Term 2 Lecture 3: Blood- RBC And Other Components

Question 1

Primary fluids of the body

Answer 1

Intracellular fluid

Extracellular fluid:
Blood, interstitial fluid, lymph

Question 2

Circulatory system

Answer 2

Blood, blood vessels, lymph, lymph vessels and heart

Question 3

Functions of blood

Answer 3

Transportation- gases, nutrients, hormones and waste

Regulation - pH (7.35-7.45), acts as a buffer, body temp, water balance

Protection - from microorganisms and cancer cells also aids in clotting and would repair

Question 4

CAR : T cell therapy for cancer treatment

Answer 4

1)Collect blood sample from patient to harvest T-cells

2) make CAR T-cells in lab by inserting CAR genes into patients T cells
CAR= chimeric antigen receptor

3) grow millions of CAR T cells in lab

4) infuse CAR T cells into patients circulatory system

5) the CAR T cells bind to cancer cells and kill them

Question 5

General facts about blood

Answer 5

Blood is a specialised connective tissue
Makes up 1/4 of all extracellular fluid
Adult human blood vol is 7% of total body weight e.g.
in a 58kg female ~4litres
In a 70kg male ~5litres (2 plasma 3 blood cells)

Question 6

Plasma

Answer 6

Is ECM
92% water
7% proteins (Albumin ~60% Globulins ~35% Fibrinogen ~4%)
~1% enzymes/proenzymes/hormones

All plasma proteins except gamma globulins are produced by the liver

1% is organic molecules (aa, glucose, lipids, nitrogenous waste), ions (Na+, K+, Cl-, H+, Ca²+ & HCO3-), trace elements, vitamins, dissolved O2 & CO2

Question 7

Blood vessels

Answer 7

Mean diameter/wall thickness

Artery: 4.0 / 1.0mm
Endothelium, elastic tissue, smooth muscle and fibrous tissue (all)

Arteriole: 30.0/ 6.0 micrometre
Endo and smooth

Capillary: 3.0/ 0.5 micrometres
Just endo

Venule: 20.0/ 1.0 micrometre
Endo and fibrous

Vein: 5.0/0.5mm
All

Question 8

Capillaries

Answer 8

On average 10 billion in the adult human body and therefore cover the largest total cross sectional area of all vessels and have the lowest velocity due to being the narrowest type of vessel

Question 9

Capillaries are exchange vessels between blood and interstitial fluid

Answer 9

• in systemic capillaries net pressure = hydrostatic pressure - colloid osmotic pressure
• colloid osmotic pressure within the capillary pulls fluid into the capillary
  -excess water and solutes that filter out of the capillary are picked up by the lymph vessels and returned to the circulation
• plasma proteins are responsible for colloid pressure
• colloid osmotic pressure is constant
  -hydrostatic pressure drops from arterial to venous side

Question 10

Capillary exchange takes place by diffusion, bulk flow and transcytosis

Answer 10

Diffusion - directly through endothelium
Transcytosis- vesicle transport through endothelium
Bulk flow: through channel in endothelium, between cells of endothelium*

• Endothelium normally forms continuous flat membrane but some have holes aka fenestrations or pores these are referred to as fenestrated or discontinuous epithelium

Question 11

Cellular elements of blood

Answer 11

RBC (erythrocytes)
Platelets
WBC: lymphocytes, monocytes, neutrophils, eosinophils and basophils

Question 12

Blood count: % of each component

Answer 12

Measured using a hematocrit tube (centrifuged)
RBC~ 42% WBC ~<1% Plasma ~58%
Read by a micro-capillary reader

The hematocrit (hct) is the % of total blood volume occupied by packed RBC (after centrifuge)

The hemoglobin (Hb) content of RBC is measured as total Hb content of blood (gHb/dl)

Mean RBC vol (MVC) in some disease states may be abnormally large or small e.g. abnormally small in those w/anaemia

RBC count in millions per microlitre a machine counts the cells as they stream through a beam of light

Morphology of RBC can give clues to diseases - sometimes cells lose their flat disc shape and become spherical (spherocytosis) or sickle shaped (sickle cell anaemia)

Total WBC count tells total no. of all types

Differential WBC count estimates rel no. of all 5 types of white cell, carried out by medical technologists using a blood smear

Platelet count- suggestive of bloods ability to clot

Question 13

Hematocrit average result:

Answer 13

Male/ female

Hematocrit: 40-54% / 37-47%
Hemoglobin (gHb/dl): 14-17% / 12-16%
RBC count (cells/microlitre):
4.5-6.5x10⁶ / 3.9-5.6x10⁶
Total WBC count (cells/microlitre)
4-11x10³ / 4-11x10³

Differential WBC count same proportions in males and females
Neutrophils 50-70%
Eosinophils 1-4%
Basophils <1%
Lymphocytes 20-40%
Monocytes (per microlitre) 15-45x10⁴

Question 14

Plasma Vs serum

Answer 14

Plasma: ~ 52% contains albumins, immunoglobulins, lipids (lipoproteins), hormones, vitamins and salts
Buffy coat: leukocytes and platelets (1%)
Hematocrit: RBC 42-47% blood collected and centrifuged in presence of an anticoagulant (heparin or sodium citrate)

Serum
Majority, protein rich fluid lacking fibrinogen but containing albumin, immunoglobulins and other components
Blood collected w/out anticoagulant and left to coagulate

Question 15

Blood components: cells

Answer 15

RBC- O2 & CO2 transport, most abundant cell in the body

WBC - 5 types, immune defence and phagocytosis

Platelets - clotting

Wright’s stain is used to reveal nuclear shape and cytoplasmic colour

Question 16

Methods of RBC analysis

Answer 16

Blood smear
Cross section
Super resolution imaging
Electron microscopy (SEM)
In blood vessel imaging

Question 17

RBC notes

Answer 17

• most abundant cell type in blood
• lose their nuclei in maturation stage
• lack mitochondria and ER so are unable to renew plasma membrane components, carry out protein synth or aerobic metabolism, also cannot renew enzymes therefore RBCs are short-lived usually lasting 4 months
• have an elaborate cytoskeleton
• mature cells are biconcave discs
• remarkably flexible
• contain haemoglobin
• bind O2 in heme group
• bind some CO2 on globulin
• generate ATP by glycolysis

Question 18

Old RBCs are broken down and bilirubin is detoxified

Answer 18

Old RBCs lose their flexibility so become more fragile and likely to rupture
They are engulfed by scavenging macrophages in the spleen, liver, lymph nodes and bone marrow

Senescent RBC engulfed by phagocyte
Broken down to globulin, bilirubin, heme and iron

Bilirubin is released and binds to albumin

Bilirubin is detached from albumin and taken into a hepatocyte as free bilirubin where it is conjugated to glucuronic acid to form conjugated bilirubin a soluble detoxified form that is released into bile.

Question 19

Bilirubin: yellow byproduct of haemoglobin metabolism

Answer 19

Not water soluble - toxic and causes brain damage
Cannot be directly removed from the body
Hepatocytes bind bilirubin to glucuronic acid using UDP-glucoronyl transferase to form bilirubin diglucoronide
^aka conjugated bilirubin
Which is water soluble and expelled into the intestine where it is further metabolised to stercobilin, urobilinogen and urobilin (uros make plasma yellow)

Bilirubin build up results in jaundice (icterus) it also causes gall stones, alcoholic liver disease, pancreatitis, hepatitis and sickle cell anaemia

Question 20

Lifecycle of erythrocytes (RBCs)

Answer 20

Bone marrow: RBCs are generated (erythropoiesis) in the bone marrow then enter the blood stream

Spleen: When they become senescent the spleen digests them to AAs and haemoglobin which is then split into iron and bilirubin

Liver: Bilirubin is converted to bile in the liver and excreted in faeces. The liver also converts Ferritin (F-iron) is converted to transferrin (T - Iron) which returns to the bone marrow through the circulatory system.

Kidneys: Circulating bile is excreted by kidneys in urine. EPO (erythroprotein) returned to bone marrow.

Question 21

RBC cytoskeleton

Answer 21

Anion transporter channel allows HCO3- to exit through the plasma membrane in exchange for intake of Cl-. This exchange facilitates release of CO2 in the lungs.

Junctional complex - spectrin* tetramers are linked to a 3 protein complex: a short actin filament of actin monomers linked to tropomyosin and protein 4.1. binding
between actin and spectrin is stimulated by adducin (a calmodulin binding protein)

*Spectrin- large dimeric protein consisting of 2 polypeptide chains alpha spectrin (240kd) and beta spectrin (220kd). The 2 polypeptides associate in antiparallel pairs to form a rod 100nm long. Two chains join head to head to form a tetramer found in the cortical region of the RBC.

Question 22

Erythrocyte pathology: Hereditary spherocytosis (HS)

Answer 22

RBCs are spheroidal, less rigid, of variable diameters and subject to destruction in the spleen. This alteration is caused by cytoskeletal abnormalities involving site interactions between spectrin alpha/beta and protein 4.1

Question 23

Erythrocyte pathology: membrane/metabolic/haemoglobin defects

Answer 23

Membrane cytoskeletal defects

Elliptocytosis: AD, EPB41, SPTA1 or SPTB genes (EPB41 codes protein 4.1)

SPTA1 & SPTB encode alpha/beta spectrins

Spherocytosis occurs due to AD spectrin deficiency.
Clinical features: jaundice, anaemia, splenogematic

Metabolic defects:
Glucose 6 phosphate dehydrogenase (G6PD) or pyruvate kinase deficiency

Haemoglobin defects:
Alpha or beta globin chain defects

Question 24

Erythrocyte pathology: sickle cell anaemia

Answer 24

Sickle cell anaemia:
Hemoglobins HbA (alpha 2, beta 2 96%) HBA2(alpha 2 and delta 2 3%) fetal Hb, HBF(alpha 2 gamma 2 1%)

Incidence: 8% afro Americans - heterozygous mutation up to 30% in parts of Africa where malaria is endemic

Cause: beta globulin chain carries a E6V point mutation causing a glutamine to be substituted with valine. Therefore RBCs have a lifespan of only 20 days as their membranes are more susceptible to damage and dehydration with more rigidity and sickled form.

Leads to:
-extravascular chronic anaemia - sickled cells are broken down by spleen

-microvascular obstructions

-hemolysis - on deoxygenation Hb aggregates affecting RBC cell shape which can damage plasma membrane causing hemolysis

Infarction- Cells may regain normal shape with O2 but repeated rounds of deformation lead to aggregation causing an infarction - death of an area of tissue e.g. in lung or bone marrow

Question 25

Erythrocyte pathology: Beta- Thalassemia

Answer 25

(treated by bone marrow transplant at an early age)

Cause: a defect in transcription/RNA splicing/translation

More than 100 diff mutations - most are single base substitutions

Effect: reduced beta globulin synthesis and in some cases complete absence of beta globulin

Summary of defects:
-inadequate HbA formation
-RBCs hypochromic and microcytic
- unpaired alpha chains aggregate and damage the membrane
- extravascular hemolysis
- growth retardation
- skeletal deformities
- cardiac failure due to iron overload

Summary: mutation in Hb peptide chain leads to lack of beta globulin and aggregation of alpha globulin. This leads to erythrocyte deformation resulting in destruction in the spleen. As less RBCs are present in the circulation this leads to anaemia. Expansion of bone marrow to produce enough erythrocytes to compensate for losses leads to skeletal deformation