Who Protects the Liver When it Faces Disease?

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00:00:12:05 Hi, I'm Chrystelle Vilfranc.
00:00:13:24 And today I would like to explore the question,
00:00:16:16 Who protects the liver when it faces a threat?
00:00:20:15 But first I'd like to talk about the Brita pitcher that I
00:00:22:29 purchased back in college.
00:00:25:02 It was pretty simple.
00:00:26:17 Whenever the filter in the pitcher needed to be
00:00:29:01 switched out,
00:00:29:29 an indicator would come on and tell me that
00:00:32:17 I needed to switch it out.
00:00:34:20 However, after having the pitcher for years,
00:00:37:11 It started malfunctioning.
00:00:39:09 Similarly, the second largest organ in our body,
00:00:42:12 the liver,
00:00:43:09 it acts as its own form of a filtering system.
00:00:46:17 See a healthy liver is able to assist in
00:00:49:07 breaking down a number of different substances,
00:00:51:10 that enter into our body.
00:00:52:28 From foods to medications and other things.
00:00:56:13 However, if the liver is threatened,
00:00:59:05 by a form of injury,
00:01:00:27 we have a different story happening.
00:01:04:06 When a form of injury threatens
00:01:05:21 a healthy liver - the fatty diet
00:01:07:28 in this case -
00:01:09:10 it can have the healthy liver transform into more
00:01:12:04 of a fatty liver.
00:01:13:18 Then if that injury persists,
00:01:16:26 that fatty liver can then progress to a more
00:01:18:29 serious form of liver disease called fibrosis.
00:01:22:12 And up until this point,
00:01:24:05 these liver disease stages are still reversible.
00:01:27:17 Meaning that our filtering system the liver,
00:01:31:00 it's able to actually repair itself.
00:01:33:05 And it can actually reduce that damage
00:01:35:29 that occurred over time,
00:01:37:21 if it's allowed the time to recover.
00:01:40:08 So the liver is actually able to regenerate itself.
00:01:44:15 Unfortunately,
00:01:45:21 if the liver continues to be threatened
00:01:48:21 with a form of injury,
00:01:50:19 What we see is that the fibrotic liver,
00:01:53:22 can then progress to a point of no return,
00:01:56:26 where it can become more diseased.
00:01:59:04 It can become a cirrhotic liver,
00:02:01:02 and that can later progress to one of the most common
00:02:03:28 forms of liver cancer,
00:02:05:13 Hepatocellular carcinoma or HCC.
00:02:09:06 See, at that point of no return,
00:02:11:20 those disease stages that follow it are irreversible.
00:02:15:12 Meaning, that our liver that's usually able to
00:02:17:22 repair damage is no longer able to do so to the
00:02:20:10 levels that it once could.
00:02:22:27 And the issue here with these irreversible diseases is that
00:02:26:20 the only form of treatment is liver transplant surgery and
00:02:31:12 being on a transplant list is not really ideal for those
00:02:34:08 patients because they're often on these lists for a very
00:02:37:22 long time.
00:02:39:05 Some of them don't actually survive to the point that they
00:02:42:03 make the top of the list.
00:02:43:22 And some of them are honestly not great candidates to even
00:02:46:13 be put on the list or to be chosen for the liver transplant
00:02:49:23 surgery.
00:02:50:28 And those of them that are good candidates or who survive to
00:02:54:05 the point that they get to that top of that list.
00:02:57:01 Not all of the time are the surgeries actually buying them
00:03:00:16 more survival time.
00:03:02:08 So there is a great need for treatment options for chronic
00:03:06:10 liver diseases,
00:03:07:12 such as these diseases that are past the point of no return.
00:03:12:10 But what if we had an indicator that warned us of our
00:03:15:12 liver's vulnerability to injury?
00:03:18:06 It's like my Brita pitcher.
00:03:20:00 What if I had an indicator that told me that my liver was
00:03:22:22 going to start malfunctioning?
00:03:25:05 And I think that I study one of those indicators or markers,
00:03:28:26 I study a very huge protein called Bruce.
00:03:32:07 And I've nicknamed Bruce, Bruce almighty.
00:03:35:00 Bruce almighty because he's involved in a number of
00:03:38:07 different cellular functions.
00:03:40:14 From inhibiting cell death to promoting the
00:03:43:27 degradation of proteins by the proteasome.
00:03:47:11 To facilitating DNA repair.
00:03:49:22 Bruce has his hands in a lot of different functions and has
00:03:53:13 proven to be very important for our bodies
00:03:55:15 and our organ systems.
00:03:58:10 But what happens when we lose Bruce?
00:04:01:00 To answer this question,
00:04:02:15 previous lab members decided to delete
00:04:05:13 both copies of Bruce in a mouse and
00:04:08:00 try to see what would happen.
00:04:10:00 Unfortunately for them,
00:04:11:24 those mice were actually embryonic lethal,
00:04:14:16 meaning that they could not be carried to term.
00:04:18:01 What they found was that the control mice,
00:04:20:23 they had normal yolk sacs and normal placentas.
00:04:24:14 However, the mice that had Bruce knocked out,
00:04:27:16 their yolk SACS and their placentas were defective.
00:04:30:29 And so they were not able to be carried to term.
00:04:34:26 And so knowing this,
00:04:36:18 those previous lab members decided,
00:04:39:00 maybe deleting both copies of Bruce
00:04:41:04 was a little bit dramatic.
00:04:43:08 And so instead of deleting both copies,
00:04:45:07 they decided to delete a single copy of Bruce.
00:04:48:13 Fortunately for them, those mice actually survived.
00:04:52:08 But unfortunately what they found was those mice develop
00:04:56:20 spontaneous liver tumors.
00:04:58:24 And that is being pointed out by these yellow arrows.
00:05:02:08 These liver tumors happened after only 10 months of age and
00:05:06:04 that's with no form of injury, no special diets.
00:05:09:10 These mice were just living and develop tumors.
00:05:12:10 This happened at an 11% incidence
00:05:14:13 compared to the wild type mice.
00:05:17:20 Those mice with a single copy of Bruce,
00:05:20:07 or heterozygous Bruce mouse,
00:05:22:10 they actually had Hepatocellular carcinoma.
00:05:26:02 And that was confirmed by our pathologist that we usually
00:05:29:09 work closely with.
00:05:30:22 And he confirmed,
00:05:31:20 these are indeed Hepatocellular carcinoma
00:05:34:12 which is interesting for us because
00:05:37:09 hepatocellular carcinoma is the fourth
00:05:39:25 most common cause of cancer related deaths worldwide.
00:05:44:00 Patients with hepatocellular carcinoma
00:05:46:03 are often asymptomatic and they can actually
00:05:49:03 be diagnosed at later stages of the disease.
00:05:52:28 And so it's usually a little bit too late for interventions
00:05:55:25 to happen.
00:05:57:08 And so with this in mind,
00:05:58:23 and knowing that Bruce seemed to be doing something in the
00:06:02:04 mouse liver, our collaborator, the pathologist,
00:06:05:09 he donated human patient samples to us.
00:06:08:25 And he asked us,
00:06:10:10 what do you guys think the Bruce levels looked like?
00:06:13:17 And so in our lab, what happened was the previous members.
00:06:17:07 They decided to incorporate a technique called
00:06:20:07 immunohistochemistry,
00:06:21:18 which is basically a staining to look at different markers.
00:06:24:26 So the brownish staining would help us detect whether or not
00:06:29:00 there was Bruce present, and what those levels looked like.
00:06:33:00 Looking at the intensity of that staining,
00:06:35:02 we can get an idea of how much Bruce is there.
00:06:38:21 And so when we stain those livers,
00:06:40:25 those liver biopsies for Bruce,
00:06:43:17 what we found in the normal tissue or the normal adjacent
00:06:46:22 tissue,
00:06:47:17 which is basically tissue that is close to disease tissue,
00:06:51:14 those normal adjacent tissues seem to have
00:06:53:27 a great level of Bruce.
00:06:56:01 Looking again at that brown.
00:06:58:01 But when we look at more chronic liver diseases,
00:07:00:13 such as hepatitis, cirrhosis, and HCC,
00:07:03:26 those livers seem to have less brown. That suggests that
00:07:07:04 there's less Bruce.
00:07:08:10 By the time we get to HCC,
00:07:10:10 we see that Bruce levels are barely detectable.
00:07:14:04 And so this told us - mouse data,
00:07:16:14 as well as this data - suggest that Bruce is probably aiding
00:07:19:28 and protecting both the mouse and the human livers.
00:07:22:25 And so it's probably pretty important.
00:07:25:18 Bruce almighty striking again.
00:07:28:12 What if we knock out Bruce in the liver?
00:07:31:08 That was the question I decided to ask when I joined the
00:07:33:28 lab, because of course,
00:07:36:08 if we had a heterozygous mouse that only had a single copy
00:07:39:12 of Bruce, they develop spontaneous liver tumors.
00:07:43:02 Of course, if I decided to knock out Bruce in the liver,
00:07:46:17 it should surely create disease.
00:07:49:26 It should be a more dramatic HCC than what those
00:07:53:13 heterozygous mice developed.
00:07:55:18 Unfortunately, for me, that didn't happen.
00:07:59:00 Those livers looked exactly like our control livers.
00:08:03:18 The Bruce liver specific knockouts,
00:08:05:11 or LKO's, to the eye
00:08:07:29 it looked like those livers were healthy livers.
00:08:10:13 They seemed normal.
00:08:12:05 And I was a bit bummed by this,
00:08:14:04 but I thought let's not panic.
00:08:17:04 What if we decide to threaten or give a form of injury to
00:08:21:15 those Bruce deficient mouse livers?
00:08:23:19 Maybe,
00:08:25:01 there could be a chance for my dissertation research.
00:08:28:23 And so I decided to subject those normal livers to a form of
00:08:32:18 liver injury.
00:08:34:09 And what I used was Diethylnitrosamine or DEN.
00:08:38:21 We decided to inject those mice, both the control mice,
00:08:42:08 as well as the Bruce deficient liver knockouts or the LKO,
00:08:46:15 with the DEN, and we decided to look at what would happen.
00:08:50:07 One of those immediate stress responses is cell
00:08:53:03 proliferation.
00:08:54:24 And what I found was that when I did another staining
00:08:58:02 of both the control, as well as the Bruce deficient livers.
00:09:02:06 I found that looking at a cell proliferation marker, in this
00:09:06:11 case Ki67,
00:09:08:08 I could determine how many cells were proliferating
00:09:11:20 in these liver tissues.
00:09:13:11 And so in the control mice,
00:09:15:13 we're seeing a little bit of cell proliferation,
00:09:18:13 but in the liver specific knockouts,
00:09:20:23 those livers that are deficient for Bruce,
00:09:22:24 have an increase of cell proliferation.
00:09:27:10 Another stress response is inflammation.
00:09:30:17 And so I decided to look at an inflammation marker and
00:09:34:18 measure its gene expression,
00:09:36:05 to see whether or not it was active or get a
00:09:39:01 suggestion of its activity.
00:09:41:04 In this case,
00:09:41:29 I used tumor necrosis factor alpha to look at its gene
00:09:45:11 expression level.
00:09:46:23 And that would give me a suggestion of how much inflammation
00:09:49:26 these livers might have.
00:09:51:19 And when I looked at the liver specific knockouts,
00:09:54:14 they had an increase of that inflammation marker,
00:09:57:08 suggesting an increase of inflammation compared to the
00:10:00:03 control.
00:10:02:03 So remember when I told you that our Bruce almighty protein
00:10:05:13 is able to help facilitate DNA repair?
00:10:09:08 The way that it's able to do this is
00:10:11:08 once there is a form of DNA damage.
00:10:13:23 If there is a form of either a double-stranded DNA break or
00:10:17:11 a single strand DNA break,
00:10:19:07 we see that the presence of Bruce is able to allow for the
00:10:23:16 recruitment of a number of different repair proteins.
00:10:27:07 So when there's a double strand DNA break,
00:10:29:04 the ATM kinase will bind to the sites of damage.
00:10:32:15 It'll be phosphorylated or activated.
00:10:35:04 And that will cause for the recruitment of the
00:10:36:26 other repair proteins.
00:10:38:20 However, when there's a single strand DNA break,
00:10:40:26 the ATR kinase mostly responds there,
00:10:43:16 and it will be phosphorylated and activated,
00:10:45:22 and its presence will allow for the recruitment of other
00:10:48:17 repair proteins.
00:10:50:10 These processes only happen when there is the presence of
00:10:54:03 Bruce.
00:10:54:28 So Bruce being present at the sites of damage allows for the
00:10:58:19 recruitment of the repair proteins and the facilitation of
00:11:01:22 efficient DNA repair.
00:11:04:28 And so I decided to look at what happens when the livers
00:11:08:21 were exposed to DEN.
00:11:11:04 I tried to look at ATM activity,
00:11:13:05 but it didn't seem that DEN was causing
00:11:15:22 any activation of ATM.
00:11:17:21 But when I looked at ATR activity,
00:11:19:20 I found that the control livers had more of an increase of
00:11:23:20 ATR activity.
00:11:25:14 This was measured again,
00:11:26:28 by looking at the brownish staining of the cells,
00:11:29:26 the brownish cells would suggest to us
00:11:32:04 that there is ATR activity.
00:11:34:07 It seemed to be increased in the control livers.
00:11:36:25 But when we take a look at the liver specific knockouts
00:11:39:02 where the liver is deficient of Bruce,
00:11:41:11 there seems to be less ATR activity indicated by less
00:11:45:07 brownish cells.
00:11:47:02 Even in livers,
00:11:48:12 Bruce is proving that it's necessary for facilitating DNA
00:11:51:16 repair.
00:11:53:08 And so since there's not much DNA repair happening in the
00:11:55:29 liver specific knockouts,
00:11:57:28 I wanted to measure the levels of DNA damage.
00:12:02:12 And what I found was that when I looked at a global DNA
00:12:05:22 damage marker, and I counted all of those positive cells,
00:12:09:11 those positive brownish cells that would tell me,
00:12:12:20 or give me an idea that there is DNA damage
00:12:15:16 happening in these cells.
00:12:17:04 What I found is that in the control mice,
00:12:19:25 of course, they get a form of DNA damage.
00:12:22:04 DEN causes DNA damage.
00:12:24:04 However,
00:12:24:29 because they have that intact ATR activity to help
00:12:28:16 facilitate DNA repair,
00:12:30:21 that DNA damage,
00:12:31:28 those levels in the control livers go down over time.
00:12:36:12 However, with the liver specific knockouts,
00:12:38:25 we get that spike of DNA damage eventually at day three.
00:12:43:21 But over time,
00:12:45:02 those livers are not able to resolve the DNA damage to the
00:12:48:22 levels of the control mice.
00:12:51:04 And that proved that Bruce liver deficiency increases DNA
00:12:54:21 damage in mice that
00:12:57:21 are exposed to DEN.
00:13:01:07 And so let's go a little bit further.
00:13:04:04 I wanted to see what would happen to those mice when they
00:13:06:15 were exposed to DEN for such a long period of time.
00:13:12:10 So we asked the question,
00:13:13:21 Are Bruce deficient livers more susceptible
00:13:16:07 to chronic liver diseases?
00:13:18:06 And so let's take a look at that last stage of liver disease
00:13:23:07 where the liver damage is still reversible.
00:13:25:22 Let's take a look at liver fibrosis and see what happens,
00:13:29:11 but let's first explain what liver fibrosis is.
00:13:32:18 Again,
00:13:33:13 the liver can get threatened with injury because it's taking
00:13:36:10 on a number of different things that are entering our
00:13:38:20 bodies.
00:13:39:21 And so when we have hepatocytes,
00:13:42:01 which are the most common cells in the liver,
00:13:45:07 they make up about 85% or more of the liver.
00:13:49:12 And so when hepatocytes are threatened by injury and they
00:13:52:04 get injured, they will release pro inflammation signals.
00:13:56:13 Those pro inflammation signals will cause for the
00:13:59:02 recruitment of immune cells.
00:14:01:17 Some of those cells are called hepatic stellate cells and
00:14:05:28 they're normally quiet or inactive. However,
00:14:09:06 when we get the recruitment of all of those immune cells,
00:14:12:00 the normally quiet hepatic stellate cells become activated.
00:14:16:14 And that activation of hepatic stellate cells or HSCs is
00:14:19:29 important because that's actually the biggest contributing
00:14:22:23 factor to liver fibrosis because activated hepatic
00:14:26:10 stellate cells,
00:14:27:13 they overproduce the extracellular matrix components.
00:14:31:04 We're talking about collagen, etcetera.
00:14:35:00 All of those different components that are in the
00:14:37:08 extracellular matrix,
00:14:38:22 the overproduction of them is what causes for the scarring
00:14:42:02 of the liver that we often find in liver fibrosis.
00:14:46:16 So when we looked at the livers of the mice that were
00:14:49:00 exposed to DEN we found that as early as six months of
00:14:52:14 exposure to DEN the liver specific knockouts
00:14:55:15 seemed to have an increase of collagen deposition.
00:14:58:21 I measured collagen deposition by staining those livers
00:15:02:09 with a technique called Sirius Red staining.
00:15:05:03 So the Sirius Red staining allows for the collagen that's
00:15:09:09 in the liver to be stained with this reddish color.
00:15:13:11 And so when I looked at the reddish color,
00:15:15:14 I see that there is more in the liver specific knockouts
00:15:18:13 indicating more of an increase of collagen deposition.
00:15:24:18 When we looked at collagen deposition,
00:15:26:22 in the mice that were exposed to DEN for eight months.
00:15:30:22 We see that it's also increased compared to the controls.
00:15:35:27 But let's look at those activated HSCs or
00:15:38:23 Hepatic Stellate Cells.
00:15:40:13 Remember those are the cells that contribute to the
00:15:43:01 overproduction of the extracellular matrix or ECM.
00:15:47:18 And that is what is usually promoting fibrosis.
00:15:51:14 And so when we stained liver tissue samples of the mice that
00:15:55:25 were exposed to DEN for activated HSCs,
00:15:59:25 we find that there's more brownish staining in the liver
00:16:02:17 specific knockouts that brownish staining is letting us know
00:16:05:24 that there's more of an activation of hepatic stellate cells
00:16:08:27 in this case.
00:16:11:04 And so I decided to stay at that time point at eight months
00:16:14:13 of exposure,
00:16:15:18 and I took some of the livers and I prepared RNA from them.
00:16:20:00 And I sent their RNA to be sequenced because I wanted to
00:16:23:12 look at, what genes might be up-regulated or down-regulated?
00:16:28:00 In those livers at that specific time point.
00:16:30:21 And I found that there was an increase of both inflammation,
00:16:34:00 as well as extracellular matrix or ECM related markers in
00:16:38:07 the liver specific knockouts.
00:16:40:23 When I looked at more fibrosis related markers,
00:16:43:09 I found that there was also an increase of those markers in
00:16:46:10 the livers that were deficient for Bruce.
00:16:50:25 And so I took it a step further and looked at that final
00:16:53:17 time point of 14 months and found that there was still this
00:16:57:06 activation of the hepatic stellate cells.
00:16:59:28 And so that's likely what's contributing to fibrosis
00:17:03:08 susceptibility in Bruce liver deficient mice.
00:17:07:06 And so those mice seem to be more susceptible to fibrosis
00:17:10:06 than their controls.
00:17:12:10 And so let's move past that
00:17:14:04 reversible disease and get to the point of no return.
00:17:17:16 Let's look specifically at tumor formation and getting into
00:17:21:28 HCC development.
00:17:25:00 And what I found was that after eight months of exposure to
00:17:28:20 DEN the Bruce liver specific knockouts
00:17:31:23 develop tumors,
00:17:33:13 and what happened was that the control mice did not develop
00:17:37:08 tumors until about 10 to 12 months of exposure to DEN,
00:17:41:03 and that happened at about a 50% incidence.
00:17:44:03 But by then all of the Bruce liver specific knockouts
00:17:47:02 had already developed tumors.
00:17:49:22 By 14 months,
00:17:50:22 which was the final time point,
00:17:52:08 only 80% of the wild type mice develop tumors,
00:17:55:14 and 100% of those Bruce liver specific
00:17:57:28 knockouts had tumors.
00:17:59:17 And when we had our trusty pathologist
00:18:02:17 take a look at those livers,
00:18:03:25 he confirmed that all of the livers in the
00:18:06:02 14 month time point were actually HCC,
00:18:09:00 but this is what it looked like.
00:18:11:16 If you take a look by your eye,
00:18:13:19 the liver specific knockout livers
00:18:15:21 look more engrossed with tumors.
00:18:18:06 It looks a little bit dramatic or exacerbated if you will.
00:18:23:13 And so I decided to look at the histology of those livers.
00:18:27:05 And so I did a special staining.
00:18:28:28 In this case,
00:18:29:23 I'm doing a Hematoxylin and Eosin staining.
00:18:32:26 And that would give me an idea of just what the liver
00:18:35:18 histology looks like in these two genotypes.
00:18:39:01 What I found was that the liver specific knockouts,
00:18:41:20 they had more signs of fatty liver
00:18:44:07 and that's shown by those white balloonish looking figures.
00:18:48:20 Additionally, they also had something called
00:18:51:14 a trabecular staining pattern.
00:18:53:17 Which is a disruption of the hepatic plate.
00:18:56:28 And that is demonstrated by that dark purple
00:19:00:17 lining that's going out.
00:19:02:18 And what we found was that that trabecular staining pattern,
00:19:06:08 that disruption was actually a key component that healthcare
00:19:11:14 providers use when they're looking at human HCC and it's
00:19:15:09 actually correlated with poor prognosis in humans.
00:19:20:04 And so I decided to look at one of those
00:19:21:25 stress responses again: cell proliferation.
00:19:24:19 I found that as early as three months of DEN exposure,
00:19:29:06 this is before tumors form,
00:19:31:03 there's already this increase of cell proliferation in the
00:19:33:28 liver specific knockouts compared to the controls.
00:19:37:16 At 8 months,
00:19:38:15 which again is the time of tumor onset
00:19:40:29 in the liver specific knockouts,
00:19:42:20 we're still seeing this increase of cell proliferation.
00:19:47:10 When we look further to the mice that already have HCC,
00:19:51:00 and we decided to stay in those livers, we saw again,
00:19:54:04 there was this increase of cell proliferation.
00:19:58:11 So, so far I've shown you that when there is Bruce liver
00:20:01:15 deficiency in the mice, those mice develop tumors faster.
00:20:06:08 And it seems that they are having more of an exacerbated HCC
00:20:10:16 phenotype.
00:20:11:26 But, what other factors are contributing to their HCC?
00:20:16:07 And so,
00:20:17:04 one of the factors I wanted to look at was the presence or
00:20:20:04 the mutations of different oncogenes.
00:20:22:19 Oncogenes are genes that usually drive cancers.
00:20:26:18 One of the oncogenes that I wanted to look at in this case
00:20:29:13 was beta-catenin because beta-catenin is found to be mutated
00:20:34:06 in up to 50% of human HCC patient samples.
00:20:38:27 And that mutation is normally an activating mutation of
00:20:42:15 Beta-catenin.
00:20:45:26 And so when I looked at the livers that were exposed to DEN
00:20:49:13 and I stained those livers for beta-catenin,
00:20:52:17 I found that there was an increase of nuclear positive
00:20:55:10 beta-catenin cells.
00:20:57:04 That told me that there was a suggestion that the
00:21:00:12 beta-catenin is active
00:21:02:21 because when beta-catenin is activated,
00:21:04:29 it can be stabilized in the cytoplasm
00:21:07:20 and it can also move to the nucleus.
00:21:10:14 When I looked at the time of tumor onset for the liver
00:21:14:03 specific knockouts,
00:21:15:04 I found that there was still this increase of nuclear beta
00:21:17:26 catenin in the liver specific knockouts compared to the
00:21:21:07 controls.
00:21:23:23 And when we get to the final time point at 14 months,
00:21:27:09 when those controls,
00:21:28:20 as well as the liver specific knockouts have already
00:21:31:04 developed HCC,
00:21:32:28 we're still seeing that there's more of nuclear positivity
00:21:36:12 of beta-catenin in the liver specific knockout tumors
00:21:39:16 compared to the control tumors.
00:21:43:14 And so I wanted to look at the gene expression of beta
00:21:46:09 catenin and I found that there was an increase of gene
00:21:49:11 expression in the liver specific knockouts,
00:21:51:24 by the time those mice have HCC.
00:21:55:07 But I also looked at the protein levels.
00:21:57:11 So it wasn't just happening at the mRNA level.
00:22:00:16 It's also happening at the protein level.
00:22:02:11 There's more beta-catenin protein being produced
00:22:05:14 in the liver specific knockouts compared to their controls.
00:22:09:19 Meaning that beta-catenin gets to the nucleus.
00:22:12:05 It's able to act as a transcription factor and it's able to
00:22:15:21 promote the transcription of other genes.
00:22:18:16 And so one of its targets that it's able to promote its
00:22:21:14 transcription is cyclin D1.
00:22:23:22 And I measured the protein levels of cyclin D1,
00:22:26:16 and found that those protein levels were
00:22:29:05 seen to be increased in the liver specific knockouts
00:22:32:14 compared to the controls.
00:22:35:02 And so I took it a step further - instead of just looking at
00:22:37:26 the mice and what happens with them.
00:22:40:10 I decided to look at a human HCC cell line.
00:22:45:07 And I decided to look at the phosphorylation of beta-catenin
00:22:47:23 at serine 675.
00:22:49:23 This phosphorylation site is important because that is what
00:22:53:01 allows for beta-catenin to be stabilized as well as moved to
00:22:56:08 the nucleus.
00:22:57:08 And it gives us more of a suggestion that beta-catenin is
00:23:00:05 activated.
00:23:01:08 So when we measure the levels of phosphorylation of beta
00:23:03:28 catenin at serine 675,
00:23:06:06 we see that it is increased in those knockout cells,
00:23:10:01 those human HCC knockouts,
00:23:12:14 and that tells us that without Bruce,
00:23:15:08 there is this increase of beta-catenin being stabilized and
00:23:19:02 that's causing it to be hyperactive compared to the control
00:23:22:25 cells.
00:23:24:18 And so I wanted to ask,
00:23:26:23 Is Bruce a good marker for measuring chronic
00:23:29:04 liver disease susceptibility?
00:23:31:16 And so when we look at the highlights of this study,
00:23:34:12 we find that as early as 3 to 4 months of exposure to DEN,
00:23:38:25 the liver specific knockouts already have an increase of
00:23:41:19 cell proliferation.
00:23:44:00 Additionally,
00:23:45:04 the liver specific knockouts seem to have an increase of
00:23:48:07 nuclear beta-catenin,
00:23:49:19 which is the oncogene that we know promotes HCC development
00:23:53:13 in humans.
00:23:55:26 As early as six months of exposure to DEN,
00:23:58:24 the liver specific knockouts also developed fibrosis.
00:24:03:16 By 8 months,
00:24:04:20 they develop tumors and we did not see tumors in the
00:24:08:19 control mice until 10 months of exposures to DEN.
00:24:12:26 By the end time point of 14 months, both the
00:24:15:23 normal mice or control mice had HCC,
00:24:19:03 as well as the liver specific knockouts.
00:24:22:03 However,
00:24:23:07 I hope you remembered the control mice only developed HCC at
00:24:27:15 80% incidence compared to the 100% incidence we
00:24:31:03 found in those livers that were deficient of Bruce.
00:24:34:28 Not only did they all develop HCC,
00:24:38:08 but they seem to have more of an exacerbated HCC phenotype
00:24:41:27 compared to the controls.
00:24:44:28 And so what's next? What did we learn from this study?
00:24:48:28 So not only did this study show us or demonstrate to us that
00:24:53:04 our Bruce almighty protein is able to help
00:24:55:23 and protect in the liver.
00:24:57:26 We should all consider Bruce
00:24:58:24 to be a marker or an indicator.
00:25:01:12 It could potentially help us tackle the issues that we have
00:25:05:00 with liver disease in this country.
00:25:08:19 With that I'd like to thank the iBiology team.
00:25:11:27 And I'd like to thank my lab both past and present, my
00:25:15:04 graduate program, as well as all of my funding. Thank you.

This Talk

Speaker: Chrystelle Vilfranc

Audience:

Student
Educators of Adv. Undergrad / Grad
Researcher

Recorded: November 2020

Talk Overview

Chronic liver diseases affect millions of people worldwide. By understanding how liver disease progresses, we may be able to identify new therapies that can protect the liver. Dr. Chrystelle Vilfranc studied the role of BRUCE, a protein that is known to be important in several cellular processes in our bodies, in liver disease. She found that the absence of BRUCE in mouse livers led to accelerated liver disease and higher rates of liver cancer when combined with a liver damaging compound. Furthermore, hepatocellular carcinomas that develop in the absence of BRUCE in the liver appear to have increased β-catenin activity. Loss of BRUCE may be a marker of early liver disease in humans, and rescuing BRUCE expression or activity may help stop or reverse disease in the liver.

Speaker Bio

Chrystelle Vilfranc

Dr. Chrystelle Vilfranc received her PhD in Cancer and Cell Biology from the University of Cincinnati. After graduating, she was a 2021 AAAS Mass Media Fellow at the Indianapolis Star. She is now a postdoctoral fellow at Columbia Mailman School of Health, investigating cancer-related population sciences. Her Blog Continue Reading

Credits

Eric Kornblum (iBiology): Videographer, editor
Brittany Anderton (iBiology): Producer
Chris George (iBiology): Graphics
Maggie Hubbard (iBiology): Graphics

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Chrystelle Vilfranc

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