The nucleus houses, organizes, and protects chromatin to ensure genome integrity and proper gene expression, but how the nucleus adapts mechanically to changes in the extracellular environment is poorly understood. Recent studies have revealed that extracellular physical stresses induce chromatin compaction via mechanotransductive processes. We report that increased extracellular multivalent cations lead to increased heterochromatin levels through activation of mechanosensitive ion channels (MSCs), without large-scale cell stretching. In cells with perturbed chromatin or lamins, this increase in heterochromatin suppresses nuclear blebbing associated with nuclear rupture and DNA damage. Through micromanipulation force measurements, we show that this increase in heterochromatin increases chromatin-based nuclear rigidity, which protects nuclear morphology and function. In addition, transduction of elevated extracellular cations rescues nuclear morphology in model and patient cells of human diseases, including progeria and the breast cancer model cell line MDA-MB-231. We conclude that nuclear mechanics, morphology, and function can be modulated by cell sensing of the extracellular environment through MSCs and consequent changes to histone modification state and chromatin-based nuclear rigidity.

Background Veno‐venous extracorporeal membrane oxygenation (V‐V ECMO) support is increasingly used in the management of COVID‐19‐related acute respiratory distress syndrome (ARDS). However, the clinical decision‐making to initiate V‐V ECMO for severe COVID‐19 still remains unclear. In order to determine the optimal timing and patient selection, we investigated the outcomes of both COVID‐19 and non‐COVID‐19 patients undergoing V‐V ECMO support. Methods Overall, 138 patients were included in this study. Patients were stratified into two cohorts: those with COVID‐19 and non‐COVID‐19 ARDS. Results The survival in patients with COVID‐19 was statistically similar to non‐COVID‐19 patients ( p = .16). However, the COVID‐19 group demonstrated higher rates of bleeding ( p = .03) and thrombotic complications ( p < .001). The duration of V‐V ECMO support was longer in COVID‐19 patients compared to non‐COVID‐19 patients (29.0 ± 27.5 vs 15.9 ± 19.6 days, p < .01). Most notably, in contrast to the non‐COVID‐19 group, we found that COVID‐19 patients who had been on a ventilator for longer than 7 days prior to ECMO had 100% mortality without a lung transplant. Conclusions These findings suggest that COVID‐19‐associated ARDS was not associated with a higher post‐ECMO mortality than non‐COVID‐19‐associated ARDS patients, despite longer duration of extracorporeal support. Early initiation of V‐V ECMO is important for improved ECMO outcomes in COVID‐19 ARDS patients. Since late initiation of ECMO was associated with extremely high mortality related to lack of pulmonary recovery, it should be used judiciously or as a bridge to lung transplantation.

Genetically engineering cells to perform customizable functions is an emerging frontier with numerous technological and translational applications. However, it remains challenging to systematically engineer mammalian cells to execute complex functions. To address this need, we developed a method enabling accurate genetic program design using high-performing genetic parts and predictive computational models. We built multifunctional proteins integrating both transcriptional and posttranslational control, validated models for describing these mechanisms, implemented digital and analog processing, and effectively linked genetic circuits with sensors for multi-input evaluations. The functional modularity and compositional versatility of these parts enable one to satisfy a given design objective via multiple synonymous programs. Our approach empowers bioengineers to predictively design mammalian cellular functions that perform as expected even at high levels of biological complexity.

The nucleus houses, organizes, and protects chromatin to ensure genome integrity and proper gene expression, but how the nucleus adapts mechanically to changes in the extracellular environment is poorly understood. Recent studies have revealed that extracellular physical stresses induce chromatin compaction via mechanotransductive processes. We report that increased extracellular multivalent cations lead to increased heterochromatin levels through activation of mechanosensitive ion channels, without large-scale cell stretching. In cells with perturbed chromatin or lamins, this increase in heterochromatin suppresses nuclear blebbing associated with nuclear rupture and DNA damage. Through micromanipulation force measurements, we show that this increase in heterochromatin increases chromatin-based nuclear rigidity, which protects nuclear morphology and function. In addition, transduction of elevated extracellular cations rescues nuclear morphology in model and patient cells of human diseases, including progeria and the breast cancer model cell line MDA-MB-231. We conclude that nuclear mechanics, morphology, and function can be modulated by cell sensing of the extracellular environment through mechanosensitive ion channels and consequent changes to histone modification state and chromatin-based nuclear rigidity.

Intro Force measurements of the nucleus, the strongest organelle, have propelled the field of mechanobiology to understand the basic mechanical components of the nucleus and how these components properly support nuclear morphology and function. Micromanipulation force measurement provides separation of the relative roles of nuclear mechanical components chromatin and lamin A. Methods To provide access to this technique, we have developed a universal micromanipulation apparatus for inverted microscopes. We outline how to engineer and utilize this apparatus through dual micromanipulators, fashion and calibrate micropipettes, and flow systems to isolate a nucleus and provide force vs. extensions measurements. This force measurement approach provides the unique ability to measure the separate contributions of chromatin at short extensions and lamin A strain stiffening at long extensions. We then investigated the apparatus’ controllable and programmable micromanipulators through compression, isolation, and extension in conjunction with fluorescence to develop new assays for nuclear mechanobiology. Results Using this methodology, we provide the first rebuilding of the micromanipulation setup outside of its lab of origin and recapitulate many key findings including spring constant of the nucleus and strain stiffening across many cell types. Furthermore, we have developed new micromanipulation-based techniques to compress nuclei inducing nuclear deformation and/or rupture, track nuclear shape post-isolation, and fluorescence imaging during micromanipulation force measurements. Conclusion We provide the workflow to build and use a micromanipulation apparatus with any inverted microscope to perform nucleus isolation, force measurements, and various other biophysical techniques.