Successfully reported this slideshow.
We use your LinkedIn profile and activity data to personalize ads and to show you more relevant ads. You can change your ad preferences anytime.

Pizza club Zoé 28.09.16

93 views

Published on

Pizza club Zoé 28.09.16

Published in: Science
  • Be the first to comment

  • Be the first to like this

Pizza club Zoé 28.09.16

  1. 1. Pizza Club Zoé Hanss 28.09.2016
  2. 2. What are prions ? • Kuru, Creutzfeldt–Jakob disease, Scrapie… → Caused by a new pathogenic element • Non viral or bacteriologic • Resistant to standard decontamination • Transmission process unclear β-sheetsα-helix Native protein Alternative shape SOLUBLE Aggregates PrP PrPSc
  3. 3. Misconformed protein Fragmentation Propagation Nucleus Aggregates New nucleus Native Protein Limited step Spontaneous aggregation Aggregatesnumber Temps Prion mechanism Latency Nucleus formation
  4. 4. Misconformed protein Fragmentation Propagation Nucleus Aggregates New nucleus Native Protein Limited step Spontaneous aggregation Aggregatesnumber Time Exogenous nucleus Decreased latency Exogenous nucleus Prion mechanism
  5. 5. Parkinson’s disease Multiple system atrophyAlzehimer’s disease Prion Disease Aβ Amyloïdes plaques Tau Tangles Prion PrPsc aggregates α-synuclein Lewy Body α-synuclein Glial cytoplasmic inclusions
  6. 6. Multiple System Atrophy (MSA) • Sporadic, adult-onset, neurodegenerative disorder • Incidence 3/100 000 (>50 Yo) • Progressive loss of autonomic nervous system function • Signs of parkinsonism • Glial cytoplasmic inclusions (GCI) of filaments of α-synuclein Prion behaviour ?
  7. 7. Previous experiment (2013) 2 MSA cases TgM83+/- α-syn A53T 120 days Progressive CNS dysfunction Phosphorylated α–syn in neurons Are these 2 cases real “transmission” ? 2) Transmission to TgM83+/- :1) Transmission to HEK cells - 1 control brain - 6 PD brains - 14 MSA brains Spontaneous aggregation Aggregatesnumber TimeDecreased latency Exogenous nucleus Brain homogenates Mouse model prone to form α-syn aggregates
  8. 8. Characterisation of samples 1) Neuropathological characteristic of patients samples SN depigmentation Phosphorylated α-syn staining GCILB
  9. 9. By ELISA Characterisation of samples 2) Total α-synuclein amount
  10. 10. FC: Frontal cortex SN: Substancia nigra BG: Basal ganglia P: Pons MSA/PD : higher amounts of phosphorylated α-syn than controls Characterisation of samples 3) Level of insoluble α-synuclein
  11. 11. Experiment 1 : Transmission of MSA prions to cultured HEK cells HEK cells expressing full length α-synuclein containing the A53T mutation fused to yellow fluorescent protein : exposed for 4 days to precipitated sample from brains 4 days Sodium phosphotunstic acid (PTA) Precipitation of α-syn Imaging Confocal fluorescence microscopy A53T mut YFP - 1 control brain - 6 PD brains - 14 MSA brains
  12. 12. MSA samples induce aggregate formation in HEK cells Experiment 1 : Transmission of MSA prions to cultured HEK cells No aggregates with control brains No aggregates with PD brains Aggregates with all MSA brains
  13. 13. 30 µl of 1% homogenate Right parietal lobe 2 month old Checked for neurological illness twice a week Experiment 2 : Transmission of MSA prions to TgM83+/- mice Brain homogenates - 1 control brain - 6 PD brains - 14 MSA brains
  14. 14. Inoculation with all MSA samples caused CNS dysfunction at 100-150 dpi: dysmetria and circling behavior PD samples : no sign of neurological dysfunction at >360 dpi Aggregates of phosphorylated a-syn Astrocytic gliosis Experiment 2 : Transmission of MSA prions to TgM83+/- mice
  15. 15. Contralateral hemisphere De novo formation of α-synuclein aggregates in contro-lateral hemisphere of mouse brain Experiment 2 : Transmission of MSA prions to TgM83+/- mice
  16. 16. TgM83+/- MSA TgM83+/+ Primary transmission Experiment 3 : Serial transmission of MSA prions to TgM83+/- mice Mouse model prone to form α-syn aggregates Mouse model spontaneously forming α-syn aggregates
  17. 17. Shorter incubation time with MSA inoculated mouse brain Experiment 3 : Serial transmission of MSA prions to TgM83+/- mice
  18. 18. TgM83+/- MSA TgM83+/+ Primary transmission Secondary transmission TgM83+/- TgM83+/- Experiment 3 : Serial transmission of MSA prions to TgM83+/- mice
  19. 19. Same incubation time: replication of MSA prions at same level Experiment 3 : Serial transmission of MSA prions to TgM83+/- mice
  20. 20. TgM83+/- WT Tg(SNCA) Snca0/0 Experiment 4 : Role of endogenous α-syn
  21. 21. A53T mutation facilitate prion replication TgM83+/- WT Tg(SNCA) Snca0/0 Experiment 4 : Role of endogenous α-syn
  22. 22. IC IM Intraglossaly Incubation time 133 ± 6 days 136 ± 6 days > 220 days MSA Experiment 4 : Role of inoculation routes
  23. 23. Discussion • All 14 human brain samples from MSA patients could be transmitted to both HEK culture and Tg mice expressing α-syn A53T mutation • At least 2 strains of α-synuclein prions : MSA and TgM83+/+ • MSA and PD prion strains are different • Important to note that transmission of MSA prions requires A53T α- synuclein mutation • Health care issues
  24. 24. Thank you for your attention !

×