scholarly journals Interleukin-1β Induced Matrix Metalloproteinase Expression in Human Periodontal Ligament-Derived Mesenchymal Stromal Cells under In Vitro Simulated Static Orthodontic Forces

2021 ◽  
Vol 22 (3) ◽  
pp. 1027
Author(s):  
Christian Behm ◽  
Michael Nemec ◽  
Alice Blufstein ◽  
Maria Schubert ◽  
Xiaohui Rausch-Fan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Periodontal Ligament ◽  
Induced Expression ◽  
Gene And Protein Expression ◽  
Tensile Strains ◽  
Orthodontic Forces ◽  
Low Magnitude

The periodontal ligament (PDL) responds to applied orthodontic forces by extracellular matrix (ECM) remodeling, in which human periodontal ligament-derived mesenchymal stromal cells (hPDL-MSCs) are largely involved by producing matrix metalloproteinases (MMPs) and their local inhibitors (TIMPs). Apart from orthodontic forces, the synthesis of MMPs and TIMPs is influenced by the aseptic inflammation occurring during orthodontic treatment. Interleukin (IL)-1β is one of the most abundant inflammatory mediators in this process and crucially affects the expression of MMPs and TIMPs in the presence of cyclic low-magnitude orthodontic tensile forces. In this study we aimed to investigate, for the first time, how IL-1β induced expression of MMPs, TIMPs and how IL-1β in hPDL-MSCs was changed after applying in vitro low-magnitude orthodontic tensile strains in a static application mode. Hence, primary hPDL-MSCs were stimulated with IL-1β in combination with static tensile strains (STS) with 6% elongation. After 6- and 24 h, MMP-1, MMP-2, TIMP-1 and IL-1β expression levels were measured. STS alone had no influence on the basal expression of investigated target genes, whereas IL-1β caused increased expression of these genes. In combination, they increased the gene and protein expression of MMP-1 and the gene expression of MMP-2 after 24 h. After 6 h, STS reduced IL-1β-induced MMP-1 synthesis and MMP-2 gene expression. IL-1β-induced TIMP-1 gene expression was decreased by STS after 6- and 24-h. At both time points, the IL-1β-induced gene expression of IL-1β was increased. Additionally, this study showed that fetal bovine serum (FBS) caused an overall suppression of IL-1β-induced expression of MMP-1, MMP-2 and TIMP-1. Further, it caused lower or opposite effects of STS on IL-1β-induced expression. These observations suggest that low-magnitude orthodontic tensile strains may favor a more inflammatory and destructive response of hPDL-MSCs when using a static application form and that this response is highly influenced by the presence of FBS in vitro.

scholarly journals Preconditioning of mesenchymal stromal cells with low-intensity ultrasound: influence on chondrogenesis and directed SOX9 signaling pathways

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Neety Sahu ◽  
Gaurav Budhiraja ◽  
Anuradha Subramanian
Keyword(s):  
Gene Expression ◽  
Protein Expression ◽  
Signaling Pathways ◽  
Mesenchymal Stromal Cells ◽  
Nuclear Localization ◽  
Stromal Cells ◽  
Actin Reorganization ◽  
Low Intensity ◽  
Gene And Protein Expression ◽  
Low Intensity Ultrasound

Abstract Background Continuous low-intensity ultrasound (cLIUS) facilitates the chondrogenic differentiation of human mesenchymal stromal cells (MSCs) in the absence of exogenously added transforming growth factor-beta (TGFβ) by upregulating the expression of transcription factor SOX9, a master regulator of chondrogenesis. The present study evaluated the molecular events associated with the signaling pathways impacting SOX9 gene and protein expression under cLIUS. Methods Human bone marrow-derived MSCs were exposed to cLIUS stimulation at 14 kPa (5 MHz, 2.5 Vpp) for 5 min. The gene and protein expression of SOX9 was evaluated. The specificity of SOX9 upregulation under cLIUS was determined by treating the MSCs with small molecule inhibitors of select signaling molecules, followed by cLIUS treatment. Signaling events regulating SOX9 expression under cLIUS were analyzed by gene expression, immunofluorescence staining, and western blotting. Results cLIUS upregulated the gene expression of SOX9 and enhanced the nuclear localization of SOX9 protein when compared to non-cLIUS-stimulated control. cLIUS was noted to enhance the phosphorylation of the signaling molecule ERK1/2. Inhibition of MEK/ERK1/2 by PD98059 resulted in the effective abrogation of cLIUS-induced SOX9 expression, indicating that cLIUS-induced SOX9 upregulation was dependent on the phosphorylation of ERK1/2. Inhibition of integrin and TRPV4, the upstream cell-surface effectors of ERK1/2, did not inhibit the phosphorylation of ERK1/2 and therefore did not abrogate cLIUS-induced SOX9 expression, thereby suggesting the involvement of other mechanoreceptors. Consequently, the effect of cLIUS on the actin cytoskeleton, a mechanosensitive receptor regulating SOX9, was evaluated. Diffused and disrupted actin fibers observed in MSCs under cLIUS closely resembled actin disruption by treatment with cytoskeletal drug Y27632, which is known to increase the gene expression of SOX9. The upregulation of SOX9 under cLIUS was, therefore, related to cLIUS-induced actin reorganization. SOX9 upregulation induced by actin reorganization was also found to be dependent on the phosphorylation of ERK1/2. Conclusions Collectively, preconditioning of MSCs by cLIUS resulted in the nuclear localization of SOX9, phosphorylation of ERK1/2 and disruption of actin filaments, and the expression of SOX9 was dependent on the phosphorylation of ERK1/2 under cLIUS.

Lenalidomide Differentially Modifies the Genomic Profile and miRNA Expression of Mesenchymal Stromal Cells From Patients with 5q- Syndrome,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3810-3810
Author(s):  
Sandra Muntión ◽  
Carlos Santamaría ◽  
Beatriz Roson ◽  
Carlos Romo ◽  
Olga López-Villar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Profile ◽  
Mesenchymal Stromal Cells ◽  
Expression Profile ◽  
Stromal Cells ◽  
Microrna Expression ◽  
Advisory Committees ◽  
Healthy Donors ◽  
Lower Expression

Abstract Abstract 3810 Mesenchymal stromal cells (MSC) are a non-hematopoietic BM cell population considered to be not only the osteoblastic progenitors, but also a key component of the hematopoietic microenvironment. Raaijmakers et al (Nature, 2010) have recently shown that deletion of Dicer1 in MSC-derived osteoprogenitors as well as its target gene SBDS resulted in myelodysplasia (MDS) in a murine model. We have previously confirmed these results in human MSC from MDS patients (ASH 2010, # 397). In a previous paper (Leukemia, 2009) we showed that MSC from 5q- syndrome patients were different from MSC from other types of MDS and could be involved in their development. We have hypothesized that lenalidomide, the standard treatment of 5q- patients could act not only on hematopoietic progenitors but also on the BM microenvironment. For this purpose BM-MSC from healthy donors (HD) (n=7) and 5q- syndrome patients (n=5) were expanded in vitro and treated with 50 uM lenalidomide or its solvent (DMSO) as control. RNA was obtained from MSC and DICER1, DROSHA and SBDS relative gene expression was assessed by real-time PCR using TaqMan® assay as well as several microRNAs with known role in hematopoiesis and immune system regulation. In addition, MSC gene expression profile was studied. Labeled samples were hybridized to affymetrix of oligonucleotide HU 1.OST arrays in 5q- patients (n=4) and compared with MSCs from HD (n=3). For this purpose the ratio lenalidomide-treated sample and its paired DMSO control was calculated and markers with a fold change >1.5 were selected for hierarchical clustering analysis (HCA). MSCs from 5q-syndrome showed lower expression of DICER1 when compared with those from HD (.35 x10−3 vs.20 x10−3 p=0.03) but this expression was recovered when 5q-MSCs were treated with Lenalidomide (0.32 x10−3 p= 0.34). By contrast, no differences in DROSHA expression were observed. In addition, 5q-MSC showed SBDS lower expression than HD-MSC and in both groups the expression increased when they were treated with lenalidomide fig1). When microRNAs were analyzed, we observed a lower microRNA expression in lenalidomide-treated MSC from healthy donors when was compared to paired non-treated cells, especially for miRNA-155 (p=0.028), miRNA-222 (p=0.028),and miRNA-181a (p=0.075; Table 1). By contrast, lenalidomide-treated MSC from MDS showed a trend towards higher microRNA expression in comparison to paired non-treated MSC.Table 1.HD-MSC DMSO vs LENA5q-MSC DMSO vs LENAmiRNA 1460.50 vs 0.30p=0.2490.07 vs 0.10p=0.7miRNA 1500.004 vs 0.0065p=0.60.001 vs 0.006p=0.07miRNA 1550.90 vs 0.58p=0.0280.80 vs 0.96p=0.7miRNA 181a2.47 vs 1,83p=0.0751.66 vs 2.32p=0.07miRNA 22286.2 vs 68.0p=0.02843.2 vs 56.2p=0.07 When the gene expression profile was carried out based in 421 selected probes including 306 known genes, MSC-treated cells from 5q- were separated from HD MSC by HCA (Fig2). We can conclude that Lenalidomide not only acts on HPC from 5q- patients but also on microenvironment by modifying the expression of DICER-1 and SBDS as well as the expression of some microRNAs and genes. Disclosures: San Miguel: Celgene Corp.: Membership on an entity's Board of Directors or advisory committees. del Cañizo:Celgene Corp.: Spanishn Adviory committee.

scholarly journals Growth Factor-Mediated Tenogenic Induction of Multipotent Mesenchymal Stromal Cells Is Altered by the Microenvironment of Tendon Matrix

2018 ◽  
Vol 27 (10) ◽  
pp. 1434-1450 ◽  
Author(s):  
Susanne Pauline Roth ◽  
Susanna Schubert ◽  
Patrick Scheibe ◽  
Claudia Groß ◽  
Walter Brehm ◽  
...  
Keyword(s):  
Gene Expression ◽  
Growth Factor ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Multipotent Mesenchymal Stromal Cells ◽  
Histological Evaluation ◽  
Tendon Tissue ◽  
Tenascin C ◽  
Tenogenic Differentiation

Age-related degenerative changes in tendon tissue represent a common cause for acute tendon pathologies. Although the regenerative potential of multipotent mesenchymal stromal cells (MSC) was reported to restore functionality in injured tendon tissue, cellular mechanisms of action remain partly unclear. Potential tenogenic differentiation of applied MSC is affected by various intrinsic and extrinsic factors. The current study presents an in vitro model to evaluate the combined extrinsic effects of decellularized equine tendon matrix, transforming growth factor beta 3 (TGFβ3) and bone morphogenetic protein 12 (BMP12) on the tenogenic fate of equine adipose tissue-derived MSC. Monolayer MSC cultures supplemented with TGFβ3 and BMP12 as well as MSC cultured on tendon matrix scaffolds preloaded with the growth factors were incubated for 3 and 5 days. Histological evaluation and real time reverse transcription polymerase chain reaction (RT-PCR) revealed that growth factor-mediated tenogenic induction of MSC was modified by the conditions of the surrounding microenvironment. While the gene expression pattern in monolayer cultures supplemented with TGFβ3 or TGFβ3 and BMP12 revealed an upregulation for collagen 1A2, collagen 3A1, tenascin c, scleraxis and mohawk ( p < 0.05 ), the presence of tendon matrix led to an upregulation of decorin and osteopontin as well as to a downregulation of smad8 ( p < 0.05). Preloading of scaffolds with either TGFβ3, or with TGFβ3 and BMP12 promoted a tenocyte-like phenotype and improved cell alignment. Furthermore, gene expression in scaffold culture was modulated by TGFβ3 and/or BMP12, with downregulation of collagen 1A2, collagen 3A1, decorin, scleraxis, smad8 and osteopontin, whereas gene expression of tenascin c was increased. This study shows that growth factor-induced tenogenic differentiation of equine MSC is markedly altered by topographical constraints of decellularized tendon tissue in vitro. While TGFβ3 represents an effective mediator for tenogenic induction, the role of BMP12 in tenogenesis may be of modulatory character and needs further evaluation.

scholarly journals Human bone marrow-derived, pooled, allogeneic mesenchymal stromal cells manufactured from multiple donors at different times show comparable biological functions in vitro, and in vivo to repair limb ischemia

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Charan Thej ◽  
Sudha Balasubramanian ◽  
Mathiyazhagan Rengasamy ◽  
Ankita Walvekar ◽  
Priyanka Swamynathan ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mesenchymal Stromal Cells ◽  
Nude Mice ◽  
Stromal Cells ◽  
Limb Ischemia ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Cell Populations ◽  
Gene And Protein Expression

Abstract Background We have previously demonstrated that a pooled population of bone marrow-derived, allogeneic mesenchymal stromal cells (BMMSC), Stempeucel®-1, produced under good manufacturing practices (GMP) conditions, showed clinical efficacy and safety in patients suffering from critical limb ischemia (CLI) due to Buerger’s disease. While Stempeucel®-1 is currently used for CLI and other clinical indications, we wanted to ensure that the product’s continuity is addressed by developing and characterizing a second generation of pooled product (Stempeucel®-1A), manufactured identically from second BM aspirates of the same three donors after a 2-year interval. Methods The two versions of Stempeucel® were manufactured and subjected to gene and protein expression analysis. The nature of various growth factors/cytokines secreted and immunomodulatory activity of these two cell populations were compared directly by various in vitro assays. The preclinical efficacy of these two cell types was compared in an experimental model of hind limb ischemia (HLI) in BALB/c nude mice. The reversal of ischemia, blood flow, and muscle regeneration were determined by functional scoring, laser Doppler imaging, and immunohistochemical analyses. Results Qualitative and quantitative analyses of genes and proteins involved in promoting angiogenic activity and immune regulatory functions revealed high levels of correlation between Stempeucel®-1 and Stempeucel®-1A cell populations. Moreover, intramuscular (i.m) administration of these two cell products in the ischemic limbs of BALB/c nude mice showed significant repair (≥ 70%) of toe and foot necrosis, leading to improved ambulatory function and limb salvage. Furthermore, a biodistribution kinetics study showed that Stempeucel®-1 was mostly localized in the ischemic muscles of mice for a significantly longer time compared to normal muscles, thus playing an essential role in modulating and reversing HLI damage. Conclusions This study shows that with a reproducible manufacturing procedure, it is possible to generate large numbers of pooled mesenchymal stromal cells from human bone marrow samples to establish product equivalence. We conclude from these results that, for the first time, two pooled, allogeneic BMMSC products can be repeatedly manufactured at different time intervals using a two-tier cell banking process with robust and comparable angiogenic properties to treat ischemic diseases.

25-Hydroxyvitamin D3 inhibits cytokine induced-expression of immunomediators in human periodontal ligament derived mesenchymal stromal cells

Cytotherapy ◽  
2021 ◽  
Vol 23 (5) ◽  
pp. S61
Author(s):  
C. Behm ◽  
A. Blufstein ◽  
J. Gahn ◽  
A. Moritz ◽  
X. Rausch-Fan ◽  
...  
Keyword(s):  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Periodontal Ligament ◽  
Induced Expression

scholarly journals RNA-sequencing reveals positional memory of multipotent mesenchymal stromal cells from oral and maxillofacial tissue transcriptomes

2020 ◽  
Author(s):  
Satoru Onizuka ◽  
Yasuharu Yamazaki ◽  
Sung-Joon Park ◽  
Takayuki Sugimoto ◽  
Yumiko Sone ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Sequencing ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Periodontal Ligament ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Multipotent Mesenchymal Stromal Cells ◽  
Human Mscs ◽  
Minimum Criteria

Abstract Background Multipotent mesenchymal stromal cells (MSCs) can be isolated from numerous tissues and are attractive candidates for therapeutic clinical applications due to their immunomodulatory and pro-regenerative capacity. Although the minimum criteria for defining MSCs have been defined, their characteristics are known to vary depending on their tissue of origin.Results We isolated and characterized human MSCs from three different bones (ilium (I-MSCs), maxilla (Mx-MSCs) and mandibular (Md-MSCs)) and proceeded with next generation RNA-sequencing. Furthermore, to investigate the gene expression profiles among other cell types, we obtained RNA-seq data of human embryonic stem cells (ESCs) and several types of MSCs (periodontal ligament-derived MSCs, bone marrow-derived MSCs, and ESCs-derived MSCs) from the Sequence Reads Archive and analyze the transcriptome profile. We found that MSCs derived from tissues of the maxillofacial region, such as the jaw bone and periodontal ligament, were HOX-negative, while those derived from other tissues were HOX-positive. We also identified that MSX1, LHX8, and BARX1 , an essential regulator of craniofacial development, were strongly expressed in maxillofacial tissue-derived MSCs. Although MSCs may be divided into two distinct groups, the cells originated from over the neck or not, on the basis of differences in gene expression profile, the expression patterns of all CD antigen genes were similar among different type of MSCs, except for ESCs.Conclusions Our findings suggest that MSCs from different anatomical locations, despite meeting general characterization criteria, have remarkable differences in gene expression and positional memory. Although stromal cells from different anatomical sources are generally categorized as MSCs, their differentiation potential and biological functions vary. We suggested that MSCs may retain an original tissue memory about the developmental process, including gene expression profiles. This could have an important impact when choosing an appropriate cell source for regenerative therapy using MSCs.

scholarly journals Membrane particles from mesenchymal stromal cells reduce the expression of fibrotic markers on pulmonary cells

PLoS ONE ◽  
2021 ◽  
Vol 16 (3) ◽  
pp. e0248415
Author(s):  
Ana Merino ◽  
Martin J. Hoogduijn ◽  
Maria Molina-Molina ◽  
Elena G. Arias-Salgado ◽  
Sander S. Korevaar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pulmonary Fibrosis ◽  
Telomere Length ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Treatment Options ◽  
A549 Cells ◽  
Lung Fibroblasts ◽  
Pai 1

Background Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease with limited treatment options in which the telomere shortening is a strong predictive factor of poor prognosis. Mesenchymal stromal cells (MSC) administration is probed in several experimental induced lung pathologies; however, MSC might stimulate fibrotic processes. A therapy that avoids MSC side effects of transformation would be an alternative to the use of living cells. Membranes particles (MP) are nanovesicles artificially generated from the membranes of MSC containing active enzymes involved in ECM regeneration. We aimed to investigate the anti-fibrotic role of MP derived from MSC in an in vitro model of pulmonary fibrosis. Methods Epithelial cells (A549) and lung fibroblasts, from IPF patients with different telomere length, were co-cultured with MP and TGF-β for 48h and gene expression of major pro-fibrotic markers were analyzed. Results About 90% of both types of cells effectively took up MP without cytotoxic effects. MP decreased the expression of profibrotic proteins such as Col1A1, Fibronectin and PAI-1, in A549 cells. In fibroblasts culture, there was a different response in the inhibitory effect of MP on some pro-fibrotic markers when comparing fibroblast from normal telomere length patients (FN) versus short telomere length (FS), but both types showed an inhibition of Col1A1, Tenascin-c, PAI-1 and MMP-1 gene expression after MP treatment. Conclusions MP conserve some of the properties attributed to the living MSC. This study shows that MP target lung cells, via which they may have a broad anti-fibrotic effect.

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