Progenitor Bone Marrow Stromal Cells Research Articles

Antiosteoporosis Effects, Pharmacokinetics, and Drug Delivery Systems of Icaritin: Advances and Prospects.

Osteoporosis is a systemic skeletal disorder affecting over 200 million people worldwide and contributes dramatically to global healthcare costs. Available anti-osteoporotic drug treatments including hormone replacement therapy, anabolic agents, and bisphosphonates often cause adverse events which limit their long-term use. Therefore, the application of natural products has been proposed as an alternative therapy strategy. Icaritin (ICT) is not only an enzyme-hydrolyzed product of icariin but also an intestinal metabolite of eight major flavonoids of the traditional Chinese medicinal plant Epimedium with extensive pharmacological activities, such as strengthening the kidney and reinforcing the bone. ICT displays several therapeutic effects, including osteoporosis prevention, neuroprotection, antitumor, cardiovascular protection, anti-inflammation, and immune-protective effect. ICT inhibits bone resorption activity of osteoclasts and stimulates osteogenic differentiation and maturation of bone marrow stromal progenitor cells and osteoblasts. As for the mechanisms of effect, ICT regulates relative activities of two transcription factors Runx2 and PPARγ, determines the differentiation of MSCs into osteoblasts, increases mRNA expression of OPG, and inhibits mRNA expression of RANKL. Poor water solubility, high lipophilicity, and unfavorable pharmacokinetic properties of ICT restrict its anti-osteoporotic effects, and novel drug delivery systems are explored to overcome intrinsic limitations of ICT. The paper focuses on osteogenic effects and mechanisms, pharmacokinetics and delivery systems of ICT, and highlights bone-targeting strategies to concentrate ICT on the ideal specific site of bone. ICT is a promising potential novel therapeutic agent for osteoporosis.

Changes in Bone Marrow Stromal Progenitor Cells in Patients with Hematoblastosis at the Onset of the Disease.

Multipotent mesenchymal stromal cells (MSC) are the key regulators of hematopoiesis. We studied changes in MSC characteristics in patients with myeloid leukemia and patients with lymphoproliferative diseases. MSC were obtained from the bone marrow of patients at the time of diagnostic puncture using a standard technique. Their proliferative potential and expression of genes associated with differentiation and regulation of hematopoiesis were studied. The total cell production of MSC in patients with leukemia at the onset of the disease did not differ from that in the group of healthy donors. The relative expression of the IL6, TGFb1 and TGFb2, PPARG genes was similar in all patients. The relative expression of the JAG1, LIF, IGF1, CSF1, IL1b, and IL1bR1 genes in MSC of patients with leukemia was enhanced and the relative expression of SDF1 was unchanged in comparison with MSC from donors. MSC from patients with leukemia were characterized by enhanced relative expression of PDGFRA and PDGFRB, and reduced expression of SOX9. Changes functions of the stromal microenvironment in patients with hemoblastoses attested to the role of stromal cells in the maintenance and spread of tumor cells.

Roles of MicroRNAs in Osteogenesis or Adipogenesis Differentiation of Bone Marrow Stromal Progenitor Cells.

Bone marrow stromal cells (BMSCs) are multipotent cells which can differentiate into chondrocytes, osteoblasts, and fat cells. Under pathological stress, reduced bone formation in favour of fat formation in the bone marrow has been observed through a switch in the differentiation of BMSCs. The bone/fat switch causes bone growth defects and disordered bone metabolism in bone marrow, for which the mechanisms remain unclear, and treatments are lacking. Studies suggest that small non-coding RNAs (microRNAs) could participate in regulating BMSC differentiation by disrupting the post-transcription of target genes, leading to bone/fat formation changes. This review presents an emerging concept of microRNA regulation in the bone/fat formation switch in bone marrow, the evidence for which is assembled mainly from in vivo and in vitro human or animal models. Characterization of changes to microRNAs reveals novel networks that mediate signalling and factors in regulating bone/fat switch and homeostasis. Recent advances in our understanding of microRNAs in their control in BMSC differentiation have provided valuable insights into underlying mechanisms and may have significant potential in development of new therapeutics.

Metabolic programming determines the lineage-differentiation fate of murine bone marrow stromal progenitor cells

Enhanced bone marrow adipogenesis and impaired osteoblastogenesis have been observed in obesity, suggesting that the metabolic microenvironment regulates bone marrow adipocyte and osteoblast progenitor differentiation fate. To determine the molecular mechanisms, we studied two immortalized murine cell lines of adipocyte or osteoblast progenitors (BMSCsadipo and BMSCsosteo, respectively) under basal and adipogenic culture conditions. At baseline, BMSCsadipo, and BMSCsosteo exhibit a distinct metabolic program evidenced by the presence of specific global gene expression, cellular bioenergetics, and metabolomic signatures that are dependent on insulin signaling and glycolysis in BMSCsosteo versus oxidative phosphorylation in BMSCsadipo. To test the flexibility of the metabolic program, we treated BMSCsadipo with parathyroid hormone, S961 (an inhibitor of insulin signaling) and oligomycin (an inhibitor of oxidative phosphorylation). The treatment induced significant changes in cellular bioenergetics that were associated with decreased adipocytic differentiation. Similarly, 12 weeks of a high-fat diet in mice led to the expansion of adipocyte progenitors, enhanced adipocyte differentiation and insulin signaling in cultured BMSCs. Our data demonstrate that BMSC progenitors possess a distinct metabolic program and are poised to respond to exogenous metabolic cues that regulate their differentiation fate.

Bone Marrow Multipotent Mesenchymal Stromal Cells in Patients with Diffuse Large B-Cell Lymphoma.

In diffuse large B-cell lymphoma, bone marrow involvement is rarely diagnosed. We compared the properties of bone marrow stromal progenitor cells and the concentration of fibroblast CFU in patients with diffuse large B-cell lymphoma without bone marrow involvement and in healthy donors. It was found that the properties of multipotent mesenchymal stromal cells in patients in the debut of the disease differed considerably from those in healthy donors. In particular, the total cell production in patients was significantly higher than in donors. In multipotent mesenchymal stromal cells of patients, some cell parameters were changes; the mean fluorescence intensity of the adhesion molecule ICAM1 on the cell surface was increased. The mean fluorescence intensity of mesenchymal stromal cell markers (HLA-ABC, CD73 and CD90) was significantly elevated. The relative expression of BMP4, MMP2, FGFR1, and ICAM1 genes in mesenchymal stromal cell was reduced, while the expression of FGFR2 gene was enhanced. Despite the absence of proven involvement of the bone marrow, the properties of mesenchymal stromal cells, the components in the stromal microenvironment niche regulating hemopoiesis are altered in patients with diffuse large B-cell lymphoma.

Regulation Of Hematopoietic Stem Cell Trafficking By The Coagulation Pathway

Hematopoeitic stem and progenitor cells (HSPC) dynamically switch between a quiescent, non-motile mode in the bone marrow (BM), to an active state, in which they proliferate, differentiate and egress to the circulation. Injection of the coagulation protease thrombin induced rapid HSPC mobilization to the blood via activation of its major receptor, protease activated receptor 1 (PAR1) on BM hematopoietic and stromal cells. We hypothesized that coagulation factors control stem cells fate in the BM. We examined if thrombin is generated in the murine BM and found by immunohistochemistry prothrombin associated with bone lining osteoblasts in the endosteum region. These cells also highly express osteopontin which induces stem cell quiescence and retention. Cleavage of osteopontin by thrombin or by osteoclast derived cathepsin K induces stem cell mobilization. In addition, a unique structure of multinucleated CD45+ cell clusters in the trabecular-rich area of the murine femoral metaphysis express the cell surface receptor Tissue Factor (TF), a potent initiator of the coagulation cascade leading to thrombin generation. These clusters were found adjacent to multinucleated TRAP (tatrate resistant acid phosphate) positive active osteoclasts. In vitro, we found that immature osteoclasts expressed TF in cell fusion areas, suggesting that osteoclast maturation also activates the coagulation thrombin/PAR1 axis, thus mediating HSPC recruitment to the circulation. Supporting this notion, bleeding which prompts a hemostatic response and thrombin production, is a strong inducer of osteoclasts activation and HSPC mobilization. In addition, injection of bacterial lipopolysaccharides (LPS) is known to activate osteoclasts and induce HSPC mobilization (Kollet et al Nat Med 06). We found that LPS injection upregulated TF expression by CD45+ myeloid cells in the murine BM. LPS treatment provoked massive HSPC mobilization, which was attenuated by PAR1 inhibition. To further address the role of thrombin in stem cell maintenance, we targeted prothrombin in vivo by applying Antisense Oligonucleotides (ASO) knockdown technology, previously shown to induce a dose- and time-dependent up to 90% reduction of prothrombin mRNA levels in the murine liver (Monia et al Blood 2010). Prothrombin depletion altered the BM niche microenvironment by expanding the mesenchymal stem and progenitor (MSPC) population and the long-term repopulating CD34-/ROSlow/LSK HSPC population in the BM. In untreated mice, TF was also expressed by a small MSPC population, suggesting that the bone stomal compartment may also contribute to the regulation of HSPC mobilization upon demand. To further asses the role of thrombin generation in HSPC development, we examined the involvement of the endothelial cell receptor Thrombomodulin (TM) that is pivotal for the anticoagulant pathway which mediates activation of protein C. TM protein is expressed by BM small blood vessels resembling sinusoids and by neighboring MSPC. By immunohistochemistry, we also detected activated protein C on the same blood vessels. A mouse model with a mutation in the TM gene (TMPro/Pro) is characterized by reduced capacity for activated protein C generation which in turn increases thrombin levels in these mice. We found increased circulating hematopoietic stem cells in TMPro/Pro mice, suggesting that chronically increased basal levels of thrombin generation can promote HSC egress. Conversely, short term (5 day) intermittent treatment of mice with low dose thrombin that mainly causes activated protein C formation in vivo, display higher levels of CD34-/ROSlow/LSK and EPCR+LSK stem cells in the BM, indicating additional roles for the anticoagulant pathway in BM stem cell pool maintenance. In summary, our results provide evidence that the activator of the coagulation cascade, TF, and coagulation factors Thrombin and activated protein C are present in the BM and regulate and integrate functions of hematopoietic stem and progenitor cells and BM stromal progenitor cells. Disclosures: Crosby: Isis pharmaceuticals: the ASO for prothrombin was obtained from Isis pharmaceuticals Other. Monia:Isis pharmaceuticals: the ASO for prothrombin was obtained from Isis pharmaceuticals Other.

CXCL12/CXCR4 signal axis plays an important role in mediating bone morphogenetic protein 9-induced osteogenic differentiation of mesenchymal stem cells.

Mesenchymal progenitor stem cells (MPCs) are a group of bone marrow stromal progenitor cells processing osteogenic, chondrogenic, adipogenic and myogenic lineages differentiations. Previous studies have demonstrated that bone morphogeneic protein 9(BMP9) is one of the most osteogenic BMPs both in vitro and in vivo, however, the underlying molecular mechanism of osteogenesis induced by BMP9 is needed to be deep explored. Here, we used the recombinant adenoviruses assay to introduce BMP9 into C3H10T1/2 mesenchymal stem cells to elucidate the role of CXCL12/CXCR4 signal axis during BMP9-incuced osteogenic differentiation. The results showed that CXCL12 and CXCR4 expressions were down-regulated at the stage of BMP9-induced osteogenic differentiation, in a dose- and time-dependent. Pretreatment of C3H10T1/2 cells with CXCL12/CXCR4 could significantly affect the early and mid osteogenic markers alkaline phosphatase (ALP), osteocalcin (OCN), the transcription factors of Runx2, Osx, Plzf and Dlx5 expression, through activating the Smad, MAPK signaling pathway. Addition of exogenous CXCL12 did not affect the changes of the late osteogenic marker calcium deposition. Thus, our findings suggest a co-requirement of the CXCL12/CXCR4 signal axis in BMP9-induced the early- and mid-process of osteogenic differentiation of MSCs.

Functions and action mechanisms of flavonoids genistein and icariin in regulating bone remodeling

Increasingly natural products particularly flavonoids are being explored for their therapeutic potentials in reducing bone loss and maintaining bone health. This study has reviewed previous studies on the two better known flavonoids, genistein and icariin, their structures, functions, action mechanisms, relative potency, and potential application in regulating bone remodeling and preventing bone loss. Genistein, an isoflavone abundant in soy, has dual functions on bone cells, able to inhibit bone resorption activity of osteoclasts and stimulate osteogenic differentiation and maturation of bone marrow stromal progenitor cells (BMSCs) and osteoblasts. Genistein is an estrogen receptor (ER)-selective binding phytoestrogen, with a greater affinity to ERβ. Genistein inhibits tyrosine kinases and inhibits DNA topoisomerases I and II, and may act as an antioxidant. Genistein enhances osteoblastic differentiation and maturation by activation of ER, p38MAPK-Runx2, and NO/cGMP pathways, and it inhibits osteoclast formation and bone resorption through inducing osteoclastogenic inhibitor osteoprotegerin (OPG) and blocking NF-κB signaling. Icariin, a prenylated flavonol glycoside isolated from Epimedium herb, stimulates osteogenic differentiation of BMSCs and inhibits bone resorption activity of osteoclasts. Icariin, whose metabolites include icariside I, icariside II, icaritin, and desmethylicaritin, has no estrogenic activity. However, icariin is more potent than genistein in promoting osteogenic differentiation and maturation of osteoblasts. The existence of a prenyl group on C-8 of icariin molecular structure has been suggested to be the reason why icariin is more potent than genistein in osteogenic activity. Thus, the prenylflavonoids may represent a class of flavonoids with a higher osteogenic activity.

A Comprehensive Analysis of the Dual Roles of BMPs in Regulating Adipogenic and Osteogenic Differentiation of Mesenchymal Progenitor Cells

Pluripotent mesenchymal stem cells (MSCs) are bone marrow stromal progenitor cells that can differentiate into osteogenic, chondrogenic, adipogenic, and myogenic lineages. Several signaling pathways have been shown to regulate the lineage commitment and terminal differentiation of MSCs. Here, we conducted a comprehensive analysis of the 14 types of bone morphogenetic protein (BMPs) for their abilities to regulate multilineage specific differentiation of MSCs. We found that most BMPs exhibited distinct abilities to regulate the expression of Runx2, Sox9, MyoD, and PPARgamma2. Further analysis indicated that BMP-2, BMP-4, BMP-6, BMP-7, and BMP-9 effectively induced both adipogenic and osteogenic differentiation in vitro and in vivo. BMP-induced commitment to osteogenic or adipogenic lineage was shown to be mutually exclusive. Overexpression of Runx2 enhanced BMP-induced osteogenic differentiation, whereas knockdown of Runx2 expression diminished BMP-induced bone formation with a decrease in adipocyte accumulation in vivo. Interestingly, overexpression of PPARgamma2 not only promoted adipogenic differentiation, but also enhanced osteogenic differentiation upon BMP-2, BMP-6, and BMP-9 stimulation. Conversely, MSCs with PPARgamma2 knockdown or mouse embryonic fibroblasts derived from PPARgamma2(-/-) mice exhibited a marked decrease in adipogenic differentiation, coupled with reduced osteogenic differentiation and diminished mineralization upon BMP-9 stimulation, suggesting that PPARgamma2 may play a role in BMP-induced osteogenic and adipogenic differentiation. Thus, it is important to understand the molecular mechanism behind BMP-regulated lineage divergence during MSC differentiation, as this knowledge could help us to understand the pathogenesis of skeletal diseases and may lead to the development of strategies for regenerative medicine.

Hey1 Basic Helix-Loop-Helix Protein Plays an Important Role in Mediating BMP9-induced Osteogenic Differentiation of Mesenchymal Progenitor Cells

Pluripotent mesenchymal stem cells (MSCs) are bone marrow stromal progenitor cells that can differentiate into osteogenic, chondrogenic, adipogenic, and myogenic lineages. We previously demonstrated that bone morphogenetic protein (BMP) 9 is one of the most potent and yet least characterized BMPs that are able to induce osteogenic differentiation of MSCs both in vitro and in vivo. Here, we conducted gene expression-profiling analysis and identified that Hey1 of the hairy/Enhancer of split-related repressor protein basic helix-loop-helix family was among the most significantly up-regulated early targets in BMP9-stimulated MSCs. We demonstrated that Hey1 expression was up-regulated at the immediate early stage of BMP9-induced osteogenic differentiation. Chromatin immunoprecipitation analysis indicated that Hey1 may be a direct target of the BMP9-induced Smad signaling pathway. Silencing Hey1 expression diminished BMP9-induced osteogenic differentiation both in vitro and in vivo and led to chondrogenic differentiation. Likewise, constitutive Hey1 expression augmented BMP9-mediated bone matrix mineralization. Hey1 and Runx2 were shown to act synergistically in BMP9-induced osteogenic differentiation, and Runx2 expression significantly decreased in the absence of Hey1, suggesting that Runx2 may function downstream of Hey1. Accordingly, the defective osteogenic differentiation caused by Hey1 knockdown was rescued by exogenous Runx2 expression. Thus, our findings suggest that Hey1, through its interplay with Runx2, may play an important role in regulating BMP9-induced osteoblast lineage differentiation of MSCs.

Enhanced epithelial gap closure and increased angiogenesis in wounds of diabetic mice treated with adult murine bone marrow stromal progenitor cells

The direct application of bone marrow (BM) can accelerate the healing of chronic wounds. We hypothesized that this effect is due to the presence of stromal progenitor cells (SPCs) found within whole BM preparations. To test this hypothesis, we isolated adult murine SPCs from whole BM and examined their ability to enhance impaired wound healing compared with ficoll separated BM cells in the diabetic (db/db) mouse model. SPCs significantly enhanced reepithelialization, granulation tissue formation, and neovascularization compared with control wounds treated with BM or PBS alone. Higher frequencies of donor SPC cells compared with donor BM cells were observed in treated wounds at 7 days. Transdifferentiation into GFP-positive mature endothelial cells was not observed. These observations suggest that SPCs improve wound healing through indirect mechanisms which lead to enhanced vascularization rather than through direct participation and incorporation into tissue. We conclude that topical application of BM-derived SPCs may represent an effective strategy for the treatment of chronic diabetic wounds.

Negative regulation of BMP signaling by the ski oncoprotein.

The bone morphogenetic proteins (BMPs) play important roles in the regulation of multiple aspects of vertebrate development. BMPs signal through the cell surface receptors and downstream Smad molecules. Upon stimulation with BMP, Smad1, Smad5, and Smad8 are phosphorylated by the activated BMP receptors, form a complex with Smad4, and translocate into the nucleus, where they regulate the expression of BMP target genes. The activity of this signal pathway can be modulated both by extracellular factors that regulate the binding of BMPs to the receptor and by intracellular proteins that interact with the Smad proteins. We have shown that Ski is an important negative regulator of the Smad proteins. Ski can bind to the BMP-Smad protein complexes in response to BMP and repress their ability to activate BMP target genes through disruption of a functional Smad complex and through recruitment of transcriptional co-repressors. The antagonism of BMP signaling by Ski results in neural specification in Xenopus embryos and inhibition of osteoblast differentiation in mouse bone-marrow stromal progenitor cells. This ability to modulate BMP signaling by Ski may play an important role in the regulation of craniofacial, neuronal, and skeletal muscle development.

Silk matrix for tissue engineered anterior cruciate ligaments

A silk-fiber matrix was studied as a suitable material for tissue engineering anterior cruciate ligaments (ACL). The matrix was successfully designed to match the complex and demanding mechanical requirements of a native human ACL, including adequate fatigue performance. This protein matrix supported the attachment, expansion and differentiation of adult human progenitor bone marrow stromal cells based on scanning electron microscopy, DNA quantitation and the expression of collagen types I and III and tenascin-C markers. The results support the conclusion that properly prepared silkworm fiber matrices, aside from providing unique benefits in terms of mechanical properties as well as biocompatibility and slow degradability, can provide suitable biomaterial matrices for the support of adult stem cell differentiation toward ligament lineages. These results point toward this matrix as a new option for ACL repair to overcome current limitations with synthetic and other degradable materials.