Red blood cells

Question 1

What are some properties of a red blood cell?

Answer 1

• most abundant cell in the body
• transport gases predominantly O2 and CO2
• biconcave shape to increase surface area
• no internal organelles to maximize space for gas transport
• strong yet flexible

Question 2

What is erythropoesis?

Answer 2

The production of RBCs. From haematopoietic stem cells to myeloid progenitors to precursors to RBCs. Amplification occurs throughout this process to form large numbers of RBCs in comparison to stem cell reservoirs

Question 3

Where do erythropoiesis and RBC differentiation occur?

Answer 3

• differentiation occurs on erythroblastic islands in the bone marrow
• these contain central macrophages on the surface of which erythropoiesis occurs
• in the final stages, the RBC becomes anucleate as the macrophage removes its nucleus

Question 4

Why might we need an alternate stem cell source of RBCs other than donations?

Answer 4

• reduces risk of blood borne infection
• reduces immune mismatch
• transfusion-dependent patients can develop antibodies to blood group types
• those with rare blood groups
• cells are younger and better
• supply inefficieny

Question 5

What are blood groups?

Answer 5

• antigenic determinants important in transfusion medicine
• can be polymorphisms in membrane proteins like the urea transporter or chemokine receptors
• or polymorphisms in glycosyltransferases

Question 6

What kinds of cytokines and factors are required to develop in vitro erythroid culture systems?

Answer 6

• stem cell factor
• interleukin 3
• erythropoetin
• transferin

Question 7

What are stage 1, 2 and 3 feeder free culture systems?

Answer 7

Feeder 3 = no other cell types present
Stage 3 is the most effective at producing RBCs with a 60-90% enucleation rate

Question 8

What are some hurdles to generating red blood cells in vitro?

Answer 8

• yield
• cells don’t divide indefinitely so need to keep collecting stem cells
• but can immortalise early stem cells to keep producing blood cells

Question 9

What are BEL-A cells?

Answer 9

• immortalised adult erythroid stem cells
• do not require repeat SC collection
• recaptulate normal erythropoesis
• scaleable, sustainable source of red blood cells

Question 10

How are BEL-A cells made?

Answer 10

• put bone marrow, peripheral blood or umbilical cord HSCs in a plasmid with HPV E6 and E7
• culture cells to proerythroids and add doxycylin to turn off E6/7 and keep the cells dividing at this stage
• removal of doxycylin will cause cells to differentiate again if needed

Question 11

How can we genetically edit BEL-A cells?

Answer 11

• CRISPR
• KOs and overexpressions are retained in differentiation
• can make blood more universally compatible by adding null phenotypes
• can make cells last longer in circulation or increase their oxygen-carrying capacity and delivery

Question 12

How could RBCs be used as drug delivery and targeting agents?

Answer 12

• modify cells to break down toxins, replenish enzymes, deliver proteins or reduce drug toxicity
• modify or add surface proteins to recognise pathogens, improve targetting to wounds or cancer and to enhance drug delivery