scholarly journals Human Platelet Lysate as an Alternative to Autologous Serum for Human Chondrocyte Clinical Use

Cartilage ◽  
2021 ◽  
pp. 194760352110354
Author(s):  
Virginie Philippe ◽  
Alexis Laurent ◽  
Philippe Abdel-Sayed ◽  
Nathalie Hirt-Burri ◽  
Lee Ann Applegate ◽  
...  
Keyword(s):  
Human Platelet ◽  
Platelet Lysate ◽  
Cartilage Tissue ◽  
Autologous Serum ◽  
Blue Staining ◽  
Alcian Blue Staining ◽  
Clinical Use ◽  
3 Dimensional ◽  
Human Platelet Lysate

Objective A pivotal aspect of cartilage tissue engineering resides in cell culture medium supplementation, in view of maximizing in vitro cell proliferation and preserving cellular functionality. Autologous human serum (aHS) is commonly used as an inducive supplement for safe human articular chondrocyte (HAC) proliferation prior to clinical implantation. However, practical clinical use of aHS is hindered by constraining manufacturing requirements and quality assurance-driven downstream processing. The present study investigated potential alternative use of commercial human platelet lysate (hPL) supplements in HAC manufacturing workflows related to clinical therapeutic pathways. Design Differential effects of hPL, aHS, and fetal bovine serum were assessed on primary cultured HAC parameters (viability, proliferative rates, and morphology) in 2-dimensional in vitro systems. A 3-dimensional HAC pellet model served for postexpansion assessment of cellular functionality, by visualizing proteoglycan production (Alcian blue staining), and by using qRT-PCR relative quantification of chondrogenic marker ( SOX9, COL2-A1, and ACAN) genetic expression. Results We found that monolayer HAC culture with hPL or aHS supplements presented similar characteristics (elongated cell morphology and nearly identical growth kinetics). Chondrogenic activity appeared as conserved in HACs expanded with human or bovine supplements, wherein histologic analysis indicated a progressive sGAG accumulation and SOX9, COL2-A1, ACAN gene expression was upregulated in 3-dimensional HAC pellet models. Conclusion This study therefore supports the use of hPL as a functional equivalent and alternative to aHS for cultured HAC batch preparation, with the potential to effectively alleviate pressure on clinical and manufacturing bottlenecks in cell therapy approaches for cartilage regeneration.

scholarly journals Silencing Smad7 Potentiates BMP2-induced Chondrogenic Differentiation and Inhibits Endochondral Ossification in Human Synovial Derived Mesenchymal Stem Cells

2020 ◽  
Author(s):  
pengcheng xiao ◽  
Zhenglin Zhu ◽  
Chengcheng Du ◽  
Yongsheng Zeng ◽  
junyi Liao ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Chondrogenic Differentiation ◽  
Endochondral Ossification ◽  
Cartilage Tissue Engineering ◽  
Cartilage Tissue ◽  
Blue Staining ◽  
Alcian Blue Staining ◽  
Cartilage Injuries

Abstract Background: Cartilage injuries pose formidable challenges for effective clinical management. Autologous stem cell-based therapies and transgene-enhanced cartilage tissue engineering may open new avenues for the treatment of cartilage injuries. Bone morphogenetic protein 2 (BMP2) is a promising chondrogenic growth factors for transgene-enhanced cartilage tissue engineering. However the BMP2 is failed to maintain a stable chondrogenic phenotype as it also induces robust endochondral ossification. Recently, human synovial derived mesenchymal stem cells (hSMSCs) arouse interested through the poor differentiation potential into osteogenic lineage. Smad7, a protein to antagonizes TGF-β/BMP signaling pathway has been discovered significant in the endochondral ossification. In the present study ,we further explore the effect of downregulate Smad7 in BMP2-induced chondrogenic differentiation of hSMSCs. Methods: hSMSCs were isolated from synovium of human knee joint through adhesion growth. In vitro and in vivo chondrogenic differentiation models of hSMSCs were constructed . Transgenes of BMP2, silencing Smad7 and Smad7 were expressed by adenoviral vectors. The osteogenic differentiation was detected by alkaline phosphatase staining, alizarin red staining. The chondrogenic differentiation was detected by alcian blue staining. Gene expression was determined by reverse transcription and quantitative real-time PCR (RT-qPCR), Immunofluorescence and immunohistochemistry. The subcutaneous stem cell implantation model was established and evaluated by micro-CT , h＆e staining, alcian blue staining and immunohistochemistry assay.Results: Compared to other MSCs, hSMSCs performed less of capability to osteogenic differentiation. But the occurrence of endochondral ossification is still inevasible during BMP2 induced cartilage formation. We found that silencing Smad7 enhanced the BMP2-induced chondrogenic differentiation of hSMSCs in vitro. Also, it leading to much less of hypertrophic differentiation. The subcutaneous stem cells implantation assays demonstrated silencing Smad7 potentiates BMP2-induced cartilage formation and inhibits endochondral ossification. Conclusion: This study strongly suggests that application of hSMSCs , cell scaffolds and silencing Smad7 can potentiate BMP2-induced chondrogenic differentiation and inhibit endochondral ossification. Thus, inhibit the expression of Smad7 in BMP2-induced hSMSCs differentiation may be a new strategy for cartilage tissue engineering.

scholarly journals The Effect of Human Platelet-Rich Plasma and L-Ascorbic Acid on Morphology, Proliferation, and Chondrogenesis Ability towards Human Adipose-Derived Stem Cells

2019 ◽  
Vol 3 (1) ◽  
pp. 26 ◽  
Author(s):  
Imam Rosadi ◽  
Karina Karina ◽  
Iis Rosliana ◽  
Siti Sobariah ◽  
Irsyah Afini ◽  
...  
Keyword(s):  
Stem Cells ◽  
Ascorbic Acid ◽  
Alcian Blue ◽  
Human Platelet ◽  
Platelet Rich Plasma ◽  
Cartilage Tissue ◽  
Population Doubling ◽  
Blue Staining ◽  
Alcian Blue Staining ◽  
Adipose Derived Stem Cells

Background: Microtia is a congenital malformation in the external ear due to cartilage defect. Adipose-derived stem cells (ADSC) is promising cells to develop cartilage tissue engineering for microtia. In this study, we focused on proliferation and chondrogenesis of ADSC in three different media, which consist of 10% fetal bovine serum (FBS), 10% FBS with L-ascorbic acid, and 10% human platelet rich plasma (PRP). Methods: ADSC were induced to differentiate into adipocytes, chondrocyte and osteocytes. ADSC morphology, proliferation and population doubling time was compared in three different media and analysed. Observation and alcian blue staining were done every 7 days to assess chondrogenic potency of ADSC from each treatment.Results: Isolated ADSC were able to differentiate into adipocytes, osteocytes and chondrocytes. ADSC in all group have fibroblast-like morphology, but cells in 10% FBS and 10% FBS with LAA group were flattened and larger. ADSC in 10% PRP group proliferates faster than 10% FBS with and without LAA. PDT values of ADSC were 34 hours, 44 hours and 48 hours, respectively for 10% PRP, 10% FBS with LAA and 10% FBS group. Alcian blue staining revealed that ADSC in 10% FBS with LAA and 10% PRP were able to proceed to chondrogenesis when cultured time were prolong up to 21 days, but not with ADSC in 10% FBS. Conclusion: We conclude that adding 10% FBS with LAA or 10% PRP into medium culture can support proliferation and chondrogenesis of ADSC. Key words: human ADSC, PRP, L-ascorbic acid, proliferation, chondrogenesis

scholarly journals Human Platelet Lysate Acts Synergistically With Laminin to Improve the Neurotrophic Effect of Human Adipose-Derived Stem Cells on Primary Neurons in vitro

2021 ◽  
Vol 9 ◽  
Author(s):  
Martino Guiotto ◽  
Wassim Raffoul ◽  
Andrew M. Hart ◽  
Mathis O. Riehle ◽  
Pietro G. di Summa
Keyword(s):  
Stem Cells ◽  
Neurite Outgrowth ◽  
Animal Studies ◽  
Human Platelet ◽  
Platelet Lysate ◽  
Neural Cells ◽  
Adipose Derived Stem Cells ◽  
Peripheral Nerve Lesions ◽  
Human Platelet Lysate

BackgroundDespite the advancements in microsurgical techniques and noteworthy research in the last decade, peripheral nerve lesions have still weak functional outcomes in current clinical practice. However, cell transplantation of human adipose-derived stem cells (hADSC) in a bioengineered conduit has shown promising results in animal studies. Human platelet lysate (hPL) has been adopted to avoid fetal bovine serum (FBS) in consideration of the biosafety concerns inherent with the use of animal-derived products in tissue processing and cell culture steps for translational purposes. In this work, we investigate how the interplay between hPL-expanded hADSC (hADSChPL) and extracellular matrix (ECM) proteins influences key elements of nerve regeneration.MethodshADSC were seeded on different ECM coatings (laminin, LN; fibronectin, FN) in hPL (or FBS)-supplemented medium and co-cultured with primary dorsal root ganglion (DRG) to establish the intrinsic effects of cell–ECM contact on neural outgrowth. Co-cultures were performed “direct,” where neural cells were seeded in contact with hADSC expanded on ECM-coated substrates (contact effect), or “indirect,” where DRG was treated with their conditioned medium (secretome effect). Brain-derived nerve factor (BDNF) levels were quantified. Tissue culture plastic (TCPS) was used as the control substrate in all the experiments.ResultshPL as supplement alone did not promote higher neurite elongation than FBS when combined with DRG on ECM substrates. However, in the presence of hADSC, hPL could dramatically enhance the stem cell effect with increased DRG neurite outgrowth when compared with FBS conditions, regardless of the ECM coating (in both indirect and direct co-cultures). The role of ECM substrates in influencing neurite outgrowth was less evident in the FBS conditions, while it was significantly amplified in the presence of hPL, showing better neural elongation in LN conditions when compared with FN and TCPS. Concerning hADSC growth factor secretion, ELISA showed significantly higher concentrations of BDNF when cells were expanded in hPL compared with FBS-added medium, without significant differences between cells cultured on the different ECM substrates.ConclusionThe data suggest how hADSC grown on LN and supplemented with hPL could be active and prone to support neuron–matrix interactions. hPL enhanced hADSC effects by increasing both proliferation and neurotrophic properties, including BDNF release.

scholarly journals Developing a Glyoxal-Crosslinked Chitosan/Gelatin Hydrogel for Sustained Release of Human Platelet Lysate to Promote Tissue Regeneration

2021 ◽  
Vol 22 (12) ◽  
pp. 6451
Author(s):  
Ching-Cheng Tsai ◽  
Tai-Horng Young ◽  
Guang-Shih Chen ◽  
Nai-Chen Cheng
Keyword(s):  
Growth Factor ◽  
Sustained Release ◽  
Tissue Regeneration ◽  
Transforming Growth Factor ◽  
Human Platelet ◽  
Umbilical Vein ◽  
Platelet Lysate ◽  
Release Pattern ◽  
Human Platelet Lysate

The clinical application of human platelet lysate (HPL) holds promise for tissue regeneration, and the development of an efficient vehicle for its delivery is desired. Chitosan-based hydrogels are potential candidates, but they often exhibit weak mechanical properties. In this study, a chitosan/gelatin (CS-GE) hydrogel crosslinked by glyoxal was fabricated for sustained release of HPL. The influence of HPL on Hs68 fibroblast and human umbilical vein endothelial cell (HUVEC) culture was evaluated, and we found that supplementing 5% HPL in the medium could significantly improve cell proliferation relative to supplementing 10% fetal bovine serum (FBS). Moreover, HPL accelerated the in vitro wound closure of Hs68 cells and facilitated the tube formation of HUVECs. Subsequently, we fabricated CS-GE hydrogels crosslinked with different concentrations of glyoxal, and the release pattern of FITC-dextrans (4, 40 and 500 kDa) from the hydrogels was assessed. After an ideal glyoxal concentration was determined, we further characterized the crosslinked CS-GE hydrogels encapsulated with different amounts of HPL. The HPL-incorporated hydrogel was shown to significantly promote the proliferation of Hs68 cells and the migration of HUVECs. Moreover, the release pattern of transforming growth factor-β1 (TGF-β1) and platelet-derived growth factor-BB (PDGF-BB) from hydrogel was examined in vitro, demonstrating a sustained release profile of the growth factors. Finally, the chick chorioallantoic membrane assay revealed that HPL encapsulation in the hydrogel significantly stimulated angiogenesis in ovo. These results demonstrate the great potential of the crosslinked CS-GE hydrogel to serve as an effective delivery system for HPL to promote tissue regeneration.

Exosomes derived from human platelet lysate affect MSC functions in vitro

2014 ◽  
Author(s):  
Elena Torreggiani ◽  
Francesca Perut ◽  
Laura Roncuzzi ◽  
Nicola Baldini
Keyword(s):  
Human Platelet ◽  
Platelet Lysate ◽  
Human Platelet Lysate

The effect of human platelet lysate on corneal nerve regeneration

2019 ◽  
pp. bjophthalmol-2019-314408 ◽  
Author(s):  
Chin-Te Huang ◽  
Hsiao-Sang Chu ◽  
Kuo-Chi Hung ◽  
Lily Wei Chen ◽  
Mei-Yun Chen ◽  
...  
Keyword(s):  
Wound Healing ◽  
Nerve Regeneration ◽  
Neurotrophic Factors ◽  
Ganglion Cells ◽  
Human Platelet ◽  
Platelet Lysate ◽  
Human Platelet Lysate ◽  
Corneal Nerve

AimThis study aimed to test whether human platelet lysate (HPL) has neurotrophic ability for corneal nerve regeneration.MethodsWe measured the neurotrophic factors in human peripheral serum (HPS) and two commercially available HPLs, UltraGRO and PLTMax. In vitro, we compared the growth rates, neuronal differentiation and immunostaining of neuron markers in mouse neuroblastoma cell line (Neuro-2a) and primary culture of mouse trigeminal ganglion cells that were cultivated in different concentrations of fetal bovine serum, HPS and HPL. In vivo, we created corneal wounds on Sprague Dawley rats with a rotating burr and evaluated the effects of topical HPL on wound healing and corneal nerve regeneration by in vivo confocal microscopy and corneal aesthesiometry.ResultsHPLs had significantly higher concentrations of various neurotrophic factors compared with HPS (p<0.05). In Neuro-2a cells, 3% HPL was better at promoting neuronal growth and differentiation compared with HPS at the same concentration. HPL was also found to have superior neurotrophic effects compared with HPS in primary cultures of mouse trigeminal ganglion cells. In vivo, HPL-treated eyes had better corneal epithelial wound healing rate, nerve regeneration length and corneal touch threshold compared with eyes treated with artificial tears (p<0.05).ConclusionHPL has significantly higher concentrations of neurotrophic factors compared with HPS. It showed not only in vitro but also in vivo corneal neurotrophic abilities. Our results suggest that HPL may have a potential role in the treatment of diseases related to corneal nerve damage or degeneration.

Sign in / Sign up

Export Citation Format

Share Document