What is blood made up of?
55% plasma- proteins, water, solutes 45% formed elements- platelets RBCs, WBCs
What are the types of white blood cells?
Neutrophils 60-70% Lymphocytes 20-25% Monocytes 3-8% Eosinophils 2-4% Basophils 0.5-1%
What is in blood plasma?
water 91.5% Proteins 7%- albumin- regulates osmotic pressure, golbulins- antibodies or immunoglobins, fibrinogen- blood clotting Solutes 1.5%- electrolytes, gases, vitamins etc.
How long do platelets, RBCs and WBCs live?
PLatelets - 10 days Erythrocytes- 120 days Lymphocytes- up to a few years (memory cells)
What is the process by which blood cells are formed?
Haemopoiesis. (100 billion new blood cells formed each day!)
Where does haemopoiesis occur?
First few weeks of gestation- in yolk sac
2-7 months- liver, spleen
5-9 months- bone marrow
Haemopoiesis is derived from...
A population of stem cells found on the dorsal aorta which is known as the AGM. The embryonic stem cells give rise to a multi-potent cell called haemangioblasts, which give rise to endothelial cells and haemopoietic cells.
What are the first blood cells made?
What happens in weeks 2-8?
At 2-8 weeks, primitive nucleated erythroid cells are found in the yolk sac (contain haemoglobin but don't mature to fully developed RBCs.) Liver spleen and bone marrow are derived from haemangioblasts amongst other cells.
First blood cells develop with the earliest blood vessels. Mesenchyme cells cluster into blood islands. Late in 2nd month- liver and spleen take over blood formation. Bone becomes major haematopoietic organ at month 7
What are the types of bone marrow?
Red marrow= actively generates new blood cells. 0.05-0.1% of red marrow are pluripotent stem cells. Yellow marrow= contains many fat cells. Not part of haemopoiesis. Located in long bones of adults. IN infants bone marrow from most bones take part. In adults just ribs, pelvis, sternum and a few.
Describe bone marrow at the site of haemopoiesis
Central sinus in the middle, with a medullary artery. Then a medullary cavity around that, then the cortex all the way round like a ring, with endosteal capillaries and medullary vascular sinuses. Developing blood cells form outside of bone marrow sinuses and mature cells are released in these spaces. Bone marrow is in the central medullary cavity. Haematopoiesis occurs in the extravascular spaces between the sinuses.
What do all blood cells originate from?
Haemopoietic stem cells. Two types of this= 1. Lymphoid stem cell- gives rise to lymphocytes 2. Myeloid stem cell- gives rise to all other blood cells
How are stem cells identified?
By using immunological testing. CD34+, CD38. CD34+ glycosylated surface antigen preferentially expressed on haemopoietic stem cells. Plays a role in cell to cell adhesion and maybe regulating other haemopoiesis genes. Monoclonal antibodies to CD34+ have been raised and these have allowed identification and characterization of a whole range of haemopoietic progenitor cells. CD34+ is expressed most strongyl on the most primitive cells and is progressively lost as cells differentiate. CD38- haemopoietic cells dont express this protein.
Pulripotent stem cell lineage
What is the bone marrow environment that allows stem cells to grow?
Bone marrow is made up of stromal cells and microvascular network.
Stromal cells are: fibroblasts, endothelial cells, fat cells, reticulum cells and macrophages.
Stromal cells secrete extracellular proteins which are needed by stem cells to grow and differentiate. Also secrete growth factors vital for stem cell survival.
Romanowsky stain blood cells why is it used?
Stained on slides to highlight features of each cell and identify different blood cells. Contains Azure B and Eosin Y.
Azure B binds anionic molecules like nucleic acids. Eosin Y binds cationic sites on proteins like haemoglobin.
Stages of differentiation of blood cells
What is CFU?
Colony Forming Units.
Given to cells based on what lineage they give rise to. e.g. myeloid cells are CFU GEMM because they give rise to granulocytes, erythroctyes, monocytes and megakaryocytes.
Lymphoid stem cell= CFU L. as it makes t and b cell lymphocytes.
What is the differentation of cells regulated by?
By transcription factors:
PU-1: cells are differentiated along the myeloid lineage.
GATA-1: differentiates cells along the erythropoietic and megakaryotic cell lineages.
Glycoproteins. can increase production of haemopoietic cell lines. Regulate progenitor cells. Regulate function of mature blood cells. They are present in the extracellular environment. Bind to ECM and act on cells by binding to growth factor receptors. Exert effects at low concentrations. Stimulation of the growth factor can increase further synthesis of the growth factor or receptor.
Which growth factors play a role in haemopoiesis?
G-CSF: granulocyte colony-stimulating factor.
Interleukin 3: works with scf and gm-csf to increase proliferation of stem cells.
Interleukin 5: produced by T cells. role in eosinophil growth and differentiation
GM-CSF: Granulocyte macrophage colony-stimulating factor. Growth and development of granulocyte and macrophage progenitor cells. Stimulates myeloblasts and monoblasts.
M-CSF: macrophage colony stimulating factor. Monocytes and macrophages.
SCF: stem cell factor. Works with IL3 and gm-csf to increase proliferation of stem cells.
Erythropoietin- increases red blood cells.
Thrombopoietin- more megakaryocytes and platelet.
Main steps of erythropoiesis?
Proerythroblast- large cell with cytoplasm that stains dark blue
Give rise to normoblasts- smaller cells, cytoplasm start to stain lighter blue
Late normoblasts have extruded chromatin
Reticulocyte- contains some ribosomal RNA, circulates in peripheral blood (1-2 days)
Mature RBC, RNA lost
Growth factor receptors
•Growth factors signal via their receptors
•Many receptors such as GM-CSFR dimerise after binding of the growth factor
•Activation of the receptor results in the initiation of a signal transduction pathway
•This can cause activation of transcription factors and changes in gene expression
•MAPK kinase pathways control proliferation
•P13 pathways block apoptosis
B lymphocyte differentiation in fetus occurs in liver but in adults occurs in bone marrow
B lymphocytes mature in to plasma cells
B cells - in birds the B cells leave marrow and mature in an organ called Bursa (means purse or pocket) of Fabricius
T Lymphocyte tend to mature in thymus
Promonocyte-large cell with indented nucleus,only found in bone marrow
Monocyte- contains nucleus (‘c’ or ‘s’ shape) with clumped chromatin
Monocytes migrate to tissues and mature into macrophages
Monocytes stay for 20-40 hours in peripheral blood circulation
Macrophages are phagocytic cells which ingest bacteria such as e.coli
Process of endomitosis to produce megakaryoblasts
Replication of chromosomes occurs but cells don’t divide (Endomitotic replication)
Cells become larger and polyploid (64 n DNA content)
Mature megakaryocytes contain 64n DNA content
Endomitosis ceases resulting in granulated cytoplasm
Process takes 2-3 days
Each Megakaryocyte produces ~ 4000 platelets
Extra reading about growth factors/PRP
The most widely used modality is the use of platelet-rich plasma (PRP), i.e. of the portion of plasma that, after centrifugation, is rich in platelets.
PRP contains numerous growth factors, the main ones being: platelet-derived growth factor (PDGF), transforming growth factor (TGF), platelet-derived endothelial growth factor (PDEGF), interleukin 1, insulin-like growth factor (IGF), osteocalcin and osteonectin, although there are many others.
PRP has been used successfully in orthopaedics for the biological regeneration of cartilage, tendons, ligaments and, of course, bone tissue