Precision 3D printed meniscus scaffolds to facilitate hMSCs proliferation and chondrogenic differentiation for tissue regeneration

Abstract Background The poor regenerative capability and structural complexity make the reconstruction of meniscus particularly challenging in clinic. 3D printing of polymer scaffolds holds the promise of precisely constructing complex tissue architecture, however the resultant scaffolds usually lack of sufficient bioactivity to effectively generate new tissue. Results Herein, 3D printing-based strategy via the cryo-printing technology was employed to fabricate customized polyurethane (PU) porous scaffolds that mimic native meniscus. In order to enhance scaffold bioactivity for human mesenchymal stem cells (hMSCs) culture, scaffold surface modification through the physical absorption of collagen I and fibronectin (FN) were investigated by cell live/dead staining and cell viability assays. The results indicated that coating with fibronectin outperformed coating with collagen I in promoting multiple-aspect stem cell functions, and fibronectin favors long-term culture required for chondrogenesis on scaffolds. In situ chondrogenic differentiation of hMSCs resulted in a time-dependent upregulation of SOX9 and extracellular matrix (ECM) assessed by qRT-PCR analysis, and enhanced deposition of collagen II and aggrecan confirmed by immunostaining and western blot analysis. Gene expression data also revealed 3D porous scaffolds coupled with surface functionalization greatly facilitated chondrogenesis of hMSCs. In addition, the subcutaneous implantation of 3D porous PU scaffolds on SD rats did not induce local inflammation and integrated well with surrounding tissues, suggesting good in vivo biocompatibility. Conclusions Overall, this study presents an approach to fabricate biocompatible meniscus constructs that not only recapitulate the architecture and mechanical property of native meniscus, but also have desired bioactivity for hMSCs culture and cartilage regeneration. The generated 3D meniscus-mimicking scaffolds incorporated with hMSCs offer great promise in tissue engineering strategies for meniscus regeneration. Graphical Abstract

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Fabrication of hyaline-like cartilage constructs using mesenchymal stem cell sheets

Scientific Reports ◽

10.1038/s41598-020-77842-0 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Hallie Thorp ◽

Kyungsook Kim ◽

Makoto Kondo ◽

David W. Grainger ◽

Teruo Okano

Keyword(s):

Chondrogenic Differentiation ◽

Cell Sheet ◽

Cartilage Regeneration ◽

Cell Sheets ◽

Chondrogenic Potential ◽

Cell Sheet Technology ◽

Cell Functionality ◽

Articular Cartilage Regeneration

AbstractCell and tissue engineering approaches for articular cartilage regeneration increasingly focus on mesenchymal stem cells (MSCs) as allogeneic cell sources, based on availability and innate chondrogenic potential. Many MSCs exhibit chondrogenic potential as three-dimensional (3D) cultures (i.e. pellets and seeded biomaterial scaffolds) in vitro; however, these constructs present engraftment, biocompatibility, and cell functionality limitations in vivo. Cell sheet technology maintains cell functionality as scaffold-free constructs while enabling direct cell transplantation from in vitro culture to targeted sites in vivo. The present study aims to develop transplantable hyaline-like cartilage constructs by stimulating MSC chondrogenic differentiation as cell sheets. To achieve this goal, 3D MSC sheets are prepared, exploiting spontaneous post-detachment cell sheet contraction, and chondrogenically induced. Results support 3D MSC sheets’ chondrogenic differentiation to hyaline cartilage in vitro via post-contraction cytoskeletal reorganization and structural transformations. These 3D cell sheets’ initial thickness and cellular densities may also modulate MSC-derived chondrocyte hypertrophy in vitro. Furthermore, chondrogenically differentiated cell sheets adhere directly to cartilage surfaces via retention of adhesion molecules while maintaining the cell sheets’ characteristics. Together, these data support the utility of cell sheet technology for fabricating scaffold-free, hyaline-like cartilage constructs from MSCs for future transplantable articular cartilage regeneration therapies.

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Articular chondrocyte-derived extracellular vesicles promote cartilage differentiation of human umbilical cord mesenchymal stem cells by activation of autophagy

Journal of Nanobiotechnology ◽

10.1186/s12951-020-00708-0 ◽

2020 ◽

Vol 18 (1) ◽

Author(s):

Ke Ma ◽

Bo Zhu ◽

Zetao Wang ◽

Peian Cai ◽

Mingwei He ◽

...

Keyword(s):

Stem Cells ◽

Umbilical Cord ◽

Extracellular Vesicles ◽

Chondrogenic Differentiation ◽

Phenotypic Expression ◽

Rabbit Model ◽

Cartilage Regeneration ◽

Human Umbilical Cord ◽

Articular Chondrocyte

Abstract Background Umbilical cord mesenchymal stem cell (HUCMSC)-based therapies were previously utilised for cartilage regeneration because of the chondrogenic potential of MSCs. However, chondrogenic differentiation of HUCMSCs is limited by the administration of growth factors like TGF-β that may cause cartilage hypertrophy. It has been reported that extracellular vesicles (EVs) could modulate the phenotypic expression of stem cells. However, the role of human chondrogenic-derived EVs (C-EVs) in chondrogenic differentiation of HUCMSCs has not been reported. Results We successfully isolated C-EVs from human multi-finger cartilage and found that C-EVs efficiently promoted the proliferation and chondrogenic differentiation of HUCMSCs, evidenced by highly expressed aggrecan (ACAN), COL2A, and SOX-9. Moreover, the expression of the fibrotic marker COL1A and hypertrophic marker COL10 was significantly lower than that induced by TGF-β. In vivo, C-EVs induced HUCMSCs accelerated the repair of the rabbit model of knee cartilage defect. Furthermore, C-EVs led to an increase in autophagosomes during the process of chondrogenic differentiation, indicating that C-EVs promote cartilage regeneration through the activation of autophagy. Conclusions C-EVs play an essential role in fostering chondrogenic differentiation and proliferation of HUCMSCs, which may be beneficial for articular cartilage repair.

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Articular chondrocyte-derived exosomes promote cartilage differentiation of human umbilical cord mesenchymal stem cells by activation of autophagy

10.21203/rs.3.rs-23463/v2 ◽

2020 ◽

Author(s):

Ke Ma ◽

Bo Zhu ◽

Zetao Wang ◽

Peian Cai ◽

Mingwei He ◽

...

Keyword(s):

Stem Cells ◽

Umbilical Cord ◽

Cartilage Repair ◽

Chondrogenic Differentiation ◽

Phenotypic Expression ◽

Cartilage Regeneration ◽

Human Umbilical Cord ◽

Articular Chondrocyte ◽

Differentiation And Proliferation

Abstract Background Umbilical cord mesenchymal stem cell (HUCMSC)-based therapies were previously utilised for cartilage regeneration because of the chondrogenic potential of MSCs. However, chondrogenic differentiation of HUMSCs is limited by the administration of growth factors like TGF-β that may cause cartilage hypertrophy. It has been reported that exosomes could modulate the phenotypic expression of stem cells. However, the role of human chondrogenic-derived exosomes (C-EXOs) in chondrogenic differentiation of HUCMSCs has not been reported. Results In this study, we successfully isolated chondrocyte-derived exosomes (C-EXO) from human multi-finger cartilage and found that C-EXO efficiently promoted the proliferation and chondrogenic differentiation of HUCMSCs, evidenced by highly expressed aggrecan (ACAN), COL2A and SOX-9. Also, the expression of the fibrotic marker, COL1A and hypertrophic marker, COL10, was significantly lower than that induced by TGF-β. In vivo, stimulation of C-EXO accelerated HUCMSCs-mediated cartilage repair in rabbit models. Furthermore, C-EXO led to increasing autophagosomes during the process of chondrogenic differentiation, indicating that C-EXO promoted cartilage regeneration might be through the activation of autophagy. Conclusions C-EXOs play an essential role in fostering chondrogenic differentiation and proliferation of HUCMSCs, which may be beneficial for articular cartilage repair.

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Application of robotic-assisted in situ 3D printing in cartilage regeneration with HAMA hydrogel: An in vivo study

Journal of Advanced Research ◽

10.1016/j.jare.2020.01.010 ◽

2020 ◽

Vol 23 ◽

pp. 123-132 ◽

Cited By ~ 9

Author(s):

Kaiwei Ma ◽

Tianzheng Zhao ◽

Longfei Yang ◽

Peng Wang ◽

Jing Jin ◽

...

Keyword(s):

3D Printing ◽

Cartilage Regeneration ◽

In Vivo Study ◽

Robotic Assisted

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The Role of Physical Stimuli on Calcium Channels in Chondrogenic Differentiation of Mesenchymal Stem Cells

International Journal of Molecular Sciences ◽

10.3390/ijms19102998 ◽

2018 ◽

Vol 19 (10) ◽

pp. 2998 ◽

Cited By ~ 11

Author(s):

Ilona Uzieliene ◽

Paulius Bernotas ◽

Ali Mobasheri ◽

Eiva Bernotiene

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Calcium Channels ◽

Chondrogenic Differentiation ◽

Human Mesenchymal Stem Cells ◽

Cartilage Regeneration ◽

Cellular Processes ◽

Stem Cell Source ◽

Physical Stimuli ◽

The Impact

Human mesenchymal stem cells (hMSC) are becoming increasingly popular in tissue engineering. They are the most frequently used stem cell source for clinical applications due to their high potential to differentiate into several lineages. Cartilage is known for its low capacity for self-maintenance and currently there are no efficient methods to improve cartilage repair. Chondrogenic differentiation of hMSC isolated from different tissues is widely employed due to a high clinical demand for the improvement of cartilage regeneration. Calcium channels that are regulated by physical stimuli seem to play a pivotal role in chondrogenic differentiation of MSCs. These channels increase intracellular calcium concentration, which leads to the initiation of the relevant cellular processes that are required for differentiation. This review will focus on the impact of different physical stimuli, including electrical, electromagnetic/magnetic and mechanical on various calcium channels and calcium signaling mechanisms during chondrogenic differentiation of hMSC.

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The Overview of Porous, Bioactive Scaffolds as Instructive Biomaterials for Tissue Regeneration and Their Clinical Translation

Pharmaceutics ◽

10.3390/pharmaceutics12070602 ◽

2020 ◽

Vol 12 (7) ◽

pp. 602

Author(s):

Gaëtan Lutzweiler ◽

Albana Ndreu Halili ◽

Nihal Engin Vrana

Keyword(s):

Tissue Regeneration ◽

Degrees Of Freedom ◽

Cell Activity ◽

Tissue Growth ◽

Porous Scaffolds ◽

Bioactive Scaffolds ◽

Cell Functions ◽

Tissue Repair And Regeneration

Porous scaffolds have been employed for decades in the biomedical field where researchers have been seeking to produce an environment which could approach one of the extracellular matrixes supporting cells in natural tissues. Such three-dimensional systems offer many degrees of freedom to modulate cell activity, ranging from the chemistry of the structure and the architectural properties such as the porosity, the pore, and interconnection size. All these features can be exploited synergistically to tailor the cell–material interactions, and further, the tissue growth within the voids of the scaffold. Herein, an overview of the materials employed to generate porous scaffolds as well as the various techniques that are used to process them is supplied. Furthermore, scaffold parameters which modulate cell behavior are identified under distinct aspects: the architecture of inert scaffolds (i.e., pore and interconnection size, porosity, mechanical properties, etc.) alone on cell functions followed by comparison with bioactive scaffolds to grasp the most relevant features driving tissue regeneration. Finally, in vivo outcomes are highlighted comparing the accordance between in vitro and in vivo results in order to tackle the future translational challenges in tissue repair and regeneration.

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Gelatin reduced Graphene Oxide Nanosheets as Kartogenin Nanocarrier Induces Rat ADSCs Chondrogenic Differentiation Combining with Autophagy Modification

Materials ◽

10.3390/ma14051053 ◽

2021 ◽

Vol 14 (5) ◽

pp. 1053

Author(s):

Delong Jiao ◽

Jing Wang ◽

Wenting Yu ◽

Ning Zhang ◽

Ke Zhang ◽

...

Keyword(s):

Stem Cells ◽

Graphene Oxide ◽

Reduced Graphene Oxide ◽

Chondrogenic Differentiation ◽

Drug Loading ◽

Pcr Analysis ◽

Promoting Effect ◽

Reduced Graphene

Biocompatible reduced graphene oxide (rGO) could deliver drugs for synergistically stimulating stem cells directed differentiation with influences on specific cellular activities. Here, we prepared a biodegradable gelatin reduced graphene oxide (rGO@Ge) to evaluate its functions in promoting rat adipose derived mesenchymal stem cells (ADSCs) chondrogenic differentiation through delivering kartogenin (KGN) into the stem cell efficiently. The optimum KGN concentration (approximately 1 μM) that promoted the proliferation and chondrogenic differentiation of ADSCs was clarified by a series of experiments, including immunofluorescent (IF) staining (Sox-9, Col II), alcian blue (Ab) staining, toluidine blue (Tb) staining and real-time quantitative PCR analysis of the chondrogenic markers. Meanwhile, the biocompatibility of rGO@Ge was evaluated to clearly define the nonhazardous concentration range, and the drug loading and releasing properties of rGO@Ge were tested with KGN for its further application in inducing ADSCs chondrogenic differentiation. Furthermore, the mechanism of rGO@Ge entering ADSCs was investigated by the different inhibitors that are involved in the endocytosis of the nanocarrier, and the degradation of the rGO@Ge in ADSCs was observed by transmission electron microscopy (TEM). The synergistic promoting effect of rGO@Ge nanocarrier on ADSCs chondrogenesis with KGN was also studied by the IF, Ab, Tb stainings and PCR analysis of the chondrogenic markers. Finally, the intracellular Reactive Oxygen Species (ROS) and autophagy induced by KGN/rGO@Ge complex composites were tested in details for clarification on the correlation between the autophagy and chondrogenesis in ADSCs induced by rGO@Ge. All the results show that rGO@Ge as a biocompatible nanocarrier can deliver KGN into ADSCs for exerting a pro-chondrogenic effect and assist the drug to promote ADSCs chondrogenesis synergistically through modification of the autophagy in vitro, which promised its further application in repairing cartilage defect in vivo.

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The dependence of in vivo stable ectopic chondrogenesis by human mesenchymal stem cells on chondrogenic differentiation in vitro

Biomaterials ◽

10.1016/j.biomaterials.2008.01.021 ◽

2008 ◽

Vol 29 (14) ◽

pp. 2183-2192 ◽

Cited By ~ 75

Author(s):

Kai Liu ◽

Guang Dong Zhou ◽

Wei Liu ◽

Wen Jie Zhang ◽

Lei Cui ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Chondrogenic Differentiation ◽

Human Mesenchymal Stem Cells

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Articular chondrocyte derived exosomes promote cartilage differentiation of human umbilical cord mesenchymal stem cells by activation of autophagy

10.21203/rs.3.rs-23463/v1 ◽

2020 ◽

Author(s):

Ke Ma ◽

Bo Zhu ◽

Zetao Wang ◽

Peian Cai ◽

Mingwei He ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Umbilical Cord ◽

Cartilage Repair ◽

Chondrogenic Differentiation ◽

Phenotypic Expression ◽

Cartilage Regeneration ◽

Human Umbilical Cord ◽

Articular Chondrocyte

Abstract Background Umbilical cord mesenchymal stem cells (HUCMSCs)-based therapies were previously predicated in cartilage regeneration due to the chondrogenic potential of MSCs. However, chondrogenic differentiation of HUMSCs is limited by administration of growth factors like TGF-β that may cause cartilage hypertrophy. It has been reported the exosomes could modulate phenotypic expression of stem cells. However, the role of human chondrogenic derived exosomes (C-EXO) in chondrogenic differentiation of HUCMSCs has not been reported. Results In this study, we successfully isolated chondrocyte-derived exosomes (C-EXO) from human multi-finger cartilage and found that C-EXO efficiently promoted the proliferation and chondrogenic differentiation of HUCMSCs, evidenced by highly expressed aggrecan (ACAN), COL2A and SOX-9. Also, the expression of the fibrotic marker, COL1A and hypertrophic marker, COL10, was significantly lower than that induced by TGF-β. In vivo, stimulation of C-EXO accelerated HUCMSCs-mediated cartilage repair in rabbit models. Furthermore, C-EXO led to increasing autophagosomes during the process of chondrogenic differentiation, indicating that C-EXO promoted cartilage regeneration might be through the activation of autophagy. Conclusions This study suggests that C-EXO has an essential role in fostering chondrogenic differentiation and proliferation of HUCMSCs, which may be a stable supply for articular cartilage repair.

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A pilot study on the use of 3D printers in veterinary medicine

Brazilian Journal of Veterinary Pathology ◽

10.24070/bjvp.1983-0246.v14i3p159-164 ◽

2021 ◽

Vol 14 (3) ◽

pp. 159-164

Author(s):

Leonardo Leonardi ◽

◽

Roberto Marsili ◽

Enrico Bellezza ◽

Giovanni Angeli ◽

...

Keyword(s):

Additive Manufacturing ◽

3D Printing ◽

Bone Tissue ◽

Three Dimensional ◽

The Body ◽

Porous Scaffolds ◽

Layer By Layer ◽

Three Dimensional Objects ◽

Cellular Attachment

Additive manufacturing (AM) is the process of joining materials to create layer-by-layer three-dimensional objects using a 3D printer from a digital model. The great advantage of Additive Manufacturing is to allow a freer design than traditional processes. The development of additive manufacturing processes has permitted to optimize the production of the customized product through the modeling of the geometry and the knowledge of the morphometric parameters of the body structures. 3D printing has revolutionized the field of Regenerative Medicine because, starting from computerized tomography (CT) images and using traditional materials such as plastic and metals, it can provide customized prostheses for each patient, which adapt perfectly to the needs of the subject and act as structures support. 3D printing allows you to print three-dimensional porous scaffolds with a precise shape and chemical composition suitable for medical and veterinary use. Some of these scaffolds are biodegradable and appear to be ideal for bone tissue engineering. In fact, they are able to simulate extracellular matrix properties that allow mechanical support, favoring mechanical interactions and providing a model for cellular attachment and in vivo stimulation of bone tissue formation.

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