scholarly journals A Hyaluronan and Platelet-Rich Plasma Hydrogel for Mesenchymal Stem Cell Delivery in the Intervertebral Disc: An Organ Culture Study

2021 ◽  
Vol 22 (6) ◽  
pp. 2963
Author(s):  
Fabrizio Russo ◽  
Luca Ambrosio ◽  
Marianna Peroglio ◽  
Wei Guo ◽  
Sebastian Wangler ◽  
...  
Keyword(s):  
Gene Expression ◽  
Intervertebral Disc ◽  
Organ Culture ◽  
Platelet Rich Plasma ◽  
Cell Activity ◽  
Human Mesenchymal Stem Cells ◽  
Cell Phenotype ◽  
Cytokeratin 19 ◽  
Stem Cell Delivery ◽  
Normal Metabolism

The purpose of the present pilot study was to evaluate the effect of a hydrogel composed of hyaluronic acid (HA) and platelet-rich plasma (PRP) as a carrier for human mesenchymal stem cells (hMSCs) for intervertebral disc (IVD) regeneration using a disc organ culture model. HA was mixed with batroxobin (BTX) and PRP to form a hydrogel encapsulating 1 × 106 or 2 × 106 hMSCs. Bovine IVDs were nucleotomized and filled with hMSCs suspended in ~200 μL of the PRP/HA/BTX hydrogel. IVDs collected at day 0 and nucleotomized IVDs with no hMSCs and/or hydrogel alone were used as controls. hMSCs encapsulated in the hydrogel were also cultured in well plates to evaluate the effect of the IVD environment on hMSCs. After 1 week, tissue structure, scaffold integration, hMSC viability and gene expression of matrix and nucleus pulposus (NP) cell markers were assessed. Histological analysis showed a better preservation of the viability of the IVD tissue adjacent to the gel in the presence of hMSCs (~70%) compared to the hydrogel without hMSCs. Furthermore, disc morphology was maintained, and the hydrogel showed signs of integration with the surrounding tissues. At the gene expression level, the hydrogel loaded with hMSCs preserved the normal metabolism of the tissue. The IVD environment promoted hMSC differentiation towards a NP cell phenotype by increasing cytokeratin-19 (KRT19) gene expression. This study demonstrated that the hydrogel composed of HA/PRP/BTX represents a valid carrier for hMSCs being able to maintain a good cell viability while stimulating cell activity and NP marker expression.

scholarly journals An organ culture system to model early degenerative changes of the intervertebral disc II: profiling global gene expression changes

2013 ◽  
Vol 15 (5) ◽  
pp. R121 ◽  
Author(s):  
Dessislava Z Markova ◽  
Christopher K Kepler ◽  
Sankar Addya ◽  
Hallie B Murray ◽  
Alexander R Vaccaro ◽  
...  
Keyword(s):  
Gene Expression ◽  
Intervertebral Disc ◽  
Organ Culture ◽  
Culture System ◽  
Global Gene Expression ◽  
Degenerative Changes ◽  
Organ Culture System

scholarly journals Regenerative Potential of Blood-Derived Products in 3D Osteoarthritic Chondrocyte Culture System

2021 ◽  
Vol 43 (2) ◽  
pp. 665-675
Author(s):  
Olga Kuten-Pella ◽  
Andrea De Luna ◽  
Karina Kramer ◽  
Markus Neubauer ◽  
Stefan Nehrer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Platelet Rich Plasma ◽  
Cell Activity ◽  
Blood Products ◽  
Tissue Quality ◽  
Xtt Assay ◽  
Matrix Deposition ◽  
Genes Expression ◽  
Significant Difference

Intra-articular injection of different types of blood-derived products is gaining popularity and clinical importance in the treatment of degenerative cartilage disorders such as osteoarthritis. The regenerative potential of two types of platelet-rich plasma (PRP), prepared in the presence of EDTA (EPRP) and citrate (CPRP) and an alternative blood product-hyperacute serum (hypACT) was evaluated using a 3D osteoarthritic chondrocyte pellet model by assessing the metabolic cell activity, cartilage-related gene expression and extracellular matrix deposition within the pellets. Chondrocyte viability was determined by XTT assay and it revealed no significant difference in metabolic activity of OA chondrocyte pellets after supplementation with different blood products. Nevertheless, the selection of blood products influenced the cartilage-related genes expression, ECM morphology and the tissue quality of pellets. Both PRP types had a different biological effect depending upon concentration and even though CPRP is widely used in clinics our assessment did not reveal good results in gene expression either tissue quality. HypACT supplementation resulted in superior cartilage-related genes expression together with tissue quality and seemed to be the most stable product since no remarkable changes were observed between the two different concentrations. All in all, for successful regenerative therapy, possible molecular mechanisms induced by blood-derived products should be always carefully investigated and adapted to the specific medical indications.

DNA Demethylation Regulates Anabolic and Catabolic Gene Expression in Intervertebral Disc Cells

2014 ◽  
Vol 4 (1_suppl) ◽  
pp. s-0034-1376587-s-0034-1376587
Author(s):  
N. Chutkan ◽  
R. Sangani ◽  
H. Zhou ◽  
S. Fulzele
Keyword(s):  
Gene Expression ◽  
Intervertebral Disc ◽  
Dna Demethylation ◽  
Catabolic Gene ◽  
Disc Cells

Anti-Inflammatory Effects of Novel Standardized Platelet Rich Plasma Releasates on Knee Osteoarthritic Chondrocytes and Cartilage in vitro

2019 ◽  
Vol 20 (11) ◽  
pp. 920-933 ◽  
Author(s):  
Lucía Gato-Calvo ◽  
Tamara Hermida-Gómez ◽  
Cristina R. Romero ◽  
Elena F. Burguera ◽  
Francisco J. Blanco
Keyword(s):  
Gene Expression ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Cartilage Explants ◽  
Ex Vivo Model ◽  
Chondrocyte Viability ◽  
Platelet Concentration ◽  
The Absolute ◽  
Anti Inflammatory

Background: Platelet Rich Plasma (PRP) has recently emerged as a potential treatment for osteoarthritis (OA), but composition heterogeneity hampers comparison among studies, with the result that definite conclusions on its efficacy have not been reached. Objective: 1) To develop a novel methodology to prepare a series of standardized PRP releasates (PRP-Rs) with known absolute platelet concentrations, and 2) To evaluate the influence of this standardization parameter on the anti-inflammatory properties of these PRP-Rs in an in vitro and an ex vivo model of OA. Methods: A series of PRPs was prepared using the absolute platelet concentration as the standardization parameter. Doses of platelets ranged from 0% (platelet poor plasma, PPP) to 1.5·105 platelets/µl. PRPs were then activated with CaCl2 to obtain releasates (PRP-R). Chondrocytes were stimulated with 10% of each PRP-R in serum-free culture medium for 72 h to assess proliferation and viability. Cells were co-stimulated with interleukin (IL)-1β (5 ng/ml) and 10% of each PRP-R for 48 h to determine the effects on gene expression, secretion and intra-cellular content of common markers associated with inflammation, catabolism and oxidative stress in OA. OA cartilage explants were co-stimulated with IL-1β (5 ng/ml) and 10% of either PRP-R with 0.75·105 platelets/µl or PRP-R with 1.5·105 platelets/µl for 21 days to assess matrix inflammatory degradation. Results: Chondrocyte viability was not affected, and proliferation was dose-dependently increased. The gene expression of all pro-inflammatory mediators was significantly and dose-independently reduced, except for that of IL-1β and IL-8. Immunoblotting corroborated this effect for inducible NO synthase (NOS2). Secreted matrix metalloproteinase-13 (MMP-13) was reduced to almost basal levels by the PRP-R from PPP. Increasing platelet dosage led to progressive loss to this anti-catabolic ability. Safranin O and toluidine blue stains supported the beneficial effect of low platelet dosage on cartilage matrix preservation. Conclusion: We have developed a methodology to prepare PRP releasates using the absolute platelet concentration as the standardization parameter. Using this approach, the composition of the resulting PRP derived product is independent of the donor initial basal platelet count, thereby allowing the evaluation of its effects objectively and reproducibly. In our OA models, PRP-Rs showed antiinflammatory, anti-oxidant and anti-catabolic properties. Platelet enrichment could favor chondrocyte proliferation but is not necessary for the above effects and could even be counter-productive.

scholarly journals Resveratrol promotes osteogenesis of human mesenchymal stem cells by upregulating RUNX2 gene expression via the SIRT1/FOXO3A axis

2011 ◽  
Vol 26 (10) ◽  
pp. 2552-2563 ◽  
Author(s):  
Pei-Chi Tseng ◽  
Sheng-Mou Hou ◽  
Ruey-Jien Chen ◽  
Hsiao-Wen Peng ◽  
Chi-Fen Hsieh ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Human Mesenchymal Stem Cells

scholarly journals A50 MODULATION OF GOBLET CELL ACTIVITY DURING GIARDIA DUODENALIS INFECTION: A ROLE FOR PAR2

2021 ◽  
Vol 4 (Supplement_1) ◽  
pp. 6-7
Author(s):  
E Fekete ◽  
C B Amat ◽  
T Allain ◽  
M Hollenberg ◽  
K Mihara ◽  
...  
Keyword(s):  
Gene Expression ◽  
Goblet Cell ◽  
Cysteine Protease ◽  
Protease Activity ◽  
Calcium Release ◽  
Cell Activity ◽  
Goblet Cells ◽  
Giardia Infection ◽  
Mucus Production ◽  
Mucin Gene

Abstract Background Giardia duodenalis has been shown to alter the structure of the intestinal mucus layers during infection via obscure mechanisms. We hypothesize that goblet cell activity may be disrupted in part due to proteolytic activation of protease-activated receptor 2 (PAR2) by Giardia proteases, resulting in disruption of mucus production and secretion by intestinal goblet cells. Aims Characterize alterations in goblet cell activity during Giardia infection, focusing on the roles of Giardia protease activity and PAR2. Methods Chinese hamster ovary cells transfected with nano-luciferase tagged PAR2 were incubated with Giardia NF or GSM trophozoites. Cleavage within the activation domain results in release of enzymes into the supernatant. Luminescence in the supernatant was measured as an indication of PAR cleavage by Giardia. LS174T, a human colonic mucus-producing cell line, was infected with Giardia trophozoites (isolates NF, WB, S2, and GSM). Prior to infection, trophozoites were treated with E64, a broad-spectrum cysteine protease inhibitor, and LS174T were treated with a PAR2 antagonist, a calcium chelator, or an ERK1/2 inhibitor. Quantitative PCR (qPCR) was performed for the MUC2 mucin gene. Wild-type (WT) and PAR2 knockout (KO) mice were infected with Giardia. Colonic mucus was stained using fluorescein-coupled wheat-germ agglutinin (WGA), and qPCR was performed for Muc2 and Muc5ac. Results Giardia trophozoites cleaved PAR2 within the N-terminal activation domain in a cysteine protease-dependent manner. Cleavage was isolate dependent, with isolates that show higher protease activity cleaving at a higher rate. High protease activity Giardia isolates increased MUC2 gene expression in LS714T. This increase was attenuated by inhibition of Giardia cysteine protease activity, and by antagonism of PAR2, inhibition of calcium release, or inhibition of ERK1/2 activity in LS174T cells. Both Muc2 and Muc5ac expression were upregulated in the colons of WT mice in response to Giardia infection, while in the jejunum Muc2 expression decreased and Muc5ac expression increased. In KO, no changes in gene expression were seen in the colon in response to Giardia infection, while in the jejunum, Muc2 expression was unchanged and Muc5ac expression decreased. Both WT infected and KO noninfected mice showed thinning of the colonic mucus layer compared to WT controls. There was some recovery in thickness in KO infected mice. Conclusions PAR2 plays a significant role in the regulation of mucin gene expression in mice and in a human colonic cell line. Results suggest that Giardia cysteine proteases cleave and activate PAR2, leading to calcium release and activation of the MAPK pathway in goblet cells, ultimately leading to altered mucin gene expression. Findings identify a novel regulatory pathway for mucus production by intestinal goblet cells. Funding Agencies CAG, CCC

In vitro and in vivo Effect of Platelet-Rich Plasma-Based Autologous Topical Serum on Cutaneous Wound Healing

2021 ◽  
pp. 1-13
Author(s):  
Eduardo Anitua ◽  
Victoria Muñoz ◽  
Libe Aspe ◽  
Roberto Tierno ◽  
Adrian García-Salvador ◽  
...  
Keyword(s):  
Wound Healing ◽  
Tissue Damage ◽  
Collagen Synthesis ◽  
Dermal Fibroblast ◽  
Platelet Rich Plasma ◽  
Cell Activity ◽  
Cutaneous Wound Healing ◽  
Cutaneous Wound ◽  
Fibroblast Migration ◽  
Edge Contraction

<b><i>Introduction:</i></b> Skin injury and wound healing is an inevitable event during lifetime. However, several complications may hamper the regeneration of the cutaneous tissue and lead to a chronic profile that prolongs patient recovery. Platelet-rich plasma is rising as an effective and safe alternative to the management of wounds. However, this technology presents some limitations such as the need for repeated blood extractions and health-care interventions. <b><i>Objective:</i></b> The aim of this study was to assess the use of an endogenous and storable topical serum (ES) derived from plasma rich in growth factors promoting wound healing, and to obtain preliminary data regarding its clinical and experimental effect over ulcerated skin models and patient care. <b><i>Methods:</i></b> Human dermal fibroblast and 3D organotypic ulcerated skin models were used to assess ES over the main mechanisms of wound healing including cell migration, edge contraction, collagen synthesis, tissue damage, extracellular matrix remodeling, cell death, metabolic activity, and histomorphometry analysis. Additionally, 4 patients suffering from skin wounds were treated and clinically assessed. <b><i>Results:</i></b> ES promoted dermal fibroblast migration, wound edge contraction, and collagen synthesis. When topically applied, ES increased collagen and elastin deposition and reduced tissue damage. The interstitial edema, structural integrity, and cell activity were also maintained, and apoptotic levels were reduced. Patients suffering from hard-to-heal wounds of different etiologies were treated with ES, and the ulcers healed completely within few weeks with no reported adverse events. <b><i>Conclusion:</i></b> This preliminary study suggests that ES might promote cutaneous wound healing and may be useful for accelerating the re-epithelization of skin ulcers.

scholarly journals Influence of Angiopoietin Treatment with Hypoxia and Normoxia on Human Intervertebral Disc Progenitor Cell’s Proliferation, Metabolic Activity, and Phenotype

2021 ◽  
Vol 11 (15) ◽  
pp. 7144
Author(s):  
Muriel C. Bischof ◽  
Sonja Häckel ◽  
Andrea Oberli ◽  
Andreas S. Croft ◽  
Katharina A. C. Oswald ◽  
...  
Keyword(s):  
Gene Expression ◽  
Intervertebral Disc ◽  
Metabolic Activity ◽  
Cell Metabolism ◽  
Trauma Patients ◽  
Low Back ◽  
Relative Gene Expression ◽  
Progenitor Cell Population ◽  
Ivd Degeneration ◽  
Relative Gene

Increasing evidence implicates intervertebral disc (IVD) degeneration as a major contributor to low back pain. In addition to a series of pathogenic processes, degenerated IVDs become vascularized in contrast to healthy IVDs. In this context, angiopoietin (Ang) plays a crucial role and is involved in cytokine recruitment, and anabolic and catabolic reactions within the extracellular matrix (ECM). Over the last decade, a progenitor cell population has been described in the nucleus pulposus (NP) of the IVD to be positive for the Tie2 marker (also known as Ang-1 receptor). In this study, we investigated the influence of Ang-1 and Ang-2 on human NP cell (Tie2+, Tie2- or mixed) populations isolated from trauma patients during 7 days in normoxia (21% O2) or hypoxia (≤ 5% O2). At the end of the process, the proliferation and metabolic activity of the NP cells were analyzed. Additionally, the relative gene expression of NP-related markers was evaluated. NP cells showed a higher proliferation depending on the Ang treatment. Moreover, the study revealed higher NP cell metabolism when cultured in hypoxia. Additionally, the relative gene expression followed, with an increase linked to the oxygen level and Ang concentration. Our study comparing different NP cell populations may be the start of new approaches for the treatment of IVD degeneration.

Cell shape and gene expression in human intervertebral disc cells: in vitro tissue engineering studies

2003 ◽  
Vol 78 (2) ◽  
pp. 109-117 ◽  
Author(s):  
He Gruber ◽  
Ja Ingram ◽  
K Leslie ◽  
Hj Norton ◽  
En Hanley Jr
Keyword(s):  
Gene Expression ◽  
Tissue Engineering ◽  
Intervertebral Disc ◽  
Cell Shape ◽  
Engineering Studies ◽  
Disc Cells
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