lecture 20: stem cells and the eye Flashcards Preview

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Flashcards in lecture 20: stem cells and the eye Deck (17):
1

 What is Jane's problem ?

  • 44 year old female 
  • tripping over children's toys (night vision was not a problem at the time) 
  • a lot of car accidents 
  • vision seems good
  • can't see at night 

2

What is retinitis pigmentosa?

  • optic nerve
  • blood vessels that come from optic nerve 
  • macula is part of eye that allows us to see centrally 
  • black stuff and white stuff 
  • gradual loss of photoreceptors
  • tunnel receptors 
  • can still see centrally because that area is not affected 
  • unless central vision is affected most people don't complain 
  • retinitis pigmentosa is an inherited condition that leads to complete loss of photoreceptors

3

What are the basics of the retina?

  • contains neurons and glia
    • 6 types of neurons 
  • laminated structure
    • complex synapses 
  • sits on a layer of support cells called the Retinal Pigment Epithelium (RPE) (keep photoreceptors alive) 
  • immune privileged (the way foreign material is recognised is completely different) 
  • originates from neural stem cells very early in development 
  • retinitis pigmentosa is a condition that affects a particular protein found in rods  → they die → peripheral loss of vision → loss of cones → complete blindness 
  • no treatment 
  • people are really interested in working out how vision can be restored 

4

What are retinal degenerations?

  • photoreceptor death
    • genetic mutations affecting proteins important for photoreceptor function
    • loss of RPE cells
    • barrier to nutrient flow from underlying vasculature 
    • also seen in age related loss of vision → very applicable treatments 
    • age-related macular degeneration = the commonest cause of blindness in the western world

5

What are types of "stem cell" technology?

  • regenerative medicine
    • stem cells to replace lost cells (e.g. photoreceptors or RPE) 
    • ESCs or iPSCs 
  • trophic support of neurons 
    • provide growth factors or other environmental support to reduce loss  

6

What is the potential of embryonic stem cells in the eye?

  • can ESCs replace lost photoreceptors?
  • MacLaren et al (2006) Nature 
  • P1 cells incorporated into P1 littermate retinas 
  • age of donor plays a role in whether cells incorporated into the adult retina 
  • embryonic stem cells didn't really do anything → basically formed balls, no connections
  • want the progenitor cells as opposed to the ESCs 
  • translated P1 Nrl-gfp +/+ cells into three mouse models of retinal degeneration → were able to form photoreceptors 
  • still kind of blind
  • tested pupils - showed a response 
  • ganglion cells → demonstrated a response, not the same as normal but not the same as the blind mice  

7

What is the effectiveness of transplanting precursors?

  • restores vision
  • have to inject heaps of precursors into the retina 
  • you have 120 million photorecpetors in any eye - potentially need to replace all of these
  • progenitors formed terminals 
  • one of the biggest problems is getting connections to second order neurons 
  • increase the number of these cells → increase the amount this animal can see 

8

Do progenitor cells provide useful vision?

  • these animals have normal retina but the photoreceptors don't work 
  • make mouse choose between a side with lines and a side with no lines 
  • can test how thin/wide lines are etc 
  • more progenitor cells you put into the retina the more likely you are to restore vision 
  • very different when trying to restore vision in animal that has lost vision because of a disease process  → have had some success in these kinds of mice 

9

How do we produce retinal progenitors en masse?

  • originally got progenitor cells from embryonic stem cells 
  • can produce a whole retina in a dish
  • do over and over again
  • harvest progenitor cells from the dish 
  • take a range of ESCs → put in a suspension of matrigel → wait → embryoid body → eye field → optic vesicle → optic cup → harvest 
  • more ethically acceptable and feasible way of harvesting the type and number of cells required to perform such treatments 

10

What has been done with stem cells to restore vision?

  • ESCs and RPE cells
  • RPE cells are much easier to transplant because they are only a monolayer 
  • have been shown to restore some vision in clinical trials 
  • sits right under photoreceptors 
  • crucial for supplying photoreceptors with nutrients and vitamins (particularly vitamin A) to allow them to detect light
  • w/o them photoreceptors die 
  • recently clinical trials testing whether stem cell replacement of RPE cells can restore vision 
  • Schwatz et al, The Lancet 2012
    • two patients: stargardt's disease; Dry AMD 
    • blind in both eyes
    • implanted 50,000 cells (low number) 
    • vision: Patient 1: 0 to 5 letters; 6/240; patient 2: 21 to 28 letters 
    • proved that it was at least safe to try and use ESCs to create RPEs to try and restore vision 
    • larger clinical trial now underway 
    • perhaps more possible than  

11

How can iPSCs be used?

  • harvested skin cells
  • reprogramming 
  • iPSCs
  • ex-vivo genetic repair
  • directed differentiation 
    • photoreceptors
    • RPE
  • photoreceptors and RPE can be derived from iPSCs
  • positives
    • overcomes ethical issues
    • no issues with graft rejection
    • shown to be functional
  • drawback
    • for genetic diseases, the photoreceptors derived from iPSCs carry the same mutation as the host

12

What is regeneration of the adult retina?

  • in fish, amphibians and some reptiles, if you completely ablate the retina, within a month it will grow back 
  • why can lower vertebrates do this while we can't?
  • what are the cellular and biochemical mechanisms?

13

What cells have been implicated in repair following intense light treatment?

  • glial cells
  • they are the support cells for neurons 
  • after light induced damage, glial cells proliferate 
  • 49% of MCs re-enter cell cycle 
  • when proliferation of glial cells is prevented, regeneration does not occur 

14

How do glia form photoreceptors?

  • Wnt and Notch signalling pathways instrumental in instigating this proliferation and then sending down stem cell route 
  • what separates us from fish?
  • maybe there are inhibitory factors in humans 

15

Do mammalian glia de-differentiate?

  • answer: yes
  • karl et al (PNAS 2008) 
  • toxic treatment: intraviteal NMDA and/or GFs into mouse vitreous 
  • caused many cells to proliferate 
  • these are glial cells 
  • not many of the cells do this however → don't have enough cells dividing, less than 5%
  • don't go down the full route of dedifferentiation 
  • glial cells respond to loss of photoreceptors
    • proliferate (~30% proliferate)
    • express early markers of stem cell route
      • pax6, Hes5
    • can we trick them to keep going?

16

What is an example of trying to get mammalian glial cells to keep dedifferentiating/differentiating to photoreceptors?

  • activated retinal glial cells express opsin
  • activated muller cells (glial cells) take on a neural progenitor phenotype
    • label for pax6
  • some activated muller cells express opsin
  • you can tweak them to change into photoreceptors
  • still very crude 
  • a lot more needs to be done before this becomes viable 
  • this one is perhaps a bit dodgy 

17

summary

  • ESCs can be used to restore vision
    • issues: formation of correct connections
    • clinical trials are promising 
  • glial cells undergo de-differentiation and form photoreceptors 
    • need to understand mechanisms so as to exploit further