US 11,926,849 B2
Biosensors in human gut organoids
Alessio Fasano, Boston, MA (US); Timothy K. Lu, Cambridge, MA (US); Stefania Senger, Melrose, MA (US); and Maria Eugenia Inda, Cambridge, MA (US)
Assigned to The General Hospital Corporation, Boston, MA (US); and Massachusetts Institute Of Technology, Cambridge, MA (US)
Filed by The General Hospital Corporation, Boston, MA (US); and MASSACHUSETTS INSTITUTE OF TECHNOLOGY, Cambridge, MA (US)
Filed on Mar. 30, 2020, as Appl. No. 16/834,727.
Claims priority of provisional application 62/826,352, filed on Mar. 29, 2019.
Prior Publication US 2020/0308549 A1, Oct. 1, 2020
Int. Cl. C12N 5/071 (2010.01); G01N 33/50 (2006.01); G01N 33/68 (2006.01)
CPC C12N 5/0679 (2013.01) [G01N 33/5044 (2013.01); G01N 33/6803 (2013.01); C12N 2501/998 (2013.01); C12N 2503/02 (2013.01); C12N 2506/03 (2013.01); C12N 2513/00 (2013.01); G01N 2500/10 (2013.01)] 7 Claims
 
1. An ex vivo model system, comprising: a monolayer consisting of differentiated mammalian epithelial cells, wherein the monolayer consists of enterocytes with microvilli, mucus-producing goblet cells, and Microfold (M) cells, having an apical surface and a basolateral surface, and wherein cells of the monolayer express one or more exogenous biosensors responsive to an inflammation-related signal comprising at least one promoter that binds to the inflammation-related signal and drives expression of a detectable output protein upon binding of the inflammation-related signal to the promoter wherein the differentiated mammalian epithelial cells are generated by a method comprising: obtaining primary stem cells from a mammalian subject; culturing the primary cells under conditions sufficient to allow proliferation of the cells and formation of organoids; seeding the cells of the organoid into a culture plate comprising a permeable support device; maintaining the cells underconditions to allow for sufficient proliferation to form a monolayer having an apical and basolateral surface; contacting the apical surface with DAPT and the basolateral surface with RANKL, in amounts sufficient to induce differentiation of the cells to form enterocytes with microvilli, mucus-producing goblet cells, and M cells.