Human Bone Marrow-derived Mesenchymal Cells Research Articles

Ca2+/Calmodulin-dependent Protein Kinase II Inhibitor KN-93 Enhances Chondrogenesis of Bone Marrow Mesenchymal Stem Cells and Delays Chondrogenic Hypertrophy.

Cartilage tissue engineering has been popularly applied in the treatment of articular cartilage defect because it is more effective in generating functional engineered cartilage than traditional methods. Although the chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells (BM-MSCs) is well established, it is often accompanied by undesired hypertrophy. Ca2+/calmodulin-dependent protein kinase II (CaMKII) is a crucial mediator in the ion channel pathway which is known to be involved in chondrogenic hypertrophy. Therefore, this study aimed to reduce the hypertrophy of BM-MSCs by inhibiting CaMKII activation. BM-MSCs were cultured in three-dimensional (3D) scaffold under chondrogenic induction with and without CaMKII inhibitor, KN-93. After cultivation, markers of chondrogenesis and hypertrophy were investigated. KN-93 at a concentration of 2.0 μM had no effect on the viability of BM-MSCs, while the activation of CaMKII was suppressed. A long period of KN-93 treatment significantly up-regulated the expression of SRY-box transcription factor 9 and aggrecan on day 28 compared to untreated BM-MSCs. Furthermore, KN-93 treatment significantly down-regulated the expression of RUNX family transcription factor 2 and collagen type X alpha 1 chain on days 21 and 28. Immunohistochemistry showed increased expression of aggrecan and type II collagen while the expression of type X collagen was reduced. A CaMKII inhibitor, KN-93 is able to enhance chondrogenesis of BM-MSCs and suppress chondrogenic hypertrophy, suggesting its potential applicability in cartilage tissue engineering.

Evaluating material-driven regeneration in a tissue engineered human in vitro bone defect model.

Advanced in vitro human bone defect models can contribute to the evaluation of materials for in situ bone regeneration, addressing both translational and ethical concerns regarding animal models. In this study, we attempted to develop such a model to study material-driven regeneration, using a tissue engineering approach. By co-culturing human umbilical vein endothelial cells (HUVECs) with human bone marrow-derived mesenchymal stromal cells (hBMSCs) on silk fibroin scaffolds with in vitro critically sized defects, the growth of vascular-like networks and three-dimensional bone-like tissue was facilitated. After a model build-up phase of 28days, materials were artificially implanted and HUVEC and hBMSC migration, cell-material interactions, and osteoinduction were evaluated 14days after implantation. The materials physiologically relevant for bone regeneration included a platelet gel as blood clot mimic, cartilage spheres as soft callus mimics, and a fibrin gel as control. Although the in vitro model was limited in the evaluation of immune responses, hallmarks of physiological bone regeneration were observed in vitro. These included the endothelial cell chemotaxis induced by the blood clot mimic and the mineralization of the soft callus mimic. Therefore, the present in vitro model could contribute to an improved pre-clinical evaluation of biomaterials while reducing the need for animal experiments.

Non-hypertrophic chondrogenesis of mesenchymal stem cells through mechano-hypoxia programing

Cartilage tissue engineering aims to generate functional replacements to treat cartilage defects from damage and osteoarthritis. Human bone marrow-derived mesenchymal stem cells (hBM-MSC) are a promising cell source for making cartilage, but current differentiation protocols require the supplementation of growth factors like TGF-β1 or −β3. This can lead to undesirable hypertrophic differentiation of hBM-MSC that progress to bone. We have found previously that exposing engineered human meniscus tissues to physiologically relevant conditions of the knee (mechanical loading and hypoxia; hence, mechano-hypoxia conditioning) increased the gene expression of hyaline cartilage markers, SOX9 and COL2A1, inhibited hypertrophic marker COL10A1, and promoted bulk mechanical property development. Adding further to this protocol, we hypothesize that combined mechano-hypoxia conditioning with TGF-β3 growth factor withdrawal will promote stable, non-hypertrophic chondrogenesis of hBM-MSC embedded in an HA-hydrogel. We found that the combined treatment upregulated many cartilage matrix- and development-related markers while suppressing many hypertrophic- and bone development-related markers. Tissue level assessments with biochemical assays, immunofluorescence, and histochemical staining confirmed the gene expression data. Further, mechanical property development in the dynamic compression treatment shows promise toward generating functional engineered cartilage through more optimized and longer culture conditions. In summary, this study introduced a novel protocol to differentiate hBM-MSC into stable, cartilage-forming cells.

Abstract 2341: Phenotypic and quality assessments of continuously expanded human bone marrow-derived mesenchymal stem cells from multiple donors

Abstract 2341: Phenotypic and quality assessments of continuously expanded human bone marrow-derived mesenchymal stem cells from multiple donors

Comparison of the Effects of Tissue Processing on the Physicochemical Properties of Bone Allografts.

Purpose: To address the hypothesis that the tissue processing methods of solvent dehydration and freeze-drying would differentially affect the physicochemical characteristics of four commercially available bone allografts and the adhesion and differentiation of human bone marrow-derived mesenchymal stromal cells (hBMSCs) on such substrates in vitro. Materials and Methods: The surface morphology, surface area, and elemental composition of four commercially available cancellous bone allografts were examined using SEM, Brunauer-Emmett-Teller (BET) gas adsorption, and inductively coupled plasma (ICP) analyses. SEM was also employed to compare the allograft surfaces to that of human bone exposed by in vitro osteoclastic resorption. The allografts were seeded with hBMSCs, and the number of adhered cells was assessed at 3 and 7 days. Alkaline phosphatase (ALP) activity was quantified as a measure of osteogenic differentiation after 21 days. Results: Marked differences were seen between the physicochemical characteristics of the solvent-dehydrated and freeze-dried allografts, as well as between their resulting bone microarchitectures and that of osteoclast-resorbed human bone. Increased hBMSC adhesion and differentiation were observed on the solvent-dehydrated allografts compared to freeze-dried allografts, which suggests a higher putative osteogenic potential. The latter was attributed to better preservation of the bone collagen microarchitecture integrity, which may provide not only a more complex substrate architecture, but also a more favorable microenvironment to allow nutrients and oxygen to flow to the adhered cells. Conclusion: Commercially available cancellous bone allografts significantly differ in their physicochemical characteristics, stemming from differences in tissue processing and sterilization methods undertaken by tissue banks. These differences impact the response of MSCs in vitro and may alter the biologic performance of the grafts in vivo. Therefore, it is important to consider these characteristics when choosing a bone substitute for clinical application, as the physicochemical properties of the grafts play a crucial role in their interactions with the biologic environment and subsequent incorporation into the native bone.

Periplaneta americana extract promotes osteoblast differentiation of human alveolar bone marrow mesenchymal stem cells.

This study aims to investigate the effects of Traditional Chinese medicine, Periplaneta americana extract (PAE), on osteoblast differentiation of human alveolar bone marrow-derived mesenchymal stem cells (hABMMSCs). Human alveolar bone marrow-derived mesenchymal stem cells were treated with different concentrations of PAE. Cell Counting Kit-8 (CCK-8) assay and transwell migration assay were conducted to evaluate cell proliferation and migration, respectively. Alkaline phosphatase (ALP) staining, ALP activity assay, and Alizarin red S staining were performed to detect osteogenesis in hABMMSCs. In addition, real-time quantitative polymerase chain reaction (RT-qPCR) and western blot (WB) assay were performed to evaluate expression levels of osteogenic markers. Finally, RNA sequencing analysis and WB were carried out to elucidate the underlying mechanism. A total of 0.1 mg/ml PAE promoted cell proliferation and migration. PAE also increased ALP activity and mineralized nodule formation of hABMMSCs. In addition, PAE upregulated the expression of osteogenesis-related genes (RUNX2, COL1A1, and BGLAP). RNA-sequencing analysis revealed that PAE activated the focal adhesion signaling pathway. Treatment with Defactinib, an inhibitor of FAK, attenuated the effects induced by PAE. PAE could enhance osteoblast differentiation of hABMMSCs through focal adhesion signaling pathway, suggesting a therapeutic potential for the alveolar bone defect.

Anti-Inflammatory Effect of Specialized Proresolving Lipid Mediators on Mesenchymal Stem Cells: An In Vitro Study

An interconnection between tissue inflammation and regeneration has been established through the regulation of defense and repair mechanisms within diseased dental tissue triggered by the release of immune-resolvent mediators. To better our understanding of the role of specific pro-resolving mediators (SPMs) in inflamed human bone marrow-derived mesenchymal stem cells (hBMMSCs), we studied the effects of Resolvin E1 (RvE1) and Maresin 1 (MaR1) in lipopoly-saccharide (LPS) stimulated hBMMSCs. The hBMMSCs were divided into five different groups, each of which was treated with or without SPMs. Group-1: negative control (no LPS stimulation), Group-2: positive control (LPS-stimulated), Group-3: RvE1 100 nM + 1 μg/mL LPS, Group-4: MaR1 100 nM + 1 µg/mL LPS, and Group-5: RvE1 100 nM + MaR1100 nM + 1 μg/mL LPS. Cell proliferation, apoptosis, migration, colony formation, Western blotting, cytokine array, and LC/MS analysis were all performed on each group to determine the impact of SPMs on inflammatory stem cells. According to our data, RvE1 plus MaR1 effectively reduced inflammation in hBMMSCs. In particular, IL-4, 1L-10, and TGF-β1 activation and downregulation of RANKL, TNF-α, and IFN-γ compared to groups receiving single SPM were shown to be significantly different (Group 3 and 4). In addition, the LC/MS analysis revealed the differentially regulated peptide's role in immunological pathways that define the cellular state against inflammation. Inflamed hBMMSCs treated with a combination of Resolvin E1 (RvE1) and Maresin 1 (MaR1) promoted the highest inflammatory resolution compared to the other groups; this finding suggests a potential new approach of treating bacterially induced dental infections.

Libanoridin Isolated from Corydalis heterocarpa Inhibits Adipogenic Differentiation of Bone Marrow-Derived Mesenchymal Stromal Cells.

Bone marrow adiposity is a complication in osteoporotic patients. It is a result of the imbalance between adipogenic and osteogenic differentiation of bone marrow cells. Phytochemicals can alleviate osteoporotic complications by hindering bone loss and decreasing bone marrow adiposity. Corydalis heterocarpa is a biennial halophyte with reported bioactivities, and it is a source of different coumarin derivatives. Libanoridin is a coumarin isolated from C. heterocarpa, and the effect of libanoridin on adipogenic differentiation of human bone marrow-derived mesenchymal stromal cells (hBM-MSCs) was evaluated in the present study. Cells were induced to undergo adipogenesis, and their intracellular lipid accumulation and expression of adipogenic markers were observed under libanoridin treatment. Results showed that 10 μM libanoridin-treated adipocytes accumulated 44.94% less lipid compared to untreated adipocytes. In addition, mRNA levels of PPARγ, C/EBPα, and SREBP1c were dose-dependently suppressed with libanoridin treatment, whereas only protein levels of PPARγ were decreased in the presence of libanoridin. Fluorescence staining of adipocytes also revealed that cells treated with 10 μM libanoridin expressed less PPARγ compared to untreated adipocytes. Protein levels of perilipin and leptin, markers of mature adipocytes, were also suppressed in adipocytes treated with 10 μM libanoridin. Analysis of MAPK phosphorylation levels showed that treatment with libanoridin inhibited the activation of p38 and JNK MAPKs observed by decreased levels of phosphorylated p38 and JNK protein. It was suggested that libanoridin inhibited adipogenic differentiation of hBM-MSCs via suppressing MAPK-mediated PPARγ signaling. Future studies revealing the anti-adipogenic effects of libanoridin in vivo and elucidating its action mechanism will pave the way for libanoridin to be utilized as a nutraceutical with anti-osteoporotic properties.

An osteogenic magnesium alloy with improved corrosion resistance, antibacterial, and mechanical properties for orthopedic applications.

The aim of this study was to develop a novel biodegradable magnesium (Mg) alloy for bone implant applications. We used scandium (Sc; 2wt%) and strontium (Sr; 2wt%) as alloying elements due to their high biocompatibility, antibacterial efficacy, osteogenesis, and protective effects against corrosion. In the present work, we also examined the effect of a heat treatment process on the properties of the Mg-Sc-Sr alloy. Alloys were manufactured using a metal casting process followed by heat treatment. The microstructure, corrosion, mechanical properties, antibacterial activity, and osteogenic activity of the alloy were assessed in vitro. The results showed that the incorporation of Sc and Sr elements controlled the corrosion, reduced the hydrogen generation, and enhanced mechanical properties. Furthermore, alloying with Sc and Sr demonstrated a significantly enhanced antibacterial activity and decreased biofilm formation compared to control Mg. Also, culturing Mg-Sc-Sr alloy with human bone marrow-derived mesenchymal stromal cells showed a high degree of biocompatibility (>90% live cells) and a significant increase in osteoblastic differentiation in vitro shown by Alizarin red staining and alkaline phosphatase activity. Based on these results, the Mg-Sc-Sr alloy heat-treated at 400°C displayed optimal mechanical properties, corrosion rate, antibacterial efficacy, and osteoinductivity. These characteristics make the Mg-Sc-Sr alloy a promising candidate for biodegradable orthopedic implants in the fixation of bone fractures such as bone plate-screws or intramedullary nails.

Simultaneous late-gadolinium enhancement and T1 mapping of fibrosis and a novel cell-based combination therapy in hypertensive mice

Fibrosis is a hallmark of chronic hypertension and disrupts the viability of human bone marrow-derived mesenchymal stromal cells (BM-MSCs) post-transplantation. This study thus, determined whether the anti-fibrotic drug, serelaxin (RLX), could enhance the therapeutic effects of BM-MSCs or BM-MSC-derived exosomes (BM-MSC-EXO) in hypertensive mice. Left ventricular (LV) fibrosis in particular was assessed using conventional histological staining and non-invasive cardiac magnetic resonance imaging (CMRI). CMRI was employed using a novel magnetisation prepared 2 rapid acquisition gradient echo (MP2RAGE) sequence to simultaneously perform late gadolinium enhancement imaging and T1 mapping. Adult male C57BL/6 mice were uninephrectomised, received deoxycorticosterone acetate and saline to drink (1 K/DOCA/salt) for 21 days, whilst control mice were given normal drinking water for the same time-period. On day 14 post-injury, subgroups of 1 K/DOCA/salt-hypertensive mice were treated with RLX alone or in combination with BM-MSCs or BM-MSC-EXO; or the mineralocorticoid receptor antagonist, spironolactone. At day 21 post-injury, LV and kidney histopathology was assessed, whilst LV fibrosis and function were additionally analysed by CMRI and echocardiography. 1 K/DOCA/salt-hypertensive mice developed kidney tubular injury, inflammation, fibrosis, and more moderate LV hypertrophy, fibrosis and diastolic dysfunction. RLX and BM-MSCs combined provided optimal protection against these pathologies and significantly reduced picrosirius red-stained organ fibrosis and MP2RAGE analysis of LV fibrosis. A significant correlation between MP2RAGE analysis and histologically-stained interstitial LV fibrosis was detected. It was concluded that the MP2RAGE sequence enhanced the non-invasive CMRI detection of LV fibrosis. Furthermore, combining RLX and BM-MSCs may represent a promising treatment option for hypertensive cardiorenal syndrome.

PEG Reinforced Scaffold Promotes Uniform Distribution of Human MSC-Created Cartilage Matrix.

Previously, we used a gelatin/hyaluronic acid (GH)-based scaffold to induce chondrogenic differentiation of human bone marrow-derived mesenchymal stromal cells (hBMSC). The results showed that hBMSCs underwent robust chondrogenesis and facilitated in vivo cartilage regeneration. However, it was noticed that the GH scaffolds display a compressive modulus that is markedly lower than native cartilage. In this study, we aimed to enhance the mechanical strength of GH scaffolds without significantly impairing their chondrosupportive property. Specifically, polyethylene glycol diacrylate (PEGDA) and photoinitiators were infiltrated into pre-formed hBMSC-laden GH scaffolds and then photo-crosslinked. Results showed that infiltration of PEG at the beginning of chondrogenesis significantly increased the deposition of glycosaminoglycans (GAGs) in the central area of the scaffold. To explore the mechanism, we compared the cell migration and proliferation in the margin and central areas of GH and PEG-infiltrated GH scaffolds (GH+PEG). Limited cell migration was noticed in both groups, but more proliferating cells were observed in GH than in GH+PEG. Lastly, the in vitro repairing study with bovine cartilage explants showed that PEG- impregnated scaffolds integrated well with host tissues. These results indicate that PEG-GH hybrid scaffolds, created through infiltrating PEG into pre-formed GH scaffolds, display good integration capacity and represent a new tool for the repair of chondral injury.

Binucleated human bone marrow-derived mesenchymal cells can be formed during neural-like differentiation with independence of any cell fusion events

Although it has been reported that bone marrow-derived cells (BMDCs) can transdifferentiate into neural cells, the findings are considered unlikely. It has been argued that the rapid neural transdifferentiation of BMDCs reported in culture studies is actually due to cytotoxic changes induced by the media. While transplantation studies indicated that BMDCs can form new neurons, it remains unclear whether the underlying mechanism is transdifferentiation or BMDCs-derived cell fusion with the existing neuronal cells. Cell fusion has been put forward to explain the presence of gene-marked binucleated neurons after gene-marked BMDCs transplantation. In the present study, we demostrated that human BMDCs can rapidly adopt a neural-like morphology through active neurite extension and binucleated human BMDCs can form with independence of any cell fusion events. We also showed that BMDCs neural-like differentiation involves the formation of intermediate cells which can then redifferentiate into neural-like cells, redifferentiate back to the mesenchymal fate or even repeatedly switch lineages without cell division. Furthermore, we have discovered that nuclei from intermediate cells rapidly move within the cell, adopting different morphologies and even forming binucleated cells. Therefore, our results provide a stronger basis for rejecting the idea that BMDCs neural transdifferentiation is merely an artefact.

Inhibition of the Menin-MLL1 Interaction in MLL-Rearranged Primary Mixed-Phenotype Acute Leukemia Samples Promotes Leukemic Differentiation

Mixed phenotype acute leukemias (MPALs) simultaneously present features typical for more than one hematological cell lineage. The rarity of such patients combined with the lack of well-defined diagnostic criteria have so far prevented the design of optimal therapy regimens resulting in the occurrence of lineage switch and therewith associated treatment failures. Rearrangements involving MLL gene (also known as KMT2A) occur in around 15% of MPAL patients (typically infants) and are associated with a very poor outcome. Moreover, several studies demonstrated dependency of MLL-rearranged (MLL-r) leukaemias on the interaction between N-terminus of MLL with Menin and its successful therapeutic targeting by small molecular weight compounds, that are currently validated in clinical trials. Here we investigated potential application of the Menin-MLL inhibitor SNDX-5613 in MLL/AF4 MPAL infants who presented with the bilineage B/Myeloid phenotype. Using a co-culture approach with human bone marrow-derived mesenchymal stromal cells (MSCs) and media optimized for growth of lymphoid and myeloid lineages, we were able to expand and monitor primary MLL-r patient material for more than three weeks. Primary material from two MPAL patients and their lymphoid and myeloid subpopulations were analyzed for their drug responses including immunophenotypic and global transcriptomic characterization. We observed a dramatic loss of cell numbers in response to SNDX-5613, with the IC50 value ranging from 80 to 800nM. Similar analyses of the sorted fractions revealed that the observed loss of cell numbers is mainly caused by the reduction of the lymphoid subfraction. Immunophenotypic analyses showed that surviving cells within both lymphoid and myeloid fractions are losing CD34 with simultaneous gain of the CD38 marker, suggesting cellular maturation. Treatment of the lymphoid fraction with SNDX-5613 caused a dose-dependent loss of the CD19+CD33+ cell population and a simultaneous gain of CD19+CD33-, and CD19-CD33+ cells. These data suggest the cellular origin of the myeloid fraction to be residing within the lymphoid counterpart. Interestingly, myeloid cells responded to Menin-MLL inhibition with increased surface expression of monocytic markers (e.g. CD11b, CD14, CD15), elevated RNA expression of genes involved in cell adhesion and morphological changes, indicating myeloid differentiation of the surviving cells. Moreover, transcriptomic analysis of the lymphoid subfraction revealed dose-dependent increase of the MS4A1 gene expression (encoding for CD20) suggesting simultaneous lymphoid maturation of the CD19+ proB-like blasts. In conclusion, our study shows that Menin-MLL inhibition drives both myeloid differentiation and lymphoid maturation in MLL-r MPALs. These myeloid data are consistent with observations of differentiation syndrome as an on-target effect in current clinical trials while the phenotypic changes in the lymphoid cells are new findings. Together, these data strongly suggests that MLL-r MPAL patients could benefit from the inclusion of the Menin-MLL1 inhibitors into their treatment regimens.

Isolation and In Vitro Chondrogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stromal Cells.

Bone marrow-derived mesenchymal stromal cells (BM-MSC) are widely studied in the field of cartilage regeneration due to their capacity to differentiate into chondrocytes under specific in vitro culture conditions. This chapter describes the isolation of MSC from bone marrow aspirate, their expansion in monolayer, and the chondrogenic differentiation in pellet culture.

Phosphate promotes osteogenic differentiation through non-canonical Wnt signaling pathway in human mesenchymal stem cells.

Phosphate is indispensable in osteogenesis and mineralization. However, mechanisms by which phosphate enhances osteogenic differentiation are not fully understood. In this study, we studied the effect of phosphate on osteogenic differentiation as well as signaling pathways induced by phosphate in the process. Induced human bone marrow-derived mesenchymal stem cells differentiation into osteoblasts by the change of media containing β-glycerophosphate (GP), 1mM inorganic phosphate, or 3mM inorganic phosphate (Pi). The differentiation of osteoblasts was verified by the expression of osteoblast differentiation markers and calcium deposition. RNA sequencing was performed to assess transcriptome in the early stage of osteogenic differentiation. Osteogenic differentiation and mineralization were promoted in the 3mM Pi group compared to those in the GP and 1mM Pi groups on day 7 of culture. RNA sequencing revealed that the gene expressions involved in osteogenesis and the components in the Wnt signaling pathway was increased in 3mM Pi group compared with those in the GP on day 7. Analysis with qPCR and Western blot suggested upregulation of components in the non-canonical Wnt signaling pathway, including WNT5b and phosphorylated-c-Jun in the 3mM Pi group on day 7. WNT11 mRNA expression was increased in the 2 induction groups on day 7. Inhibition of WNT5b by siRNA experiment attenuated the components in non-canonical Wnt signaling expression, including WNT5b, WNT11 and ROR2 mRNA expression and phosphorylated-c-Jun protein expression. In addition, osteogenic differentiation and mineralization were partly decreased in 3mM Pi group on day 7 by the inhibition of WNT5b. Pi promoted osteogenic differentiation through the up-regulation of the non-canonical Wnt signaling pathway, including WNT5b, WNT11, p-c-Jun/c-Jun, in the early stage of differentiation. These findings provide a new perspective into the association of Pi and the non-canonical Wnt signaling pathway during osteogenic differentiation.

Biocompatible Properties and Mineralization Potential of Premixed Calcium Silicate-Based Cements and Fast-Set Calcium Silicate-Based Cements on Human Bone Marrow-Derived Mesenchymal Stem Cells.

Premixed calcium silicate-based cements (CSCs) and fast-set CSCs were developed for the convenience of retrograde filling during endodontic microsurgery. The aim of this study was to analyze the biocompatible properties and mineralization potential of premixed CSCs, such as Endocem MTA Premixed (EM Premixed) and EndoSequence BC RRM putty (EndoSequence), and fast-set RetroMTA on human bone marrow-derived mesenchymal stem cells (BMSCs) compared to ProRoot MTA. Using CCK-8, a significantly higher proliferation of BMSCs occurred only in the EM Premixed group on days 2 and 4 (p < 0.05). On day 6, the ProRoot MTA group had significantly higher cell proliferation than the control group (p < 0.05). Regardless of the experimental materials, all groups had complete cell migration by day 4. Alizarin Red-S staining and alkaline phosphatase assay demonstrated higher mineralization potential of all CSCs similar to ProRoot MTA (p < 0.05). The EndoSequence group showed more upregulation of SMAD1 and OSX gene expression than the other experimental groups (p < 0.05), and all experimental cements upregulated osteogenic gene expression more than the control group (p < 0.05). Therefore, using premixed CSCs and fast-set CSCs as retrograde filling cements may facilitate satisfactory biological responses and comparable osteogenic potential to ProRoot MTA.

Dysregulated transforming growth factor-beta mediates early bone marrow dysfunction in diabetes

Diabetes affects select organs such as the eyes, kidney, heart, and brain. Our recent studies show that diabetes also enhances adipogenesis in the bone marrow and reduces the number of marrow-resident vascular regenerative stem cells. In the current study, we have performed a detailed spatio-temporal examination to identify the early changes that are induced by diabetes in the bone marrow. Here we show that short-term diabetes causes structural and molecular changes in the marrow, including enhanced adipogenesis in tibiae of mice, prior to stem cell depletion. This enhanced adipogenesis was associated with suppressed transforming growth factor-beta (TGFB) signaling. Using human bone marrow-derived mesenchymal progenitor cells, we show that TGFB pathway suppresses adipogenic differentiation through TGFB-activated kinase 1 (TAK1). These findings may inform the development of novel therapeutic targets for patients with diabetes to restore regenerative stem cell function.

Stem Cell-Secreted Allogeneic Elastin-Rich Matrix with Subsequent Decellularization for the Treatment of Critical Valve Diseases in the Young.

Critical valve diseases in infants have a very poor prognosis for survival. Particularly challenging is for the valve replacement to support somatic growth. From a valve regenerative standpoint, bio-scaffolds have been extensively investigated recently. While bio-scaffold valves facilitate acute valve functionality, their xenogeneic properties eventually induce a hostile immune response. Our goal was to investigate if a bio-scaffold valve could be deposited with tissues derived from allogeneic stem cells, with a specific dynamic culture protocol to enhance the extracellular matrix (ECM) constituents, with subsequent stem cell removal. Porcine small intestinal submucosa (PSIS) tubular-shaped bio-scaffold valves were seeded with human bone marrow-derived mesenchymal stem cells (hBMMSCs), cultured statically for 8 days, and then exposed to oscillatory fluid-induced shear stresses for two weeks. The valves were then safely decellularized to remove the hBMMSCs while retaining their secreted ECM. This de novo ECM was found to include significantly higher (p < 0.05) levels of elastin compared to the ECM produced by the hBMMSCs under standard rotisserie culture. The elastin-rich valves consisted of ~8% elastin compared to the ~10% elastin composition of native heart valves. Allogeneic elastin promotes chemotaxis thereby accelerating regeneration and can support somatic growth by rapidly integrating with the host following implantation. As a proof-of-concept of accelerated regeneration, we found that valve interstitial cells (VICs) secreted significantly more (p < 0.05) collagen on the elastin-rich matrix compared to the raw PSIS bio-scaffold.

Hyunganol II Exerts Antiadipogenic Properties via MAPK-Mediated Suppression of PPARγ Expression in Human Bone Marrow-Derived Mesenchymal Stromal Cells.

Bone marrow adiposity has been associated with several metabolic syndromes such as diabetes and osteoporosis. Imbalance in adipogenic and osteoblastogenic differentiation of human bone marrow mesenchymal stromal cells (hBM-MSCs) was suggested to be the cause of elevated bone marrow adiposity. There are several drugs, of both natural and synthetic origin, to treat bone loss. In this study, as a part of a recent trend to discover natural products with more biocompatibility and fewer side effects to treat bone loss, the effect of hyunganol II (HNG), a coumarin isolated from Corydalis heterocarpa, on hBM-MSC adipogenesis was investigated. Cells treated with HNG showed decreased lipid accumulation indicating a diminished adipocyte phenotype. Treatment with HNG also suppressed the mRNA and protein expressions of PPARγ, C/EBPα, and SREBP1c, and three adipogenic marker genes. Further analysis of MAPK signaling pathway exhibited that HNG treatment elevated ERK activation and suppressed the JNK-mediated cFos and cJun phosphorylation, which inhibits PPARγ transcriptional activity. Taken together, HNG treatment was shown to inhibit adipogenesis via suppressed PPARγ expression as a result of altered MAPK signaling. Therefore, it was suggested that HNG might prevent bone marrow adiposity by inhibiting hBM-MSC adipogenesis and can be utilized as a drug or nutraceutical with beneficial effects on bone. Thus, further studies should be conducted to analyze its effect in vivo.

Hydroxyapatite-collagen- carboxylic carbon quantum dot composite loaded with chrysin supported the proliferation and differentiation of human bone marrow derived mesenchymal stem cells

Developing a bioactive scaffold with biocompatible material is a substantial approach to bone regeneration and functional healing. Hydroxyapatite (HAP) is the main component in bone formation as an inorganic component and regeneration due to its osteoconductive properties. In this study, we prepared a scaffold material composed of HAP and collagen (COL) cross-linked via carboxylic carbon quantum dots (CCQD) with a chrysin (CRN) molecule. CRN is a flavonoid that has been shown to encourage the bone development of bone marrow-derived mesenchymal stem cells. It is loaded for enhancing bone regeneration and HAP’s growth ability. XRD, FT-IR, SEM, and TEM analysis have characterized the prepared composites for their crystalline nature, functional behavior, and morphological evaluations. The HAP has retained its original crystalline lattice confirmed from XRD analysis in the prepared composites. The addition of CRN molecule has decreased the length of HAP rods from ∼932 nm to ∼459 nm, as confirmed by TEM images. The increased particle sizes have been observed for the prepared composites. It reaches the maximum at 938.0 nm for the final HAP/COL/CCQD/CRN composite, which was confirmed by particle size analysis. The in-vitro CRN release behavior shows that the CRN molecule has controlled release up to 23% for 48 h. The biocompatibility of prepared material was investigated and confirmed on human bone marrow-derived mesenchymal stem cells (hBMSCs). This examination has proven that the prepared material is good for bone cell regeneration. The material may apply for bone regeneration applications after in-vivo and clinical investigations.