Case 11

Doping
Plasma, 55% of blood
• What is blood?
Connective tissue, composed of cellular elements suspended in fluid
matrix called plasma, responsible for carrying the materials from one
part of the body to another.
• Plasma / yellowish fluid matrix of blood, where its cellular elements
are suspended.
• What is the main component of plasma?
Water accounts for 92% of its weight.
• Proteins? 7%
• Ions and dissolved organic molecules? 1%
Blood Functions
• What’s the average volume of blood?
5/6 liters in male, 4 /5 in female
• What are the 3 main blood functions?
Distribution, regulation, protection.
• Distribution includes
- Delivering oxygen and nutrients from to body tissues
- Transporting metabolic waste products for elimination
- Transporting hormones from endocrine organs to targets
Cont.
• Regulation includes
- maintaining appropriate body temperature, by distributing the heat
through the body and to skin to encourage heat loss.
- maintaining pH, many blood proteins work as buffers
- preventing blood loss, via clot formation
- preventing infection
Cont.
• What is the difference between the composition of plasma and
interstitial fluid?
The presence of plasma proteins.
• What are the most abundant type of proteins?
Albumins, 60% of the total.
• Plasma proteins are synthesized where?
In liver.
• What synthesizes immunoglobulins?
Specialized blood cells.
Cont.
• The presence of proteins makes the osmotic pressure of the blood
higher than that of interstitial fluid.
• What are some functions of plasma proteins?
Cellular Elements
• Erythrocytes, leukocytes, and platelets (thrombocytes) are the 3 main
cellular elements found in blood.
• What are the only fully functional cells in circulation?
WBC.
• Platelets are cell fragments that have split off from?
Megakaryocytes.
• Most blood cells don’t divide, how are they replaced?
Stem cells divide continuously to replace them.
• Erythrocytes are the most abundant cellular elements.
Erythrocytes
• Small cells, 7.5 um, biconcave, appear lighter in color at center.
• RBCs proteins
- HB – gas transport
- antioxidant enzymes – get rid of harmful free radicals
- spectrin – structural protein, allow erythrocytes to change shape when
necessary and resume their concave shape
• What are the structural characteristics of RBC that contribute to its gas
transport function?
- its small size & biconcave shape provides huge surface area relative to
volume
- it’s composed of 97% Hb
- they lack mitochondria, thus don’t consume any of the oxygen the carry
Cont.
• Hemoglobin is made up from?
4 Red heme pigment bound to protein globin.
Iron is bound to heme
• Globin consists of?
4 polypeptide chains, 2 alpha, 2 beta
• Hemoglobin molecule can transport 4 Oxygen only?
Because each iron can combine reversibly with one O.
• Single RBC contains 250 m HB.
• How can the binding of HB to RBC be beneficial?
It prevents it from breaking into fragments, and raising blood osmotic
pressure.
Cont.

• Where does CO2 bind in HB?

It binds to globin amino acids, it’s
called carbaminohemoglobin.
Hematopoiesis Production of erythrocytes
• Red bone marrow is composed largely of reticular soft connective
tissue, bordering on sinusoidal capillaries. As blood cells mature, they
migrate via … the sinusoidal capillaries..
• In adults, RBM is found chiefly in?
proximal epiphysis of humerus and femur.
• All blood cells arise from? Hematopoietic stem cells.
• The commitment of stem cell to specific blood cell type is signaled
by?
Membrane surface receptor.
Cont.
• The process consists of 2 phases:
1- EPO dependent: maturation & proliferation of progenitors
2- Iron dependent: differentiation of erythrocytes
• Basophilic erythroblasts : produces huge number of ribosomes.
• During which phases the cells divides many times?
Proerythroblasts and basophilic erythroblasts.
• Hemoglobin is synthesized and iron accumulate as the cell transform
into?
From basophilic to Polychromatic erythroblasts and then into
orthochromatic erythroblast.
Cont.
• The orthochromatic erythrocyte becomes red because it has accumulated
all of its hemoglobin.
• What happens to orthochromatic erythroblast when it degenerate its
nucleus?
Cell is collapsed inward, resulting in reticulocyte. 1-2% of all EC
• Reticulocyte: contains reticulum of clumped ribosomes.
• The entire process from hematopoietic stem cell till reticulocytes takes
about 15 days.
• As reticulocytes enter the bloodstream, their ribosomes are degraded by
intracellular enzymes and they become mature erythrocytes within 2 days
of release.
Cont. PB PORE
Regulation and requirements of
erythropoiesis
• Regulation of the number of erythrocytes is very important. Why?
Because having too much RBCs makes the blood viscus, while too little
causes hypoxia.
• The process of erythropoiesis is controlled by hormones,
depends on adequate supply of iron, amino acids, certain B vitamins.
• What is erythropoietin?
Glycoprotein hormone, stimulate production of RBC. Produced by
kidneys.
• What happens when certain renal cells become hypoxic?
Oxygen sensitive enzymes won’t be able to degrade hypoxic inducible
factor HIF, which accelerate the production of EPO.
Cont.
• What are the triggers of EPO formation?
- reduced number of RBCs due to hemorrhage
- insufficient HB per RBC
- reduced availability of O2
• Too many erythrocytes can depress erythropoietin production.
• Thus what controls the rate of erythrocytes production?
The ability of RBCs to transport oxygen.
• The blood carried EPO stimulate which cell types to become RBCs?
Red marrow cells that are already commited to become RBCs
Cont.
• Renal failure can lead to reduction in EPO production, thus less
erythrocytes.
• What is the consequence of injecting EPO?
Increase of normal hematocrit from 45% up to 65%, then with
dehydration, the blood concentrate and become viscus which can
lead to clotting stroke and heart failure.
• Testosterone also increases the kidneys’ production of EPO.
• What are the raw materials required for erythropoiesis?
The usual nutrients and structural materials( a.a, carbs, lipids)
• Iron is essential for hemoglobin synthesis.
Iron Recycling
• 65% of the body’s iron supply is in? HB, reminder is stored in liver,
spleen, BM.
• Iron is stored within the cells as…?
Protein- iron complexes, e.g. ferritin, hemosiderin.
• Which protein mediate iron transport in blood?
Transferrin.
• In women, menstrual flow accounts for additional loss of iron ( daily
average of 1.7mg in comparison with males 0.9 mg
Iron recycling
• Human have approximately 4-5 grams of iron in the body.
• Where is iron found?
Most of it is found in HB (2.5 g), 2 g bound to ferritin, in BM spleen
and liver, 0.4 in muscles with myoglobin.
• Thus, most of iron is found in blood.
• Blood plasma of iron:
neonates : 17-24
women : 10 – 25
men : 14 – 28
• higher values iron accumulation (haemochromatosis )
Cont.
• Where is iron stored?
Bound to ferritin and hemosiderin in the macrophages of liver. Spleen
and BM
• What’s the daily need of iron for men?
1 mg , while women 1-2 mg and pregnant 2.5 mg.
• Iron is present in 2 forms in food which are:
- heme in meat and trivalent iron complex in vege Fe3.
• How is the non-heme iron absorbed?
First it is reduced to divalent iron by ascorbic acid (vitamin C), and this
form can be absorbed.
Cont.
• The divalent iron ( ferric iron) is absorbed in?
duodenum, where special receptors for the divalent iron are present.
• What is the fate of the absorbed iron?
Bind with ferritin or exported to the blood stream.
• Your note + toto.
Fate and destruction of RBCs
• Life span of RBCs?
100 to 120 days.
• When does RBCs become old?
When they lose their flexibility, become rigid and fragile, and their HB
begins to destroy.
• Where do they become trapped and fragmented?
In small circulatory channels, i.e. spleen.
• Bilirubin?
Yellow pigment that is release to blood and bind to albumin for
transport.
Cont.
• RBCs are broken down by?
Kupffer cells in liver and
spleen.
• Components are split :
- Heme / iron and bilirubin
- globin
Erythrocytes disorders
• Anemia
blood oxygen carrying capacity is too low, sign of other disorder and
its hallmark is inadequate O2 levels.
• Causes of anemia can be divided into 3 gp:
blood loss, not enough RBCs, vigorous destruction of RBCs.
1- Hemorrhagic anemia:
caused by blood loss, in acute/rapid = it is treated by replacing lost
blood.
In chronic blood loss, once the primary cause is treated, normal
erythropoietic mechanisms restore the lost RBCs
Cont.
2-Inadequate no. of RBCs
can be caused by numerous problems, ranging from lack of essential
raw material and complete failure of RBM.

- iron deficiency anemia:

second result of hemorrhagic anemia, can also be result from
inadequate iron- food intake & impaired absorption.
The erythrocytes produced are called? microcytes
small and pale because the cannot synthesize HB.
Cont.
• Pernicious anemia
autoimmune disease that most often affect the elderly.
- Caused by?
Destruction of stomach mucosa that contains substance called
intrinsic factor, which must be present for production of B12.
- what is the importance of B12?
Without it, the developing erythrocytes grow but cannot divide,
which result in pale macrocytes
Cont.
• Hemolytic anemia
Erythrocytes rupture, or lyse prematurely.
- Caused by?
Production of abnormal hemoglobin
* Thalassemia :
- occurs in Mediterranean people
- one of the B chains are lost or faulty which cause deficiency in HB &
erythrocytes are thin and delicate.
- classified according to which HB chain is affected.
Cont.
• Sickle-cell anemia
- caused by?
*Abnormal hemoglobin, resulting from change in one of the 146 a.a in
beta chain of globin.
* This alteration causes the 2 beta chain to link together- under low
O2 levels- , forming sharp HB, which in turn cause RBC to become
crescent in shape.
• How can this affect the RBCs?
When oxygen content if blood is low, RBCs become crescent shape,
these deformed RBCs rupture easily and obstruct small blood vessels
and the O2 delivery is interrupted.
Cont.
• Why would such a dangerous genetic trait persist in a population?
Individuals with 1 copy of (sickle- cell trait) have a better chance in
surviving malaria.
- How?
Their RBCs becomes sickle shaped only under vigorous condition such
malaria. Sickling reduces the parasite ability to survive and enhances
macrophages ability to destroy infected RBCs.
• Treatment? Not an ILO.
Cont.
• Polycythemia
abnormal excess of RBCs, increases blood viscosity.
1- polycythemia vera: bone narrow cancer, characterized by
increased RBCs count and hematocrit as high as 80%, which impair
the circulation.
2- secondary polycythemia : results when less O2 is available , e.g.
high altitude.
3-blood doping : artificially induced polycythemia.
Some of athlete RBCs are drawn off and stored, which induces
erythropoiesis to replace them, then the stored blood is reinfused
again which result in temporary polycythemia.
• Blood doping consists of any means used to increase and optimize
oxygen delivery to the muscles
- includes blood transfusions, administration of
erythropoiesis-stimulating substances, blood substitutes, natural or
artificial altitude facilities, and innovative gene therapies
Leukocytes
• The only formed elements that are complete cells with nuclei and
usual organelles, form mobile army.
• Less than 1% of blood volume.
• WBC count for over than 11.000 cells/ul is called?
Leukocytosis, normal homeostatic response to infection.
• Leukocytes are grouped under 2 major categories on basis of
chemical and structural characteristics:
1- Granulocytes: contains membrane-bound granules
2- agranulocytes: no membrane bound granules
Granulocytes
• (Neutrophils, Eosinophils,
basophil) Spherical in shape.
• Larger, have much shorter life
span
• What is the general
characteristic of their nuclei?
They have lobed nuclei, which
are rounded nuclear masses
connected by thinner masses of
nuclear materials.
Cont.
• PMN Neutrophils
the most numerous WBCs, 50-70% (62%)
- why are they called neutrophils?
Because they take up both (acidic) red and (basic)
blue stain, which gives them their lilac color.
- Their granules contain what?
Lysosomal enzymes and antimicrobial proteins called
defensis.
- they defense against?
Fungi and bacteria
via process called respiratory burst, where the cell
metabolize oxygen to produces potent-germ
oxidazing agent.
Cont.
• Eosinophils 2.3%
- shape of nucleus?
Two lobes connected by band of nuclear
materials.
- cause of name?
large coarse(harsh) granules, stain with eosin
acid that forms brick red color.
- granules filled with?
Digestive enzymes.
- Role?
Defend against parasitic worms that are too
large to be phagocytized.
Cont.
• Basophil
- the rarest, 0.5-1%
- purple nucleus with U or S shape
cause of name?
their cytoplasm contain large histamine
containing granules, have affinity for basic
dyes. Baso – phil.
- mast cells are granulated cells similar to
basophil, exist in connective tissues
Agranulocytes ( lymphocytes and monocytes)
• Lymphocytes
30%
- shape?
Has large dark purple nucleus that
occupies most of its volume ,
surrounded by thin pale cytoplasm.
- cause of naming?
Because they’re closely associated
with lymphoid tissues.
T lymphocytes : acts directly on
viruses
B lymphocytes : give rise to plasma
cells which produces antibodies
Cont.
• Monocytes
3-8%, the largest leukocytes.
- Nucleus?
U or kidney shaped
- Role?
When they leave the bloodstream and
enter tissues, they differentiate into
macrophages.
- Macrophages?
- Phagocytes that are crucial to body
defense against viruses, bacterial
parasites and chronic infections.
- activate lymphocytes
Leukopoiesis
• What are the 2 groups of hematopoietic factors that stimulate
leukopoiesis? ( glycoproteins)
interleukins and colony- stimulating factors ( CSFs).
• Interleukins are numbered ( IL-3, IL-5), CSFs are named according to the
population that they stimulate ( Granulocytes- colony stimulating factors
(G-CSFs).
• What give rise to all of the formed elements in blood?
Hematopoietic stem cell, which branches into (lymphoid stem cell and
myeloid stem cell).
• Lymphoid stem cell: produce lymphocytes.
• Myeloid stem cell: produce all other formed elements.
Cont.
• The committed cells in each granulocytes line are called?
Myoblasts.
• Then, they accumulate lysosomes and become Promyelocytes.
• When does the distinctive granules of each granulocyte appear?
In the myelocyte stage, cell division stops.
• Then the nucleus constrict producing band cell stage.
• Why does the bone marrow contain huge reservoir of granulocytes?
Because it stores them , and they have short life span, most die
combatting invading microorganisms.
• Monocytes share common precursor with? Neutrophils.
Platelets
• Cytoplasmic fragments of large cells called megakaryocytes.
• Under microscope, how do they appear?
Their outer area stains with blue, inner area that contains granules
stain with purple.
• What does theses granules contain?
Chemical that act in blood clotting, e.g. PDGF, enzymes, ADP.
• Role of platelets?
Blood clotting.
• They degenerate quickly if they weren’t involved in clotting, because
they’re anucleate.
Cont.
• Which hormone regulate the formation of platelets?
Thrombopoietin.
• Repeated mitosis of the megakaryoblats occur without cytokinesis
which result in?
mature Megakaryoblasts.
• How are the megakaryocytes fragmented?
As they mature, they send extensions via the sinusoid wall, theses
extensions rupture, releasing platelet fragment to blood stream.
Glycolysis
• Glycolysis is a catabolic reaction. What do we mean by that?
It serve to capture chemical energy in the form of ATP from
degeneration of energy-rich molecule.
• Gluconeogenesis is anabolic reaction, what do we mean by this?
It is endergonic, requires energy which is provided by break down of
ATP to ADP and Pi.
• How can glucose enter to the cells?
1- facilitated diffusion transport system
2- Na- cotransport monosaccharide system
Reactions of Glycolysis
• The conversion of glucose to pyruvate occur in 2 stages:
1- energy investment stage ( first 5 stages)
2- energy generation stage
• What happens in energy investment stage?
ATP is used to phosphorylate the intermediates.
• What happens in energy generation stage?
Net of 2 ATP molecules are generated per 1 glucose molecule by
substrate – level phosphorylation.
1- Phosphorylation of glucose
• Irreversible reaction, traps the
glucose inside the cell.
• Phosphorylation of glucose is
catalyzed by?
Hexokinase, in most tissue.
Glucokinase in liver parenchyma
and beta cells of pancreases
• 1 ATP molecule is used.
2- Isomerization of glucose -6- phosphate

• Reversible reaction, catalyzed

by phosphoglucose isomerase
• Formation from aldose sugar to
ketose sugar
3- Phosphorylation of fructose -6-
phosphate
• Irreversible reaction, catalyzed by froctukinase 1 ( PFK-1)
• The most important control point and the rate limiting step.
• What is the meaning of rate limiting step?
It is the slowest step of chemical pathway, that determines the rate of
other reactions in the pathway.
• the PFK-1 activity is controlled by what?
The available concentration of ATP and fructose – 6 – phosphate and
regulatory substances.
Cont.
• What is the effect of energy
levels within a cell on PFK-1?
Elevated levels of ATP and citrate
inhibits it, while elevated levels of
AMP activates it.
• What is the effect of fructose
2,6-bisphosphate on PFK-1?
Activate PFK-1
4- Cleavage of fructose 1,6-bisphosphate
• Reaction is catalyzed by ? Aldolase
• Reversible, not regulated.
• Fructose-1,6-bisphosphate cleavage yields?
G3P ( Glyceraldehyde -3- phosphate) and DHAP (
Dihydroxyaceton-3-phosphate)
5 - Isomerization of DHAP
- reaction catalyzed by ? Triose phosphate isomerase
DHAP is converted into G3P.
6- Oxidation of G3P
• What is the first oxidation - reduction reaction of glycolysis ?
The conversion of G3P into 1,3-bisphosphoglycerate (1,3-BPG).
• This reaction is catalyzed by?
Glyceraldehide-3-phosphate dehydrogenase, needs 1 NAD+.
• Mutase enzyme
found in RBCs, catalyze the conversion of 1,3-bisphosphoglycerate
into 2,3-bisphosphoglycerate (2,3 BPG).
7- synthesis of 3-phosphoglycerate
• Substrate level phosphorylation.
• 1,3-BPG is converted to 3-phosphoglycerate.
• Reaction is catalyzed by?
Phosphoglycerate kinase , reversible reaction.
• 2 ATP are produced.
Gluconeogenesis
• Some tissues need continues supply of glucose as metabolic fuel.
• In the absence of dietary carbs intake, liver glucagon can meet those
needs only for 10 to 18 hours.
• Thus, glucose will be formed from precursor such as:
lactate, pyruvate, glycerol.
• During overnight fasting, 90% of gluconeogenesis occur in liver, 10%
in kidneys, while during prolonged fasting, the kidneys become the
major glucose producers.
Strategies for gluconeogenesis
• Glycerol
- released from?
Hydrolysis of triacylglyceroles in adipose tissues, delivered via blood
to liver.
1- Glycerol ( phosphorylation by glycerol kinase) Glycerol phosphate
2- Glycerol phosphate ( oxidation by glycerol phosphate dehydrogenase) DHAP
• Lactate
- released into blood by?
Exercising skeletal muscles, and cells that lack mitochondria, I,e. RBCs.
Lactate is taken up by liver, converted to glucose again.
Cont.
• Amino acids
- derived from?
Hydrolysis of tissue proteins, ( glucogenic amino acids) the major
source of glucose during fast.
• They enter the TCA cycle and form oxaloacetate, direct precursor of
phosphoenolpyruvate
Reactions of gluconeogenesis
• 7 reactions of glycolysis are reversible and are used, however 3 of
them are irreversible.
• What is the first roadblock to overcome the synthesis of glucose?
The irreversible conversion of PEP to pyruvate via pyruvate kinase.
• Carboxylation of pyruvate
Pyruvate ( carboxylation by pyruvate carboxylase) oxaloacetate
• Transport of oxaloacetate to the cytosol
OA cannot directly pass the inner mitochondrial membrane.
OA ( reduction by malate dehydrogenase) malate – transported-
malate ( reoxidation malate dehydrogenase) OA
Cont.
• Decarboxylation of oxaloacetate
Oxaloacetate ( decarboxilated and phosphorylated by PEP
carboxikinase PEPCK ) Phosphoenolpyruvate
Then, PEP is acted to the reaction of glycolysis till it becomes
1,6-bisphosphate.
• Dephosphorylation of fructose 1,6-bisphosphate
Fructose 1,6-bisphosphate ( hydrolysis by
fructose-1,6-bisphosphatase) fructose 6-phoaphate. **
Cont.
• Dephosphorylation of glucose 6-phosphate
Glucose -6- phosphate ( hydrolysis by glocuse-6-phosphatase)glucose
- only liver and kidney can do it
- this process requires 2 enzymes:
Glucose -6-phosphate translocase, to transport it to ER membrane,
and the aforementioned enzyme.
Cori Cycle
• Gluconeogenesis recycles the lactate produced, occur in liver, then
glucose produced is transported to blood.
• When cell needs ATP, Glycolysis is more active, it is exergonic reaction
• When there is little need for ATP, gluconeogenesis is more active,
exogenic reaction.