4 ms·

For my masters’ degree I studied extracellular free mitochondria and extracellular vesicles containing mitochondria. It’s a wild world.

by epgui 2y ago

breck 2y ago
Very interesting. Can you tell more? What was that like? What was your day to day? Wet lab/dry lab, etc? What % are extracellular? Did you study mitoribosomes at all? No detail is too small!
- epgui 2y ago
  Wet lab. It was horrible. 6 years of dishwashing and monotony. I write software now and am much happier. The numbers were very difficult to reproduce and sensitive to tiny variations in experimental conditions, to the point of mystery. My MSc took me 6 years rather than 2, and that’s with a >90% scope cut. I can say that there was a “comparable” proportion of freeMitos to mitoMPs in the samples I was studying, which were induced from 786-O and 786-O/VHL cell cultures using a variety of agonists (atypically low concentrations of A23187, compared to literature, and serum-starved cell culture medium, seemed to work best; usual concentrations of A23187 seemed very violent on the cells, which is relevant if you’re trying to ascertain anything about physiologic conditions). Shockingly, a very sizeable proportion (10-50% of EVs above a certain detection threshold, probably near 100-150nm) of total EVs were either mitoMPs or freeMitos. I did not study mitoribosomes. I was trying to optimize conditions for the generation of EVs (literature is lacking here), enrichment of samples, and flow cytometric techniques for measuring EV subpopulations. Mitochondria make everything much more complicated and even gum up the tubes, requiring special care. Experiments that measure EVs without controlling for the presence of mitochondria are IMO inherently suspect. Their prevalence will depend on sample types, but from what I’ve seen, I believe the default assumption should be that they are present in relevant amounts. I’ve observed similar subpopulations in other cell lines, but with fewer controls and validations (I was focused on 786-O cells). There is a really great paper published by L Boudreau in the journal Blood with better results than anything I ever achieved, it’s very interesting stuff: https://pubmed.ncbi.nlm.nih.gov/25082876/ https://pubmed.ncbi.nlm.nih.gov/25082876/
comboy 2y ago
So what do you think about the paper?
bilsbie 2y ago
Could we ever get mitochondria injections? Is anyone researching that?
- TaylorAlexander 2y ago
  Sure I bet you could find someone to inject you with mitochondria…
- vixen99 2y ago
  I don't know about that but mitochondria can and are modulated by a variety of factors and of course, they include dietary components. Mitochondrial Modulators: The Defender https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9953029/ https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9953029/ This paper refers to '61 mitochondrial modulators which are able to protect the mitochondria from toxic insults and/or improve mitochondrial function.'
- epgui 2y ago
  That sounds rather dangerous. Free mitochondria provoke quite intense inflammatory reactions. I’m not saying there’s nothing to see there, but if there’s anything it’s not obvious.
inglor_cz 2y ago
It is my impression that extracellular vesicles are now considered to be one of the prime suspects in the aging process, including its potential reversibility.
- epgui 2y ago
  Calling them a “prime suspect” is both true and the wrong way around. They are ubiquitous and necessary to maintain normal physiological conditions / normal function, across all cell types and in all systems. This is the reason why we are discovering that they play important roles in pathology: it’s because they’re an important part of the machinery, and things can go wrong with any part of machinery.
agumonkey 2y ago
Are they still recognized as part of the self or are they attacked by immune cells ?
- epgui 2y ago
  I would say yes to both questions, in some sense. In many ways, they don't merely get attacked by immune cells, but also participate in immune function (and pathology). At the same time, intracellular components found outside the cell, generally, tend to activate the innate immune system, in a similar way that foreign bodies do.

For my masters’ degree I studied extracellular free mitochondria and extracellular vesicles containing mitochondria. It’s a wild world.

by epgui 2y ago

For my masters’ degree I studied extracellular free mitochondria and extracellular vesicles containing mitochondria. It’s a wild world.

breck 2y ago

Very interesting. Can you tell more? What was that like? What was your day to day? Wet lab/dry lab, etc? What % are extracellular? Did you study mitoribosomes at all? No detail is too small!

epgui 2y ago
Wet lab. It was horrible. 6 years of dishwashing and monotony. I write software now and am much happier. The numbers were very difficult to reproduce and sensitive to tiny variations in experimental conditions, to the point of mystery. My MSc took me 6 years rather than 2, and that’s with a >90% scope cut. I can say that there was a “comparable” proportion of freeMitos to mitoMPs in the samples I was studying, which were induced from 786-O and 786-O/VHL cell cultures using a variety of agonists (atypically low concentrations of A23187, compared to literature, and serum-starved cell culture medium, seemed to work best; usual concentrations of A23187 seemed very violent on the cells, which is relevant if you’re trying to ascertain anything about physiologic conditions). Shockingly, a very sizeable proportion (10-50% of EVs above a certain detection threshold, probably near 100-150nm) of total EVs were either mitoMPs or freeMitos. I did not study mitoribosomes. I was trying to optimize conditions for the generation of EVs (literature is lacking here), enrichment of samples, and flow cytometric techniques for measuring EV subpopulations. Mitochondria make everything much more complicated and even gum up the tubes, requiring special care. Experiments that measure EVs without controlling for the presence of mitochondria are IMO inherently suspect. Their prevalence will depend on sample types, but from what I’ve seen, I believe the default assumption should be that they are present in relevant amounts. I’ve observed similar subpopulations in other cell lines, but with fewer controls and validations (I was focused on 786-O cells). There is a really great paper published by L Boudreau in the journal Blood with better results than anything I ever achieved, it’s very interesting stuff: https://pubmed.ncbi.nlm.nih.gov/25082876/ https://pubmed.ncbi.nlm.nih.gov/25082876/

comboy 2y ago

So what do you think about the paper?

bilsbie 2y ago

Could we ever get mitochondria injections? Is anyone researching that?

TaylorAlexander 2y ago
Sure I bet you could find someone to inject you with mitochondria…
vixen99 2y ago
I don't know about that but mitochondria can and are modulated by a variety of factors and of course, they include dietary components. Mitochondrial Modulators: The Defender https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9953029/ https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9953029/ This paper refers to '61 mitochondrial modulators which are able to protect the mitochondria from toxic insults and/or improve mitochondrial function.'
epgui 2y ago
That sounds rather dangerous. Free mitochondria provoke quite intense inflammatory reactions. I’m not saying there’s nothing to see there, but if there’s anything it’s not obvious.

inglor_cz 2y ago

It is my impression that extracellular vesicles are now considered to be one of the prime suspects in the aging process, including its potential reversibility.

epgui 2y ago
Calling them a “prime suspect” is both true and the wrong way around. They are ubiquitous and necessary to maintain normal physiological conditions / normal function, across all cell types and in all systems. This is the reason why we are discovering that they play important roles in pathology: it’s because they’re an important part of the machinery, and things can go wrong with any part of machinery.

agumonkey 2y ago

Are they still recognized as part of the self or are they attacked by immune cells ?

epgui 2y ago
I would say yes to both questions, in some sense. In many ways, they don't merely get attacked by immune cells, but also participate in immune function (and pathology). At the same time, intracellular components found outside the cell, generally, tend to activate the innate immune system, in a similar way that foreign bodies do.