LIFESCI 2A03 Lecture Notes - Fibroblast, Cyclic Adenosine Monophosphate, Epithelium

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2 Feb 2013
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Summary of Experiments
1st Experiment
Purpose: Determining if MSC can differentiate into AECs
Hypo: If created in a particular condition as described in method, the airway-epithelium culture condition may be useful
to assay for differentiation of MSCs to AECs
Fig ab&c shows the state of primary human AECs after being cultured for 2 weeks & how closely they mimic the
morphology & function of surface epithelium conducting airways
Found: MSCs can appear differently in different conditions
1) Plastic surface Normal condition GFP-MSCs have slender fibro-blast like or flat polygonal cell shape
2) In mixed culture MSCs assume epithelia like cuboidal or columnar cell shape
2nd Experiment
Purpose: Determining whether the cocultured MSCs had important epithelia-specific marker genes: CK-18 & Occludin &
CFTR
1st Hypo: If it is cocultured with CK-18 antibody then MSCs should differentiate into AECs
Result: Only some (as seen in fig i-k)
Fig a&b shows CK-18 effect on normal AECs
Fig cd, gh shows CK-18 present on GFP-MSCs
2nd Hypo: If the cocultured MSCs are mixed with primary it should show Occludin
Result: 10% showed presence of occluding also under fluroscent microscopy GFP-MSCs expressed occluding Fig (o-q)
3rd Hypo Determining whether CFTR is expressed after it is cocultured with human AECs
Hypo: If MSCs are cocultured with human AECs then CFTR is expressed
Result: Using FACS the GFP-MSCs are purified (fig a&b)
3rd Hypo part 2: If WT MSCs differentiate into AECs, WT CFTR should be expressed
3rd Experiment
Purpose: See if MSCs can be:
i) Amenable to gene correction
ii) Maintain their pluripotency after gene correction
As seen through deltaF508 CF patients Bone marrow has been aspirated from them
Result: Gene corrected MSCs from CF patients have the same development potentials as the MSCs from normal
individuals as seen in Fig 4 a-f
Result: 1) CF MSCs also underwent differentiation as seen fro mthe table
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