skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

Explore Research Products in the PAR
It may take a few hours for recently added research products to appear in PAR search results.


Title: Using light to establish and measure stiffness gradients in three-dimensional engineered tissues
Studies of cell-extracellular matrix (ECM) interactions within fibrous systems such as collagen or fibrin are challenging, particularly if peri-cellular stiffness cannot be monitored. Here we present our light-based method for non-invasive patterning of molecular crosslinking combined with multi-axes optical tweezers active microrheology to map ECM stiffness landscapes. This method allows us to generate prescribed stiffness gradients and associated anisotropies, which model stiffness of the natural peri-cellular ECM. Patterned crosslinking induces strain hardening and measured stiffness gradients are in agreement with predicted strain fields. Migratory cells respond to these gradients as assessed by change in F-actin distribution and morphological properties.  more » « less
Award ID(s):
1953410
PAR ID:
10321482
Author(s) / Creator(s):
; ;
Date Published:
Journal Name:
spie: Optical Trapping and Optical Micromanipulation XVIII
Volume:
11798
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. ; ; (, APS March Meeting 2021)
    Bulk measurements of ECM stiffness are commonly used in mechanobiology. However, peri-cellular stiffness can be quite heterogenous and divergent from the bulk properties. Here, we use optical tweezers active microrheology (AMR) to quantify how two different cell lines embedded in 1.0 and 1.5 mg/ml type 1 collagen (T1C) establish dissimilar patterns of peri-cellular stiffness. We found that dermal fibroblasts (DFs) increase local stiffness of 1.0 mg/ml T1C hydrogels, but surprisingly do not alter stiffness of 1.5 mg/ml T1C hydrogels. In contrast, MDA-MB-231 cells (MDAs) predominantly do not stiffen T1C hydrogels, as compared to cell-free controls. Results suggest that MDAs adapt to the bulk ECM stiffness, while DFs regulate local stiffness to levels they intrinsically “prefer”. Further, cells were subjected to treatments, that were previously shown to alter migration, proliferation and contractility of DFs and MDAs. Following treatment, both cell lines established different levels of stiffness magnitude and anisotropy, which were dependent on the cell line, T1C concentration and treatment. In summary, our findings demonstrate that AMR reveals otherwise masked mechanical properties such as spatial gradients and anisotropy, which are known to affect cell behavior at the macro-scale. 
    more » « less
  2. ; ; (, Scientific Reports)
    Abstract Cells are known to continuously remodel their local extracellular matrix (ECM) and in a reciprocal way, they can also respond to mechanical and biochemical properties of their fibrous environment. In this study, we measured how stiffness around dermal fibroblasts (DFs) and human fibrosarcoma HT1080 cells differs with concentration of rat tail type 1 collagen (T1C) and type of ECM. Peri-cellular stiffness was probed in four directions using multi-axes optical tweezers active microrheology (AMR). First, we found that neither cell type significantly altered local stiffness landscape at different concentrations of T1C. Next, rat tail T1C, bovine skin T1C and fibrin cell-free hydrogels were polymerized at concentrations formulated to match median stiffness value. Each of these hydrogels exhibited distinct fiber architecture. Stiffness landscape and fibronectin secretion, but not nuclear/cytoplasmic YAP ratio differed with ECM type. Further, cell response to Y27632 or BB94 treatments, inhibiting cell contractility and activity of matrix metalloproteinases, respectively, was also dependent on ECM type. Given differential effect of tested ECMs on peri-cellular stiffness landscape, treatment effect and cell properties, this study underscores the need for peri-cellular and not bulk stiffness measurements in studies on cellular mechanotransduction. 
    more » « less
  3. ; ; (, Research square)
    Cells are known to continuously remodel their local extracellular matrix (ECM) and in a reciprocal way, they can also respond to mechanical and biochemical properties of their fibrous environment. In this study, we measured how stiffness around dermal fibroblasts (DFs) and human fibrosarcoma HT1080 cells differs with concentration of rat tail type 1 collagen (T1C) and type of ECM. Peri-cellular stiffness was probed in four directions using multi-axes optical tweezers active microrheology (AMR). First, we found that neither cell type significantly altered local stiffness landscape at different concentrations of T1C. Next, rat tail T1C, bovine skin T1C and fibrin cell-free hydrogels were polymerized at concentrations formulated to match median stiffness value. Each of these hydrogels exhibited distinct fiber architecture. Stiffness landscape and fibronectin secretion, but not nuclear/cytoplasmic YAP ratio differed with ECM type. Further, cell response to Y27632 or BB94 treatments, inhibiting cell contractility and activity of matrix metalloproteinases, respectively, was also dependent on ECM type. Given differential effect of tested ECMs on peri-cellular stiffness landscape, treatment effect and cell properties, this study underscores the need for peri-cellular and not bulk stiffness measurements in studies on cellular mechanotransduction. 
    more » « less
  4. ; ; (, EUROPEAN BIOPHYSICS JOURNAL WITH BIOPHYSICS LETTERS)
    Bulk ECM stiffness measurements are often used in research on cell mechanobiology. However, past studies by our group have shown that peri-cellular stiffness can span few orders of magnitude and diverges from the bulk properties. Us- ing optical tweezers active microrheology (AMR) we can de- scribe stiffness landscape around individual cells. In this study, we show how different cell lines cultured in 1.0 and 1.5 mg/ml type 1 collagen (T1C) create disparate patterns of peri-cellular stiffness. Dermal fibroblasts (DFs) increase peri-cellular stiffness, when embedded in 1.0 mg/ml T1C hy- drogels, but do not alter stiffness in 1.5 mg/ml T1C hydro- gels. In contrast, invasive human breast cancer MDA-MB- 231 cells (MDAs) do not significantly change the stiffness of T1C hydrogels, as compared to cell-free controls. Results indicate that while MDAs adapt to the bulk ECM stiffness, DFs regulate local stiffness to levels they intrinsically “fa- vor”. Further, cells were also subjected to treatments that were previously shown to regulate their migration, prolifera- tion and contractility. Following each treatment, cells estab- lished dissimilar stiffness patterns. Stiffness magnitude and extent of anisotropy varied with the cell line, T1C concen- tration and treatment. In summary, we demonstrate that AMR can reveal otherwise masked mechanical properties of the local ECM, which are known to affect cell behavior. 
    more » « less
  5. ; ; (, Current Protocols)
    Abstract Physical properties of the extracellular matrix (ECM) affect cell behaviors ranging from cell adhesion and migration to differentiation and gene expression, a process known as mechanotransduction. While most studies have focused on the impact of ECM stiffness, using linearly elastic materials such as polyacrylamide gels as cell culture substrates, biological tissues and ECMs are viscoelastic, which means they exhibit time‐dependent mechanical responses and dissipate mechanical energy. Recent studies have revealed ECM viscoelasticity, independent of stiffness, as a critical physical parameter regulating cellular processes. These studies have used biomaterials with tunable viscoelasticity as cell‐culture substrates, with alginate hydrogels being one of the most commonly used systems. Here, we detail the protocols for three approaches to modulating viscoelasticity in alginate hydrogels for 2D and 3D cell culture studies, as well as the testing of their mechanical properties. Viscoelasticity in alginate hydrogels can be tuned by varying the molecular weight of the alginate polymer, changing the type of crosslinker—ionic versus covalent—or by grafting short poly(ethylene‐glycol) (PEG) chains to the alginate polymer. As these approaches are based on commercially available products and simple chemistries, these protocols should be accessible for scientists in the cell biology and bioengineering communities. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Tuning viscoelasticity by varying alginate molecular weight Basic Protocol 2: Tuning viscoelasticity with ionic versus covalent crosslinking Basic Protocol 3: Tuning viscoelasticity by adding PEG spacers to alginate chains Support Protocol 1: Testing mechanical properties of alginate hydrogels Support Protocol 2: Conjugating cell‐adhesion peptide RGD to alginate 
    more » « less
  • Citation Formats
  • MLA
  • APA
  • Chicago
  • BibTeX
  • Save / Share this Record
  • Send to Email