Suppression of discoidin domain receptor 1 expression enhances the chondrogenesis of adipose-derived stem cells

Effectively directing the chondrogenesis of adipose-derived stem cells (ADSCs) to engineer articular cartilage represents an important challenge in ADSC-based articular cartilage tissue engineering. The discoidin domain receptor 1 (DDR1) has been shown to affect cartilage homeostasis; however, little is known about the roles of DDR1 in ADSC chondrogenesis. In this study, we used the three-dimensional culture pellet culture model system with chondrogenic induction to investigate the roles of DDR1 in the chondrogenic differentiation of human ADSCs (hADSCs). Real-time polymerase chain reaction and Western blot were used to detect the expression of DDRs and chondrogenic genes. Sulfated glycosaminoglycan (sGAG) was detected by Alcian blue and dimethylmethylene blue (DMMB) assays. Terminal deoxy-nucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining was used to assess cell death. During the chondrogenesis of hADSCs, the expression of DDR1 but not DDR2 was significantly elevated. The depletion of DDR1 expression in hADSCs using short hairpin RNA increased the expression of chondrogenic genes (SOX-9, collagen type II, and aggrecan) and cartilaginous matrix deposition (collagen type II and sGAG) and only slightly increased cell death (2–8%). DDR1 overexpression in hADSCs decreased the expression of chondrogenic genes (SOX-9, collagen type II, and aggrecan) and sGAG and enhanced hADSC survival. Moreover, DDR1-depleted hADSCs showed decreased expression of the terminal differentiation genes runt-related transcription factor 2 (Runx2) and matrix metalloproteinase 13 (MMP-13). These results suggest that DDR1 suppression may enhance ADSC chondrogenesis by enhancing the expression of chondrogenic genes and cartilaginous matrix deposition. We proposed that the suppression of DDR1 in ADSCs may be a candidate strategy of genetic modification to optimize ADSC-based articular cartilage tissue engineering.

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Simvastatin Enhances the Chondrogenesis but Not the Osteogenesis of Adipose-Derived Stem Cells in a Hyaluronan Microenvironment

Biomedicines ◽

10.3390/biomedicines9050559 ◽

2021 ◽

Vol 9 (5) ◽

pp. 559

Author(s):

Shun-Cheng Wu ◽

Chih-Hsiang Chang ◽

Ling-Hua Chang ◽

Che-Wei Wu ◽

Jhen-Wei Chen ◽

...

Keyword(s):

Stem Cells ◽

Articular Cartilage ◽

Mrna Expression ◽

Collagen Type ◽

Cartilage Regeneration ◽

Bone Morphogenetic Protein 2 ◽

Adipose Derived Stem Cells ◽

Type Ii ◽

Collagen Type Ii ◽

Articular Cartilage Regeneration

Directing adipose-derived stem cells (ADSCs) toward chondrogenesis is critical for ADSC-based articular cartilage regeneration. Simvastatin (SIM) was reported to promote both chondrogenic and osteogenic differentiation of ADSCs by upregulating bone morphogenetic protein-2 (BMP-2). We previously found that ADSC chondrogenesis is initiated and promoted in a hyaluronan (HA) microenvironment (HAM). Here, we further hypothesized that SIM augments HAM-induced chondrogenesis but not osteogenesis of ADSCs. ADSCs were treated with SIM in a HAM (SIM plus HAM) by HA-coated wells or HA-enriched fibrin (HA/Fibrin) hydrogel, and chondrogenic differentiation of ADSCs was evaluated. SIM plus HAM increased chondrogenesis more than HAM or SIM alone, including cell aggregation, chondrogenic gene expression (collagen type II and aggrecan) and cartilaginous tissue formation (collagen type II and sulfated glycosaminoglycan). In contrast, SIM-induced osteogenesis in ADSCs was reduced in SIM plus HAM, including mRNA expression of osteogenic genes, osteocalcin and alkaline phosphatase (ALP), ALP activity and mineralization. SIM plus HAM also showed the most effective increases in the mRNA expression of BMP-2 and transcription factors of SOX-9 and RUNX-2 in ADSCs, while these effects were reversed by CD44 blockade. HAM suppressed the levels of JNK, p-JNK, P38 and p-P38 in ADSCs, and SIM plus HAM also decreased SIM-induced phosphorylated JNK and p38 levels. In addition, SIM enhanced articular cartilage regeneration, as demonstrated by implantation of an ADSCs/HA/Fibrin construct in an ex vivo porcine articular chondral defect model. The results from this study indicate that SIM may be an enhancer of HAM-initiated MSC-based chondrogenesis and avoid osteogenesis.

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Auricular Cartilage Regeneration with Adipose-Derived Stem Cells in Rabbits

Mediators of Inflammation ◽

10.1155/2018/4267158 ◽

2018 ◽

Vol 2018 ◽

pp. 1-8 ◽

Cited By ~ 2

Author(s):

Se-Joon Oh ◽

Hee-Young Park ◽

Kyung-Un Choi ◽

Sung-Won Choi ◽

Sung-Dong Kim ◽

...

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Cartilage Defect ◽

Collagen Type ◽

Cartilage Regeneration ◽

Control Group ◽

Adipose Derived Stem Cells ◽

Type Ii ◽

Collagen Type Ii ◽

Auricular Cartilage

Tissue engineering cell-based therapy using induced pluripotent stem cells and adipose-derived stem cells (ASCs) may be promising tools for therapeutic applications in tissue engineering because of their abundance, relatively easy harvesting, and high proliferation potential. The purpose of this study was to investigate whether ASCs can promote the auricular cartilage regeneration in the rabbit. In order to assess their differentiation ability, ASCs were injected into the midportion of a surgically created auricular cartilage defect in the rabbit. Control group was injected with normal saline. After 1 month, the resected auricles were examined histopathologically and immunohistochemically. The expression of collagen type II and transforming growth factor-β1 (TGF-β1) were analyzed by quantitative polymerase chain reaction. Histopathology showed islands of new cartilage formation at the site of the surgically induced defect in the ASC group. Furthermore, Masson’s trichrome staining and immunohistochemistry for S-100 showed numerous positive chondroblasts. The expression of collagen type II and TGF-β1 were significantly higher in the ASCs than in the control group. In conclusion, ASCs have regenerative effects on the auricular cartilage defect of the rabbit. These effects would be expected to contribute significantly to the regeneration of damaged cartilage tissue in vivo.

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A 3D Porous Gelatin-Alginate-Based-IPN Acts as an Efficient Promoter of Chondrogenesis from Human Adipose-Derived Stem Cells

Stem Cells International ◽

10.1155/2015/252909 ◽

2015 ◽

Vol 2015 ◽

pp. 1-17 ◽

Cited By ~ 12

Author(s):

Sorina Dinescu ◽

Bianca Galateanu ◽

Eugen Radu ◽

Anca Hermenean ◽

Adriana Lungu ◽

...

Keyword(s):

Stem Cells ◽

Collagen Type ◽

Cartilage Tissue Engineering ◽

Superior Performance ◽

Cartilage Tissue ◽

Adipose Derived Stem Cells ◽

Interpenetrating Network ◽

Cell Scaffold ◽

New Strategies

Cartilage has limited regeneration potential. Thus, there is an imperative need to develop new strategies for cartilage tissue engineering (CTE) amenable for clinical use. Recent CTE approaches rely on optimal cell-scaffold interactions, which require a great deal of optimization. In this study we attempt to build a novel gelatin- (G-) alginate- (A-) polyacrylamide (PAA) 3D interpenetrating network (IPN) with superior performance in promoting chondrogenesis from human adipose-derived stem cells (hADSCs). We show that our G-A-PAA scaffold is capable of supporting hADSCs proliferation and survival, with no apparent cytotoxic effect. Moreover, we find that after exposure to prochondrogenic conditions a key transcription factor known to induce chondrogenesis, namely, Sox9, is highly expressed in our hADSCs/G-A-PAA bioconstruct, along with cartilage specific markers such as collagen type II, CEP68, and COMP extracellular matrix (ECM) components. These data suggest that our G-A-PAA structural properties and formulation might enable hADSCs conversion towards functional chondrocytes. We conclude that our novel G-A-PAA biomatrix is a good candidate for prospectivein vivoCTE applications.

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The Antihypertensive Drug Nifedipine Modulates the Metabolism of Chondrocytes and Human Bone Marrow-Derived Mesenchymal Stem Cells

10.21203/rs.2.10478/v1 ◽

2019 ◽

Author(s):

Ilona Uzieliene ◽

Eiva Bernotiene ◽

Greta Urbonaite ◽

Jaroslav Denkovskij ◽

Edvardas Bagdonas ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Antihypertensive Drug ◽

Mitochondrial Respiration ◽

Collagen Type ◽

Cell Types ◽

Cartilage Tissue ◽

Type Ii ◽

Collagen Type Ii

Abstract Aging is associated with the development of various chronic diseases, in which both hypertension and osteoarthritis (OA) are dominant. Currently, there is no effective treatment for OA, whereas hypertension is often treated using L-type voltage-operated calcium channel (VOCC) blocking drugs, nifedipine being among the most classical ones. Although nifedipine together with other VOCC inhibitors plays an important role in people wellbeing, there are unresolved questions on its possible effect on cartilage tissue homeostasis and the development of OA. Due to that, the aim of this study was to analyse the effects of nifedipine on metabolic processes in human chondrocytes and bone marrow mesenchymal stem cells (BMMSCs). To analyze whether those events were mediated specifically through VOCC, agonist BayK8644 was used. Our results demonstrate that nifedipine downregulated chondrocyte proliferation rate as well as mitochondrial respiration and ATP production (Agilent Seahorse) in both cell types. Analysis of cartilage explant histological sections by electron microscopy also suggested that part of mitochondria lose their activity in response to nifedipine.However, switch of energetic metabolic pathway towards glycolytic was observed only in chondrocytes. Stimulation with either nifedipine or BayK8644 resulted in elevated production of collagen type II and proteoglycans in micromass cultures under chondrogenic condition, although the effects of VOCC inhibitor Bay8466 were less expressed. Nitric oxide (NO) activity, as measured by flow cytometry, was upregulated by nifedipine in BMMSCs and particularly chondrocytes, suggesting that NO at least in part may account for the effects of nifedipine on metabolism in both tested cell types.Taken together, we conclude that antihypertensive drug nifedipine inhibits mitochondrial respiration in both chondrocytes and BMMSCs and that these effects may be associated with increased NO accumulation and pro-inflammatory activity. Glycolytic capacity was enhanced only in chondrocytes, suggesting that these cells have the capacity to switch from oxidative phosphorylation to glycolysis and alter their metabolic activity in response to VOCC inhibition. Finally, nifedipine stimulated production of collagen type II and proteoglycans in both cell types, implying its potentially beneficial anabolic effects on articular cartilage. These results highlight a potential link between consumption of antihypertensive drugs and cartilage health

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Chondrogenic Differentiation of Human Adipose-Derived Stem Cells: A New Path in Articular Cartilage Defect Management?

BioMed Research International ◽

10.1155/2014/740926 ◽

2014 ◽

Vol 2014 ◽

pp. 1-7 ◽

Cited By ~ 17

Author(s):

Jan-Philipp Stromps ◽

Nora Emilie Paul ◽

Björn Rath ◽

Mahtab Nourbakhsh ◽

Jürgen Bernhagen ◽

...

Keyword(s):

Stem Cells ◽

Articular Cartilage ◽

Cartilage Tissue Engineering ◽

Cartilage Tissue ◽

Cartilage Defects ◽

Adipose Derived Stem Cells ◽

Age Related ◽

Healthy Cartilage ◽

Control And Prevention ◽

Articular Cartilage Defects

According to data published by the Centers for Disease Control and Prevention, over 6 million people undergo a variety of medical procedures for the repair of articular cartilage defects in the U.S. each year. Trauma, tumor, and age-related degeneration can cause major defects in articular cartilage, which has a poor intrinsic capacity for healing. Therefore, there is substantial interest in the development of novel cartilage tissue engineering strategies to restore articular cartilage defects to a normal or prediseased state. Special attention has been paid to the expansion of chondrocytes, which produce and maintain the cartilaginous matrix in healthy cartilage. This review summarizes the current efforts to generate chondrocytes from adipose-derived stem cells (ASCs) and provides an outlook on promising future strategies.

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Infrapatellar Fat Pads–Derived Stem Cell Is a Favorable Cell Source for Articular Cartilage Tissue Engineering: An In Vitro and Ex Vivo Study Based on 3D Organized Self-Assembled Biomimetic Scaffold

Cartilage ◽

10.1177/1947603520988153 ◽

2021 ◽

pp. 194760352098815

Author(s):

Chen-Chie Wang ◽

Ing-Ho Chen ◽

Ya-Ting Yang ◽

Yi-Ru Chen ◽

Kai-Chiang Yang

Keyword(s):

Tissue Engineering ◽

Adipose Tissue ◽

Collagen Type ◽

Ex Vivo ◽

Cartilage Tissue Engineering ◽

Cartilage Regeneration ◽

Cartilage Tissue ◽

Type Ii ◽

Collagen Type Ii ◽

Fat Pads

Objective Adipose tissue–derived stem cells (ASCs) are a promising source of cells for articular cartilage regeneration. However, ASCs isolated from different adipose tissue depots have heterogeneous cell characterizations and differentiation potential when cultured in 3-dimensional (3D) niches. Design We compared the chondrogenicity of ASCs isolated from infrapatellar fat pads (IPFPs) and subcutaneous fat pads (SCFPs) in 3D gelatin-based biomimetic matrix. Results The IPFP-ASC-differentiated chondrocytes had higher ACAN, COL2A1, COL10, SOX6, SOX9, ChM-1, and MIA-3 mRNA levels and lower COL1A1 and VEGF levels than the SCFP-ASCs in 3D matrix. The difference in mRNA profile may have contributed to activation of the Akt, p38, RhoA, and JNK signaling pathways in the IPFP-ASCs. The chondrocytes differentiated from IPFP-ASCs had pronounced glycosaminoglycan and collagen type II production and a high chondroitin-6-sulfate/chondroitin-4-sulfate ratio with less polymerization of β-actin filaments. In an ex vivo mice model, magnetic resonance imaging revealed a shorter T2 relaxation time, indicating that more abundant extracellular matrix was secreted in the IPFP-ASC–matrix group. Histological examinations revealed that the IPFP-ASC matrix had higher chondrogenic efficacy of new cartilaginous tissue generation as evident in collagen type II and S-100 staining. Conclusion. ASCs isolated from IPFPs may be better candidates for cartilage regeneration, highlighting the translational potential of cartilage tissue engineering using the IPFP-ASC matrix technique.

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Hyaluronan initiates chondrogenesis mainly via CD44 in human adipose-derived stem cells

Journal of Applied Physiology ◽

10.1152/japplphysiol.01132.2012 ◽

2013 ◽

Vol 114 (11) ◽

pp. 1610-1618 ◽

Cited By ~ 45

Author(s):

Shun-Cheng Wu ◽

Chung-Hwan Chen ◽

Je-Ken Chang ◽

Yin-Chih Fu ◽

Chih-Kuang Wang ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Collagen Type ◽

Cell Surface Receptor ◽

Marker Genes ◽

Adipose Derived Stem Cells ◽

Type Ii ◽

Collagen Type Ii ◽

Sulfated Glycosaminoglycan ◽

Chondrogenic Marker

Cell-matrix adhesion is one of the important interactions that regulates stem cell survival, self-renewal, and differentiation. Our previous report (Wu SC, Chang JK, Wang CK, Wang GJ, Ho ML. Biomaterials 31: 631–640, 2010) indicated that a microenvironment enriched with hyaluronan (HA) initiated and enhanced chondrogenesis in human adipose-derived stem cells (hADSCs). We further hypothesize that HA-induced chondrogenesis in hADSCs is mainly due to the interaction of HA and CD44 (HA-CD44), a cell surface receptor of HA. The HA-CD44 interaction was tested by examining the mRNA expression of hyaluronidase-1 (Hyal-1) and chondrogenic marker genes (SOX-9, collagen type II, and aggrecan) in hADSCs cultured on HA-coated wells. Cartilaginous matrix formation, sulfated glycosaminoglycan, and collagen productions by hADSCs affected by HA-CD44 interaction were tested in a three-dimensional fibrin hydrogel. About 99.9% of hADSCs possess CD44. The mRNA expressions of Hyal-1 and chondrogenic marker genes were upregulated by HA in hADSCs on HA-coated wells. Blocking HA-CD44 interaction by anti-CD44 antibody completely inhibited Hyal-1 expression and reduced chondrogenic marker gene expression, which indicates that HA-induced chondrogenesis in hADSCs mainly acts through HA-CD44 interaction. A 2-h preincubation and coculture of cells with HA in hydrogel (HA/fibrin hydrogel) not only assisted in hADSC survival, but also enhanced expression of Hyal-1 and chondrogenic marker genes. Higher levels of sulfated glycosaminoglycan and total collagen were also found in HA/fibrin hydrogel group. Immunocytochemistry showed more collagen type II, but less collagen type X, in HA/fibrin than in fibrin hydrogels. Our results indicate that signaling triggered by HA-CD44 interaction significantly contributes to HA-induced chondrogenesis and may be applied to adipose-derived stem cell-based cartilage regeneration.

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Collagen Type I and Type II Expression Evaluation on Cartilage Defect Regeneration Treated with Dwikora–Ferdiansyah–Lesmono–Purwati (DFLP) Scaffold Supplemented with Adipose–Derived Stem Cells (ASCs) or Secretome: An In-Vivo Study

Qanun Medika - Medical Journal Faculty of Medicine Muhammadiyah Surabaya ◽

10.30651/jqm.v4i2.4377 ◽

2020 ◽

Vol 4 (2) ◽

pp. 225

Author(s):

Adrianto Prasetyo Perbowo ◽

Dwikora Novembri Utomo ◽

Lukas Widhiyanto ◽

Primadenny Ariesa Airlangga ◽

Purwati Purwati

Keyword(s):

Stem Cells ◽

Collagen Type ◽

Collagen Type I ◽

Cartilage Defects ◽

Type I ◽

Adipose Derived Stem Cells ◽

Type Ii ◽

Collagen Type Ii ◽

Significant Difference

Abstract Cell-based therapies such as Scaffold, stem cells, and secretome, are one of the alternatives to enhance the regeneration of hyaline-like cartilage in cases of cartilage defects. This study is an in-vivo experiment using animal models, in which we apply a composite of DFLP (Dwikora-Ferdiansyah-Lesmono-Purwati) Scaffold and Adipose-Derived Stem Cells (ASCs) or Secretome to an injury model on the distal femoral trochlea of New Zealand White Rabbits. The animals were divided into four groups: (1) control (K); (2) Scaffold only (S); (3) Scaffold + ASCs (SA); (4) Scaffold + Secretome (SS). Animals were terminated in the 12th week, and an immunohistochemistry (IHC) evaluation for Collagen type I and II were done. Statistical analysis shows that collagen type I IHC between groups shows no significant difference (p = 0.546). Collagen type II IHC shows significant difference between groups (p = 0,016). The findings in this study showed that Scaffold + ASCs group and Scaffold + Secretome have better collagen type II expression compared to the control group. DFLP Scaffold composite with ASCs or Secretome shows potential for cartilage regeneration therapy by increasing type II collagen expression as in hyaline-like cartilage which may be used for regenerative therapy for cartilage defects. Keywords : DFLP Scaffold; Adipose-Derived Stem Cells (ASCs); Secretome; Collagen Type I; Collagen Type IICorrespondence : [email protected]

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Electromagnetic fields enhance chondrogenesis of human adipose-derived stem cells in a chondrogenic microenvironment in vitro

Journal of Applied Physiology ◽

10.1152/japplphysiol.01216.2012 ◽

2013 ◽

Vol 114 (5) ◽

pp. 647-655 ◽

Cited By ~ 33

Author(s):

Chung-Hwan Chen ◽

Yi-Shan Lin ◽

Yin-Chih Fu ◽

Chih-Kuang Wang ◽

Shun-Cheng Wu ◽

...

Keyword(s):

Stem Cells ◽

Electromagnetic Field ◽

Cell Viability ◽

Chondrogenic Differentiation ◽

Collagen Type ◽

Cartilage Tissue ◽

Pulse Electromagnetic Field ◽

Type I ◽

Adipose Derived Stem Cells ◽

Pemf Treatment

We tested the hypothesis that electromagnetic field (EMF) stimulation enhances chondrogenesis in human adipose-derived stem cells (ADSCs) in a chondrogenic microenvironment. A two-dimensional hyaluronan (HA)-coated well (2D-HA) and a three-dimensional pellet culture system (3D-pellet) were used as chondrogenic microenvironments. The ADSCs were cultured in 2D-HA or 3D-pellet, and then treated with clinical-use pulse electromagnetic field (PEMF) or the innovative single-pulse electromagnetic field (SPEMF) stimulation. The cytotoxicity, cell viability, and chondrogenic and osteogenic differentiations were analyzed after PEMF or SPEMF treatment. The modules of PEMF and SPEMF stimulations used in this study did not cause cytotoxicity or alter cell viability in ADSCs. Both PEMF and SPEMF enhanced the chondrogenic gene expression (SOX-9, collagen type II, and aggrecan) of ADSCs cultured in 2D-HA and 3D-pellet. The expressions of bone matrix genes (osteocalcin and collagen type I) of ADSCs were not changed after SPEMF treatment in 2D-HA and 3D-pellet; however, they were enhanced by PEMF treatment. Both PEMF and SPEMF increased the cartilaginous matrix (sulfated glycosaminoglycan) deposition of ADSCs. However, PEMF treatment also increased mineralization of ADSCs, but SPEMF treatment did not. Both PEMF and SPEMF enhanced chondrogenic differentiation of ADSCs cultured in a chondrogenic microenvironment. SPEMF treatment enhanced ADSC chondrogenesis, but not osteogenesis, when the cells were cultured in a chondrogenic microenvironment. However, PEMF enhanced both osteogenesis and chondrogenesis under the same conditions. Thus the combination of a chondrogenic microenvironment with SPEMF stimulation can promote chondrogenic differentiation of ADSCs and may be applicable to articular cartilage tissue engineering.

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Spidroin Striped Micropattern Promotes Chondrogenic Differentiation of Human Wharton’s Jelly Mesenchymal Stem Cells

10.21203/rs.3.rs-445399/v1 ◽

2021 ◽

Author(s):

Anggraini Barlian ◽

Dinda Hani’ah Arum Saputri ◽

Adriel Hernando ◽

Ekavianty Prajatelistia ◽

Hutomo Tanoto

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Mesenchymal Stem Cells ◽

Chondrogenic Differentiation ◽

Spider Silk ◽

Cartilage Tissue Engineering ◽

Wharton’S Jelly ◽

Cartilage Tissue ◽

Type Ii ◽

Wharton's Jelly

Abstract Cartilage tissue engineering, particularly micropattern, can influence the biophysical properties of mesenchymal stem cells (MSCs) leading to chondrogenesis. In this research, human Wharton’s jelly MSCs (hWJ-MSCs) were grown on a striped micropattern containing spider silk protein (spidroin) from Argiope appensa. This research aims to direct hWJ-MSCs chondrogenesis using micropattern made of spidroin bioink as opposed to fibronectin that often used as the gold standard. Cells were cultured on striped micropattern of 500 µm and 1000 µm width sizes without chondrogenic differentiation medium for 21 days. The immunocytochemistry result showed that spidroin contains RGD sequences and facilitates cell adhesion via integrin β1. Chondrogenesis was observed through the expression of glycosaminoglycan, type II collagen, and SOX9. The result on glycosaminoglycan content proved that 1000 µm was the optimal width to support chondrogenesis. Spidroin micropattern induced significantly higher expression of SOX9 mRNA on day-21 and SOX9 protein was located inside the nucleus starting from day-7. COL2A1 mRNA of spidroin micropattern groups was downregulated on day-21 and collagen type II protein was detected starting from day-14. These results showed that spidroin micropattern enhances chondrogenic markers while maintains long-term upregulation of SOX9, and therefore has the potential as a new method for cartilage tissue engineering.

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