Ch.8 Regenerative Medicine Flashcards
What is PRP?
A plasma suspension containing a quantity of platelets that is increased in concentration from that of whole blood and also contains varying amounts of leukocytes, RBC and plasma proteins.
The use of PRP is based on what concept?
The delivery of growth factors and proteins of the clotting system may optimize tissue healing .
What is responsible for the antimicrobial properties of PRP?
Platelet lysates
What growth factors present in PRP are important in assisting bone and cartilage healing
Osteocalcin (Oc)
Osteonectin (On)
Fibronectin (Fn)
Thrombospondin-1 (TSP-1)
How soon after injecting PRP does the clotting cascade start?
Within minutes - 95% of the growth factors are secreted within 1 hour
Why is mutagenic activity and tumour formation not an issue with PRP application
It does not act directly on the nucleus
What is the half life of platelets
5-7 days
What should be considered when using combination therapy of suspending mesenchymal cells in PRP?
When platelet concentrates are supplemented in media at volumes >30% proliferation of mesenchymal stem cells declines.
Longterm suspension of mesenchymal cells in PRP may negatively affect mesenchymal cell viability
PRP preparation requires a “soft spin” and a “hard spin” why?
Soft spin - Leukocytes and RBC are larger and more dense than platelets so they settle faster leaving the platelets suspended in the plasma.
Hard spin - Centrifugation for platelets to be pelleted
What is P-PRP
Pure PRP or Leukocyte-poor
Contains platelets and plasma and little to no RBC or leukocyte. Produced by a soft spin where the RBC and leukocytes are pelleted and the plasma and platelets left suspended
eg Arthrex ACP Double syringe system
or in a lab by plasmapheresis
Why must Equine specific PRP kits be used to prepare equine PRP
Interspecies differences in blood viscosity, platelet density, degree of platelet activation and aggregation.
Effect of neutrophils, IL-1B and tumour necrosis factor alpha within tendons
Decreases matrix synthesis
Increases tissue catabolism
Effect of peripheral blood mononuclear cells PBMCs on repair
Enhance anabolic effects of PRP
How has IGF-1 been shown to affect tendonitis
Improved
1- cell proliferation,
2 - collagen content,
3 - mechanical stiffness and
4 - sonographic appearance during 8 weeks of tx
What is COMP (Cartilage oligomeric matrix protein) ?
A component of ligament healing response and a marker of appropriate ligament matrix remodelling.
What is the proposed use of acellular bone marrow aspirate?
Use instead of PRP on desmitis as
enhances healing by
providing anabolic factors
which stimulate matrix production
what component of PRP stimulates angiogenesis
VEGF
Vascular endothelial growth factor
Up to how long post PRP tx is neovascularisation thought to occur
23 weeks
2 primary pro inflammatory cytokines that initiate and participate in the destructive cytokine cascade of osteoarthritis
IL-1
TNF-Alpha
How can the activity of IL-1 be inhibited
IL-1 receptor antagonist protein IL-1ra prevents the interaction btw IL-1 and its receptor
Direct injection of the antagonist
Gene therapy or conditioning of the serum to stimulate production of IL-1ra
How is autologous conditioned serum ACS, produced?
Conditioning certain blood components, primarily monocytes, to increase endogenous production of IL-1ra
Accomplished by exposing whole blood to medical grade chromium sulphate treated glass beads.
What increase in IL-1ra must occur to block all of the available IL-1 receptors?
10-1000 fold increase
What is IRAP
(ACS)
InterLeukin1 Receptor Antagonist Protein
Orthokine - human product
Whole blood - medical grade chromium- sulphate treated glass beads - incubated at 37* for 24hrs, centrifuged, serum obtained and stored at -20*
6 2cc syringes given over 3 weeks
Which system Arthrex or Dechra has been found to produce a high amount of IRAP
Arthrex
What is APS, how is it produced?
Autologous Protein Serum
ProStride
20mins instead of 24 hours
Whole blood - APS separator sequesters WBC and platelets
Then transferred to an APS concentrator - filters it through Polyacramide beads and desiccates it
Result is a concentrated sol of WBC, platelets and plasma proteins
How does APS differ to ACS?
Autologous Protein Serum
Higher levels of IL-1ra and platelet derived growth factors but also highly variable levels of IL-1B
Where are stem cells most commonly harvested in equine medicine
Adipose - Tail head/sup gluteal area
Bone marrow - Sternum, ilium
Older horses >13 hard to harvest from ilium and less MSC than from sternum
What methods can be used to prepare the stem cells
1 Culture and expansion
2 Patient side centrifugation BMC - Bone marrow wconcentrate
3 - If adipose derided need to be digested first - 3 days shipping and prep
Non adult sources of stem cells
Fetal fibroblasts
Umbilical cord blood
Umbilical cord tissue/matrix
Placental tissue
Amniotic fluid and membrane
What stage are embryonic stem cells derived from
blastocyst
What are iPSCs induced pluripotent stem cells
Pluripotent stem cells generated through genetic reprogramming of adult somatic cells
Yamanka factors - mouse
What are MSCs
Mesenchymal stem cells
undiferentiated cells capable of asymmetric cell division - one daughter cell develops into a somatic cell type while the other remains as a stem cell in the “niche” tissue
How do MSCs aid in inflammation
Signal endothelial cells to promote angiogenesis
Prevent leukocyte transmigration
Stimulate fibroblasts to produce extracellular matrix
Activate and recruit tissue intrinsic stem/progenitor cells
How do MSCs aid in inflammation
Signal endothelial cells to promote angiogenesis
Prevent leukocyte transmigration
Stimulate fibroblasts to produce extracellular matrix
Activate and recruit tissue intrinsic stem/progenitor cells
Inhibit T cell activation, B-cell function and dendritic cell maturation
What is the ability of MSCs to modulate T-cell prolif and cytokine secretion dependent on
PGE-2
Cautions against using allogenic MSCs
Loss of efficacy of MSC as destroyed by host before eliciting effect
Risk of adverse rxns
How is the best quality and content of MSC obtained on aspirate
First 5cc aspirate
If need 60 such as for Bone marrow concentrate aspirate 5 to 10 from different sites/depths
What is the location for sternal aspirate
4th and 5th marrow spaces
Where is adipose tissue harvested for stem cells and how much
Subvcut Tail head/superior gluteal
15-20g
BMP-12 Bone Morphogenic Protein 12 been shown to aid in the formation of what tissue
Tendon and ligament like tissue
Which possesses a higher capacity for chondrogenic differentiation UCB-MSCs or BM-MSCs
UCB-MSCs
umbilical chord blood derived MSCs
