PRP Flashcards
Young, fecund, robust, inactivated platelets are HOW BIG
1-3 μm discoids
Humans: Normally HAVE HOW many platelets
150,000-300,000/ml of blood
life span of a platelet
one week
dog platelets amount
145,000-440,000/ml but similar size
cat platelet amount
190,000-800,000/ml Very large platelets Easily activated Much higher tendency to aggregate
horse platelets
100,000-600,000/ml
Small platelets
Uniform size and shape
cow platelets
100,000-800,000/ml Small platelets of variable size Lack the microtubular system and open canalicular system of non-ruminants
mouse platelets
LOTS (>1,000,000/ml)
Also small platelets
that are very readily
activated. Why?
α-granules contain
clotting factors, growth factors, and various other proteins
dense granules contain
ADP, ATP, Serotonin, and Calcium
young platelet size
large and heavy
old platelet size
small and light
young platelets aggregation compared to old platelets
much faster (3-5 x) than older platelets
Young platelets ATP and ADP release compared to old platelets
release dramatically more ATP (4-8 x) and ADP (4-6 x) than do older platelets
Old platelets require substantially greater amounts
ADP to be activated than do young platelets
Interestingly, old and young platelets don’t necessarily
live in the same neighborhood
Interestingly, old and young platelets don’t necessarily
live in the same neighborhood
PRIMARY HEMOSTASIS OF PLATELETS
vasoconstriction, platelet adhesion, platelet aggregation
secondary hemostasis
activation of coagulation factors. fibrin formation
fibrinolysis
activation of fibrinolytic system, clot lysis
platelet process up to aggregation
Wound
Exposes subendothelial collagen
Binds von Willebrand Factor
Platelet adhesion to blood vessel wall via glycoprotein
IIb/IIIa receptors
Platelet activation
Platelet cytoskeleton (via actin and myosin) expands
from a disc to a multi-pseudopodal sticky blob
Platelet aggregation
platelet aggregation process
Serotonin Vasoconstriction
ADP Recruits other platelets to aggregate and degranulate
Thromboxane Platelet aggregation and PGF release
…and more cytokines, chemotactic and growth factors, etc., etc. than you can imagine (more are discovered on a monthly basis)
PDGF: Platelet-Derived Growth Factor
STRONGLY mitogenic and chemotactic for leukocytes
By itself PDGF application doubles the rate of collagen
deposition in a wound
(plus a bunch of other stuff that aggressively accelerates
healing…)
TGF-β: Transforming Growth Factor-Beta
Also strongly mitogenic
Allows damaged (irradiated, corticosteroid-treated)
tissues to revert to normalized collagen deposition
Attracted neutrophils and macrophages release
a host
of other healing factors
Granulocytes (neutrophils) may be
good bad or indifferent
neutrophils Cells expressing the CD34
protein are
concentrated in
the mononuclear layer of
platelet concentrate
-These cells are stem cell “markers” and are
important for other cells’ adhesion/chemotaxis
TGF beta source and function
platelets,matrix of bone, cartilage, TH1 cells, natural killer cells, macrophages, and monocytes. mesenchymal cell proliferation, regulates endothelial fibroblasts, osteoblastic mitogenesis, inhibits macrophage and monocyte proliferation.
TGF beta source and function
platelets,matrix of bone, cartilage, TH1 cells, natural killer cells, macrophages, and monocytes. mesenchymal cell proliferation, regulates endothelial fibroblasts, osteoblastic mitogenesis, inhibits macrophage and monocyte proliferation.
PRP
“The components of whole blood remaining after the removal of (most of) the red cells”
The buffy coat (white cells and platelets) extending ??? into the top of the red cell column
PLUS all of the plasma
PPP
“Plasma layer without the buffy coat”
So…you get lots of fibrinogen & coagulation factors but no cells
PLATELET CONCENTRATE
Essentially the buffy coat +/- a small (variable) amount of plasma
Buffy coat = leukocytes + platelets
Leukocytes = neutrophils, eosinophils, basophils, macrophages, B- and T-lymphocytes
Platelet gel
Platelet Concentrate with enough fibrinogen (2-4mg/ml) to “set up” when combined with an activator
activator =
= Thrombin (bovine or human), Calcium (usually CaCl₂) or Collagen
platelet concentration in platelet gel
Platelets 2-6x over baseline
how many platelets do you need
- )Increase in multiples above baseline
- Typically 2-6 times above baseline
- Evidence suggests >6x actually delays healing…why? Theory suggests possible up-regulation of other tissues to “factors” in the presence of thrombocytopenia.
2) Absolute numeric concentration
- Typically > 1,000,000/μL
nervous tissue and platelet gel
doesn’t do crap on nervous tissue
platelet gel contraindications
Severe hypovolemia Unstable angina/LM disease Heparin therapy Post-incisional harvest Thrombocytopenia (~
Never, ever, EVER apply platelet concentrates to
coronary grafts (and don’t let surgeons do it, either!)
Cell-saver based systems are (IMHO) “maximally suboptimal because of
Dramatic differences in cost, % platelet recovery, volume accommodated, increases in platelet count above baseline, “hands on” requirements, consistency, time required for processing, technical support, portability, customization potential, RBC recovery, PPP availability, and perfusion support
platelet gel makers
arteriocyte magellan, cytomedix angel
To get more volume…
…you either need more blood initally
…or lower “Increases Above Baseline” or you add more RBCs to the mix. There IS no “free lunch” (although sales reps may try to convince you & your surgeons otherwise!)
You don’t get “something for nothing”…
If you start with less volume, the only way to get more PRP
is to have less enrichment and/or add more RBC’s (a bad thing)
How many platelets in a “unit” of whole blood
(Platelets/μL) X 1000 X (ml of whole blood
How many platelets in a “unit” of PRP?
(Platelets/μL) X 1000 X (ml of PRP
What is the percent yield of platelets in PRP?
(# platelets in PRP X 100)/(# platelets in whole blood)
Essentially two “fields” of stem cell therapy
Bone Marrow (“Mesenchymal”) Derived 2) Adipose Derived
mesenchymal stem cells
Generally found in the bone marrow but can be isolated from circulating blood, cord blood, fallopian tubes, and fetal tissue.
High capacity for pluropotentiality (what’s this?)
High capacity for self renewal
mesenchymal stem cells
Generally found in the bone marrow but can be isolated from circulating blood, cord blood, fallopian tubes, and fetal tissue.
High capacity for pluropotentiality (what’s this?)
High capacity for self renewal
bone marrow derived stem cells Problems:
Critters are “funny” about having a massive pointy needle rammed into their pelvis and having those contents removed with vigorous suction!
People are less sensitive about having fat sucked out of “fat-rich” parts of their anatomy (“Take more! TAKE MORE!!!)
Stem cells are “few and far between” in the bone marrow
adipose stem cells
Also a source of multipotent (less pluropotent?) stem cells
> 500 X more stem cells in 1 gram of fat as compared to 1 gram of aspirated bone marrow
Have similar (+/-) ability to differentiate as does B.M.-derived.
Can be extracted without anesthesia/sedative/tranquilizer
Avoids “fetal stem cell” discussion
*Technology for utilizing adipose-derived stem cells is much simpler (and more commercially available)
(Moving away from B.M. towards A.D.)
Stem cells have the
ability to
to morph
stem cells are synergistically attracted/stimulated by
activated platelets and the “factors” those platelets produce
adipose stem cells extraction systems
1) Remove the “lipids”
2) Remove the supernatant (saline, phenylephrine, lidocaine)
3) Concentrate the adipose stem cells (hopefully)
4) Maintain the Stromal Vascular Fraction (SVF) which creates a warm, fuzzy microenvironment for the stem cells and helps promote graft retention
5) +/or allow for the stem cell extract to be mixed at some ratio with platelet concentrate in an aerobic environment
platelet to adipose ratio
Still being elucidated
1:2-1:10…too early for anything resembling “evidence
procedure for horse with scrap metal
Step 1: 4X4 soaked in sterile saline, wrung out to “Just Damp” is applied
Step 2: Cotton wrap bandage is applied
Step 3: Water proof tape or Op- Site
Step 4: Coban Dressing is Top Coat
outcomes from scrap metal horse
The wound was 80% closed at 5 Day inspection
Complete healing at 15 days
Hair re-generated at 20 days
horse with right front hoof 6 day old the dressing
Step 1: 4X4 soaked in sterile saline, wrung out to “Just Damp” is applied Step 2: Cotton wrap bandage is applied Step 3: Water proof tape or Op- Site Step 4: Compression Bandage Step 5: Coban Dressing is Top Coat
outcomes for hoof injury horse
Single Treatment of PG Required No Oozing or Drainage on Day 5 No Infection During Recovery Uneventful Complete Recovery Resumed Normal Activities at 80 Days Barrel Racing at 90 Days
knee wound horse outcomes
At 5 day inspection the wound appeared to have healed to a 14 day level
The wound continued to heal over the next 3 ½ weeks until all that remained was a scabbed wound about 1” high x 2” long.
The horse then became tangled in a slot in a gate and recreated the wound to almost the original size
The horse was then sent to a special wound therapy clinic and treated with allograft growth factors. The wound healed in about 3 months
knee wound horse outcomes
At 5 day inspection the wound appeared to have healed to a 14 day level
The wound continued to heal over the next 3 ½ weeks until all that remained was a scabbed wound about 1” high x 2” long.
The horse then became tangled in a slot in a gate and recreated the wound to almost the original size
The horse was then sent to a special wound therapy clinic and treated with allograft growth factors. The wound healed in about 3 months
device considerations for animal pg
The size of the client must be taken into consideration as it relates directly to the availability of a sufficient blood draw.
The wound size is not always in proportion
Small volume device availability is a must for the treatment of dogs and cats
ATF type devices are required for some wounds in horses and cattle
The device should provide enough PG (3x-4x) to completely cover the wound. PPP may be used as a top coat.