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. 2018 Jan 10;19(1):212.
doi: 10.3390/ijms19010212.

Comparative Analysis of Different Platelet Lysates and Platelet Rich Preparations to Stimulate Tendon Cell Biology: An In Vitro Study

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Free PMC article

Comparative Analysis of Different Platelet Lysates and Platelet Rich Preparations to Stimulate Tendon Cell Biology: An In Vitro Study

Franka Klatte-Schulz et al. Int J Mol Sci. .
Free PMC article

Abstract

The poor healing potential of tendons is still a clinical problem, and the use of Platelet Rich Plasma (PRP) was hypothesized to stimulate healing. As the efficacy of PRPs remains unproven, platelet lysate (PL) could be an alternative with its main advantages of storage and characterization before use. Five different blood products were prepared from 16 male donors: human serum, two PRPs (Arthrex, (PRP-ACP); RegenLab (PRP-BCT)), platelet concentrate (apheresis, PC), and PL (freezing-thawing destruction of PC). Additionally, ten commercial allogenic PLs (AlloPL) from pooled donors were tested. The highest concentration of most growth factors was found in AlloPL, whereas the release of growth factors lasted longer in the other products. PRP-ACP, PRP-BCT, and PC significantly increased cell viability of human tenocyte-like cells, whereas PC and AlloPL increased Col1A1 expression and PRP-BCT increased Col3A1 expression. MMP-1, IL-1β, and HGF expression was significantly increased and Scleraxis expression decreased by most blood products. COX1 expression significantly decreased by PC and AlloPL. No clear positive effects on tendon cell biology could be shown, which might partially explain the weak outcome results in clinical practice. Pooled PL seemed to have the most beneficial effects and might be the future in using blood products for tendon tissue regeneration.

Keywords: cell culture; platelet lysate; platelet rich plasma; tendon healing; tenocyte-like cells.

Conflict of interest statement

The coauthors Markus Rojewski and Hubert Schrezenmeier provided AlloPl, and Axel Pruss provided PC and PL for the study. The kits to produce PRP-ACP and PRP-BCT were purchased from the companies.

Figures

Figure 1
Figure 1
Platelet (A) and leukocyte (B) concentration in Arthrex, (PRP-ACP), RegenLab (PRP-BCT), and platelet concentrate (PC) compared to whole blood. (A) Platelet concentration was significantly higher in PRP-ACP and PC group and lower in the PRP-BCT group. (B) Leukocyte concentration was significantly reduced in all groups. °,* indicate outliers, n = 16 individual donors, all blood product were produced from each donor.
Figure 2
Figure 2
Growth factor quantification in the blood products PRP-ACP, platelet rich plasma (PRP), RegenKit-Blood Cell Therapie (BCT), PC, platelet lysat (PL), Allogenic platelet lysate (AlloPL), and human serum (HS) control measured by ELISA. (A) Basic fibroblast growth factor (bFGF) concentration was higher in PC compared to HS, PRP-BCT, and PL as well as in AlloPL compared to HS, both PRPs, and PL. (B) Hepatocyte growth factor (HGF) concentration was not significantly changed. (C) Insulin-like growth factor 1 (IGF-1) concentration was decreased in the AlloPL group. (D) Platelet-derived growth factor (PDGF-AB) and (E) transforming growth factor β (TGF-β1) concentration was lower in the PRP-BCT group and higher in the PC and AlloPL group compared to all other groups and for TGF-β1 concentration except for PC and AlloPL. (F) Vascular endothelial growth factor (VEGF) concentration was increased in the AlloPL group compared to HS, PRP-BCT, and PL. °,* indicate outliers, n = 16, except for AlloPL n = 10.
Figure 3
Figure 3
Cumulative growth factor release from blood products into the medium measured after 1, 4, 24, 48, and 120 h by ELISA. IGF-1 (A), PDGF-AB (B), and TGF-β1 (C) were release only over 4 h by AlloPL but constantly over 2–5 days by the other blood products. The release experiments were performed exemplarily for n = 4 donors.
Figure 4
Figure 4
Cell viability and relative gene expression in human tenocyte-like cells (hTLCs) stimulated with blood products compared to HS control (line) measured by qPCR using ΔCt method with efficiency correction normalized to 18S rRNA. (A) Cell viability was significantly increased by both PRPs and PC compared to HS control. (B) Col1A1 expression was significantly increased by PC and AlloPL group compared to HS control and in AlloPL compared to PL. Col3A1 expression was significantly increased by PRP-BCT and scleraxis (SCX) expression decreased in all groups except PRP-ACP compared to HS control. (C) MMP-1 expression significantly increased by all blood products compared to HS control with significantly highest expression in the PC group and MMP-13 decreased by PC stimulation. (D) MMP-2 and MMP-9 expression did not change. # marks significant differences between the HS control and the blood products and the spanning line between the individual groups. °,* indicate outliers. PRP-ACP: n = 11, PRP-BCT: n = 12, PC: n = 15, PL: n = 14, AlloPL: n = 10. N-numbers varied due to clotting events in the well, which leads to the loss of cells.
Figure 5
Figure 5
Relative gene expression in hTLCs stimulated with blood products compared to HS control (line) measured by qPCR using ΔCt method with efficiency correction normalized to 18S rRNA. (A) IL-1β expression was significantly increased by all blood products with significantly highest expression in PRP-BCT group (higher outliners were cut off). TNF-α was not altered in all groups. IL-6 was decreased in the AlloPL group compared to PRP-BCT. (B) COX1 expression was significantly decreased in PC and AlloPL group and COX2 expression did not change. HGF expression was significantly increased by all products except PRP-BCT. # marks significant differences between the HS control and the blood products and the spanning line marks significant differences between the individual groups. °,* indicate outliers. PRP-ACP: n = 11, PRP-BCT: n = 12, PC: n = 15, PL: n = 14, AlloPL: n = 10. N-numbers varied due to clotting events in the well, which leads to the loss of cells.
Figure 6
Figure 6
Experimental setup for stimulation of hTLCs with platelet-rich blood products.

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