scholarly journals Evaluation of BMP-2 Enhances the Osteoblast Differentiation of Human Amnion Mesenchymal Stem Cells Seeded on Nano-Hydroxyapatite/Collagen/Poly(l-Lactide)

2018 ◽  
Vol 19 (8) ◽  
pp. 2171 ◽  
Author(s):  
Shuhong Wu ◽  
Zhili Xiao ◽  
Jinlin Song ◽  
Min Li ◽  
Wenhua Li
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Bone Regeneration ◽  
Osteogenic Differentiation ◽  
Calcium Deposition ◽  
Mineral Formation ◽  
Human Amnion ◽  
Alp Activity ◽  
Experimental Group

Background: The aim of this study is to evaluate the effects of recombinant human bone morphogenetic protein 2 (rhBMP-2), human amnion mesenchymal stem cells (hAMSCs), and nanohydroxyapatite/collagen/poly(l-lactide) (nHAC/PLA) in tissue engineering to provide potential approaches for periodontal bone regeneration. Methods: hAMSCs were isolated from discarded amniotic membrane samples and cultured in vitro. Alkaline phosphatase (ALP) staining and alizarin red staining were performed to evaluate the osteoblast (OB) differentiation ability of hAMSCs. Three groups were divided: the experimental group (cells transfected with pcDNA3.1-rhBMP-2), the blank group (cells without gene transfection), and the control group (cells transfected with empty plasmid). RT-PCR and western blot were used to examine whether rhBMP-2 has been successfully expressed. 3-(4,5)-dimethylthiahiazol(-z-y1)-3,5-di-phenytetrazo-liumromide assay (MTT) was done to detect the effect of rhBMP-2 on hAMSCs seeded on nHAC/PLA. ALP activity, mineral formation assay, calcium, phosphate and osteocalcin (OCN) content, and OCN and RUNX2 expression of hAMSCs were detected to evaluate osteogenic differentiation capability of rhBMP-2 on hAMSCs seeded on nHAC/PLA. Results: hAMSCs exhibited intense ALP staining, obvious calcium deposition, and mineralization nodules, and rhBMP-2 were highly expressed in the experimental group. The proliferation of the hAMSCs with rhBMP-2 on nHAC/PLA was significantly higher than the cells without rhBMP-2, and the cells all increased in a time-dependent manner. rhBMP-2 significantly increased the OCN and phosphate content, mineral formation, ALP activity, osteogenic biomarkers OCN, and Runx2, and decreased calcium content in hAMSCs seeded on the nHAC/PLA scaffold. Conclusions: This finding demonstrated that hAMSCs has an ideal OB differentiation ability. rhBMP-2 facilitates the proliferation and osteogenesis of hAMSCs. The nHAC/PLA could act as a good scaffold for hAMSCs seeding, proliferation, and osteogenic differentiation. The application of rhBMP-2, nHAC/PLA, and hAMSCs in tissue engineering may offer promising possibilities for periodontal bone regeneration.

scholarly journals Effect of puerarin on osteogenic differentiation of human periodontal ligament stem cells

2019 ◽  
Vol 48 (4) ◽  
pp. 030006051985164
Author(s):  
Jun Li ◽  
Youjian Peng
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Calcium Deposition ◽  
Pcr Analysis ◽  
Cell Surface Markers ◽  
Periodontal Ligament Stem Cells ◽  
Alizarin Red ◽  
Alp Activity ◽  
Experimental Group

Objective To investigate the effects of the flavonoid, puerarin, on osteogenic differentiation of human periodontal ligament stem cells (PDLSCs). Methods Human PDLSCs were isolated from patients undergoing orthodontic treatment, and the cell surface markers CD146, CD34, CD45, and STRO-1 were identified by immunofluorescence. Cell proliferation was detected by MTT assay; alkaline phosphatase (ALP) activity was measured, and calcium deposition was detected by alizarin red staining. PCR was then used to detect the distributions of COL-I, OPN, Runx2, and OCN, genes related to osteogenic differentiation. Results Staining was positive for cytokines CD146, CD34, CD45, and STRO-1 in the experimental group; staining was also positive for silk protein, but negative for keratin. After 7 days of culture, exposure to puerarin significantly promoted the level of intracellular ALP; increased puerarin concentration led to increased intracellular ALP. Red mineralized nodules appeared upon exposure to puerarin and the number of nodules was concentration-dependent. PCR analysis revealed that COL-I, OPN, Runx2, and OCN expression levels increased as puerarin concentration increased. Conclusions Exposure to puerarin can promote proliferation and ALP activity in human PDLSCs, thus promoting both molecular and osteogenic differentiation; these findings may provide a theoretical basis for the clinical treatment of periodontal disease with puerarin.

scholarly journals Dihydrotestosterone and 17-Estradiol Enhancement of In Vitro Osteogenic Differentiation of Castrated Male Rat Bone Marrow Mesenchymal Stem Cells (rBMMSCs)

2019 ◽  
Author(s):  
FAM Abo-Aziza ◽  
AA Zaki ◽  
AS Amer ◽  
RA Lotfy
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Population Doubling ◽  
Calcium Deposition ◽  
Alp Activity ◽  
Marrow Mesenchymal Stem Cells ◽  
Rat Bone

Background: In vitro impact of dihydrotestosterone (DHT) and 17-estradiol (E2) in osteogenic differentiation of castrated rat bone marrow mesenchymal stem cells (rBMMSC) still need to be clarified. Materials and Methods: The viability, proliferation and density of cultured rBMMSC isolated from sham operated (Sham) and castrated (Cast) male rats were evaluated. rBMMSC were cultured with osteogenic differentiating medium (ODM) in the presence of DHT (5,10 nM) and E2 (10,100 nM). Osteogenesis was evaluated by alizarin red staining and measurement of calcium deposition and bone alkaline phosphatase (BALP) activity. Results: Population doubling (PD) of rBMMSC isolated from Cast rats was significantly lower (P<0.05) compared to that isolated from Sham rats. rBMMSC from Cast rats showed low scattered calcified nodule after culturing in ODM and did not cause a significant increase in calcium deposition and B-ALP activity compared to rBMMSCs from Sham rats. Exposure of rBMMSC isolated from Cast rats to DHT (5 nM) or E2 (10 nM) in ODM showed medium scattered calcified nodules with significantly higher (P<0.05) calcium deposition and B-ALP activity. Moreover, exposure of rBMMSC to DHT (10 nM) or E2 (100 nM) showed high scattered calcified nodules with higher (P<0.01) calcium deposition and B-ALP activity Conclusion: These results indicated that the presence of testes might participate in controlling the in vitro proliferation and osteogenic differentiation capacity of rBMMSCs. DHT and E2 can enhance the osteogenic capacity of rBMMSCs in a dose-dependent manner. Based on these observations, optimum usage of DHT and E2 can overcome the limitations of MSCs and advance the therapeutic bone regeneration potential in the future.

scholarly journals Osteogenic differentiation of human mesenchymal stem cells from adipose tissue and Wharton’s jelly of the umbilical cord

2017 ◽  
Vol 64 (2) ◽  
Author(s):  
Alicja Zajdel ◽  
Magdalena Kałucka ◽  
Edyta Kokoszka-Mikołaj ◽  
Adam Wilczok
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Osteogenic Differentiation ◽  
Umbilical Cord ◽  
Wharton’S Jelly ◽  
Calcium Deposition ◽  
Human Umbilical Cord ◽  
Wharton's Jelly ◽  
Alp Activity

Induced osteogenesis of mesenchymal stem cells (MSCs) may provide an important tool for bone injures treatment. Human umbilical cord and adipose tissue are routinely discarded as clinical waste and may be used as uncontroversial MSCs sources. It still remains to be verified which source of MSCs is the most suitable for bone regeneration.The aim of this research was to investigate the osteogenic potential of human MSCs derived from adipose tissue (ASCs) and Wharton’s jelly of the human umbilical cord (WJ-MSCs) differentiated under the same conditions.Osteogenic differentiation of MSCs was detected and quantified by ARS staining for calcium deposition and alkaline phosphatase (ALP) activity, osteoprotegerin (OPG), and osteocalcin (OC) secretion measurements. Under osteogenic conditions the measured ALP activity and calcium deposition were significantly higher in ASCs than in WJ-MSCs, while the OPG and OC secretion were higher in WJ-MSCs vs. ASCs. Low concentrations of OPG and high levels of OC in ASCs and WJ-MSCs, prove that these cells reached an advanced stage of the osteogenic differentiation. The levels of OC secreted by ASCs were lower than by WJ-MSCs what indicates that the differentiation process of the ASCs reached the stage when the extracellular matrix is overproduced and the down-regulation of OC begins.Both cell types, ASCs and WJ-MSCs possess potential to differentiate towards the osteogenic lineage. However, the observed differences in the levels of osteogenic markers suggest that ASCs may be better candidates for cell-based osteogenesis than WJ-MSCs.

scholarly journals The mechanism of miR-889 regulates osteogenesis in human bone marrow mesenchymal stem cells

2019 ◽  
Vol 14 (1) ◽  
Author(s):  
Gang Xu ◽  
Zheng Ding ◽  
Hui-feng Shi
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Signaling Pathway ◽  
Calcium Deposition ◽  
Control Group ◽  
Related Protein ◽  
Alp Activity ◽  
Marrow Mesenchymal Stem Cells

Abstract Background Bone marrow mesenchymal stem cells (BMMSCs) can be used for bone regeneration in the specified condition. Osteogenic differentiation of BMMSCs is controlled by microRNAs (miRNAs) and other factors. This study was aimed to identify the role and mechanism of miR-889 in regulating the osteogenic differentiation of BMMSCs. Methods Osteoporosis patients and normal control bone tissues were collected and used PCR techniques to identify the change of miR-889 and WNT7A. Moreover, the dynamic change of miR-889 and WNT7A during osteogenic differentiation of BMMSCs was also measured. Bioinformatic analysis was performed to identify the target genes and potential pathways of miR-889. Then, we constructed miR-889 mimic and inhibitor, ALP staining, ARS, osteoblastic-related protein, and Wnt β-catenin signaling pathway-related protein were also measured. WNT7A siRNA was also used to verify the function of miR-889. Results In the present study, we showed that miR-889 expression was upregulated in osteoporosis patients than healthy control. However, the miR-889 expression was downregulated during osteogenic differentiation. Bioinformatics analysis found that miR-889 targets 666 genes and mainly through Wnt β-catenin signaling pathway. Administrated miR-889 mimic, the ALP activity, and calcium deposition were decreased than the control group, while miR-889 inhibitor shown the opposite trend. And miR-889 could bind the 3′UTR of WNT7A. We further used WNT7A siRNA to explore the function of miR-889, and the results revealed that co-cultured with miR-889 inhibitor and WNT7A siRNA was associated with a reduction of ALP activity and calcium deposition and osteoblastic-related proteins than miR-889 inhibitor alone. Conclusion Our results revealed that miR-889 plays a negative role in inducing osteogenic differentiation of BMSCs through Wnt β-catenin signaling pathway.

scholarly journals Thymoquinone loading into hydroxyapatite/alginate scaffolds accelerated the osteogenic differentiation of the mesenchymal stem cells

2021 ◽  
Author(s):  
Ebrahim Rahmani-Moghadam ◽  
Tahereh Talaei-Khozani ◽  
Vahideh Zarrin ◽  
Zahra Vojdani
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Physical And Mechanical Properties ◽  
Calcium Deposition ◽  
Alizarin Red ◽  
Qrt Pcr ◽  
Alp Activity ◽  
Sintering Method

Abstract Background: Hydroxyapatite (HA) can be loaded by some osteogenic inducing agents such as thymoquinone (TQ) and alginate. This study was performed to investigate the effect of TQ loading into HA/alginate scaffolds on osteogenic differentiation capability of mesenchymal stem cells.Methods: HA scaffolds were fabricated by casting and sintering method and impregnated by TQ containing alginate. The stem cells were loaded onto the scaffolds and induced to differentiate into osteoblasts. Alkaline Phosphatase (ALP) activity, Alizarin Red S, Real-Time qRT-PCR, and MTT assessments were done. Finally, the cells were examined with a light microscope, confocal microscope, and SEM.Results: The results showed that the presence of the alginate decelerates the degradation rate and reinforces the mechanical strength. while the presence of TQ had no significant influence on physical and mechanical properties of the HA/alginate scaffolds, it led to a significant increase in ALP activity and expression of collagen, osteopontin, and osteocalcin at early phase of differentiation. Also, TQ administration had no impact on calcium deposition and proliferation as well as bone-marker expression at long term differentiation.Conclusion: TQ accelerates the differentiation of the stem cells into the osteoblasts without changing the properties of the scaffolds, and the HA/alginate/TQ scaffold can be used as a scaffold with osteogenic properties in bone tissue engineering applications.

scholarly journals Effects of icariin on the proliferation and osteogenic differentiation of human amniotic mesenchymal stem cells

2020 ◽  
Vol 15 (1) ◽  
Author(s):  
Fang Wang ◽  
Zhiyong Yang ◽  
Wei He ◽  
Qinggao Song ◽  
Kun Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Immunocytochemical Staining ◽  
Control Group ◽  
Engineering Technology ◽  
Alizarin Red ◽  
Human Amnion ◽  
Alp Activity ◽  
Amniotic Mesenchymal Stem Cells

Abstract Background Tissue engineering technology has been applied extensively for clinical research and human amnion mesenchymal stem cells (hAMSCs) could cause mesenchymal stem cells to differentiate into the bone tissue. However, it is necessary to develop and identify the safer appropriate amount of osteogenic inducer. The objective of this study is to investigate the effect of icariin (ICA) on the proliferation and osteogenic differentiation of hAMSCs. Methods The morphology and phenotype of hAMSCs were discovered by flow cytometry and immunocytochemical staining. The osteogenic differentiation of hAMSCs under the influence of different concentrations of ICA were assessed by alkaline phosphatase (ALP) activity substrate assay and alizarin red staining. Results MTT assay revealed that the hAMSCs pretreated with ICA exhibited increased proliferation when compared with the control group, and the most optimum concentration of ICA was 1 × 10− 6 mol/L. The combined analysis of ALP activity and ARS staining showed that ICA could significantly promote the osteogenic differentiation of hAMSCs, and the effect was most significant when the concentration of ICA was 1 × 10− 6 mol/L. Conclusion All the above results implied that ICA could significantly increase proliferation and enhance the osteogenic differentiation of hAMSCs, especially when the concentration of ICA was 1 × 10− 6 mol/L.

scholarly journals Effects of icariin on the proliferation and osteogenic differentiation of human amniotic mesenchymal stem cells

2020 ◽  
Author(s):  
Fang Wang ◽  
Zhiyong Yang ◽  
He Wei ◽  
Qinggao Gao ◽  
Kun Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Immunocytochemical Staining ◽  
Control Group ◽  
Engineering Technology ◽  
Alizarin Red ◽  
Human Amnion ◽  
Alp Activity ◽  
Amniotic Mesenchymal Stem Cells

Abstract Background Tissue engineering technology has been applied extensively for clinical research and human amnion mesenchymal stem cells (hAMSCs) could cause mesenchymal stem cells to differentiate into the bone tissue. However, it is necessary to develop and identify the safer appropriate amount of osteogenic inducer. The objective of this study is to investigate the effect of icariin (ICA) on the proliferation and osteogenic differentiation of hAMSCs. Methods The morphology and phenotype of hAMSCs were discovered by flow cytometry and immunocytochemical staining. The osteogenic differentiation of hAMSCs under the influence of different concentrations of ICA were assessed by Alkaline phosphatase (ALP) activity substrate assay and Alizarin red Staining. Results MTT assay revealed that the hAMSCs pretreated with ICA exhibited increased proliferation when compared with the control group, and the most optimum concentration of ICA was 1 × 10− 6 mol/L. The combined analysis of ALP activity and ARS staining showed that ICA could significantly promote the osteogenic differentiation of hAMSCs, and the effect was most significant when the concentration of ICA was 1 × 10− 6 mol/L. Conclusion All above results implied that ICA could significantly increase proliferation and enhanced the osteogenic differentiation of hAMSCs, especially when the concentration of ICA was 1 × 10− 6 mol/L.

scholarly journals Engineered scaffolds based on mesenchymal stem cells/preosteoclasts extracellular matrix promote bone regeneration

2020 ◽  
Vol 11 ◽  
pp. 204173142092691 ◽  
Author(s):  
Rui Dong ◽  
Yun Bai ◽  
Jingjin Dai ◽  
Moyuan Deng ◽  
Chunrong Zhao ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Extracellular Matrix ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Mesenchymal Stem Cell ◽  
Bone Regeneration ◽  
Osteogenic Differentiation ◽  
Complex Biological Process

Recently, extracellular matrix-based tissue-engineered bone is a promising approach to repairing bone defects, and the seed cells are mostly mesenchymal stem cells. However, bone remodelling is a complex biological process, in which osteoclasts perform bone resorption and osteoblasts dominate bone formation. The interaction and coupling of these two kinds of cells is the key to bone repair. Therefore, the extracellular matrix secreted by the mesenchymal stem cells alone cannot mimic a complex bone regeneration microenvironment, and the addition of extracellular matrix by preosteoclasts may contribute as an effective strategy for bone regeneration. Here, we established the mesenchymal stem cell/preosteoclast extracellular matrix -based tissue-engineered bones and demonstrated that engineered-scaffolds based on mesenchymal stem cell/ preosteoclast extracellular matrix significantly enhanced osteogenesis in a 3 mm rat femur defect model compared with mesenchymal stem cell alone. The bioactive proteins released from the mesenchymal stem cell/ preosteoclast extracellular matrix based tissue-engineered bones also promoted the migration, adhesion, and osteogenic differentiation of mesenchymal stem cells in vitro. As for the mechanisms, the iTRAQ-labeled mass spectrometry was performed, and 608 differentially expressed proteins were found, including the IGFBP5 and CXCL12. Through in vitro studies, we proved that CXCL12 and IGFBP5 proteins, mainly released from the preosteoclasts, contributed to mesenchymal stem cells migration and osteogenic differentiation, respectively. Overall, our research, for the first time, introduce pre-osteoclast into the tissue engineering of bone and optimize the strategy of constructing extracellular matrix–based tissue-engineered bone using different cells to simulate the natural bone regeneration environment, which provides new sight for bone tissue engineering.

scholarly journals Effects of icariin on the proliferation and osteogenic differentiation of human amniotic mesenchymal stem cells

2020 ◽  
Author(s):  
Fang Wang ◽  
Zhiyong Yang ◽  
He Wei ◽  
Qinggao Gao ◽  
Kun Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Immunocytochemical Staining ◽  
Control Group ◽  
Engineering Technology ◽  
Alizarin Red ◽  
Human Amnion ◽  
Alp Activity ◽  
Amniotic Mesenchymal Stem Cells

Abstract Background: Tissue engineering technology has been applied extensively for clinical research and human amnion mesenchymal stem cells (hAMSCs) could cause mesenchymal stem cells to differentiate into the bone tissue. However, it is necessary to develop and identify the safer appropriate amount of osteogenic inducer. The objective of this study is to investigate the effect of icariin (ICA) on the proliferation and osteogenic differentiation of hAMSCs.Methods: The morphology and phenotype of hAMSCs were discovered by flow cytometry and immunocytochemical staining. The osteogenic differentiation of hAMSCs under the influence of different concentrations of ICA were assessed by Alkaline phosphatase (ALP) activity substrate assay and Alizarin red Staining.Results: MTT assay revealed that the hAMSCs pretreated with ICA exhibited increased proliferation when compared with the control group, and the most optimum concentration of ICA was 1×10-6 mol/L. The combined analysis of ALP activity and ARS staining showed that ICA could significantly promote the osteogenic differentiation of hAMSCs, and the effect was most significant when the concentration of ICA was 1×10-6mol/L.Conclusion: All above results implied that ICA could significantly increase proliferation and enhanced the osteogenic differentiation of hAMSCs, especially when the concentration of ICA was 1×10-6 mol/L.Keywords: Human amniotic mesenchymal stem cell, Icariin, Proliferation, Osteogenic differentiation

scholarly journals Fluid Flow Mechanical Stimulation-Assisted Cartridge Device for the Osteogenic Differentiation of Human Mesenchymal Stem Cells

Micromachines ◽  
2021 ◽  
Vol 12 (8) ◽  
pp. 927
Author(s):  
Ki-Taek Lim ◽  
Dinesh-K. Patel ◽  
Sayan-Deb Dutta ◽  
Keya Ganguly
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Fluid Flow ◽  
Osteogenic Differentiation ◽  
Flow Condition ◽  
Cultured Cells ◽  
Human Mesenchymal Stem Cells ◽  
Proliferation And Differentiation ◽  
Static Conditions

Human mesenchymal stem cells (hMSCs) have the potential to differentiate into different types of mesodermal tissues. In vitro proliferation and differentiation of hMSCs are necessary for bone regeneration in tissue engineering. The present study aimed to design and develop a fluid flow mechanically-assisted cartridge device to enhance the osteogenic differentiation of hMSCs. We used the fluorescence-activated cell-sorting method to analyze the multipotent properties of hMSCs and found that the cultured cells retained their stemness potential. We also evaluated the cell viabilities of the cultured cells via water-soluble tetrazolium salt 1 (WST-1) assay under different rates of flow (0.035, 0.21, and 0.35 mL/min) and static conditions and found that the cell growth rate was approximately 12% higher in the 0.035 mL/min flow condition than the other conditions. Moreover, the cultured cells were healthy and adhered properly to the culture substrate. Enhanced mineralization and alkaline phosphatase activity were also observed under different perfusion conditions compared to the static conditions, indicating that the applied conditions play important roles in the proliferation and differentiation of hMSCs. Furthermore, we determined the expression levels of osteogenesis-related genes, including the runt-related protein 2 (Runx2), collagen type I (Col1), osteopontin (OPN), and osteocalcin (OCN), under various perfusion vis-à-vis static conditions and found that they were significantly affected by the applied conditions. Furthermore, the fluorescence intensities of OCN and OPN osteogenic gene markers were found to be enhanced in the 0.035 mL/min flow condition compared to the control, indicating that it was a suitable condition for osteogenic differentiation. Taken together, the findings of this study reveal that the developed cartridge device promotes the proliferation and differentiation of hMSCs and can potentially be used in the field of tissue engineering.

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