EV1 Flashcards
1
Q
First TEM of exosomes
A
cancer/Rabies-infected cells secreting vesicles
2
Q
Reticulocytes
A
- immature RBCs produced from bone marrow haemocytoblastic stem cells
- PM TfRs bind diferric Tfs via EC domain
- FITC-125I anti-TfR antibodies
- 0 degrees: binding, no Tf/TfR internalisation
- 37 degrees: CCV internalise diferric-Tf/TfR into trypsin-resistant EE
- Fe2+ dissociation, cytosol export
- Tf/TfR dissociation at PM
- Apo-Tf released into ES
3
Q
Exosome uptake evidence?
A
- internalised TfR is acid and trypsin resistant
4
Q
Exosome evidence?
A
TfR containing EVs in cell supernatant
5
Q
Immunogold TfR labelling + TEM in sheep reticulocytes
A
Use cGold-labelled anti-TfR antibodies
1) TfR clusters invaginated in coated pits
2) at cell surface, inside endosomes
3) in ILVs; pronase-resistant
4) released post-MVB fusion
6
Q
How do they work? MHCII APCs
A
purified EVs are MHCII +ve but PM-ve
7
Q
Biological functionality: AP B-derived cells
A
- 10% nascent MHCII in recovered exosomes
- presentable MHCII complices
- incubation with T cells = immune response
8
Q
What do they contain? mast cells
A
RNAse protection: sRNAs
9
Q
EVs and RNAs
A
- 8% donor mRNA
- 248 exosome-unique transcripts
- 121 miRNAs (angiogenesis, haematopoesis, exocytosis, tumourigenesis)
10
Q
are they taken up? MC/9 and MHCI
A
- recipient mRNA is translatable
11
Q
Are they translated? HBMECs
A
- incubated with Gaussia luciferase mRNA
- translated into biologically active luciferase
12
Q
diagnostics in glioblastoma
A
- RNA mutants/variants, characteristic miRNAs
- EGFRvIII in 7/25
