Transcriptomic and Proteomic Analyses Reveal the Potential Mode of Action of Chondrocyte Sheets in Hyaline Cartilage Regeneration

Toyoda, Eiji; Sato, Masato; Takahashi, Takumi; Maehara, Miki; Okada, Eri; Wasai, Shiho; Watanabe, Masahiko; Iijima, Hiroshi; Nonaka, Ken; Kawaguchi, Yuka

doi:10.3390/ijms21010149

Cited by 23 publications

(19 citation statements)

References 48 publications

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“…A heatmap showed that genes related to angiogenesis in the AM_EC group were upregulated compared with those in the AM_EM group, including MMP1 [ 22 ], ESM1 [ 23 ], ANGPT2 [ 24 ] and CYP1B1 [ 25 ] (Fig. 3 a).…”

Section: Resultsmentioning

confidence: 99%

Single-cell transcriptomic analysis of eutopic endometrium and ectopic lesions of adenomyosis

Sun

Liu³

et al. 2021

Cell Biosci

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Background Adenomyosis (AM) is a common benign chronic gynaecological disorder; however, the precise pathogenesis of adenomyosis is still poorly understood. Single-cell RNA sequencing (scRNA-seq) can uncover rare subpopulations, explore genetic and functional heterogeneity, and reveal the uniqueness of each cell. It provides us a new approach to reveal biological issues from a more detailed and microscopic perspective. Here, we utilize this revolutionary technology to identify the changes of gene expression patterns between ectopic lesions and the eutopic endometrium at the single-cell level and explore a potential novel pathogenesis of AM. Methods A control endometrium (sample with leiomyoma excluding endometrial disorders, n = 1), eutopic endometrium and ectopic lesion (from a patient with adenomyosis, n = 1) samples were analysed by scRNA-seq, and additional leiomyoma (n = 3) and adenomyosis (n = 3) samples were used to confirm colocalization and vasculogenic mimicry (VM) formation. Protein colocalization was visualized by immunofluorescence, and CD34-periodic acid-Schiff (PAS) double staining was used to assess the formation of VM. Results The scRNA-seq results suggest that cancer-, cell motility- and inflammation- (CMI) associated terms, cell proliferation and angiogenesis play important roles in the progression of AM. Moreover, the colocalization of EPCAM and PECAM1 increased significantly in the ectopic endometrium group (P < 0.05), cell subpopulation with high copy number variation (CNV) levels possessing tumour-like features existed in the ectopic lesion sample, and VNN1- and EPCAM-positive cell subcluster displayed active cell motility in endometrial epithelial cells. Furthermore, during the transformation of epithelial cells to endothelial cells, we observed the significant accumulation of VM formation (positively stained with PAS but not CD34, P < 0.05) in ectopic lesions. Conclusions In the present study, our results support the theory of adenomyosis derived from the invasion and migration of the endometrium. Moreover, cell subcluster with high CNV level and tumour-associated characteristics is identified. Furthermore, epithelial-endothelial transition (EET) and the formation of VM in tumours, the latter of which facilitates the blood supply and plays an important role in maintaining cell growth, were also confirmed to occur in AM. These results indicated that the inhibition of EET and VM formation may be a potential strategy for AM management.

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Section: Resultsmentioning

confidence: 99%

Single-cell transcriptomic analysis of eutopic endometrium and ectopic lesions of adenomyosis

Sun

Liu³

et al. 2021

Cell Biosci

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“…ESM-1 is a proteoglycan involved in angiogenesis and is expressed in the vascular endothelium and adipocytes. DKK-1 is a protein that has an inhibitory effect on the Wnt signaling pathway and is involved in bone metabolism and differentiation [ 29 ]. In this study, the secretion of these humoral factors did not differ significantly between PD sheets and PD sheets-mini and, therefore, we consider that both sheets would have an equivalent effect of cartilage regeneration.…”

Section: Discussionmentioning

confidence: 99%

Development of Injectable Polydactyly-Derived Chondrocyte Sheets

Wasai

Toyoda

Takahashi

et al. 2021

IJMS

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We are conducting a clinical study of the use of allogeneic polydactyly-derived chondrocyte sheets (PD sheets) for the repair of articular cartilage damage caused by osteoarthritis. However, the transplantation of PD sheets requires highly invasive surgery. To establish a less invasive treatment, we are currently developing injectable fragments of PD sheets (PD sheets-mini). Polydactyly-derived chondrocytes were seeded in RepCell™ or conventional temperature-responsive inserts and cultured. Cell counts and viability, histology, enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qPCR), and flow cytometry were used to characterize PD sheets-mini and PD sheets collected from each culture. To examine the effects of injection on cell viability, PD sheets-mini were tested in four experimental conditions: non-injection control, 18 gauge (G) needle, 23G needle, and syringe only. PD sheets-mini produced similar amounts of humoral factors as PD sheets. No histological differences were observed between PD sheets and PD sheets-mini. Except for COL2A1, expression of cartilage-related genes did not differ between the two types of PD sheet. No significant differences were observed between injection conditions. PD sheets-mini have characteristics that resemble PD sheets. The cell viability of PD sheets-mini was not significantly affected by needle gauge size. Intra-articular injection may be a feasible, less invasive method to transplant PD sheets-mini.

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“…Polydactyly-derived cell characteristics, including high transforming growth factor (TGF) beta secretion, in juvenile chondrocyte sheets, has been reported [ 25 ]. Hyaline cartilage regenerative capacity using human juvenile polydactyly cartilage-derived chondrocyte sheets was confirmed in an immunosuppressed rabbit osteochondral defect model [ 97 ] and nude rat focal chondral defect model (Kondo et al, submitted). The rat chondral defect model without intentional bone marrow induction particularly demonstrated that regenerated hyaline cartilage forms from human origins with no gaps found at host–donor tissue interfaces ( Figure 2 ).…”

Section: Cell Sheet Engineering —Key Enabling Features Of Thermo-respmentioning

confidence: 99%

Novel therapies using cell sheets engineered from allogeneic mesenchymal stem/stromal cells

Kondō

Kameishi

Grainger

et al. 2020

Emerging Topics in Life Sciences

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Mesenchymal stem/stromal cells (MSCs) have long been recognized to help regenerate tissues, by exploiting their intrinsic potentials for differentiation and secretion of therapeutic paracrine factors together with feasibility for cell banking. These unique MSC properties are attractive to provide effective new cell-based therapies for unmet medical needs. Currently, the infusion of suspended MSCs is accepted as a promising therapy to treat systemic inflammatory diseases. However, low cell engraftment/retention in target organs and off-target entrapment using conventional cell infusion must be improved to provide reliable localized disease treatments. Cell sheet technology offers an alternative: three-dimensional (3D) tissue-like structures can be harvested from culture using mild temperature reduction, and transplanted directly onto target tissue sites without suturing, yielding stable cell engraftment and prolonged cell retention in situ without off-target losses. Engineered MSC sheets directly address two major cell therapy strategies based on their therapeutic benefits: (1) tissue replacements based on mult-ilineage differentiation capacities, focusing on cartilage regeneration in this review, and (2) enhancement of tissue recovery via paracrine signaling, employing their various secreted cytokines to promote neovascularization. MSCs also have production benefits as a promising allogeneic cell source by exploiting their reliable proliferative capacity to facilitate expansion and sustainable cell banking for off-the-shelf therapies. This article reviews the advantages of both MSCs as allogeneic cell sources in contrast with autologous cell sources, and allogeneic MSC sheets engineered on thermo-responsive cell dishes as determined in basic studies and clinical achievements, indicating promise to provide robust new cell therapies to future patients.

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Transcriptomic and Proteomic Analyses Reveal the Potential Mode of Action of Chondrocyte Sheets in Hyaline Cartilage Regeneration

Cited by 23 publications

References 48 publications

Single-cell transcriptomic analysis of eutopic endometrium and ectopic lesions of adenomyosis

Single-cell transcriptomic analysis of eutopic endometrium and ectopic lesions of adenomyosis

Development of Injectable Polydactyly-Derived Chondrocyte Sheets

Novel therapies using cell sheets engineered from allogeneic mesenchymal stem/stromal cells

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