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A defined mix of cytokines mimics conditioned medium from cultures of bone marrow-derived mesenchymal stem cells and elicits bone regeneration.

ABSTRACT: OBJECTIVES:We previously reported that conditioned medium from cultures of bone marrow-derived mesenchymal stem cells have strong potential to accelerate bone regeneration. We now examine in vitro and in vivo a defined cytokine cocktail that mimics the effects of conditioned medium on bone regeneration. MATERIALS AND METHODS:A cocktail of recombinant human insulin-like growth factor-1, vascular endothelial growth factor-A and transforming growth factor-?1 was prepared at concentrations similar to those in conditioned medium. Conversely, these cytokines were depleted from conditioned medium, and the effects of the cocktail, the conditioned medium and the cytokine-depleted conditioned medium on bone regeneration were evaluated in vitro and in vivo. RESULTS:The cytokine cocktail and conditioned medium enhanced cell migration, tube formation, and expression of osteogenic and angiogenic genes. Depletion of cytokines significantly decreased the effects of conditioned medium in vitro. Similarly, the cytokine cocktail and conditioned medium, but not cytokine-depleted medium, increased bone regeneration in damaged rat calvarial bone. Immunohistochemistry indicated that the cytokine cocktail and conditioned medium strongly enhanced recruitment of endogenous stem cells and endothelial cells. CONCLUSIONS:The data indicate that the cytokine cocktail and conditioned medium enhance the migration of stem cells and endothelial cells to damaged bone, and elicit osteogenesis and angiogenesis.

SUBMITTER: Katagiri W

PROVIDER: S-EPMC6529096 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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A defined mix of cytokines mimics conditioned medium from cultures of bone marrow-derived mesenchymal stem cells and elicits bone regeneration.

Katagiri Wataru W Sakaguchi Kohei K Kawai Takamasa T Wakayama Yukiko Y Osugi Masashi M Hibi Hideharu H

Cell proliferation 20170130 3

<h4>Objectives</h4>We previously reported that conditioned medium from cultures of bone marrow-derived mesenchymal stem cells have strong potential to accelerate bone regeneration. We now examine in vitro and in vivo a defined cytokine cocktail that mimics the effects of conditioned medium on bone regeneration.<h4>Materials and methods</h4>A cocktail of recombinant human insulin-like growth factor-1, vascular endothelial growth factor-A and transforming growth factor-β1 was prepared at concentra ...[more]

PMID: 28133828

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Project description:BackgroundRobust and reliable in vitro and in vivo models of primary cells are necessary to study the pathomechanisms of Myelodysplastic Neoplasms (MDS) and identify novel therapeutic strategies. MDS-derived hematopoietic stem and progenitor cells (HSPCs) are reliant on the support of bone marrow (BM) derived mesenchymal stroma cells (MSCs). Therefore, isolation and expansion of MCSs are essential for successfully modeling this disease. For the clinical use of healthy MSCs isolated from human BM, umbilical cord blood or adipose tissue, several studies showed that xeno-free (XF) culture conditions resulted in superior growth kinetics compared to MSCs cultured in the presence of fetal bovine serum (FBS). In this present study, we investigate, whether the replacement of a commercially available MSC expansion medium containing FBS with a XF medium is beneficial for the expansion of MSCs derived from BM of MDS patients which are often difficult to cultivate.MethodsMSCs isolated from BM of MDS patients were cultured and expanded in MSC expansion medium with FBS or XF supplement. Subsequently, the impact of culture media on growth kinetics, morphology, immunophenotype, clonogenic potential, differentiation capacity, gene expression profiles and ability to engraft in immunodeficient mouse models was evaluated.ResultsSignificant higher cell numbers with an increase in clonogenic potential were observed during culture of MDS MSCs with XF medium compared to medium containing FBS. Differential gene expression showed an increase in transcripts associated with MSC stemness after expansion with XF. Furthermore, immunophenotypes of the MSCs and their ability to differentiate into osteoblasts, adipocytes or chondroblasts remained stable. MSCs expanded with XF media were similarly supportive for creating MDS xenografts in vivo as MSCs expanded with FBS.ConclusionOur data indicate that with XF media, higher cell numbers of MDS MSCs can be obtained with overall improved characteristics in in vitro and in vivo experimental models.

Project description:Bone marrow-derived mesenchymal stem cells differentiate into neurons under the induction of Schwann cells. However, key microRNAs and related pathways for differentiation remain unclear. This study screened and identified differentially expressed microRNAs in bone marrow-derived mesenchymal stem cells induced by Schwann cell-conditioned medium, and explored targets and related pathways involved in their differentiation into neuronal-like cells. Primary bone marrow-derived mesenchymal stem cells were isolated from femoral and tibial bones, while primary Schwann cells were isolated from bilateral saphenous nerves. Bone marrow-derived mesenchymal stem cells were cultured in unconditioned (control group) and Schwann cell-conditioned medium (bone marrow-derived mesenchymal stem cell + Schwann cell group). Neuronal differentiation of bone marrow-derived mesenchymal stem cells induced by Schwann cell-conditioned medium was observed by time-lapse imaging. Upon induction, the morphology of bone marrow-derived mesenchymal stem cells changed into a neural shape with neurites. Results of quantitative reverse transcription-polymerase chain reaction revealed that nestin mRNA expression was upregulated from 1 to 3 days and downregulated from 3 to 7 days in the bone marrow-derived mesenchymal stem cell + Schwann cell group. Compared with the control group, microtubule-associated protein 2 mRNA expression gradually increased from 1 to 7 days in the bone marrow-derived mesenchymal stem cell + Schwann cell group. After 7 days of induction, microRNA analysis identified 83 significantly differentially expressed microRNAs between the two groups. Gene Ontology analysis indicated enrichment of microRNA target genes for neuronal projection development, regulation of axonogenesis, and positive regulation of cell proliferation. Kyoto Encyclopedia of Genes and Genomes pathway analysis demonstrated that Hippo, Wnt, transforming growth factor-beta, and Hedgehog signaling pathways were potentially associated with neural differentiation of bone marrow-derived mesenchymal stem cells. This study, which carried out successful microRNA analysis of neuronal-like cells differentiated from bone marrow-derived mesenchymal stem cells by Schwann cell induction, revealed key microRNAs and pathways involved in neural differentiation of bone marrow-derived mesenchymal stem cells. All protocols were approved by the Animal Ethics Committee of Institute of Radiation Medicine, Chinese Academy of Medical Sciences on March 12, 2017 (approval number: DWLI-20170311).

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A defined mix of cytokines mimics conditioned medium from cultures of bone marrow-derived mesenchymal stem cells and elicits bone regeneration.

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A defined mix of cytokines mimics conditioned medium from cultures of bone marrow-derived mesenchymal stem cells and elicits bone regeneration.

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