l4 met+animal models Flashcards
what is EDu and BrDu?
they are analogues of thymidine.
- They can be visualised
- incorperate into DNA during DNA synthesis
- label newly proliferating cells.
how are B cells visualised?
transgenic reporter line -Tg(Ins:GFP)
what does the paper by Hill et al show about gut bacterie?
certain gut bacteria/microbiota and protein they secrete are necessary for the pancreas to populate itself with a robust number of B cells.
what happens to the number of B cells in a microbe free environment?
In a microbe free environment the number of B cells does not increase after hatching (same before and after hatching)
(in microbe environment there is an increase in B cells after hatching)
what restored B cell numbers to normal levels from a microbe free environment?
- addition of certain bacteria which produced BeFA
- addition of BeFA rescued amount of B cells
what does BeFA do?
- restores B cell numbers
- Induces B cells to proliferate (identified by BrDU or Edu injection)
- some bacteria in humans produce similar protein to BeFA
How many small molecules were screened in a zebrafish model of diabetes to see what restored B cells?
~7000 small molecules screened
what were the double transgenic zebrafish used in B cell regenerative drug screen?
Tg (Ins:Kaede) (marks B cells)
Tg (Ins:CFP-NTR)
why was the Tg (Ins:CFP:NTR) fish used?
- when B cells produced insulin nitroreductase was also produced
- if MTZ added this converted NTR to a cytotoxic product - killed B cells (6-7 left)
WHat happened to double transgenic fish? (Ins:Kaede) (Ins:CFP-NTR)
- treated with MTZ (then washed)
- place in wells and add a different compound to each well (~7000 small molecules tested)
- see which compounds induced B cell regeneration
Which compounds identified in the zebrafish genetic screen restored the number of B cells?
- NECA
- all compounds that restored B cell numbers converged on ADENOSINE SIGNALLING
- adenosine agonsist (IB-Meca) increases B cell regeneration
- accelerated restoration of normoglycaemia
how was the proliferative B cell effect of NECA found in the zebrafish confirmed in mice models?
- when mice were treated with NECA - had a proliferative effect on B cells
what do NPY neurons stimulate?
NPY neurons stimulate food intake
what do pomc neurons stimulate?
Pomc neurons reduce food intake
what happened in genetically null leptin mice?
They were obese, hypoactive and had diabetes
- also were infertile
- enlarged fatty livers
what happened when leptin receptors were deleted in mice?
- same phenotype as leptin null mice
- obese, diabetic, infertile and hypoactive mice
- also enlarged fatty livers
using animal studies where was it shown that leptin receptors are?
in the arcuate nucleus of the hypothalamus
where NPY and pomc neurons are
how did the level of obesity in mice correspond with the amount of leptin receptors in the hypothalamus?
- the degree of obesity negativly correlated with the number of leptin receptors in the hypothalamus
- fewer receptors - more obese
what was increased in the plasma of obese leptin receptor defiecient mice?
-increased plasma glucose, leptin (less receptors to bind to), insulin and cortocosterone
what did the study where mice were leptin or leptin receptor deficient show?
It showed that the brain is a direct target for the weight reducing effects of leptin and liver abnormalities are secondary to defective leptin signalling in the brian.
how were NPY levels different in leptin null mice?
- NPY RNA levels in null leptin mice were higher than normal
