The ERα/KDM6B regulatory axis modulates osteogenic differentiation in human mesenchymal stem cells

AbstractOsteoporosis is a highly prevalent public health burden associated with an increased risk of bone fracture, particularly in aging women. Estrogen, an important medicinal component for the preventative and therapeutic treatment of postmenopausal osteoporosis, induces osteogenesis by activating the estrogen receptor signaling pathway and upregulating the expression of osteogenic genes, such as bone morphogenetic proteins (BMPs). The epigenetic regulation of estrogen-mediated osteogenesis, however, is still unclear. In this report, we found that estrogen significantly induced the expression of lysine-specific demethylase 6B (KDM6B) and that KDM6B depletion by shRNAs led to a significant reduction in the osteogenic potential of DMSCs. Mechanistically, upon estrogen stimulation, estrogen receptor-α (ERα) was recruited to the KDM6B promoter, directly enhancing KDM6B expression. Subsequently, KDM6B was recruited to the BMP2 and HOXC6 promoters, resulting in the removal of H3K27me3 marks and activating the transcription of BMP2 and HOXC6, the master genes of osteogenic differentiation. Furthermore, we found that estrogen enhanced DMSC osteogenesis during calvarial bone regeneration and that estrogen’s pro-osteogenic effect was dependent on KDM6B in vivo. Taken together, our results demonstrate the vital role of the ERα/KDM6B regulatory axis in the epigenetic regulation of the estrogen-dependent osteogenic response.

Download Full-text

In Vivo Study for Clinical Application of Dental Stem Cell Therapy Incorporated with Dental Titanium Implants

Materials ◽

10.3390/ma14020381 ◽

2021 ◽

Vol 14 (2) ◽

pp. 381

Author(s):

Hyunmin Choi ◽

Kyu-Hyung Park ◽

Narae Jung ◽

June-Sung Shim ◽

Hong-Seok Moon ◽

...

Keyword(s):

Stem Cells ◽

Bone Formation ◽

Alveolar Bone ◽

Human Mesenchymal Stem Cells ◽

Titanium Implants ◽

Osteogenic Potential ◽

Induction Medium ◽

Radiographic Analysis ◽

New Bone Formation

The aim of this study was to investigate the behavior of dental-derived human mesenchymal stem cells (d-hMSCs) in response to differently surface-treated implants and to evaluate the effect of d-hMSCs on local osteogenesis around an implant in vivo. d-hMSCs derived from alveolar bone were established and cultured on machined, sandblasted and acid-etched (SLA)-treated titanium discs with and without osteogenic induction medium. Their morphological and osteogenic potential was assessed by scanning electron microscopy (SEM) and real-time polymerase chain reaction (RT-PCR) via mixing of 5 × 106 of d-hMSCs with 1 mL of Metrigel and 20 μL of gel-cell mixture, which was dispensed into the defect followed by the placement of customized mini-implants (machined, SLA-treated implants) in New Zealand white rabbits. Following healing periods of 2 weeks and 12 weeks, the obtained samples in each group were analyzed radiographically, histomorphometrically and immunohistochemically. The quantitative change in osteogenic differentiation of d-hMSCs was identified according to the type of surface treatment. Radiographic analysis revealed that an increase in new bone formation was statistically significant in the d-hMSCs group. Histomorphometric analysis was in accordance with radiographic analysis, showing the significantly increased new bone formation in the d-hMSCs group regardless of time of sacrifice. Human nuclei A was identified near the area where d-hMSCs were implanted but the level of expression was found to be decreased as time passed. Within the limitations of the present study, in this animal model, the transplantation of d-hMSCs enhanced the new bone formation around an implant and the survival and function of the stem cells was experimentally proven up to 12 weeks post-sacrifice.

Download Full-text

Human Sex Matters: Y-linked lysine demethylase 5D drives accelerated male osteogenic differentiation

10.1101/2021.11.10.468047 ◽

2021 ◽

Author(s):

Madlen Merten ◽

Johannes F.W. Greiner ◽

Tarek Niemann ◽

Meike Grosse Venhaus ◽

Daniel Kronenberg ◽

...

Keyword(s):

Osteogenic Differentiation ◽

Molecular Mechanisms ◽

Female Rats ◽

Sexually Dimorphic ◽

Loss Of Function ◽

Calvarial Bone ◽

Sexual Dimorphisms ◽

Increased Risk ◽

Elevated Expression ◽

Lysine Demethylase

Female sex is increasingly associated to a loss of bone mass during aging and an increased risk for fractures developing nonunion. Hormonal factors and cell-intrinsic mechanisms are suggested to drive these sexual dimorphisms, although underlying molecular mechanisms are still a matter of debate. Here, we observed a decreased capacity of calvarial bone recovery in female rats and a profound sexually dimorphic osteogenic differentiation human adult neural crest-derived stem cells (NCSCs). Next to an elevated expression of pro-osteogenic regulators, global trancriptomics revealed Lysine Demethylase 5D (KDM5D) to be highly upregulated in differentiating male NCSCs. Loss of function by siRNA or pharmacological inhibition of KDM5D significantly reduced the osteogenic differentiation capacity of male NCSCs. In summary, we demonstrate craniofacial osteogenic differentiation to be sexually dimorphic with the expression of KDM5D as a prerequisite for accelerated male osteogenic differentiation, emphasizing the analysis of sex-specific differences as a crucial parameter for treating bone defects.

Download Full-text

Influence of Maitake (Grifola frondosa) Particle Sizes on Human Mesenchymal Stem Cells and In Vivo Evaluation of Their Therapeutic Potential

BioMed Research International ◽

10.1155/2020/8193971 ◽

2020 ◽

Vol 2020 ◽

pp. 1-13

Author(s):

Dinesh K. Patel ◽

Yu-Ri Seo ◽

Sayan Deb Dutta ◽

Ok Hwan Lee ◽

Ki-Taek Lim

Keyword(s):

Therapeutic Potential ◽

Human Mesenchymal Stem Cells ◽

Grifola Frondosa ◽

Osteogenic Potential ◽

Calcium Deposition ◽

Particle Sizes ◽

In Vivo Evaluation ◽

Rat Serum ◽

Gene Markers

Maitake (Grifola frondosa) mushroom has received an enormous amount of attention as a dietary supplement due to its high nutritional values. The particle sizes of G. frondosa mushrooms were monitored by a classifying mill. β-Glucans are the bioactive component of the mushroom, and it was revealed through Fourier transform infrared spectroscopy (FTIR), proton and carbon nuclear magnetic resonance (1H and 13C-NMR), matrix-assisted laser desorption/ionization, and time-of-flight (MALDI-TOF) spectrometry. The biocompatibility of G. frondosa particles, as well as induced osteogenesis of hMSCs, was evaluated through WST-1 assay and alizarin staining (ARS) technique, respectively. Notably, enhanced cell viability was noted in the presence of G. frondosa. Significantly improved calcium deposition has observed from hMSCs with G. frondosa, suggesting to their mineralization potential. The expression of osteogenic related gene markers was examined in the presence of G. frondosa through real-time polymerase chain reaction (qPCR) technique. The upregulation of osteogenic gene markers in the presence of G. frondosa particles was indicating their superior osteogenic potential. Besides, G. frondosa also activated the secretion of various kinds of proteins from the hMSCs indicating their potential for tissue engineering applications. Enhanced secretion of different immunoglobulins was observed in rat serum in the presence of G. frondosa, further demonstrating their therapeutic nature. Therefore, G. frondosa is effective for enhanced osteogenesis and can be utilized as a natural, edible, and osteogenic agent.

Download Full-text

Osteogenic Potential Using a Malleable, Biodegradable Composite Added Traditional Chinese Medicine: in vitro and in vivo Evaluations

The American Journal of Chinese Medicine ◽

10.1142/s0192415x06004351 ◽

2006 ◽

Vol 34 (05) ◽

pp. 873-886 ◽

Cited By ~ 6

Author(s):

Chun-Hsu Yao ◽

Bai-Shuan Liu ◽

Chau-Guey Liu ◽

Yueh-Sheng Chen

Keyword(s):

Bone Substitute ◽

Bone Growth ◽

In Vitro Study ◽

Osteogenic Potential ◽

Calvarial Bone ◽

Large Defect ◽

Biodegradable Composite ◽

Calvarial Bone Defect

The purpose of this investigation was to prepare and evaluate the feasibility and biocompatibility of a new composite as a large defect bone substitute. The new GTGG was mainly composed of tricalcium phosphate ceramic particles and glutaraldehyde crosslinked gelatin in which Gui-Lu-Jiao was added (a mixture of Cervi Colla Cornus and Colla Plastri Testudinis). In the in vitro study, rat's calvaria osteoblasts were used to study bone characteristics upon exposure to different concentrations of the Gui-Lu-Jiao solution. In the in vivo study, GTGG composites were implanted into the defects of calvarial bones in mature New Zealand rabbits to test their osteogenerative characteristics. As a result, we found that Gui-Lu-Jiao added to the culture could promote the proliferation of osteoblasts. In addition, GTGG could induce a large amount of new bone growth in the rabbit's calvarial bone defect. Therefore, the GTGG composite might be a potential bone substitute.

Download Full-text

Pantoprazole (PPZ) Inhibits RANKL-Induced Osteoclast Formation and Function In Vitro and Prevents Lipopolysaccharide- (LPS-) Induced Inflammatory Calvarial Bone Loss In Vivo

Stem Cells International ◽

10.1155/2020/8829212 ◽

2020 ◽

Vol 2020 ◽

pp. 1-13

Author(s):

Yu-Xi Li ◽

Fu-Chao Chen ◽

Ting Liu ◽

Zhao-Peng Cai ◽

Keng Chen ◽

...

Keyword(s):

Bone Resorption ◽

Bone Matrix ◽

Bone Destruction ◽

Mapk Signaling ◽

Marker Genes ◽

Calvarial Bone ◽

Hematopoietic Stem ◽

Increased Risk

Bone remodeling is a process delicately balanced between osteoclastic bone resorption and osteoblastic bone formation. Osteoclasts (OCs) are multinucleated giant cells formed through the fusion of monocytic precursors of the hematopoietic stem cells lineage. OCs are the exclusive cells responsible for the resorption and degradation of the mineralized bone matrix. Pantoprazole (PPZ), a proton pump inhibitor (PPI), is commonly prescribed to reduce excess gastric acid production for conditions such as gastroesophageal reflux disease and peptic ulcer disease. Studies have found contradictory effects of PPI therapy on bone metabolism due to the lack of understanding of the exact underlying mechanism. In this study, we found that PPZ inhibits receptor activator of nuclear factor-κB (NF-κB) ligand- (RANKL-) induced osteoclastogenesis from bone marrow monocytic/macrophage (BMMs) precursors and the bone-resorbing activity of mature OCs. Correspondingly, the expression of OC marker genes was also attenuated. At the molecular level, PPZ treatment was associated with reduced activation of the ERK MAPK signaling pathways crucial to OC differentiation. Additionally, the in vivo administration of PPZ protected mice against lipopolysaccharide- (LPS-) induced inflammatory calvarial bone erosion, as a result of the reduced number and activity of OCs on the calvarial bone surface. Although PPI use is associated with increased risk of osteoporosis and bone fractures, our study provides evidence for the direct inhibitory effect of PPZ on OC formation and bone resorption in vitro and in vivo, suggesting a potential therapeutic use of PPZ in the treatment of osteolytic disease with localized bone destruction.

Download Full-text

The Effect of Covalently Immobilized FGF-2 on Biphasic Calcium Phosphate Bone Substitute on Enhanced Biological Compatibility and Activity

BioMed Research International ◽

10.1155/2015/742192 ◽

2015 ◽

Vol 2015 ◽

pp. 1-10 ◽

Cited By ~ 7

Author(s):

Kyung-Suk Moon ◽

Eun-Joo Choi ◽

Seunghan Oh ◽

Sungtae Kim

Keyword(s):

Calcium Phosphate ◽

Osteogenic Differentiation ◽

Biphasic Calcium Phosphate ◽

Human Mesenchymal Stem Cells ◽

Optimal Dose ◽

Osteogenic Potential ◽

Pcr Analysis ◽

Control Group ◽

Biological Compatibility ◽

Cross Linker

The purpose of this research was to covalently graft fibroblast growth factor 2 (FGF-2) onto biphasic calcium phosphate (BCP) via a bifunctional cross-linker technique and to estimate the optimal dose of FGF-2 resulting in the best osteogenic differentiation of human mesenchymal stem cells (hMSCs). SEM observation revealed that the surface of the 100 ng FGF-2 coated BCP was completely covered with the nanoparticles expected to be from the silane coupling agent. XRD, FT-IR, and XPS analysis showed that silane treatment, bifunctional cross-linker coating, and FGF-2 covalent grafts were conducted successfully without deforming the crystalline structure of BCP. An MTT assay demonstrated that FGF-2 coated BCP had good biocompatibility, regardless of the concentration of FGF-2, after 24 or 48 h of incubation. An alkaline phosphatase (ALP) activity assay (14 days of incubation) and the ALP gene expression level of real-time PCR analysis (7 days of incubation) revealed that 50, 100, and 200 ng FGF-2 coated BCP induced the highest activities among all experimental groups and control group (P<0.05). Thus, low concentrations of FGF-2 facilitated excellent osteogenesis and were effective at enhancing osteogenic potential. Also, the bifunctional cross-linker technique is expected to be a more feasible way to induce osteogenic differentiation while minimizing the risk of FGF-2 overdose.

Download Full-text

Identification of WNT16 as a Predictable Biomarker for Accelerated Osteogenic Differentiation of Tonsil-Derived Mesenchymal Stem Cells In Vitro

Stem Cells International ◽

10.1155/2019/8503148 ◽

2019 ◽

Vol 2019 ◽

pp. 1-10 ◽

Cited By ~ 1

Author(s):

Yu-Hee Kim ◽

Kyung-Ah Cho ◽

Hyun-Ji Lee ◽

Minhwa Park ◽

Han Su Kim ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Selection Criterion ◽

Osteogenic Potential ◽

Differentiation Potential ◽

Real Time Quantitative Pcr ◽

Single Cell Cloning

The application of mesenchymal stem cells (MSCs) for treating bone-related diseases shows promising outcomes in preclinical studies. However, cells that are isolated and defined as MSCs comprise a heterogeneous population of progenitors. This heterogeneity can produce variations in the performance of MSCs, especially in applications that require differentiation potential in vivo, such as the treatment of osteoporosis. Here, we aimed to identify genetic markers in tonsil-derived MSCs (T-MSCs) that can predict osteogenic potential. Using a single-cell cloning method, we isolated and established several lines of nondifferentiating (ND) or osteoblast-prone (OP) clones. Next, we performed transcriptome sequencing of three ND and three OP clones that maintained the characteristics of MSCs and determined the top six genes that were upregulated in OP clones. Upregulation of WNT16 and DCLK1 expression was confirmed by real-time quantitative PCR, but only WNT16 expression was correlated with the osteogenic differentiation of T-MSCs from 10 different donors. Collectively, our findings suggest that WNT16 is a putative genetic marker that predicts the osteogenic potential of T-MSCs. Thus, examination of WNT16 expression as a selection criterion prior to the clinical application of MSCs may enhance the therapeutic efficacy of stem cell therapy for bone-related complications, including osteoporosis.

Download Full-text

Impact of humanised isolation and culture conditions on stemness and osteogenic potential of bone marrow derived mesenchymal stromal cells

Scientific Reports ◽

10.1038/s41598-019-52442-9 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 3

Author(s):

Salwa Suliman ◽

Hassan R. W. Ali ◽

Tommy A. Karlsen ◽

Jerome Amiaud ◽

Samih Mohamed-Ahmed ◽

...

Keyword(s):

Bone Marrow ◽

Replicative Senescence ◽

Therapeutic Potential ◽

Culture Conditions ◽

Human Cells ◽

Osteogenic Potential ◽

Delayed Differentiation ◽

Osteogenic Response

Abstract Therapeutic potential of human bone marrow stromal/stem cells (hBMSC) must be developed using well defined xenogenic-free conditions. hBMSC were isolated from healthy donors (n = 3) using different isolation and expansion methods. Donor I was isolated and expanded by either bone marrow directly seeded and cells expanded in 10% AB human serum (AB) +5 ng/ml fibroblast growth factor-2 (FGF2) [Direct(AB + FGFlow)] or Ammonium-Chloride-Potassium Lysing Buffer was used before the cells were expanded in 10% AB +5 ng/ml FGF-2 [ACK(AB + FGFlow)] or Lymphoprep density gradient medium was used before the cells were expanded in 10% AB +5 ng/ml FGF2 [Lympho(AB + FGFlow)] or bone marrow directly seeded and cells expanded in 10% pooled platelet lysate plasma (PL) + heparin (2 I/U/mL) [Direct(PL)]. Groups for donors II and III were: Direct(AB + FGFlow) or 10% AB +10 ng/ml FGF2 [Direct(AB + FGFhigh)] or Direct(PL). HBMSCs were assessed for viability, multi-potency, osteogenic, inflammatory response and replicative senescence in vitro after 1 and 3 weeks. Pre-selected culture conditions, Direct(AB + FGFhigh) or Direct(PL), were seeded on biphasic calcium phosphate granules and subcutaneously implanted in NOD/SCID mice. After 1 and 11 weeks, explants were analysed for inflammatory and osteogenic response at gene level and histologically. To identify implanted human cells, in situ hybridisation was performed. hBMSC from all conditions showed in vitro multi-lineage potency. hBMSCs expanded in PL expressed stemness markers in vitro at significantly higher levels. Generally, cells expanded in AB + FGF2 conditions expressed higher osteogenic markers after 1 week both in vitro and in vivo. After 11 weeks in vivo, Direct(AB + FGFhigh) formed mature ectopic bone, compared to immature mineralised tissues formed by Direct(PL) implants. Mouse responses showed a significant upregulation of IL-1α and IL-1β expression in Direct(PL). After 1 week, human cells were observed in both groups and after 11 weeks in Direct(AB + FGFhigh) only. To conclude, results showed a significant effect of the isolation methods and demonstrated a relatively consistent pattern of efficacy from all donors. A tendency of hBMSC expanded in PL to retain a more stem-like phenotype elucidates their delayed differentiation and different inflammatory expressions.

Download Full-text

In Vitro and In Vivo Evaluation of Commercially Available Fibrin Gel as a Carrier of Alendronate for Bone Tissue Engineering

BioMed Research International ◽

10.1155/2017/6434169 ◽

2017 ◽

Vol 2017 ◽

pp. 1-10 ◽

Cited By ~ 4

Author(s):

Beom Su Kim ◽

Feride Shkembi ◽

Jun Lee

Keyword(s):

Bone Regeneration ◽

Osteogenic Differentiation ◽

Release Kinetics ◽

Human Mesenchymal Stem Cells ◽

Calvarial Defect ◽

Defect Model ◽

Fibrin Gel ◽

In Vivo Evaluation

Alendronate (ALN) is a bisphosphonate drug that is widely used for the treatment of osteoporosis. Furthermore, local delivery of ALN has the potential to improve the bone regeneration. This study was designed to investigate an ALN-containing fibrin (fibrin/ALN) gel and evaluate the effect of this gel on both in vitro cellular behavior using human mesenchymal stem cells (hMSCs) and in vivo bone regenerative capacity. Fibrin hydrogels were fabricated using various ALN concentrations (10−7–10−4 M) with fibrin glue and the morphology, mechanical properties, and ALN release kinetics were characterized. Proliferation and osteogenic differentiation of and cytotoxicity in fibrin/ALN gel-embedded hMSCs were examined. In vivo bone formation was evaluated using a rabbit calvarial defect model. The fabricated fibrin/ALN gel was transparent with Young’s modulus of ~13 kPa, and these properties were not affected by ALN concentration. The in vitro studies showed sustained release of ALN from the fibrin gel and revealed that hMSCs cultured in fibrin/ALN gel showed significantly increased proliferation and osteogenic differentiation. In addition, microcomputed tomography and histological analysis revealed that the newly formed bone was significantly enhanced by implantation of fibrin/ALN gel in a calvarial defect model. These results suggest that fibrin/ALN has the potential to improve bone regeneration.

Download Full-text

The influence of the morphology of titania and hydroxyapatite on the proliferation and osteogenic differentiation of human mesenchymal stem cells

RSC Advances ◽

10.1039/d0ra08271f ◽

2021 ◽

Vol 11 (7) ◽

pp. 3843-3853

Author(s):

Yauheni U. Kuvyrkou ◽

Nadzeya Brezhneva ◽

Ekaterina V. Skorb ◽

Sviatlana A. Ulasevich

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Human Mesenchymal Stem Cells ◽

Osteogenic Potential ◽

Titania Surface ◽

Surface Modified ◽

First Time ◽

Ordered Porous ◽

Porous Morphology

Herein, the proliferation and osteogenic potential of human mesenchymal stem cells (hMSCs) on the disordered and ordered porous morphology of the titania surface and titania surface modified by hydroxyapatite (HA) are compared for the first time.

Download Full-text