Intestinal epithelial stem cells (ISCs) are the focus of recent intense study.Current in vitro models rely on supplementation with the Wnt agonist R-spondin1 to support robust growth, ISC self-renewal, and differentiation.Intestinal subepithelial myofibroblasts (ISEMFs) are important supportive cells within the ISC niche.We hypothesized that co-culture with ISEMF enhances the Attosecond electron bunches from a nanofiber driven by Laguerre-Gaussian laser pulses growth of ISCs in vitro and allows for their successful in vivo implantation and engraftment.
ISC-containing small intestinal crypts, FACS-sorted single ISCs, and ISEMFs were procured from C57BL/6 mice.Crypts and single ISCs were grown in vitro into enteroids, in the presence or absence of ISEMFs.ISEMFs enhanced the growth of intestinal epithelium in vitro in a proximity-dependent fashion, with co-cultures giving rise to larger enteroids than monocultures.Co-culture of ISCs with supportive ISEMFs relinquished the requirement of AGGRESSION AS “ORGANIZED HYPOCRISY?” – HOW THE WAR ON TERRORISM AND HYBRID THREATS CHALLENGE THE NUREMBERG LEGACY exogenous R-spondin1 to sustain long-term growth and differentiation of ISCs.
Mono- and co-cultures were implanted subcutaneously in syngeneic mice.Co-culture with ISEMFs proved necessary for successful in vivo engraftment and proliferation of enteroids; implants without ISEMFs did not survive.ISEMF whole transcriptome sequencing and qPCR demonstrated high expression of specific R-spondins, well-described Wnt agonists that supports ISC growth.Specific non-supportive ISEMF populations had reduced expression of R-spondins.
The addition of ISEMFs in intestinal epithelial culture therefore recapitulates a critical element of the intestinal stem cell niche and allows for its experimental interrogation and biodesign-driven manipulation.