Improving the immunosuppressive potential of articular chondroprogenitors in a three-dimensional culture setting

Abstract Cartilage repair in osteoarthritic patients remains a challenge. Identifying resident or donor stem/progenitor cell populations is crucial for augmenting the low intrinsic repair potential of hyaline cartilage. Furthermore, mediating the interaction between these cells and the local immunogenic environment is thought to be critical for long term repair and regeneration. In this study we propose articular cartilage progenitor/stem cells (CPSC) as a valid alternative to bone marrow-derived mesenchymal stem cells (BMMSC) for cartilage repair strategies after trauma. Similar to BMMSC, CPSC isolated from osteoarthritic patients express stem cell markers and have chondrogenic, osteogenic, and adipogenic differentiation ability. In an in vitro 2D setting, CPSC show higher expression of SPP1 and LEP, markers of osteogenic and adipogenic differentiation, respectively. CPSC also display a higher commitment toward chondrogenesis as demonstrated by a higher expression of ACAN. BMMSC and CPSC were cultured in vitro using a previously established collagen-chondroitin sulfate 3D scaffold. The scaffold mimics the cartilage niche, allowing both cell populations to maintain their stem cell features and improve their immunosuppressive potential, demonstrated by the inhibition of activated PBMC proliferation in a co-culture setting. As a result, this study suggests articular cartilage derived-CPSC can be used as a novel tool for cellular and acellular regenerative medicine approaches for osteoarthritis (OA). In addition, the benefit of utilizing a biomimetic acellular scaffold as an advanced 3D culture system to more accurately mimic the physiological environment is demonstrated.

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Xenogenic Implantation of Equine Synovial Fluid-Derived Mesenchymal Stem Cells Leads to Articular Cartilage Regeneration

Stem Cells International ◽

10.1155/2018/1073705 ◽

2018 ◽

Vol 2018 ◽

pp. 1-9 ◽

Cited By ~ 10

Author(s):

Mohammed Zayed ◽

Steven Newby ◽

Nabil Misk ◽

Robert Donnell ◽

Madhu Dhar

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Articular Cartilage ◽

Synovial Fluid ◽

Cartilage Repair ◽

Cartilage Regeneration ◽

Large Animal

Horses are widely used as large animal preclinical models for cartilage repair studies, and hence, there is an interest in using equine synovial fluid-derived mesenchymal stem cells (SFMSCs) in research and clinical applications. Since, we have previously reported that similar to bone marrow-derived MSCs (BMMSCs), SFMSCs may also exhibit donor-to-donor variations in their stem cell properties; the current study was carried out as a proof-of-concept study, to compare the in vivo potential of equine BMMSCs and SFMSCs in articular cartilage repair. MSCs from these two sources were isolated from the same equine donor. In vitro analyses confirmed a significant increase in COMP expression in SFMSCs at day 14. The cells were then encapsulated in neutral agarose scaffold constructs and were implanted into two mm diameter full-thickness articular cartilage defect in trochlear grooves of the rat femur. MSCs were fluorescently labeled, and one week after treatment, the knee joints were evaluated for the presence of MSCs to the injured site and at 12 weeks were evaluated macroscopically, histologically, and then by immunofluorescence for healing of the defect. The macroscopic and histological evaluations showed better healing of the articular cartilage in the MSCs’ treated knee than in the control. Interestingly, SFMSC-treated knees showed a significantly higher Col II expression, suggesting the presence of hyaline cartilage in the healed defect. Data suggests that equine SFMSCs may be a viable option for treating osteochondral defects; however, their stem cell properties require prior testing before application.

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Stem Cell Therapy for Articular Cartilage Repair: Review of the Entity of Cell Populations Used and the Result of the Clinical Application of Each Entity

The American Journal of Sports Medicine ◽

10.1177/0363546517729152 ◽

2017 ◽

Vol 46 (10) ◽

pp. 2540-2552 ◽

Cited By ~ 30

Author(s):

Yong-Beom Park ◽

Chul-Won Ha ◽

Ji Heon Rhim ◽

Han-Jun Lee

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Adipose Tissue ◽

Stem Cell ◽

Articular Cartilage ◽

Cell Therapy ◽

Cartilage Repair ◽

Stem Cell Therapy ◽

Peripheral Blood ◽

Cell Populations

Background: Following successful preclinical studies, stem cell therapy is emerging as a candidate for the treatment of articular cartilage lesions. Because stem cell therapy for cartilage repair in humans is at an early phase, confusion and errors are found in the literature regarding use of the term stem cell therapy in this field. Purpose: To provide an overview of the outcomes of cartilage repair, elucidating the various cell populations used, and thus reduce confusion with regard to using the term stem cell therapy. Study Design: Systematic review. Methods: The authors systematically reviewed any studies on clinical application of mesenchymal stem cells (MSCs) in human subjects. A comprehensive search was performed in MEDLINE, EMBASE, the Cochrane Library, CINAHL, Web of Science, and Scopus for human studies that evaluated articular cartilage repair with cell populations containing MSCs. These studies were classified as using bone marrow–derived MSCs, adipose tissue–derived MSCs, peripheral blood–derived MSCs, synovium-derived MSCs, and umbilical cord blood–derived MSCs according to the entity of cell population used. Results: Forty-six clinical studies were identified to focus on cartilage repair with MSCs: 20 studies with bone marrow–derived MSCs, 21 studies with adipose tissue–derived MSCs, 3 studies with peripheral blood–derived MSCs, 1 study with synovium-derived MSCs, and 1 study with umbilical cord blood–derived MSCs. All clinical studies reported that cartilage treated with MSCs showed favorable clinical outcomes in terms of clinical scores or cartilage repair evaluated by MRI. However, most studies were limited to case reports and case series. Among these 46 clinical studies, 18 studies erroneously referred to adipose tissue–derived stromal vascular fractions as “adipose-derived MSCs,” 2 studies referred to peripheral blood–derived progenitor cells as “peripheral blood–derived MSCs,” and 1 study referred to bone marrow aspirate concentrate as “bone marrow–derived MSCs.” Conclusion: Limited evidence is available regarding clinical benefit of stem cell therapy for articular cartilage repair. Because the literature contains substantial errors in describing the therapeutic cells used, researchers need to be alert and observant of proper terms, especially regarding whether the cells used were stem cells or cell populations containing a small portion of stem cells, to prevent confusion in understanding the results of a given stem cell–based therapy.

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The Middle Part of the Plucked Hair Follicle Outer Root Sheath Is Identified as an Area Rich in Lineage-Specific Stem Cell Markers

Biomolecules ◽

10.3390/biom11020154 ◽

2021 ◽

Vol 11 (2) ◽

pp. 154

Author(s):

Hanluo Li ◽

Federica Francesca Masieri ◽

Marie Schneider ◽

Alexander Bartella ◽

Sebastian Gaus ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hair Follicle ◽

Cell Populations ◽

Stem Cell Markers ◽

Outer Root Sheath ◽

Root Sheath ◽

Gene And Protein Expression ◽

Stem Cell Populations

Hair follicle outer root sheath (ORS) is a putative source of stem cells with therapeutic capacity. ORS contains several multipotent stem cell populations, primarily in the distal compartment of the bulge region. However, the bulge is routinely obtained using invasive isolation methods, which require human scalp tissue ex vivo. Non-invasive sampling has been standardized by means of the plucking procedure, enabling to reproducibly obtain the mid-ORS part. The mid-ORS shows potential for giving rise to multiple stem cell populations in vitro. To demonstrate the phenotypic features of distal, middle, and proximal ORS parts, gene and protein expression profiles were studied in physically separated portions. The mid-part of the ORS showed a comparable or higher NGFR, nestin/NES, CD34, CD73, CD44, CD133, CK5, PAX3, MITF, and PMEL expression on both protein and gene levels, when compared to the distal ORS part. Distinct subpopulations of cells exhibiting small and round morphology were characterized with flow cytometry as simultaneously expressing CD73/CD271, CD49f/CD105, nestin, and not CK10. Potentially, these distinct subpopulations can give rise to cultured neuroectodermal and mesenchymal stem cell populations in vitro. In conclusion, the mid part of the ORS holds the potential for yielding multiple stem cells, in particular mesenchymal stem cells.

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Isolation and Characterization of Human Epidermal Stem Cells

Clinical Science ◽

10.1042/cs0910141 ◽

1996 ◽

Vol 91 (2) ◽

pp. 141-146 ◽

Cited By ~ 24

Author(s):

P. H. Jones

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Matrix Proteins ◽

Stem Cell Markers ◽

Epidermal Stem Cells ◽

Cell Behaviour ◽

Isolation And Characterization ◽

Integrin Family

1. The keratinocytes in human epidermis are constantly turned over and replaced by a population of stem cells located in the basal epidermal layer. Until recently there were no markers allowing the isolation of viable epidermal stem cells. However, it has now been shown that epidermal stem cells can be isolated both in vitro and direct from the epidermis as they express high levels of functional β1 integrin family receptors for extracellular matrix proteins. 2. The evidence for integrins as stem cell markers and the insights that have been gained into stem cell behaviour are reviewed.

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5-Azacytidine-Induced Cardiomyocyte Differentiation of Very Small Embryonic-Like Stem Cells

Stem Cells International ◽

10.1155/2020/5162350 ◽

2020 ◽

Vol 2020 ◽

pp. 1-8

Author(s):

XiaoLin Sun ◽

HongXiao Li ◽

Ye Zhu ◽

Pei Xu ◽

QiSheng Zuo ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Ethical Issues ◽

Troponin T ◽

Embryonic Stem ◽

Stem Cell Population ◽

Pacemaker Activity ◽

Stem Cell Markers ◽

Seed Cells

The use of stem cells in generating cell-based pacemaker therapies for bradyarrhythmia is currently being considered. Due to the propensity of stem cells to form tumors, as well as ethical issues surrounding their use, the seed cells used in cardiac biological pacemakers have limitations. Very small embryonic-like stem cells (VSELs) are a unique and rare adult stem cell population, which have the same structural, genetic, biochemical, and functional characteristics as embryonic stem cells without the ethical controversy. In this study, we investigated the ability of rat bone marrow- (BM-) derived VSELs to differentiate in vitro into cardiomyocytes by 5-Azacytidine (5-AzaC) treatment. The morphology of VSELs treated with 10 μM 5-AzaC increased in volume and gradually changed to cardiomyocyte-like morphology without massive cell death. Additionally, mRNA expression of the cardiomyocyte markers cardiac troponin-T (cTnT) and α-sarcomeric actin (α-actin) was significantly upregulated after 5-AzaC treatment. Conversely, stem cell markers such as Nanog, Oct-4, and Sox2 were continuously downregulated posttreatment. On day 14 post-5-AzaC treatment, the positive expression rates of cTnT and α-actin were 18.41±1.51% and 19.43±0.51%, respectively. Taken together, our results showed that rat BM-VSELs have the ability to differentiate into cardiomyocytes in vitro. These findings suggest that VSELs would be useful as seed cells in exploring the mechanism of biological pacemaker activity.

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Stem Cells for Cartilage Repair: Preclinical Studies and Insights in Translational Animal Models and Outcome Measures

Stem Cells International ◽

10.1155/2018/9079538 ◽

2018 ◽

Vol 2018 ◽

pp. 1-22 ◽

Cited By ~ 28

Author(s):

Melissa Lo Monaco ◽

Greet Merckx ◽

Jessica Ratajczak ◽

Pascal Gervois ◽

Petra Hilkens ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Animal Models ◽

Outcome Measures ◽

Cartilage Repair ◽

Cartilage Tissue ◽

Preclinical Studies ◽

Paracrine Factors ◽

Advantages And Disadvantages

Due to the restricted intrinsic capacity of resident chondrocytes to regenerate the lost cartilage postinjury, stem cell-based therapies have been proposed as a novel therapeutic approach for cartilage repair. Moreover, stem cell-based therapies using mesenchymal stem cells (MSCs) or induced pluripotent stem cells (iPSCs) have been used successfully in preclinical and clinical settings. Despite these promising reports, the exact mechanisms underlying stem cell-mediated cartilage repair remain uncertain. Stem cells can contribute to cartilage repair via chondrogenic differentiation, via immunomodulation, or by the production of paracrine factors and extracellular vesicles. But before novel cell-based therapies for cartilage repair can be introduced into the clinic, rigorous testing in preclinical animal models is required. Preclinical models used in regenerative cartilage studies include murine, lapine, caprine, ovine, porcine, canine, and equine models, each associated with its specific advantages and limitations. This review presents a summary of recentin vitrodata and fromin vivopreclinical studies justifying the use of MSCs and iPSCs in cartilage tissue engineering. Moreover, the advantages and disadvantages of utilizing small and large animals will be discussed, while also describing suitable outcome measures for evaluating cartilage repair.

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An Efficient In Vitro Culture System To Amplify Spermatogonia Stem Cell Markers

Research in Molecular Medicine ◽

10.32598/rmm.8.3.2 ◽

2020 ◽

Vol 8 (3) ◽

pp. 117-124

Author(s):

Zeinab Narimanpour ◽

◽

Maryam Nazm Bojnordi ◽

Hatef Ghasemi ◽

◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Real Time ◽

Cultured Cells ◽

Expression Level ◽

Stem Cell Markers ◽

Testicular Cells ◽

Spermatogonia Stem Cell ◽

Alteration Pattern

Introduction: Proliferation of spermatogonial stem cells (SSCs) can be a treatment for infertile men. Here, we design an efficient method based on culturing in the presence of Sertoli cells to improve the expression level of some specific spermatogonia stem cell genes during two weeks post culture. Materials and Methods: Cells were derived from neonatal (2-6 days old) mice testes and were cultured in DMEM medium with FBS. The colonization of cultured SSCs in days 4, 7, and 14 of culture was counted via phase-contrast microscope and Image J software. Methyl thiazolyl tetrazolium (MTT) test was performed to evaluate the viability of cultured SSCs in days 3, 7, and 14 of culture. The expression level and the alteration pattern of specific spermatogonial markers, i.e., Stra8, DAZL, and Piwill2 was examined via real-time polymerase chain reaction (PCR) during two weeks post culture. Results: The number and the diameters of colonies showed a significant increase in cultured cells. MTT results proved the higher viability of testicular cells during the culture period. The results of ALP staining detected a positive reaction in spermatogonia colonies. Real-time PCR data showed that culturing SSCs in the presence of interstitial cells of the testis, amplified the level and alteration pattern of specific spermatogonia stem cells genes beneficial in the enrichment of SSCs propagation. Conclusion: Providing a similar culture environment to testicular niche increases viability, forms SSCs colonies, and regulates the level and alteration pattern of spermatogonia stem cell genes.

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Emilin-2 is a component of bone marrow extracellular matrix regulating mesenchymal stem cell differentiation and hematopoietic progenitors

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02674-2 ◽

2022 ◽

Vol 13 (1) ◽

Author(s):

Francesco Da Ros ◽

Luca Persano ◽

Dario Bizzotto ◽

Mariagrazia Michieli ◽

Paola Braghetta ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Extracellular Matrix ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Adipogenic Differentiation ◽

Stem Cell Differentiation ◽

Dependent Manner ◽

Hematopoietic Stem

Abstract Background Dissection of mechanisms involved in the regulation of bone marrow microenvironment through cell–cell and cell–matrix contacts is essential for the detailed understanding of processes underlying bone marrow activities both under physiological conditions and in hematologic malignancies. Here we describe Emilin-2 as an abundant extracellular matrix component of bone marrow stroma. Methods Immunodetection of Emilin-2 was performed in bone marrow sections of mice from 30 days to 6 months of age. Emilin-2 expression was monitored in vitro in primary and mesenchymal stem cell lines under undifferentiated and adipogenic conditions. Hematopoietic stem cells and progenitors in bone marrow of 3- to 10-month-old wild-type and Emilin-2 null mice were analyzed by flow cytometry. Results Emilin-2 is deposited in bone marrow extracellular matrix in an age-dependent manner, forming a meshwork that extends from compact bone boundaries to the central trabecular regions. Emilin-2 is expressed and secreted by both primary and immortalized bone marrow mesenchymal stem cells, exerting an inhibitory action in adipogenic differentiation. In vivo Emilin-2 deficiency impairs the frequency of hematopoietic stem/progenitor cells in bone marrow during aging. Conclusion Our data provide new insights in the contribution of bone marrow extracellular matrix microenvironment in the regulation of stem cell niches and hematopoietic progenitor differentiation.

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Mesenchymal stem cells promote glioma neovascularization in vivo by fusing with cancer stem cells

BMC Cancer ◽

10.1186/s12885-019-6460-0 ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 5

Author(s):

Chao Sun ◽

Xingliang Dai ◽

Dongliang Zhao ◽

Haiyang Wang ◽

Xiaoci Rong ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Tumor Growth ◽

Glioma Cells ◽

Stem Cell Markers ◽

Xenograft Tumor ◽

Angiogenic Effect

Abstract Background and objective Tumor angiogenesis is vital for tumor growth. Recent evidence indicated that bone marrow-derived mesenchymal stem cells (BMSCs) can migrate to tumor sites and exert critical effects on tumor growth through direct and/or indirect interactions with tumor cells. However, the effect of BMSCs on tumor neovascularization has not been fully elucidated. This study aimed to investigate whether fusion cells from glioma stem cells and BMSCs participated in angiogenesis. Methods SU3-RFP cells were injected into the right caudate nucleus of NC-C57Bl/6 J-GFP nude mice, and the RFP+/GFP+ cells were isolated and named fusion cells. The angiogenic effects of SU3-RFP, BMSCs and fusion cells were compared in vivo and in vitro. Results Fusion cells showed elevated levels of CD31, CD34 and VE-Cadherin (markers of VEC) as compared to SU3-RFP and BMSCs. The MVD-CD31 in RFP+/GFP+ cell xenograft tumor was significantly greater as compared to that in SU3-RFP xenograft tumor. In addition, the expression of CD133 and stem cell markers Nanog, Oct4 and Sox2 were increased in fusion cells as compared to the parental cells. Fusion cells exhibited enhanced angiogenic effect as compared to parental glioma cells in vivo and in vitro, which may be related to their stem cell properties. Conclusion Fusion cells exhibited enhanced angiogenic effect as compared to parental glioma cells in vivo and in vitro, which may be related to their stem cell properties. Hence, cell fusion may contribute to glioma angiogenesis.

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022. Isolation of stem cells from embryos and adult bovine tissues

Reproduction Fertility and Development ◽

10.1071/srb05abs022 ◽

2005 ◽

Vol 17 (9) ◽

pp. 67

Author(s):

P. J. Verma ◽

K. Upton ◽

H. Mc Connell ◽

I. Vassiliev

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Es Cells ◽

Cell Types ◽

Invasive Technique ◽

Stem Cell Markers ◽

Rt Pcr ◽

Differentiation Potential ◽

Cell Therapies

The isolation of stem cells has become an area of increasing interest due to their potential uses in animal reproduction, somatic cell nuclear transfer and cell therapies. The most attractive options are the isolation of stem cells from individual embryos or adult somatic tissues. In addition, for cell therapy, the use of autologous stem cells is considered to have an advantage over heterologous cell based therapies in that immune rejection issues would be circumvented. Here we report on our attempts to isolate stem cells from both these sources in a bovine model. Bovine ES-like (bES) cells were successfully isolated from embryos and maintained in vitro for up to six passages. These cells retained the morphology characteristic of bES cells: small cytoplasmic/nuclear ratio, nuclei with multiple nucleoli, and multiple lipid inclusions in cytoplasm. bES cell colonies grew as monolayers, as islands of ES cells surrounded by trophectoderm (TE) cells. Immunohistochemical detection of SSEA-1 and SSEA-4 demonstrated expression of these markers in bES cells but not in TE cells. Further, the expression of the pluripotent markers Oct-4, Rex-1 and SSEA-1 by RT-PCR was also detected in bES cells but not in TE cells. On spontaneous differentiation, these cells were able to form a variety of cell types including beating muscle with the cells displaying a propensity to differentiate in a manner reminiscent of human ES cells. (2) We also report the isolation of putative stem cells from adult bovine skin biopsies, which express the stem cell markers Oct-4 and SSEA-1 analysed by RT-PCR and are capable of forming 3-dimensional colonies. These cells are obtained from a skin biopsy, a relatively non-invasive technique that makes them useful as donors for therapeutic applications. In summary, we have identified populations of stem cells from embryonic and adult bovine tissues, which are readily isolated. Further characterization of the differentiation potential of these cells is needed to identify the suitability of this population for use in autologous stem cell therapies.

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