience.


Title: Soft Ring‐Shaped Cellu‐Robots with Simultaneous Locomotion in Batches
Abstract

Untethered mini‐robots can move single cells or aggregates to build complex constructs in confined spaces and may enable various biomedical applications such as regenerative repair in medicine and biosensing in bioengineering. However, a significant challenge is the ability to control multiple microrobots simultaneously in the same space to operate toward a common goal in a distributed operation. A locomotion strategy that can simultaneously guide the formation and operation of multiple robots in response to a common acoustic stimulus is developed. The scaffold‐free cellu‐robots comprise only highly packed cells and eliminate the influence of supportive materials, making them less cumbersome during locomotion. The ring shape of the cellu‐robot contributes to anisotropic cellular interactions which induce radial cellular orientation. Under a single stimulus, several cellu‐robots form predetermined complex structures such as bracelet‐like ring‐chains which transform into a single new living entity through cell–cell interactions, migration or cellular extensions between cellu‐robots.

 
more » « less
PAR ID:
10458753
Author(s) / Creator(s):
 ;  ;  ;  ;  
Publisher / Repository:
Wiley Blackwell (John Wiley & Sons)
Date Published:
Journal Name:
Advanced Materials
Volume:
32
Issue:
8
ISSN:
0935-9648
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract

    Multimeric cytoskeletal protein complexes orchestrate normal cellular function. However, protein-complex distributions in stressed, heterogeneous cell populations remain unknown. Cell staining and proximity-based methods have limited selectivity and/or sensitivity for endogenous multimeric protein-complex quantification from single cells. We introduce micro-arrayed, differential detergent fractionation to simultaneously detect protein complexes in hundreds of individual cells. Fractionation occurs by 60 s size-exclusion electrophoresis with protein complex-stabilizing buffer that minimizes depolymerization. Proteins are measured with a ~5-hour immunoassay. Co-detection of cytoskeletal protein complexes in U2OS cells treated with filamentous actin (F-actin) destabilizing Latrunculin A detects a unique subpopulation (~2%) exhibiting downregulated F-actin, but upregulated microtubules. Thus, some cells may upregulate other cytoskeletal complexes to counteract the stress of Latrunculin A treatment. We also sought to understand the effect of non-chemical stress on cellular heterogeneity of F-actin. We find heat shock may dysregulate filamentous and globular actin correlation. In this work, our assay overcomes selectivity limitations to biochemically quantify single-cell protein complexes perturbed with diverse stimuli.

     
    more » « less
  2. Abstract

    Integrated microfluidic cellular phenotyping platforms provide a promising means of studying a variety of inflammatory diseases mediated by cell‐secreted cytokines. However, immunosensors integrated in previous microfluidic platforms lack the sensitivity to detect small signals in the cellular secretion of proinflammatory cytokines with high precision. This limitation prohibits researchers from studying cells secreting cytokines at low abundance or existing at a small population. Herein, the authors present an integrated platform named the “digital Phenoplate (dPP),” which integrates digital immunosensors into a microfluidic chip with on‐chip cell assay chambers, and demonstrates ultrasensitive cellular cytokine secretory profile measurement. The integrated sensors yield a limit of detection as small as 0.25 pg mL−1for mouse tumor necrosis factor alpha (TNF‐α). Each on‐chip cell assay chamber confines cells whose population ranges from ≈20 to 600 in arrayed single‐cell trapping microwells. Together, these microfluidic features of the dPP simultaneously permit precise counting and image‐based cytometry of individual cells while performing parallel measurements of TNF‐α released from rare cells under multiple stimulant conditions for multiple samples. The dPP platform is broadly applicable to the characterization of cellular phenotypes demanding high precision and high throughput.

     
    more » « less
  3. Abstract

    The ability to program collective cell migration can allow us to control critical multicellular processes in development, regenerative medicine, and invasive disease. However, while various technologies exist to make individual cells migrate, translating these tools to control myriad, collectively interacting cells within a single tissue poses many challenges. For instance, do cells within the same tissue interpret a global migration ‘command’ differently based on where they are in the tissue? Similarly, since no stimulus is permanent, what are the long-term effects of transient commands on collective cell dynamics? We investigate these questions by bioelectrically programming large epithelial tissues to globally migrate ‘rightward’ via electrotaxis. Tissues clearly developed distinct rear, middle, side, and front responses to a single global migration stimulus. Furthermore, at no point poststimulation did tissues return to their prestimulation behavior, instead equilibrating to a 3rd, new migratory state. These unique dynamics suggested that programmed migration resets tissue mechanical state, which was confirmed by transient chemical disruption of cell–cell junctions, analysis of strain wave propagation patterns, and quantification of cellular crowd dynamics. Overall, this work demonstrates how externally driving the collective migration of a tissue can reprogram baseline cell–cell interactions and collective dynamics, even well beyond the end of the global migratory cue, and emphasizes the importance of considering the supracellular context of tissues and other collectives when attempting to program crowd behaviors.

     
    more » « less
  4. Abstract

    Characterizing the mechanical properties of single cells is important for developing descriptive models of tissue mechanics and improving the understanding of mechanically driven cell processes. Standard methods for measuring single‐cell mechanical properties typically provide isotropic mechanical descriptions. However, many cells exhibit specialized geometriesin vivo, with anisotropic cytoskeletal architectures reflective of their function, and are exposed to dynamic multiaxial loads, raising the need for more complete descriptions of their anisotropic mechanical properties under complex deformations. Here, we describe the cellular microbiaxial stretching (CμBS) assay in which controlled deformations are applied to micropatterned cells while simultaneously measuring cell stress. CμBS utilizes a set of linear actuators to apply tensile or compressive, short‐ or long‐term deformations to cells micropatterned on a fluorescent bead‐doped polyacrylamide gel. Using traction force microscopy principles and the known geometry of the cell and the mechanical properties of the underlying gel, we calculate the stress within the cell to formulate stress‐strain curves that can be further used to create mechanical descriptions of the cells, such as strain energy density functions. © 2022 Wiley Periodicals LLC.

    Basic Protocol 1: Assembly of CμBS stretching constructs

    Basic Protocol 2: Polymerization of micropatterned, bead‐doped polyacrylamide gel on an elastomer membrane

    Support Protocol: Cell culture and seeding onto CμBS constructs

    Basic Protocol 3: Implementing CμBS stretching protocols and traction force microscopy

    Basic Protocol 4: Data analysis and cell stress measurements

     
    more » « less
  5. Abstract

    During the blood stage of human malaria,Plasmodium falciparumparasites divide by schizogony—a process wherein components for several daughter cells are produced within a common cytoplasm and then segmentation, a synchronized cytokinesis, produces individual invasive daughters. The basal complex is hypothesized to be required for segmentation, acting as a contractile ring to establish daughter cell boundaries. Here we identify an essential component of the basal complex which we name PfCINCH. Using three-dimensional reconstructions of parasites at electron microscopy resolution, we show that while parasite organelles form and divide normally, PfCINCH-deficient parasites develop inviable conjoined daughters that contain components for multiple cells. Through biochemical evaluation of the PfCINCH-containing complex, we discover multiple previously undescribed basal complex proteins. Therefore, this work provides genetic evidence that the basal complex is required for precise segmentation and lays the groundwork for a mechanistic understanding of how the parasite contractile ring drives cell division.

     
    more » « less