Cell-Based Therapies Flashcards
What do the following mean?
Totipotent
Pluripotent
Multipotent
Unipotent
Totipotent –> Can differentiate into any cell in the body
Pluripotent –> Can differentate into most cell in the body
Multipotent –> Has the ability to change into a few different cells
Unipotent –> Can only change into one type of cell
How are Induced Pluripotent Stem Cells (iPSCs) made?
Collection of adult cells
Tranfection with fibroblasts with combinations for 24 important genes
These then need to be isolated due to the low success rate
What is the difference between Magnetic-Activated Cell Sorting (MACS®) and Fluorescent-Activated Cell Sorting (FACS)?
MACS –> Use of magnets on antibodies that bind to the cell of interest
Therefore when the magnet is removed the antibodies with the cells bound will all be deposited in a single place
FACS –> Cells are suspended in water and given a charge, which seperates them into different areas
Both used for isolation, and not expression!!
How could iPSCs be used as personalised cancer vaccines?
iPSCs express multiple tumour associated antigens
So if we made iPSCs from patients cells, irradiated them (as we can’t add in actively dividing cells into the body) and injected them back into the body to act as a treatment
Why aren’t hiPSCs immune compatible when it comes to transplants?
As they contain HLA Class I (but not Class II) antigens
These Class I HLA antigens have their expression changed when differentiated, meaning that an immune response is possible
How could we genetically alter hiPSCs to make them hypoimmunogenic?
Remove any class I or II HLA antigens
Use genetic editing (CRISPR/Cas-9) to ensure a high expression of CD47
Haematopoietic and Mesenchymal stem cells are which form of stem cells?
Adult Stem Cells
So can be made using PSCs
What is HSC transplantation?
Wiping out somebodies immune system, to then remake it with HSC stem cells
Can be autologous (self) or allogenic (donor…so needs to be HLA matched)
The first 100 days post transplant is the likely time for Graft vs Host Disease (GvHD) to occur
This can be minimised by enriching the HSCs with CD34+ cells
What are the 3 main things that Mesenchymal stem cells (MSCs) can be differentiated into?
Fat cells
Cartilage
Bone (osteocytes)
This varies from MSC to MSC as they are not functionally the same…. so depending on where you get them from you may get a different cell
What are the 3 roles of MSCs?
Providing daughter cells that differentiate, and participate in repair
Homing to distant sites of injury (so can be useful in cancer treatments)
Secretion of factors that support wound repair by recruiting other cell types and modulating the immune response
What is tissue engineering?
Cells are taken from the patient (autologus) and isolated before expanding them on a suitable 3D structure
Expose the cells to specific growth factors and conditions to grow what we want
The cells are then isolated again and implanted back into the patient
What are the 4 techniques for isolating cells?
Differential adhesion –> As different cells adhere to different surfaces
Density centrifugation –> Size
FACS –> Size, granularity and surface markers
MACS –> Surface markers
What are the 7 ideal characteristics of scaffolds?
Biocompatible
Biodegradable
Cytocompatible (adhesive)
Porous
Mechanically appropriate
Architecturally appropriate
Growth promoting (such as GF)
What are the 4 different types of scaffold material?
Polypeptides –> Collagen, gelatin etc
Polysaccharides –> Hylauronic acid, alginate and chitosan
Synthetic Polymers –> Polyesters….higher degree of control, larger scale, can be temperature responsive….but not very adhesive
Bioceramics and Bioactive glasses –> Hydroxyapatite, bioactive glass, alumia etc
How can Polyester scaffolds be modified beneficially?
NaOH or primary amines are added, allowing subsequent reactions with coupling reagents to attach bioactive motifs or whole proteins….such as laminin….to increase adhesion
These scaffolds then are mineralized with Simulated Body Fluid (SBF) which increases integration
What are the 3 different types of bioreactor?
Static –> Cells die out in the middle
Spinner-Flask –> Stress causes death of cells on the outside
Rotating Well –> Good maturation throughout
How can an artificial kidney be made?
Endothelial cells are made using hiPSCs
These are infused into the renal artery of a decellularised kidney, with a uniform distribution
How can we artificially make a vascular network for engineered tissues?
Seed the scaffold with endothelial cells
Incorportate VEGF into the scaffold
Build the scaffold around a vascular bed ex vivo
Endothelial Colony Forming Cells (ECFCs) in combination with platelet lysate (which contains GFs like VEGF) will form a vascular network
Why would we potentially want to activate Quiescent Stem Cells?
As they block signalling pathways that cause degenration and ageing
So activating them more could prevent age related diseases!!
