Apoptosis: Mechanism

by Richard Mitchell, MD

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    00:00 The Mechanism of Apoptosis.

    00:02 This is actually a really beautiful story.

    00:05 I'm only going to be able to kind of touch along the surface.

    00:08 But I would encourage you if you're very interested in this, that there are many very excellent reviews that talk about this.

    00:16 So, again just touching the surface, but so that you understand how this happens.

    00:21 There are many different ways to get into the apoptotic pathway.

    00:25 And we talked about, you know, the physiologic roles, etc.

    00:29 But there are lots of ways to get in there also via injury and other things.

    00:32 So, first we're gonna start with injury, driving apoptosis.

    00:36 So radiation or toxins or free radicals can actually, not necessarily induced necrosis but induce apoptosis, by causing DNA damage.

    00:48 One of the mechanisms that cells have a protective mechanism against cancer is that if they recognize that there's DNA damage was like, "dang" we don't want to have that DNA damage turn into permanent mutation that leads to a cancer.

    01:02 So if I have DNA damage, I'm going to activate P53.

    01:06 The guardian of the genome will come back to this when we talked about malignancy.

    01:11 But if there's DNA damage P53 gets activated, and then, if it's not able to fix the damage, it sends that cell towards apoptosis.

    01:22 It will then have activated series of what are called execution caspaces.

    01:28 Caspases, great name for a protein.

    01:31 And there's a family of these caspases that have a cysteine site that's the C, and cleave it aspartates, that's the ASP.

    01:39 So caspaces.

    01:41 There's a reason for the name.

    01:44 P53 activation in the setting of inadequate DNA repair will give rise to starting the sequence of events that will lead to that cell committing suicide.

    01:54 And it will do so bravely so that it never turns into a cancer.

    01:58 That's what's supposed to happen if we have irreversible injury.

    02:02 It may not be necrosis.

    02:03 This is now another safeguard.

    02:05 Okay, so that's one way in to starting the a apoptosis pathway, and we'll come back to the number three in a minute.

    02:14 Okay, we can withdraw growth factors.

    02:17 So remember I told you about that lactational breast.

    02:20 It's got all that estrogen, progesterone, and it's it's got all the other hormones that are driving lactation, and now we're no longer breast feeding the baby.

    02:27 We want that all to revert.

    02:29 We want the cells in that hyperplastic lactational breast to go away.

    02:33 So we withdraw those hormones.

    02:36 Another way into the system, it will act on mitochondria, to induce release of cytochrome C.

    02:46 And remember, we've talked about Cytochrome C is one of the major things that mitochondria can release that will control activation of apoptotic pathway.

    02:57 It's actually a whole lot more complicated.

    02:59 This is where the story gets really beautiful but more complicated and not necessarily worth your time.

    03:04 I've just listed some of the regulators of that cytochrome C release, and there are certain proteins in the mitochondria.

    03:13 BCL-2, BCL-x, there are others, the names they're not great names, but they will inhibit the release of cytochrome C, so I don't get apoptosis.

    03:25 And there are others that will promote release.

    03:28 And it's a nice kind of conversation between the promoters and the inhibitors, whether or not cytochrome C gets released.

    03:35 So when the mitochondria are part of the equation and they are for many of these pathways, we tightly regulate whether or not we let out that cytochrome C and start the process.

    03:46 So, the cytochrome C then will get to the actual formation of the apoptosome, which is the seven spoked wheel of death we have talked about at least two or three times, which will get us into the execution caspase pathway.

    04:01 All right, another way to get into this pathway.

    04:05 Intrinsic embryogenic signals, and this is gets very complicated as well.

    04:09 But it's that pole point about me having fingers and not a flipper.

    04:13 There are intrinsic signals that say during development, this cell, after having grown and made a tissue, will now undergo apoptosis and die.

    04:25 So there are intrinsic embryogenic signals that also thrive execution caspases.

    04:31 There are various receptor-ligand interactions.

    04:34 So there are a variety of factors, tumor necrosis factor, acting on receptors for tumor necrosis on the cell surface.

    04:42 There's FAS/FAS ligand don't need to necessarily know what that stands for, but there are ways that exogenous signals can interact with receptors on particular cells and drive their cell death.

    04:55 These are mainly used in the immune response.

    04:59 So they're going to be important players in that particular pathway, so they're all number ones getting into this pathway.

    05:07 And so this is just showing that receptor-ligand interaction.

    05:11 And they can drive either interaction with proteins that get us into the execution caspase.

    05:17 Or they can actually even have initiator caspases, things that are even more upstream.

    05:22 As they say, it gets complicated.

    05:24 Don't get too bogged down.

    05:25 But just realize there are many ways into this final pathway beginning with number three.

    05:31 Okay.

    05:32 And finally, cytotoxic T cells, they kill.

    05:36 A natural killer cells to kill by introducing Granzyme B which will start execution caspases.

    05:46 So all these things kind of come down to number three, the execution caspases.

    05:52 These are going to be a series of enzymes that have a cysteine interactive site and they cleave it aspartates, and they will cleave a whole variety of proteins.

    06:01 Okay.

    06:02 Next slide is that next step.

    06:04 So the executioner caspases will involve breaking down the cytoskeleton.

    06:11 They cleave acting polymers.

    06:13 They cleave intermediate filaments.

    06:15 They cleave a whole variety of proteins.

    06:18 They also have Endonuclease activity, so they will break down in a very rigorous way around his stones, the DNA.

    06:28 So we'll get nice bite size bits of nuclear material that has also been digested by the executioner caspases.

    06:37 The net result of breaking down the cytoskeleton and that results of breaking down the in the nucleus is that we end up with a series of signals driven again by the caspases that allow a little escape pod, if you will, of cytosol and organelles and membrane and nuclear fragments to be formed.

    06:56 That's the cytoplasmic bud, or a apoptotic bud or a apoptotic body.

    07:02 So we have a little escape pod that's taken up some bits of cytoplasm and all the organelles and some DNA, fragment of DNA, and we have that cytoplasmic bud, that is a separate now apoptotic body.

    07:16 It pinches off from the cell that's undergoing apoptosis, and there will be many of these, but it pinches off, along with its little bodies and those apoptotic bodies now have a important change from the internal side of the plasma membrane phosphatidylserine.

    07:36 Those fossil lipids that we've talked about previously in the cell biology topics will flip from the interphase to the outerphase of the plasma membrane on these apoptotic bodies.

    07:47 And that's an 'eat me' signal, for the macrophages or adjacent epithelial cells, or whatever happens to be around.

    07:54 And that adjacent cell can recognize that phosphatidylserine on the outerphase.

    07:58 And it will eat and degrade and break up this apoptotic body completely.

    08:05 And so in a very controlled passion, without a whole lot of damage, we can get rid of an entire cell, and it just gets gobbled up by its neighbors.

    08:14 And the constituents get turned into lipids and amino acids and sugars.

    08:19 So, apoptosis.

    08:23 Now you see it, and now you don't.

    08:25 This is again speaking to the fact that it's really hard to see by light microscopy.

    08:30 We have some...

    08:32 tests and acids that we could do that kind of highlighted, but it's hard.

    08:36 So anyway, it's usually single cells or small clusters of cells.

    08:40 It's not big areas of tissue, it's little tiny things.

    08:45 The cytoplasm as it gets condensed and crosslink because of the effects of the caspases will get very pyk and the nucleus becomes fragmented, so it may become pyknotic, little condensed areas of chromatin or actually fragmented karyorrhectic.

    09:02 We make cytoplasmic buds that turned into apoptotic bodies, that are rapidly phagocytosed by adjacent cells.

    09:09 And that's why it kind of, it's hard to see it.

    09:11 It just happens kind of under the radar.

    09:14 There's no inflammatory response, really important point.

    09:18 In apoptosis...

    09:20 there is no inflammation.

    09:21 In necrosis, there's lots of inflammation.

    09:24 And that's because when there's necrosis, we want to make sure that there's no infection.

    09:30 So that cell death of necrosis, we wanna make sure that we clean up everything and make sure there's no infection.

    09:36 And we'll revisit that in a subsequent series of talks.

    09:39 But in apoptosis, we want no inflammation, so it's a kinder, gentler, suicide of the cells, And it's really hard to see by light microscopy without the special studies, and we're not going to get into that.

    About the Lecture

    The lecture Apoptosis: Mechanism by Richard Mitchell, MD is from the course Cellular Injury.

    Included Quiz Questions

    1. Phosphatidylserine
    2. Phosphatidylcholine
    3. Cholesterol
    4. Phosphatidylinositol
    5. Sphingomyelin
    1. Granzyme B
    2. ...release of cytochrome C.
    3. ...activation of the BCL-2 protein family.
    4. ...release of interleukin 1.
    5. ...activation of the p53 proteins.
    1. BCL-2
    2. BAX
    3. BAD
    4. BAK
    5. TNF-alpha
    1. Lack of inflammatory response
    2. Involvement of the neutrophils
    3. Nuclear swelling
    4. Prominent histological changes on light microscopy
    5. Basophilic cytoplasm
    1. Activation of endonucleases
    2. Activation of cytoskeletal protein synthesis
    3. Activation of the apoptosome protein
    4. Release of perforin and granzyme B
    5. Recruitment of immune cells

    Author of lecture Apoptosis: Mechanism

     Richard Mitchell, MD

    Richard Mitchell, MD

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