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Biochemistry of Megaloblastic Anemia

by Carlo Raj, MD

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    00:00 Let’s take a look at causes of megaloblastic anemia.

    00:03 Now, once we have completed discussion of megaloblastic, I’m going to mention some of these non-megaloblasts.

    00:10 Megaloblastic anemia, now, I’m going to set this up for you and this is an important side, a lot of things that are occurring here.

    00:16 First, let’s take a look at the peripheral blood smear.

    00:18 And what do you end up seeing there? Our megaloblasts, look how large they are.

    00:23 Highly nucleated and lack of proper maturation.

    00:27 Those are RBCs.

    00:28 Can you believe that those are RBCs? Yeah.

    00:32 Or in other words, it’s primitive cells.

    00:34 And you should never have an RBC that looks like that, right? I mean, the nucleus shouldn’t be there.

    00:38 It should be central pallor, so on and so forth, but that isn’t occurring.

    00:42 Now let me set this up, I’m going to slow down here a little bit and make sure that you’re very, very familiar with these two pictures.

    00:48 And I’m going to tell you as to how to use these, okay? The top picture, for the most part, we’re going to go from the diet and we’re going to consume, let’s say, folate.

    01:00 And we’re going to then interact with B12.

    01:03 And when we interact with B12, then I’m going to walk you through what’s known as tetrahydrofolate and then have you -- Really, take a look that first picture here.

    01:13 The objective is to convert your dUMP into dTMP, right? What does that mean to you? Well you know from genetics, once again, that if you want to form DNA, you can’t have UMP.

    01:23 You can’t have uracil.

    01:25 You need to convert that U into what? Good, your thymidine.

    01:29 And so therefore, this is the objective.

    01:32 And what we are going to do here is we’re going to interplay both B12 and folate so we can form dTMP.

    01:38 Is that clear? Okay.

    01:40 So that’s the first picture and then on the second picture, I will elucidate that further.

    01:47 Okay, let’s begin at the top.

    01:48 You are now consuming folate.

    01:51 What you’re taking in is going to be that form that you see there, at least pay attention to the methyl, please.

    01:57 The F stands for folate.

    01:58 H stands for tetra.

    02:00 So there you go, tetrahydrofolate attached with the methyl.

    02:03 We don’t need methyl in order for proper action and functioning.

    02:08 So we need to find a way to remove that methyl and give it to an acceptor.

    02:13 And so, it will donate the methyl.

    02:15 Something is going to accept it and you will see this now.

    02:18 So what is this acceptor of the methyl? Because if you don’t accept the methyl, then you basically have inactive folate.

    02:25 Do you understand that? And where are you getting this from? The diet.

    02:28 Okay, so let it be greens and so on and so forth.

    02:32 So you’ve taken in the methyltetrahydrofolate, you’re going to then donate the methyl to the B12.

    02:38 Look at the B12 there.

    02:39 It says methyl B12.

    02:41 So we have to have B12 communicating or interacting with the folate so that it can accept the methyl.

    02:48 Now, what are you going to do? Now the B12 is going to donate that methyl to the homocysteine and the enzyme there is called methionine synthase.

    02:58 Wow! How appropriate is that? Quite.

    03:01 So the methyl will be donated with the help of methionine synthase, will then convert homocysteine into methionine, stop there.

    03:09 Clinically, you as a clinician, what are you going to look for in the urine to help you suspect or even perhaps confirm megaloblastic anemia being caused by B12 or folate.

    03:20 Now, what I must bring to attention is that you’re going to find hyperhomocysteinemia and hyperhomocysteinuria in both folate and B12.

    03:31 Say that three times fast.

    03:33 Ready? It’s quite fun.

    03:34 Hyperhomocysteinuria, hyperhomocysteinemia.

    03:37 So now you have tons of homocystein in the urine, but all that it's told you now is that I have megaloblastic anemia being caused only by B12? Only by folate? Or both? Both.

    03:53 So all of this tells you that your megaloblastic anemia being caused by either B12 or folate.

    03:57 But you don’t know which one.

    03:59 Why? Look, if you’re not taking your methyl, right? From the diet.

    04:04 If you’re not taking your methyltetrahydrofolate, how could you possibly convert your homocysteine into methionine? You cannot.

    04:11 So in folate deficiency, please listen.

    04:14 Fact.

    04:15 You do have hyperhomocysteinemia and you will have hyperhomocysteinuria.

    04:21 Number two, what if you didn’t have B12? If you didn’t have B12, first it wouldn’t accept the methyl.

    04:28 Secondly, methionine synthase really wouldn’t be working.

    04:32 You should know that B12 is a co-factor for methionine synthase.

    04:35 Interesting.

    04:36 So if that enzyme isn’t working and is not accepting methyl, B12 deficiency results in? Good.

    04:43 Hyperhomocysteinemia and hyperhomocysteinuria.

    04:46 All it tells you is megaloblastic anemia.

    04:48 Now, hyperhomocysteinemia from biochemistry should know about other things including your ornithine and stuff like that, but cystathionine synthase and I’m not going to bring all that here though.

    05:00 Biochem, look up cystathionine. That’s important.

    05:03 Next, all right, now if everything goes well, the methyl comes in, taken up by B12 and it's properly handled.

    05:12 You're left with tetrahydrofolate.

    05:14 Now what will you do? Well that tetrahydrofolate, if I were you, I would know some of this co-factors and this include your serine and glycine.

    05:21 And you’re going to then form what’s known as methylenetetrahydrofolate.

    05:25 This is a useable form, pay attention here.

    05:28 We have an interesting enzyme here.

    05:29 It’s called thymidylate synthase.

    05:31 Thymidylate synthase is then going to take that tetrahydrofolate, accept two of them.

    05:38 To what? Well, you have four, right? Tetrahydrofolate, it will take up two.

    05:43 And so take up the two folate and convert the dUMP to dTMP.

    05:47 You must know the enzyme, thymidylate synthase.

    05:49 I'll tell you in one second, many of you already know.

    05:52 So now, what are you left with? You’re left with dihydrofolate.

    05:56 I know you’ve heard of that, dihydrofolate.

    05:59 And what are you going to do in biochemistry with dihydrofolate? You’re going to reduce it, aren’t you? When you have a reduction type of formula or calculation in biochemistry it actually you’re going to increase it.

    06:11 Ridiculous! So anyhow, you’re going to reduce this dihydrofolate with the help of what enzyme? You all know about dihydrofolate reductase.

    06:19 There it is, look.

    06:20 We reduced it with reductase.

    06:22 So now, you took the dihydrofolate and you converted tetrahydrofolate.

    06:25 "Dr. Raj, is all this necessary?" More than you can imagine.

    06:28 Watch.

    06:29 So, my topic is B12 and folate deficiency.

    06:32 We’re going to put in the drugs here.

    06:34 We’re going to put in some pathology here.

    06:35 And then once we’re done with this picture, we’re going to move to another one.

    06:38 And I’m not going to bring that up now though, but it’s important.

    06:42 Let’s begin.

    06:43 Say that your patient has cancer, unfortunately.

    06:46 And you’re thinking about giving them an antineoplastic drug called methotrexate.

    06:50 Take a look at MTX, identify it here.

    06:53 MTX is methotrexate.

    06:55 High dose of methotrexate has been given.

    06:58 It’s mechanism of action is to inhibit the hydrofolate reductase.

    07:02 In what kind of cells? Yeah, cancer cells, right? And if you inhibit this enzyme, you can’t properly what? Recycle your folate.

    07:12 So therefore, you can’t form dTMP.

    07:15 What happens to this cancer cell? It dies, chemotherapy.

    07:19 Is it possible that methotrexate might bleed over into a normal cell? Of course.

    07:24 You already know this as well.

    07:25 You do? Uh-hmm.

    07:27 What’s the name of that rescue? Leucovorin rescue.

    07:31 What’s the name of that folate? Called folinic acid.

    07:34 All this come together? I hope so.

    07:37 So here’s methotrexate inhibits the enzyme dihydrofolate reductase.

    07:41 Okay, good.

    07:42 Now what? I can’t properly reduce my dihydrofolate.

    07:47 For all intent and purposes, you are folate deficient.

    07:50 If this bleeds into a normal human cell, eukaryotic, then you know that you’re folate deficient.

    07:56 What does this result in, please? Megaloblastic anemia.

    07:59 If it’s high dose, what are you going to give this patient to then replenish the folate in the human cells only without affecting the cancer cells? It’s called folinic acid.

    08:09 Give me the technical name or give me the, I guess, the trade name? That’s leucovorin or leucovorin rescue.

    08:17 Drug number one.

    08:18 What’s TMP? Trimethoprim.

    08:20 Who are we trying to combat here? Bacteria, right? This is an antibiotic.

    08:25 Same concept.

    08:27 Here’s the antibiotic that you’re giving.

    08:28 You’ve heard of cotrimoxazole, haven't you? That’s sulfamethoxazole plus your? Good, trimethoprim.

    08:34 So you have cotrimoxazole.

    08:35 So this is a drug that you’re going to give to kill off bacteria and inhibits dihydrofolate reductase in that bacteria.

    08:42 It kills off the bacteria.

    08:43 Unfortunately, may also cause human cells to die for the same reason.

    08:48 Is that clear? So both of these result in what kind of deficiency? Folate deficiency.

    08:53 What kind of anemia is your patient going to suffer from? Good.

    08:56 That’s your megaloblastic anemia.

    08:57 What do you want to do with this patient? Either, well as we said, leucovorin or remove the drug if need be.

    09:03 All right, change the drug, so on and so forth.

    09:06 Two down with drugs.

    09:07 Let’s talk about the third one.

    09:08 Here is my thymidylate synthase.

    09:10 Recognize it.

    09:11 You see it? Okay.

    09:12 That converts your dUMP to dTMP and it takes up two of your folates.

    09:20 So they form dihydrofolate.

    09:22 The name of this enzyme is 5-FU.

    09:25 And by FU, I mean fluorouracil.

    09:28 Hope I didn’t offend anyone, I’m sorry if I did.

    09:29 So 5-FU is 5-fluorouracil.

    09:33 That’s a drug also, antineoplastic.

    09:36 It inhibits the enzyme thymidylate synthase.

    09:39 Here once again, for all intent and purposes, please know that you’re going to then become folate deficient.

    09:44 It’s important.

    09:46 The only thing that I wish to bring to your attention, current day practice, that if you use FU, you don’t have to give leucovorin.

    09:53 In fact, if you give FU, it’s just the opposite.

    09:57 It is going to then enhance the activity, enhance the activity.

    10:01 Keep that in mind.

    10:02 And so therefore, there is no such thing as giving this patient leucovorin to save the overdose of 5-FU.

    10:10 Keep that in mind especially when you’re dealing with drugs later on in pharmacology.

    10:15 You’re trying to treat a patient with colorectal cancer.

    10:16 That’s a big one.

    10:17 Don’t forget that please.

    10:20 We’ll have discussions more and we have talked about this neoplasia as well.

    10:25 So those were the big drugs, those were the inhibitions, know those enzymes.

    10:28 There is interplay between folate and B12 and that’s the first picture.

    10:33 In the second picture, this entire biochemical pathway has nothing to do with these drugs.

    10:39 It has nothing to do with really dietary issue.

    10:41 And what it really deals with only is true B12 deficiency.

    10:47 You tell me about a patient clinically that is presenting a B12 deficiency, in which a fully deficient patient will never present with.

    10:55 What happens here? Walking wide stance, close my eyes.

    11:01 I don’t know where I am.

    11:03 And I have upper motor neuron lesion.

    11:08 All right, what the heck is all this? It’s called subacute combined degeneration.

    11:13 Let’s go step by step.

    11:15 Wide stance, I’m walking towards you but wide ataxia, ataxic gait.

    11:20 So what organ in your brain controls your coordination? Good, cerebellum.

    11:26 And so therefore, your spinocerebellar tract is knocked out, number one.

    11:31 Another one, I said I close my eyes and I’m moving, I’m swaying and vibrating.

    11:36 Not vibrating, I can’t vibrate.

    11:39 And this is what? Proprioception.

    11:42 So I don’t have -- loss of touch, loss of proprioception.

    11:47 I’m doing positive Romberg, right? I closed my eyes but I’ve lost all spatial orientation and, you know, I just can’t stand still.

    11:55 Stability has been lost.

    11:56 Which column is that? Good.

    11:58 Dorsal column, two down.

    12:00 And what’s this? Like rigidity almost.

    12:03 That’s upper motor lesion, spastic paralysis.

    12:06 That is going to be? Good.

    12:08 Lateral corticospinal tract.

    12:10 All three tracts, you must know.

    12:12 What’s this called? Subacute, slowly, combined, three tracts; lateral corticospinal, spinocerebellar and dorsal column degeneration.

    12:22 What’s degenerating in those columns or in those tracts.

    12:26 The myelin, right? Good.

    12:28 You only find that in B12 deficiency.

    12:30 You'll never find that in folate.

    12:32 Clear? What about this demyelinating disorder? Irreversible.

    12:37 Keep that in mind.

    12:38 If you don’t give a simple injection of B12 in your butt, meaning intramuscular, then understand you go through irreversible deterioration of your patient.

    12:49 That’s your patient.

    12:50 Let’s take a look at the biochemistry.

    12:52 Identify the B12.

    12:54 Interesting.

    12:55 So here, the B12 is also a co-factor for second enzyme beginning with the letter M.

    13:02 This is called methylmalonyl-CoA mutase.

    13:05 "What was the other one, Dr. Raj?" You tell me.

    13:08 That was called methionine synthase.

    13:11 If methionine synthase was knocked out, your patient with B12 results in hyperhomocysteinemia.

    13:18 This is methylmalonyl-CoA mutase.

    13:20 So what are you going to find here in this patient that you will never find in folate deficient? Good, Methylmalonic aciduria, right? How would you ever find this in folate deficiency? There is nothing, please.

    13:34 Take a look at this.

    13:35 There is nothing in this formula here that has folate in it, okay? Now, so what’s causing demyelination? Whenever you knock out an enzyme, would you tell me what happens to the proximal substrate always? Yeah, it increases.

    13:49 Keep going back.

    13:50 So we talked about MMA, which is methylmalonyl-CoA aciduria.

    13:54 And then keep going back.

    13:56 Are you going to increase propionyl-CoA? Of course you are.

    13:59 Okay. What you need to properly myelinate your nerve? Take a look at that what’s in bold here.

    14:05 You need acetyl-CoA for normal myelin synthesis.

    14:08 Uh-oh.

    14:09 Now you have too much propionyl-CoA.

    14:11 Guess what? It replaces the acetyl-CoA and you demyelinate.

    14:17 What are the tracts? Spinocerebellar, ataxic gait.

    14:21 Upper motor neuron lesion, that’s your lateral corticospinal tract.

    14:24 And positive Romberg and no vibration or touch, that is you dorsal column.

    14:29 Clear? Now the only thing that I’ll make sure that we’re clear here from biochemistry is that you are going to take up propionyl-CoA pathway.

    14:36 If everything is perfectly normal and you had enough of your, what’s known as your biotin, and you had enough of your B12 and you kept going, then you go into your TCA, tricyclic, or Krebs cycle, right? But what kind of fat would you be taking up? It’s called odd-chain fatty acids.

    14:56 Just keep that in mind because the boards love those questions about odd-chain.

    15:00 You’ve heard of long chain and short chain and medium chain, especially medium chain.

    15:04 They love that one.

    15:05 But keep in mind though, that it’s odd-chain fatty acids that you’re going to take up.

    15:08 That’s important.

    15:10 Now, I only have a little thing that I wish to bring to your attention here.

    15:13 It’s a nice little figure.

    15:14 It really is because it integrates a lot of stuff.

    15:18 It is that anytime that you find a carboxylase in biochemistry, any pathway, the co-factor here is biotin.

    15:25 Is that clear? Biotin.

    15:26 So they say that if I am able to consume, on a personal note -- if I am able to consume enough biotin and I rev up my biotin levels in my body, I’m supposed to have hair growth.

    15:36 You're yet to see that, right? So anyhow biotin is rather interesting.

    15:40 Know that is being one of those co-factors for carboxylase.

    15:44 Here however, if you have B12 deficiency, you undergo demyelination and you would never find methylmalonic aciduria in folate.

    15:52 We spend a lot of time on this figures.

    15:55 A lot of questions come from this and a lot of integration as you can see here, biochemistry, pharmacology, pathology.

    16:01 Ultimately, what are we dealing with? Macrocytic, greater than 100 MCV.

    16:05 Megaloblastic or non-megaloblastic? Good.

    16:08 Megaloblastic.


    About the Lecture

    The lecture Biochemistry of Megaloblastic Anemia by Carlo Raj, MD is from the course Macrocytic Anemia – Red Blood Cell Pathology (RBC).


    Included Quiz Questions

    1. UMP synthase
    2. Thymidylate synthase
    3. PRPP synthase
    4. Dihydrofolate reductase
    5. Ribonucleotide reductase
    1. Hyperammonemia
    2. Hyperglycemia
    3. Hyperphosphatemia
    4. Hyperkalemia
    5. Hypercalcemia
    1. Autosomal recessive
    2. Autosomal dominant
    3. Chromosomal inheritance
    4. X-linked dominant
    5. X-linked recessive
    1. Hb F
    2. Hb S
    3. Hb H
    4. Hb A2
    5. Hb A

    Author of lecture Biochemistry of Megaloblastic Anemia

     Carlo Raj, MD

    Carlo Raj, MD


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    Contradiction
    By GMU L. on 20. November 2019 for Biochemistry of Megaloblastic Anemia

    The name of the lecture was very contradicting to the material given. In addition, I felt that the explanation as everywhere there was no proper structure of things.

     
    Good job
    By Alexander B. on 22. June 2018 for Biochemistry of Megaloblastic Anemia

    Î really appreciate the Lectures, this whole Website helps a lot. But it´s annoying that the Professor always says " repeat it 3 times fast " , Theres no point in doing that , if nobody could understand him in the first place . If you cant properly spell it out /pronounce it one time, it´s no use to repeat it fast.