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Do Amniotic Products Have Viable
Stem Cells or Can They Help
Autologous Stem Cells?
[All slides are protected by copyright...
Marketing materials and/or communications from
multiple companies vending FDA 361 registered
amniotic tissues claim that t...
Typical communications from amniotic vendors:
Provider Web-sites…
Overall Results of Viability Testing:
Flowable Amniotic Product
Cells
Present?
Culture
Expandable?
Amniofix Injectable No ...
AmnioFix Injectable
(1:10 Dilution in CCM)
48 Hours (Pre-Wash)
48 Hours (Post-Wash)
Flowable Amniotic Product Cells Presen...
48 Hours (Pre-Wash)
48 Hours (Post-Wash)
BioD Restore
(1:10 Dilution in CCM)
Flowable Amniotic Product Cells Present? Cult...
48 Hours (Pre-Wash)
48 Hours (Post-Wash)
FloGraft Freedom
(1:10 Dilution in CCM)
Flowable Amniotic Product Cells Present? ...
48 Hours (Pre-Wash)
48 Hours (Post-Wash)
Ovation Cellular Repair Matrix
(1:10 Dilution in CCM)
Flowable Amniotic Product C...
48 Hours (Pre-Wash)
48 Hours (Post-Wash)
PalinGen Flow Sport
(1:10 Dilution in CCM)
Flowable Amniotic Product Cells Presen...
Can Amniotic Products Help Human MSCs?
Human bone marrow MSCs from several donors
were seeded with various amniotic products to
observe:
• Morphology
• Prolifera...
Complete Culture
Medium (CCM)
hMSC Morphology (CCM has 16.5% PL)
PicoGreen Assay Results (SF vs PL)
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AlamarBlue Assay Results (SF vs PL)
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AlamarBlue Assay Results (SF vs PL vs FBS)
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ROS DCFDA Assay Results (SF vs PL vs FBS)
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Conclusions:
• The amniotic products tested:
• Don’t contain viable stem cells or other viable cells once they’re
thawed p...
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Amniotic Stem Cell Products Testing Interventional Orthopedics Foundation

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Testing performed by a 501c3 non-profit on various products being advertised by manufacturers as amniotic stem cell therapy.

Published in: Health & Medicine
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Amniotic Stem Cell Products Testing Interventional Orthopedics Foundation

  1. 1. Do Amniotic Products Have Viable Stem Cells or Can They Help Autologous Stem Cells? [All slides are protected by copyright-Interventional Orthopedics Foundation-2015]
  2. 2. Marketing materials and/or communications from multiple companies vending FDA 361 registered amniotic tissues claim that these products contain viable stem cells. Many physicians who purchase these products for injection have web-sites that refer to these injections as “stem cell therapy”.
  3. 3. Typical communications from amniotic vendors:
  4. 4. Provider Web-sites…
  5. 5. Overall Results of Viability Testing: Flowable Amniotic Product Cells Present? Culture Expandable? Amniofix Injectable No No BioD Restore No No FloGraft Freedom No No Ovation Cellular Repair Matrix Yes No PalinGen Sport Flow No No Are there cells visible under inversion microscopy? Did any cells grow under ideal nutrition and other conditions after plating the product?
  6. 6. AmnioFix Injectable (1:10 Dilution in CCM) 48 Hours (Pre-Wash) 48 Hours (Post-Wash) Flowable Amniotic Product Cells Present? Culture Expandable? Amniofix Injectable No No BioD Restore No No FloGraft Freedom No No Ovation Cellular Repair Matrix Yes No PalinGen Sport Flow No No
  7. 7. 48 Hours (Pre-Wash) 48 Hours (Post-Wash) BioD Restore (1:10 Dilution in CCM) Flowable Amniotic Product Cells Present? Culture Expandable? Amniofix Injectable No No BioD Restore No No FloGraft Freedom No No Ovation Cellular Repair Matrix Yes No PalinGen Sport Flow No No
  8. 8. 48 Hours (Pre-Wash) 48 Hours (Post-Wash) FloGraft Freedom (1:10 Dilution in CCM) Flowable Amniotic Product Cells Present? Culture Expandable? Amniofix Injectable No No BioD Restore No No FloGraft Freedom No No Ovation Cellular Repair Matrix Yes No PalinGen Sport Flow No No
  9. 9. 48 Hours (Pre-Wash) 48 Hours (Post-Wash) Ovation Cellular Repair Matrix (1:10 Dilution in CCM) Flowable Amniotic Product Cells Present? Culture Expandable? Amniofix Injectable No No BioD Restore No No FloGraft Freedom No No Ovation Cellular Repair Matrix Yes No PalinGen Sport Flow No No Adhered object exhibits cellular morphology
  10. 10. 48 Hours (Pre-Wash) 48 Hours (Post-Wash) PalinGen Flow Sport (1:10 Dilution in CCM) Flowable Amniotic Product Cells Present? Culture Expandable? Amniofix Injectable No No BioD Restore No No FloGraft Freedom No No Ovation Cellular Repair Matrix Yes No PalinGen Sport Flow No No
  11. 11. Can Amniotic Products Help Human MSCs?
  12. 12. Human bone marrow MSCs from several donors were seeded with various amniotic products to observe: • Morphology • Proliferation-PicoGreen DNA Assay • Metabolic activity (Alamar Blue) • Reactive Oxygen Species (ROS)-A test of cell stress and overall health
  13. 13. Complete Culture Medium (CCM) hMSC Morphology (CCM has 16.5% PL)
  14. 14. PicoGreen Assay Results (SF vs PL) S e ru m -F re e M e d iu m A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix C o m p le te C u ltu re M e d iu m A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix 0 1 0 0 2 0 0 3 0 0 4 0 0 5 0 0 d s D N A Q u a n tific a tio n (P ic o G re e n ) FluorescenceIntensity(a.u.) P la te le t L ysa teS e ru m -F re e *** *** *** p < 0 .0 0 0 5 ** p < 0 .0 0 5 * p < 0 .0 5 If the amniotic tissues were causing the MSCs to proliferate more, then total DNA volume would be higher than CCM (Complete Culture Medium). This does happen when serum free (low growth factor) media is used. However, these effects are largely washed out when high growth factor culture media (platelet lysate) is used. This could indicate that while there are active growth factors in these products, but that these GF levels are likely less biologically active than those found in platelets.
  15. 15. AlamarBlue Assay Results (SF vs PL) C o m p le te C u ltu re M e d iu m (S F ) A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix C o m p le te C u ltu re M e d iu m A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix 0 2 0 0 4 0 0 6 0 0 8 0 0 1 0 0 0 M e ta b o lic A c tiv ity (A la m a rB lu e ) FluorescenceIntensity(a.u.) P la te le t L ysa teS e ru m -F re e ** *** *** * * *** p < 0 .0 0 0 5 ** p < 0 .0 0 5 * p < 0 .0 5 * If the amniotic tissues were helping the MSCs, then metabolic activity would be higher than CCM (Complete Culture Medium). Here it is generally lower.
  16. 16. AlamarBlue Assay Results (SF vs PL vs FBS) C o m p le te C u ltu re M e d iu m (S F ) A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix C o m p le te C u ltu re M e d iu m A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix C o m p le te C u ltu re M e d iu m (F B S ) A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix 0 2 0 0 4 0 0 6 0 0 8 0 0 1 0 0 0 A la m a r B lu e B a s e M e d ia C o m p a ris o n FluorescenceIntensity(a.u.) B o v in e S e ru mP la te le t L ysa teS e ru m -F re e ** *** *** * * ** ** ** *** p < 0 .0 0 0 5 ** p < 0 .0 0 5 * p < 0 .0 5 * If the amniotic tissues were helping the MSCs, then metabolic activity would be higher than CCM (Complete Culture Medium). Here it is generally lower, only AmnioFix helps and only when bovine serum is used as a culture media.
  17. 17. ROS DCFDA Assay Results (SF vs PL vs FBS) S e ru m -F re e M e d iu m A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix C o m p le te C u ltu re M e d iu m A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix 1 0 0 M T B H P (+ ) C o n tro l C o m p le te C u ltu re M e d iu m (F B S ) A m n io F ix S p o rts M e d B io D R e s to re F lo G ra ft F re e d o m O v a tio n R e p a ir M a trix 0 2 ,0 0 0 4 ,0 0 0 6 ,0 0 0 8 ,0 0 0 1 0 ,0 0 0 2 0 ,0 0 0 4 0 ,0 0 0 R O S A c tiv ity B a s e M e d ia C o m p a ris o n FluorescenceIntensity(a.u.) B o v in e S e ru mP la te le t L ysa teS e ru m -F re e ** *** p < 0 .0 0 0 5 ** p < 0 .0 0 5 * p < 0 .0 5 * *** * *** If the amniotic tissues were helping the MSCs, then ROS would be lower than CCM (Complete Culture Medium). Here it is generally the same or higher.
  18. 18. Conclusions: • The amniotic products tested: • Don’t contain viable stem cells or other viable cells once they’re thawed per the manufacturer's recommendations. • Do likely contain some active growth factors that can have a positive proliferation effect on MSCs, but those GFs in this experiment look like they are less effective than those found in platelets. • Don’t generally help the metabolic activity of MSCs. • Don’t generally help MSCs show less oxidative stress.

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