Hans Clevers (Hubrecht Institute, The Netherlands)
“Wnt Signaling and Cancer”
- The champion of all stem cells : crypt base paneth cells
- Wnt is required to maintain crypt stem cells in colon cancer，Wnt is locked in On state。
- Lqr5 is an unusual component that marks stem cells in the crypts。
- 15 stem cell to begin (mixed with1 Lqr5)
- Re-tracing
- Not asymmetric
- Extensive plasticity in crypts
- Lgr5-driven GFP in crypt base columnar cells
- Intestinal stem cells are not quiescent/not rare ~ 18%/not asymmetric
- Lgr5 +ve stem cells are the origin of colon cancer stem cells
- Adenomas retain Paneth stem cell
- Deleting floxed
- A single lgr5 cell in a red sermons behaves linebackers stem cells
- Lgr5 uses the receptor : R-spondin
- E3 ligase ZNRF3/RNF43
- R spondin binds Lgr5 and ZNRF3 that stabilize WNT complex
- Single LGR5 stem cell grown in vitro
- 1. R SPONDIN LGR5 2. EGF 3. Noggin (TGFB/ BMP) inhibitor
- Self renewal in stomach corpus glands Troy and Lgr5
- Human gastric organoid and Helicobacter
- Human Lung organoid
- chemotaxis toward RSV infected lung organoid
- A living cancer bank with SU2C
- Lineage-tracing: Nat 2007 Identification of stem cells in small intestine and colon by marker gene Lgr5.
From Ming-Han
Wnt signal: crosstalk between cells and shape the crypts of stem cells in gut.
- -Lgr5 and Wnt signal.
- -Wnt block B-cat be degraded by APC-E3 complex; that make tx factor TCF be activated. Once Wnt is weaken, the B-catenin can be releases from APC complex and go to nucleus.
- @Alkaline phosphatase: marks entrocytes lineages. (not mark stem cells!!)
- Lgr5+ cells re stem cells. (Lgr5CRE+ cells: can marked stem cells).
- Lgr5+ cells + R-spondin1 (Wnt Angonist) + EGF + Noggin (TGF inhibitor) + Matrigel —-> can form 3D crypts!! mini gut.
- –> Can inject these mini gut bact to mice which contain the damage of gut–> these small guts can cover the damaged place and fix the damage!!
- -> They expand the studies into
Gastric system:
- Stomach: gastric epithelial cell replace much slower than Gut. The stem cells called “chief cells from isthmus”.
- Only Troy+ cells yield stomach organdies and can be infected by Helicobacteria!!
- They apply Casp9 system to modify the gene in organdies.
Lung Organoids:
- Can produce Mucus! and can be infected by pathogens; for example: RSV
Urine:
- p63+ from bladder and Pax8+ from Urine.
Application for their study:
- Virology/Immunology
- Drug screening
- Tumor stem cells/ genomic screening.
- They found that the organdies made by cancer cells look totally different from the normal cells!! Can use single cell culture to sort them out and trace back to “tumor stem cells”.
- Apply Caspase9 system to study the genomic and the formation of organdies.
Special and attracting techniques: Lineage-tracing techniques (Identification of stem cells in small intestine and colon by marker gene Lgr5). (read)
Special experiment:
- They use Casp9 system to KO P53 or p16 or SMAD or….
- They found the organoids with these KO -> grow much slower than the normal cells.
- Certain KO don’t require EGF anymore.
- After injecting back to mice; although these KO organoids grow badly in vitro but can form tumours in vivo. The healthy ones can’t.