Real time microscopy of the lung vasculature for dissecting the role of the nuclear lamina in tumor cell extravasation and metastasis

Multiple types of circulating tumor cells (CTCs) metastasize into the lung parenchyma by extravasating the pulmonary vasculature. The space between the alveolus and the capillary (approximately 2 um) imposes a significant mechanical barrier on extravasating CTCs that need to squeeze their bulky and stiff nuclei through it with minimal nuclear damage. Using live imaging of the lung parenchyma with 2-photon microscopy set-up integrated with a thoracic window apparatus we will dissect the mechanisms by which cells transiently reduces their nucleus stiffening enabling extravasation. Genetic manipulations of several breast cancer cell lines by transient and permanent lamin A/C silencing and by CRISPR technology to eliminate a key laminin serine phosphorylation site will be utilized to unravel molecular players involved.