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2196 Articles

Published in last 50 years

Related Topics

  • Differentiation Of hMSCs
  • Differentiation Of hMSCs
  • Osteogenic Differentiation Potential
  • Osteogenic Differentiation Potential
  • Osteogenic Phenotype
  • Osteogenic Phenotype
  • Osteogenic Differentiation
  • Osteogenic Differentiation
  • Osteogenic Potential
  • Osteogenic Potential
  • Adipogenic Differentiation
  • Adipogenic Differentiation

Articles published on Osteogenic Lineage

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Electrical stimulation: Effective cue to direct osteogenic differentiation of mesenchymal stem cells?

Mesenchymal stem cells (MSCs) play a major role in bone tissue engineering (BTE) thanks to their capacity for osteogenic differentiation and being easily available. In vivo, MSCs are exposed to an electroactive microenvironment in the bone niche, which has piezoelectric properties. The correlation between the electrically active milieu and bone's ability to adapt to mechanical stress and self-regenerate has led to using electrical stimulation (ES) as physical cue to direct MSCs differentiation towards the osteogenic lineage in BTE. This review summarizes the different techniques to electrically stimulate MSCs to induce their osteoblastogenesis in vitro, including general electrical stimulation and substrate mediated stimulation by means of conductive or piezoelectric cell culture supports. Several aspects are covered, including stimulation parameters, treatment times and cell culture media to summarize the best conditions for inducing MSCs osteogenic commitment by electrical stimulation, from a critical point of view. Electrical stimulation activates different signaling pathways, including bone morphogenetic protein (BMP) Smad-dependent or independent, regulated by mitogen activated protein kinases (MAPK), extracellular signal-regulated kinases (ERK) and p38. The roles of voltage gate calcium channels (VGCC) and integrins are also highlighted according to their application technique and parameters, mainly converging in the expression of RUNX2, the master regulator of the osteogenic differentiation pathway. Despite the evident lack of homogeneity in the approaches used, the ever-increasing scientific evidence confirms ES potential as an osteoinductive cue, mimicking aspects of the in vivo microenvironment and moving one step forward to the translation of this approach into clinic.

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  • Biomaterials Advances
  • Jul 1, 2022
  • M Guillot-Ferriols + 3
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Osteogenic growth peptide enhances osteogenic differentiation of human periodontal ligament stem cells

Bone tissue engineering consists of three major components namely cells, scaffolds, and signaling molecules to improve bone regeneration. These integrated principles can be applied in patients suffered from bone resorption diseases, such as osteoporosis and periodontitis. Osteogenic growth peptide (OGP) is a fourteen-amino acid sequence peptide that has the potential to regenerate bone tissues. This study aimed to disseminate the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) with OGP treatment. OGP was elaborated for proliferation, cytotoxicity, osteogenic differentiation effects, and the involvement of osteogenic related signaling pathways in vitro. This study found that OGP at lower concentration shows better effects on cytotoxicity and proliferation. Moreover, OGP at concentration 0.01 nM had the most potential to differentiate hPDLSCs toward osteogenic lineage comparing with higher concentrations of OGP. The phenomenon was mainly involving transforming growth factor-beta (TGF-β), bone morphogenetic protein (BMP), Hedgehog, and Wingless-related (Wnt) pathways. Further, SB-431542 treatment demonstrated the partial involvement of OGP in regulating osteogenic differentiation of hPDLSCs. In conclusion, OGP at low concentration enhances osteogenic differentiation of hPDLSCs by governing TGF-β signaling pathway.

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  • Heliyon
  • Jul 1, 2022
  • Steven Dwi Purbantoro + 3
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Silymarin Effects on Ovine Fetal Bone Marrow-Derived Mesenchymal Stem Cells Differentiation into Osteogenic Lineage

هدف: سلول درمانی با استفاده از سلول­های بنیادی مزانشیمی (MSCs) می تواند ابزاری امیدوار کننده به عنوان طب بازساختی باشد. یکی از غنی ترین منابع سلول های بنیادی مزانشیمی مغز استخوان جنین است. سیلیمارین دارای فعالیت های آنتی اکسیدانی و ضد التهابی قوی با تأثیر مثبت بر تکثیر برخی از سلول ها و همچنین خاصیت ضد پوکی استخوان است. این مطالعه با هدف نشان دادن اثر سیلیمارین بر تمایز سلول های بنیادی مزانشیمی مشتق از مغز استخوان جنین گوسفند به رده استئوژنیک انجام شد. مواد و روش­ها: سلول­های بنیادی مزانشیمی از مغز استخوان جنین گوسفند جدا شدند. آزمون MTT جهت بررسی سمیت سلولی سیلیمارین روی سلول­ها درغلظت ­های مختلف در زمان­های 24 و 72 ساعت انجام شد. سپس، سلول‌ها در یکی از 8 گروه 1: شاهد منفی؛ 2: تیمار شده با 10 میکرومول بر لیتر سیلیمارین در محیط معمول، 3: تیمار شده با 20 میکرومول بر لیتر سیلیمارین در محیط معمول، 4: تیمار شده با 100 میکرومول بر لیتر استرادیول در محیط معمول، 5: شاهد مثبت، 6: تیمار شده با 10 میکرومول بر لیتر سیلیمارین در محیط تمایزی، 7: تیمار شده با 20 میکرومول بر لیتر سیلیمارین در محیط تمایزی، 8: تیمار شده با 100 میکرومول بر لیتر در محیط تمایزی، به‌مدت 21 روز کشت شدند.جهت تعیین تمایز استئوژنیک سلول­ها، با استفاده از رنگ آمیزی آلیزارین رد رسوب یون هیدروکسی آپاتیت بررسی شد و همچنین میزان ترشح آنزیم ALP در گروه های مورد مطالعه اندازه گیری شد. نتایج: مقایسه میانگین جذب نوری سلول­ها در غلظت­های مختلف بین زمان­های 24 و 72 ساعت پس از تیمار نشان داد که میانگین جذب نوری سلول­ها در غلظت صفر سیلیمارین، 72 ساعت پس از تیمار نسبت به زمان 24 ساعت پس از تیمار کاهش نشان داد(05/0P<) اما در سایر غلظت­ها تفاوت معنی دار مشاهده نشد(05/0P>). بررسی میزان ترشح آنزیم ALP ، 21 روز پس از تیمار در گروه­های مورد مطالعه نشان داد که در مجموع بیشترین میزان ترشح آنزیم در گروه 8 بود (05/0P≤). کمترین میزان ترشح آنزیم در گروه 1 (کنترل منفی) و پس از آن به ترتیب در گروه­ 2 و گروه 3 مشاهد شد(05/0P<). بین گروه­های 4، 5 و 6 تفاوت معنی دار مشاهده نشد(P>0.05). بر اساس نتایج حاصل از رنگ آمیزی آلیزارین رد، رسوب املاح کلسیم در تمام گروه­های مربوط به محیط کشت تمایزی مشاهده شد که به ترتیب در گروه­های 8، 7، 6 و 5 افزایش یافت. در گروه­های کشت شده در محیط معمول، در گروه 1 رسوبی مشاهده نشد و میزان رسوب به ترتیب در گروه­های 2، 3 و 4 افزایش نشان داد. در مجموع میزان رسوب در گروه­های محیط تمایزی بیشتر از محیط معمول بود. نتیجه گیری: سیلیمارین در غلظت­های مورد بررسی روی سلولهای بنیادی مزانشیمی مشتق از مغز استخوان جنین گوسفند، اثر سمیت ندارد و همچنین در غلظت­های مورد بررسی باعث افزایش تمایز سلول­ها به رده ی استخوانساز می شود که این افزایش وابسته به غلظت است. از این رو، به نظر می رسد با مطالعات بیشتر و شناسایی مسیرهای مولکولی اثر گذاری سیلیمارین، می­توان از آن در سلول درمانی جهت ترمیم ضایعات استخوانی استفاده کرد.

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  • Journal of Cell and Tissue
  • Jun 22, 2022
  • I Morovati + 3
Open Access
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Silymarin Effects on Ovine Fetal Bone Marrow-Derived Mesenchymal Stem Cells Differentiation into Osteogenic Lineage

Silymarin Effects on Ovine Fetal Bone Marrow-Derived Mesenchymal Stem Cells Differentiation into Osteogenic Lineage

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  • Cell and Tissue Journal
  • Jun 22, 2022
  • I Morovati + 3
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Lipid nanoparticle-mediated silencing of osteogenic suppressor GNAS leads to osteogenic differentiation of mesenchymal stem cells in vivo

Approved drugs for the treatment of osteoporosis can prevent further bone loss but do not stimulate bone formation. Approaches that improve bone density in metabolic diseases are needed. Therapies that take advantage of the ability of mesenchymal stem cells (MSCs) to differentiate into various osteogenic lineages to treat bone disorders are of particular interest. Here we examine the ability of small interfering RNA (siRNA) to enhance osteoblast differentiation and bone formation by silencing the negative suppressor gene GNAS in bone MSCs. Using clinically validated lipid nanoparticle (LNP) siRNA delivery systems, we show that silencing the suppressor gene GNAS invitro in MSCs leads to molecular and phenotypic changes similar to those seen in osteoblasts. Further, we demonstrate that these LNP-siRNAs can transfect a large proportion of mice MSCs in the compact bone following intravenous injection. Transfection of MSCs in various animal models led to silencing of GNAS and enhanced differentiation of MSCs into osteoblasts. These data demonstrate the potential for LNP delivery of siRNA to enhance the differentiation of MSCs into osteoblasts, and suggests that they are a promising approach for the treatment of osteoporosis and other bone diseases.

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  • Molecular Therapy
  • Jun 22, 2022
  • Genc Basha + 7
Open Access
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Aged Callus Skeletal Stem/Progenitor Cells Contain an Inflammatory Osteogenic Population With Increased IRF and NF-κB Pathways and Reduced Osteogenic Potential.

Skeletal stem/progenitor cells (SSPCs) are critical for fracture repair by providing osteo-chondro precursors in the callus, which is impaired in aging. However, the molecular signatures of callus SSPCs during aging are not known. Herein, we performed single-cell RNA sequencing on 11,957 CD45-CD31-Ter119- SSPCs isolated from young and aged mouse calluses. Combining unsupervised clustering, putative makers, and DEGs/pathway analyses, major SSPC clusters were annotated as osteogenic, proliferating, and adipogenic populations. The proliferating cluster had a differentiating potential into osteogenic and adipogenic lineages by trajectory analysis. The osteoblastic/adipogenic/proliferating potential of individual clusters was further evidenced by elevated expression of genes related to osteoblasts, adipocytes, or proliferation. The osteogenic cluster was sub-clustered into house-keeping and inflammatory osteogenic populations that were decreased and increased in aged callus, respectively. The majority of master regulators for the inflammatory osteogenic population belong to IRF and NF-κB families, which was confirmed by immunostaining, RT-qPCR, and Western blot analysis. Furthermore, cells in the inflammatory osteogenic sub-cluster had reduced osteoblast differentiation capacity. In conclusion, we identified 3 major clusters in callus SSPCs, confirming their heterogeneity and, importantly, increased IRF/NF-κB-mediated inflammatory osteogenic population with decreased osteogenic potential in aged cells.

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  • Frontiers in molecular biosciences
  • Jun 9, 2022
  • X Lin + 6
Open Access
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FACS-isolation and Culture of Fibro-Adipogenic Progenitors and Muscle Stem Cells from Unperturbed and Injured Mouse Skeletal Muscle.

Fibro-adipogenic progenitor cells (FAPs) are a population of skeletal muscle-resident mesenchymal stromal cells (MSCs) capable of differentiating along fibrogenic, adipogenic, osteogenic, or chondrogenic lineage. Together with muscle stem cells (MuSCs), FAPs play a critical role in muscle homeostasis, repair, and regeneration, while actively maintaining and remodeling the extracellular matrix (ECM). In pathological conditions, such as chronic damage and muscular dystrophies, FAPs undergo aberrant activation and differentiate into collagen-producing fibroblasts and adipocytes, leading to fibrosis and intramuscular fatty infiltration. Thus, FAPs play a dual role in muscle regeneration, either by sustaining MuSC turnover and promoting tissue repair or contributing to fibrotic scar formation and ectopic fat infiltrates, which compromise the integrity and function of the skeletal muscle tissue. A proper purification of FAPs and MuSCs is a prerequisite for understanding the biological role of these cells in physiological as well as in pathological conditions. Here, we describe a standardized method for the simultaneous isolation of FAPs and MuSCs from limb muscles of adult mice using fluorescence-activated cell sorting (FACS). The protocol describes in detail the mechanical and enzymatic dissociation of mononucleated cells from whole limb muscles and injured tibialis anterior (TA) muscles. FAPs and MuSCs are subsequently isolated using a semi-automated cell sorter to obtain pure cell populations. We additionally describe an optimized method for culturing quiescent and activated FAPs and MuSCs, either alone or in coculture conditions.

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  • Journal of Visualized Experiments
  • Jun 8, 2022
  • Giulia Riparini + 2
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Pulsed Electric Fields for Valorization of Platelets with No Therapeutic Value towards a High Biomedical Potential Product—A Proof of Concept

Nowadays, the standard media used in clinical-scale mesenchymal stem cell (MSC) production to supply hundreds of clinical trials uses animal-derived components as supplements, which raises several health concerns. Consequently, the development of xeno-free media supplements has emerged. In the current study, the effect of pulse electric field (PEF) application to platelet concentrates (PC) with no therapeutic value for producing platelet releasates (PR) able to sustain the ability of bone marrow-MSCs (BM-MSCs) to self-renew and differentiate was tested. It was demonstrated that PEF application to PC induces platelet activation and growth factor (GF) release, namely PDGF, FGF, IGF, and TGF-β. The highest GF release was observed for TGF-β, achieving similar levels to those attained in platelet lysates (PL). BM-MSCs expanded in the presence of PR obtained by the application of PEF (7 pulses of 10 and 12.5 kV/cm) to PC (PR PEF) retained the characteristic MSC cell-surface markers, and the ability to proliferate and differentiate into osteogenic, adipogenic, and chondrogenic lineages. In this study, evidence is provided that PR PEF represents a suitable alternative to fetal bovine serum (FBS) for use in MSC production.

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  • Applied Sciences
  • Jun 7, 2022
  • Daniela Salvador + 6
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Effects of Different Basal Cell Culture Media upon the Osteogenic Response of hMSCs Evaluated by 99mTc-HDP Labeling.

The osteogenic differentiation of mesenchymal stem cells is now a standard procedure in modern bone tissue engineering. As this is a promising field for future clinical applications, many cell culture media exist to promote osteogenic differentiation. Prior to differentiation, cells must be expanded to obtain sufficient numbers for experiments. Little evidence is available regarding the optimal media combination for expansion and differentiation to maximize the osteogenic response. Therefore, human BM-MSCs (n = 6) were expanded in parallel in DMEM (Dulbecco’s Modified Eagle Medium) LG (Low Glucose) and α-MEM (Minimum Essential Media alpha-modification), followed by simultaneous monolayer differentiation toward the osteogenic lineage in: 1. DMEM LG (Low Glucose), 2. DMEM HG (High Glucose), 3. α-MEM, 4. “Bernese medium”, and 5. “Verfaillie medium”, with a corresponding negative control (total 20 groups). As a marker for osteogenic differentiation, hydroxyapatite was accessed using radioactive 99mTc-HDP labeling and quantitative alizarin red staining. The results indicate that all media except “Bernese medium” are suitable for osteogenic differentiation, while there was evidence that DMEM LG is partly superior when used for expansion and differentiation of BM-hMSCs. Using “Verfaillie medium” after DMEM LG expansion led to the highest grade of osteogenic differentiation. Nevertheless, the difference was not significant. Therefore, we recommend using DMEM LG for robust osteogenic differentiation, as it is highly suitable for that purpose, economical compared to other media, and requires little preparation time.

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  • International Journal of Molecular Sciences
  • Jun 3, 2022
  • Tobias Grossner + 3
Open Access
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Mesenchymal stem cell culture in aligned porous hydroxyapatite scaffolds using a multiwell plate bioreactor for bone tissue engineering

Regeneration of bone lost by trauma, diseases and aging, and restoration of its load-bearing function are major clinical challenges. Hydroxyapatite (HA) is a clinically proven scaffold material for bone grafting, but the random-pore structure limits the homing of the cells inside the graft and the bone regeneration progresses with the resorption of the graft material. This work is based on the hypothesis that aligned through pores in the graft will lead to a faster healing by homing the local cells inside and provide a better environment for new bone formation through the graft structure. The investigation was done using aligned porous HA scaffolds seeded with human Wharton's jelly-derived mesenchymal stem cells (hWJ-MSCs) and cultured in a multiwell format bioreactor setup. The cell adhesion was studied by microscopy, cell proliferation was evaluated by Alamar blue assay and osteogenic differentiation was confirmed by biochemical and molecular assays. The results indicate that the hWJ-MSCs infiltrated through the aligned porous network of the scaffold, proliferated well when cultured in the expansion medium, and differentiated into osteogenic lineage when cultured in the differentiation medium.

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  • MedComm – Future Medicine
  • Jun 1, 2022
  • Gayathry G + 6
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Autogenous Bone is Still the Gold Standard of Graft Materials in 2022.

I read with interest the recent editorial entitled “Is Autogenous Bone Grafting Still the “Gold Standard” in Oral Bone Grafting?”.1 As the authors pointed out, I wrote a similar editorial in 2010 and concluded that indeed autogenous bone was the gold standard of graft materials.2 However, the authors have challenged my conclusion. To support their opinion, they referenced two publications on sinus bone grafting evaluating histomorphometric results and implant survival.3,4 They claimed these studies found that “bone substitutes either performed in a like manner or superior to autogenous bone alone”. However, the histomorphometric analysis showed autogenous bone resulted in the highest amount of new bone formation compared to bone substitutes, but bone substitutes seem to be a good alternative.3 The systematic review on sinus bone grafting concluded implant survival rates using bone substitutes were as effective as autogenous bone when used alone or in combination with autogenous bone.4 Although the implant survival was lower with 100% autogenous bone (87.7%) compared to autograft with bone substitutes (94.9%) or bone substitutes alone (96%), the data clearly shows that the majority of machined implants were placed in sinuses grafted with autogenous bone. The review found rough surface implants had superior survival rates compared to machined implants (96% vs. 85.6%), so this explains the lower implant survival rate in autogenous grafts. Pjetursson et al5 performed a systematic review evaluated the grafting of pneumatized sinuses that had 6.0 mm or less residual bone height. Focusing on outcomes using only rough surface implants they found high implant survival rates for all types of grafts (> 96%). However, rough implants placed in particulate autogenous bone had a significantly higher estimated three-year survival (99.8%). There are limitations in using a secondary outcome measure, such as implant survival, to evaluate graft success. The assumption is that if the implant survived the graft was successful. The influence of the amount of native bone supporting the implant is unknown in some studies unless the residual bone limit was defined, such as in the Pjetursson et al study.Another important distinction is the relationship of the augmentation to the osseous defect. Bone defects inside the bony contour, such as sockets and the maxillary sinus floor, are much easier and predictable to reconstruct than horizontal and vertical augmentation outside the bone contour. Intrabony defects have a greater number of bone walls with a higher regenerative capacity. There is also greater ease in achieving soft tissue coverage, space maintenance, graft stability and protection. As such bone substitutes perform well in treating these types of contained defects. However, horizontal and vertical bone augmentation outside the bone contour is more biologically challenging. The greater the distance from the native bone, the more difficult it may become for vascular ingrowth, cell migration and bone formation at the outer limits of the graft. As such a graft material with greater regenerative capacity may be needed to obtain the required bone gains. Autogenous bone has superior biologic qualities compared to osteoconductive bone substitutes. It is the only bone graft material that fulfills all three aspects of the tissue engineering triad - osteogenesis, osteoinduction and osteoconduction. Autogenous bone contains osteocompetent cells and osteoconductive growth factors that can positively influence bone formation. Although cancellous bone contains a higher amount of osteoblasts and mesenchymal stem cells, cortical bone grafts contain cells that have the ability to proliferate and differentiate into the osteogenic lineage, suggesting that these cells can also contribute to bone regeneration following transplantation.6 Research has found cortical bone chips contain over 40 different growth factors that can modulate the cellular aspects of bone regeneration.7 The healing time of autogenous bone grafts also is shorter than bone substitutes.8Rather than focus on implant survival, it would be more clinically relevant to evaluate horizontal and vertical bone gains produced by different graft materials. When reviewing studies measuring bone gains it becomes evident that autogenous bone is needed for larger bone augmentations and vertical gains.9,10,11,12 While I agree with my colleagues that bone substitutes may be used effectively in many clinical situations, I still maintain that autogenous bone is the gold standard of graft materials. This does not mean it is the first choice option, but it does offer biologic advantages unmatched by bone substitutes. The use of a local donor site and bone scrapers can decrease the morbidity of bone harvest. In addition, some donor sites for block grafting, such as the mandibular ramus, have a lower incidence of postoperative complications.13 Surgeons should weigh the advantages and disadvantages of each material and select an approach that has the highest likelihood of clinical success.14

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  • Journal of Oral Implantology
  • Jun 1, 2022
  • Craig M Misch
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Autophagy reprogramming stem cell pluripotency and multiple-lineage differentiation.

The cellular process responsible for the degradation of cytosolic proteins and subcellular organelles in lysosomes was termed "autophagy." This process occurs at a basal level in most tissues as part of tissue homeostasis that redounds to the regular turnover of components inside cytoplasm. The breakthrough in the autophagy field is the identification of key players in the autophagy pathway, compounded under the name "autophagy-related genes" (ATG) encoding for autophagy effector proteins. Generally, the function of autophagy can be classified into two divisions: intracellular clearance of defective macromolecules and organelles and generation of degradation products. Therapeutic strategies using stem cell-based approach come as a promising therapy and develop rapidly recently as stem cells have high self-renewability and differentiation capability as known as mesenchymal stem cells (MSCs). They are defined as adherent fibroblast-like population with the abilities to self-renew and multi-lineage differentiate into osteogenic, adipogenic, and chondrogenic lineage cells. To date, they are the most extensively applied adult stem cells in clinical trials. The properties of MSCs, such as immunomodulation, neuroprotection, and tissue repair pertaining to cell differentiation, processes to replace lost, or damaged cells, for aiding cell repair and revival. Autophagy has been viewed as a remarkable mechanism for maintaining homeostasis, ensuring the adequate function and survival of long-lived stem cells. In addition, authophagy also plays a remarkable role in protecting stem cells against cellular stress when the stem cell regenerative capacity is harmed in aging and cellular degeneration. Understanding the under-explored mechanisms of MSC actions and expanding the spectrum of their clinical applications may improve the utility of the MSC-based therapeutic approach in the future.

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  • Journal of the Chinese Medical Association
  • Jun 1, 2022
  • Yi-Ping Yang + 12
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Cell energy metabolism and bone formation

Energy metabolism plays an important role in cell and tissue ability to effectively function, maintain homeostasis, and perform repair. Yet, the role of energy metabolism in skeletal tissues in general and in bone, in particular, remains understudied. We, here, review the aspects of cell energy metabolism relevant to bone tissue, such as: i) availability of substrates and oxygen; ii) metabolism regulatory mechanisms most active in bone tissue, e.g. HIF and BMP; iii) crosstalk of cell bioenergetics with other cell functions, e.g. proliferation and differentiation; iv) role of glycolysis and mitochondrial oxidative phosphorylation in osteogenic lineage; and v) most significant changes in bone energy metabolism observed in aging and other pathologies. In addition, we review available methods to study energy metabolism on a subcellular, cellular, tissue, and live animal levels.

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  • Bone Reports
  • May 27, 2022
  • Rubens Sautchuk + 1
Open Access
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Superior processability of Antheraea mylitta silk with cryo-milling: Performance in bone tissue regeneration

Non-mulberry silk polymers have a promising future in biomedical applications. However, the dissolution of non-mulberry silk fiber is a still challenge and this poor processability has limited the use of this material. Here, we report a unique protocol to process the Antheraea mylitta (AM) silk fiber. We have shown that the cryo-milling of silk fiber reduces the beta sheet content by more than 10% and results in an SF powder that completely dissolves in routine solvents like trifluoroacetic acid (TFA) within few hours to form highly concentrated solutions (~20 wt%). Further, these solutions can be processed using conventional processing techniques such as electrospinning to form 3D scaffolds. Bombyx mori (BM) silk was used as a control sample in the study. In-vitro studies were also performed to monitor cell adhesion and proliferation and hMSCs differentiation into osteogenic lineage. Finally, the osteogenic potential of the scaffolds was also evaluated by a 4-week implantation study in rat calvarial model. The in-vitro and in-vivo results show that the processing techniques do not affect the biocompatibility of the material and the AM scaffolds support bone regeneration. Our results, thus, show that cryo-milling facilitates enhanced processability of non-mulberry silk and therefore expands its potential in biomedical applications.

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  • International Journal of Biological Macromolecules
  • May 21, 2022
  • Nimisha Parekh + 6
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Single-cell RNA sequencing unravels heterogeneity of skeletal progenitors and cell-cell interactions underlying the bone repair process.

IntroductionActivation of skeletal progenitors upon tissue injury and the subsequent cell fate specification are tightly coordinated in the bone repair process. Although known osteoimmunological signaling networks play important roles in the microenvironment of the bone defect sites, the molecular mechanism underlying the bone repair process has not been fully understood.MethodsTo better understand the behavior of the skeletal progenitors and the heterogeneity of the cells during bone repair at the microenvironmental level, we performed a combinatorial analysis consisting of lineage tracing for skeletal progenitors using the Sox9-CreERT2;R26RtdTomato mouse line followed by single-cell RNA sequencing (scRNA-seq) analysis using a mouse model of calvarial bone repair. To identify a therapeutic target for bone regeneration, further computational analysis was performed focusing on the identification of the cell–cell interactions, followed by pharmacological assessments with a critical-size calvarial bone defect mouse model.ResultsLineage tracing analysis showed that skeletal progenitors marked by Sox9 were activated upon bone injury and contributed to bone repair by differentiating into osteoblasts. The scRNA-seq analysis characterized heterogeneous cell populations at the bone defect sites; the computational analysis predicted a bifurcated lineage from skeletal progenitors toward osteogenic and adipogenic lineages. Chemokine C–C motif ligand 9 (Ccl9) was identified as a signaling molecule that regulates bone regeneration in the mouse model, possibly through the regulation of adipogenic differentiation at the bone defect site.ConclusionMultipotential skeletal progenitors and the direction of the cell differentiation were characterized at single cell resolution in a mouse bone repair model. The Ccl9 signaling pathway may be a key factor directing osteogenesis from the progenitors in the model and may be a therapeutic target for bone regeneration.

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  • Regenerative therapy
  • May 18, 2022
  • Mika Nakayama + 6
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3D bioprinting of multilayered scaffolds with spatially differentiated ADMSCs for rotator cuff tendon-to-bone interface regeneration

Regeneration of the gradient structure of the tendon-to-bone interface is still a significant clinical challenge. This study reports a novel therapeutic method combining three-dimensional (3D) bioprinting and melt electrospinning writing techniques to regenerate a functional tendon-to-bone interface. We generated biomimetic multilayered scaffolds with 3D-bioprinted pre-differentiated autologous adipose-derived mesenchymal stem cells (ADMSC), which recapitulated compositional and cellular structures of the interface. The hydrogel-based bioinks offered high cell viability and proliferative capability for rabbit ADMSCs. The hydrogels with pre-differentiated (into tenogenic, chondrogenic, and osteogenic lineages) or undifferentiated rabbit ADMSCs were 3D-bioprinted into zonal-specific constructs to mimic the structure of the tendon-to-bone interface. These scaffolds were tested in a rabbit rotator cuff injury model and the histological, radiological, and biomechanical changes were analyzed. The in vivo studies demonstrated that the scaffold with spatially differentiated autologous ADMSCs had a superior histological score and improved collagen organization when compared to acellular scaffolds and similar T2 value as the normal interface tissue. The biomechanical characterization demonstrated that the application of multilayered scaffolds improved the biomechanical properties of the tendon-to-bone interface at 12 weeks after rotator cuff reconstruction surgery, but the incorporation of autologous ADMSCs within the multilayered scaffolds showed a limited contribution. Thus, our work provides a 3D-bioprinting-based strategy with the application of autologous ADMSCs to reconstruct massive rotator cuff tendon tears.

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  • Applied Materials Today
  • May 11, 2022
  • Xiping Jiang + 12
Open Access
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LncMIR181A1HG is a novel chromatin-bound epigenetic suppressor of early stage osteogenic lineage commitment

Bone formation requires osteogenic differentiation of multipotent mesenchymal stromal cells (MSCs) and lineage progression of committed osteoblast precursors. Osteogenic phenotype commitment is epigenetically controlled by genomic (chromatin) and non-genomic (non-coding RNA) mechanisms. Control of osteogenesis by long non-coding RNAs remains a largely unexplored molecular frontier. Here, we performed comprehensive transcriptome analysis at early stages of osteogenic cell fate determination in human MSCs, focusing on expression of lncRNAs. We identified a chromatin-bound lncRNA (MIR181A1HG) that is highly expressed in self-renewing MSCs. MIR181A1HG is down-regulated when MSCs become osteogenic lineage committed and is retained during adipogenic differentiation, suggesting lineage-related molecular functions. Consistent with a key role in human MSC proliferation and survival, we demonstrate that knockdown of MIR181A1HG in the absence of osteogenic stimuli impedes cell cycle progression. Loss of MIR181A1HG enhances differentiation into osteo-chondroprogenitors that produce multiple extracellular matrix proteins. RNA-seq analysis shows that loss of chromatin-bound MIR181A1HG alters expression and BMP2 responsiveness of skeletal gene networks (e.g., SOX5 and DLX5). We propose that MIR181A1HG is a novel epigenetic regulator of early stages of mesenchymal lineage commitment towards osteo-chondroprogenitors. This discovery permits consideration of MIR181A1HG and its associated regulatory pathways as targets for promoting new bone formation in skeletal disorders.

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  • Scientific Reports
  • May 11, 2022
  • Coralee E Tye + 10
Open Access
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Temporomandibular Joint Fibrocartilage Contains CD105 Positive Mouse Mesenchymal Stem/Progenitor Cells with Increased Chondrogenic Potential.

A specific type of mesenchymal stem/progenitor cells (MSPCs), CD105+ is reported to aid in cartilage regeneration through TGF-β/Smad2-signalling. The purpose of this study was to identify and characterize CD105+ MSPCs in temporomandibular joint (TMJ) cartilage. MSPCs were isolated from mouse TMJ condyle explants and evaluated for their clonogenicity and pluripotential abilities. MSPC were examined for CD105 antigen using immunohistochemistry and flow cytometry. Immunohistochemistry revealed presence of CD105+ MSPCs in the proliferative zone of condyle's cartilage. Only 0.2% of isolated MSPCs exhibited CD105, along with the stem cell surface markers CD44 and Sca-1. In CD105+ MSPCs, intracellular immunostaining revealed significantly higher (p < 0.05) protein levels of collagen type 1, 2, proteoglycan 4. Ability for chondrogenic differentiation was found to be significantly higher (p < 0.05) after 4weeks compared to CD105- cells, using alcian blue staining. CD105+ cells were found to resemble an early MSPC subgroup with significantly higher gene expression of biglycan, proteoglycan 4, collagen type 2, Gli2, Sox5 (p < 0.001) and Sox9 (p < 0.05). In contrast, significantly lower levels of Runx2 (p < 0.05), Osterix, Trps1, Col10a1 (p < 0.01), Ihh (p < 0.001) related to chondrocyte senescence and commitment to osteogenic lineage, were observed compared to CD105- cells. The study showed the existence of a CD105+ MSPC subgroup within TMJ fibrocartilage that may be activated to aid in fibrocartilage repair.

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  • Journal of maxillofacial and oral surgery
  • May 9, 2022
  • Janis R Thamm + 5
Open Access
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A comprehensive analysis of the circRNA-miRNA-mRNA network in osteocyte-like cell associated with Mycobacterium leprae infection.

BackgroundBone formation and loss are the characteristic clinical manifestations of leprosy, but the mechanisms underlying the bone remodeling with Mycobacterium leprae (M. leprae) infection are unclear.Methodology/Principal findingsOsteocytes may have a role through regulating the differentiation of osteogenic lineages. To investigate osteocyte-related mechanisms in leprosy, we treated osteocyte-like cell with N-glycosylated muramyl dipeptide (N.g MDP). RNA-seq analysis showed 724 differentially expressed messenger RNAs (mRNAs) and 724 differentially expressed circular RNA (circRNAs). Of these, we filtered through eight osteogenic-related differentially expressed genes, according to the characteristic of competing endogenous RNA, PubMed databases, and bioinformatic analysis, including TargetScan, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes. Based on these results, we built a circRNA–microRNA (miRNA)–mRNA triple network. Quantitative reverse-transcription polymerase chain reaction and western blots analyses confirmed decreased Clock expression in osteocyte-like cell, while increased in bone mesenchymal stem cells (BMSCs), implicating a crucial factor in osteogenic differentiation. Immunohistochemistry showed obviously increased expression of CLOCK protein in BMSCs and osteoblasts in N.g MDP–treated mice, but decreased expression in osteocytes.Conclusions/SignificanceThis analytical method provided a basis for the relationship between N.g MDP and remodeling in osteocytes, and the circRNA–miRNA–mRNA triple network may offer a new target for leprosy therapeutics.

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  • PLOS Neglected Tropical Diseases
  • May 2, 2022
  • Zheng-Rong Gao + 10
Open Access
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Comprehensive Characterization of Biological Properties of Human Urine‐Derived Stem Cells

Mesenchymal (stromal) stem cells (MSCs) represent a unique tool in the field of regenerative medicine. However, their harvesting often requires invasive medical procedures. Urine‐derived stem cells (UDSCs) display similar properties to MSCs, and their sampling and further processing is non‐invasive for the donors. Here, we offer a comprehensive analysis of their biological properties. The goal of this study was to analyze their morphology, stemness, differentiation potential and cytokine profile. We have successfully isolated UDSCs from 25 urine samples. First colonies emerged up to 9 days after the initial seeding. Cell doubling time was 45 ± 0.24 SD, and when seeded at the density of 100 cells/cm2, they formed 42 ± 6.5 SD colonies within 10 days. Morphological analyzes revealed that two distinct types of the cell populations have been present. The first type had a rice‐grain shape and the second one was characterized by a polyhedral shape. All examined UDSCs expressed typical MSC‐like surface markers, CD73, CD90 and CD105. Moreover, conditioned media from UDSCs were harvested, and cytokine profile has been evaluated showing a significantly higher secretory rate of IL‐8, IL‐6 and chemokines MCP‐1 and GM‐CSF. We have also successfully induced human UDSCs into chondrogenic, osteogenic and myogenic cell lineages. Our findings indicate that UDSCs might have immense potential in the regeneration of the damaged tissues.

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  • The FASEB Journal
  • May 1, 2022
  • Lubos Danisovic + 8
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