# Cell mechanobiology in confinement using an integration of bioengineering, materials systems and in vivo models

> **NIH NIH R01** · JOHNS HOPKINS UNIVERSITY · 2024 · $385,771

## Abstract

Summary- Cells in vivo travel through confining three-dimensional (3D) pores between fibrillar extracellular
matrix (ECM) networks or channel-like tracks bordered by ECM bundles, vessels, myofibers or nerves. The
mechanisms enabling cell locomotion in diverse microenvironments are adaptive in response to the physical and
biochemical cues, such as confinement, stiffness, viscoelastic properties and composition of ECM. Adaptive
systems/modules include cell-ECM interactions, the actomyosin cytoskeleton and cell volume regulation.
Recently, we and others have also identified the key role of the nucleus in confined migration. However,
numerous fundamental and translational questions remain unanswered on the crosstalk between nuclear
mechanosensing, cytoskeleton and cell volume regulation, and their contributions to confined migration in health
and disease. The overarching goal of this project is to employ state-of-the-art bioengineering, materials and
imaging tools as well as in vivo models to provide a novel unified framework for efficient migration in confinement
by deciphering the interplay between nuclear mechanics, cytoskeleton and ion channels. This R01 application
will test the hypothesis that the nucleus senses and responds to physical confinement by exquisitely regulating
the spatial activation of RhoA along the longitudinal cell axis in confined spaces via the synergistic roles of
confinement-induced nuclear stiffening and anillin/Ect2 nuclear exit to the cytoplasm. This hypothesis is
supported by intriguing preliminary data showing that cell entry into confining µ-channels induces nuclear
stiffening which activates RhoA and supports ion channel-dependent nuclear blebbing and rupture. Nuclear
rupture induces the exit of anillin and the RhoGEF Ect2 from the nucleus to the cytoplasm. Anillin accumulates
specifically at the cell poles, where it locally bridges Ect2, RhoA and actomyosin, thereby exacerbating RhoA-
myosin II contractility. In Aim 1, we will decipher the mechanisms of anillin exit to the cytoplasm, and demonstrate
its critical role as a scaffolding protein, which bridges Ect2, RhoA and actomyosin at the cell poles, thereby
regulating the spatial activation of RhoA and bleb-based migration in confinement. We will also elucidate the
novel crosstalk between cell volume regulation and anillin/Ect2/RhoA in nuclear blebbing and rupture in
confinement. Lastly, we will decipher the contributions of nuclear pushing from the cell rear versus nuclear pulling
from the cell front (i.e., nuclear piston model) to migration as a function of the degree of confinement. In Aim 2,
we will extend the applicability of our findings to 3D gels and confining µ-channels of prescribed physiologically
relevant mechanical properties in vitro. We will also visualize the distinct localization patterns of anillin, Ect2 and
key ion channels in natural tissue tracks of different dimensions in vivo, and test how perturbations of these
molecules impact local and dis...

## Key facts

- **NIH application ID:** 10795830
- **Project number:** 5R01GM142175-04
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Konstantinos Konstantopoulos
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $385,771
- **Award type:** 5
- **Project period:** 2021-04-01 → 2025-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10795830

## Citation

> US National Institutes of Health, RePORTER application 10795830, Cell mechanobiology in confinement using an integration of bioengineering, materials systems and in vivo models (5R01GM142175-04). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10795830. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*