The Generation of Programmable Cells of Monocytic Origin Involves Partial Repression of Monocyte/Macrophage Markers and Reactivation of Pluripotency Genes

Clinic for Applied Cellular Medicine, Department of Neurosurgery, Kiel, Germany.
Stem cells and development (Impact Factor: 3.73). 03/2010; 19(11):1769-80. DOI: 10.1089/scd.2009.0351
Source: PubMed


We have recently demonstrated that peripheral blood monocytes can be differentiated in vitro into hepatocyte-like cells using appropriate differentiation media. Phenotype conversion required prior in vitro culture in the presence of M-CSF, IL-3, and human serum, during which the cells acquired a state of plasticity, so were termed "programmable cells of monocytic origin" (PCMO). Here, we have further characterized the process of PCMO generation with respect to markers of monocyte-to-macrophage transition and pluripotency. During a 6-day culture period, various monocyte/macrophage differentiation markers were down-regulated being indicative of a process of partial dedifferentiation. Dedifferentiation and hepatic redifferentiation also proceeded in highly purified monocyte preparations, albeit with different kinetics, suggesting that the presence of nonmonocytes, or soluble factors derived from them, is not essential in order for monocytes to acquire a multipotent state. PCMOs expressed various markers of human embryonic stem cells with early induction of NANOG and OCT4. Expression of the pluripotency-associated OCT4A isoform was paralleled by a global rise in histone H3 methylation on Lys-4, a marker of active chromatin, and coincided with peak sensitivity to tissue-specific differentiation. These results show that peripheral blood monocytes can be induced in vitro to transiently acquire stem cell-like properties and concomitantly a state of increased differentiation potential toward the hepatocytic phenotype.

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    • "Monocytes have also been used by our group (12) and by Hur et al (13) to generate insulin-producing cells. Our protocol includes growth factors treatment for subsequently undergoing dedifferentiation followed by programmability (12). "
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    ABSTRACT: Ob­jec­ti­ve: The vision of potential autologous cell therapy for the cure of diabetes encourages ongoing research. According to a previously published protocol for the generation of insulin-producing cells from human monocytes, we analyzed whether the addition of growth factors could increase insulin production. This protocol was then transferred to a non-human primate model by using either blood- or spleen-derived monocytes. Methods: Human monocytes were treated to dedifferentiate into programmable cells of monocytic origin (PCMO). In addition to the published protocol, PCMOs were then treated with either activin A, betacellulin, exendin 3 or 4. Cells were characterized by protein expression of insulin, Pdx-1, C-peptide and Glut-2. After identifying the optimal protocol, monocytes from baboon blood were isolated and the procedure was repeated. Spleen monocytes following splenectomy of a live baboon were differentiated and analyzed in the same manner and calculated in number and volume. Results: Insulin content of human cells was highest when cells were treated with activin A and their insulin content was 13 000 µU/1 million cells. Insulin-producing cells form primate monocytes could successfully be generated despite using human growth factors and serum. Expression of insulin, Pdx-1, C-peptide and Glut-2 was comparable to that of human neo-islets. Total insulin content of activin A-treated baboon monocytes was 16 000 µU/1 million cells. Conclusion: We were able to show that insulin-producing cells can be generated from baboon monocytes with human growth factors. The amount generated from one spleen could be enough to cure a baboon from experimentally induced diabetes in an autologous cell transplant setting.
    Full-text · Article · Jun 2014 · Journal of Clinical Research in Pediatric Endocrinology
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    • "We have previously characterized stem cell marker expression in PCMOs and have demonstrated similar expression profiles of Nanog and Oct3/4 during PCMO generation [6]. Moreover, the expression of Nanog and Oct3/4 was paralleled by a global rise in histone H3 methylation on Lys-4, a marker of active chromatin, and coincided with peak sensitivity to hepatocyte-specific differentiation [6]. "
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    ABSTRACT: Hepatocyte-like cells (NeoHepatocytes) generated from a peripheral blood monocyte-derived stem cell-like cell (the PCMO) are a promising alternative for primary hepatocytes in cell transplantation studies to cure liver diseases. However, to be therapeutically effective NeoHepatocytes are needed in large quantities. It was the aim of the present study to investigate i) whether the proportion of actively proliferating NeoHepatocytes can be enhanced by supplementing the PCMO differentiation medium (containing M-CSF, IL-3, and human serum) with either EGF or HB-EGF and ii) which signaling pathway underlies the promitotic effect. EGF and HB-EGF enhanced cell proliferation of PCMOs as demonstrated by increased expression of cycle control genes (ABL, ANAPC2, CDC2, CDK4, CDK6), phosphorylation of the retinoblastoma protein, and increased PCMO cell numbers after stimulation with EGF or HB-EGF. EGF also raised the number of monocytes expressing the proliferation marker Ki67. PCMOs expressed the EGF receptors EGFR (ERBB1) and ERBB3, and expression of both increased during PCMO generation. Phosphoimmunoblotting of PCMOs indicated that both EGF and HB-EGF activated MEK-1/2 and ERK1/2 in a concentration-dependent fashion with the effect of EGF being more prominent. EGF treatment further decreased expression of p47phox and increased that of Nanog indicating enhanced dedifferentiation and pluripotency, respectively. Treatment with both EGF and HB-EGF resulted in NeoHepatocytes with improved functional parameters. The results suggested that the addition of EGF or HB-EGF to PCMO differentiation medium superactivates MEK/ERK signaling which then increases both PCMO proliferation, number, and functional differentiation of PCMO-derived NeoHepatocytes.
    Full-text · Article · Aug 2012 · Cell Communication and Signaling
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    • "Ungefroren et al. confirmed a change of the monocytes phenotype towards a more stem-cell-like appearance during a 6-day dedifferentiation treatment. The cells downregulated some monocyte-defining gens and started to express stem cell markers over time [19]. Recently, our group demonstrated that dedifferentiation of monocytes towards PCMOs with subsequent hepatogenic differentiation can be significantly improved by the use of the patient's autologous serum instead of human AB serum or fetal calf serum (FCS) [3]. "
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    ABSTRACT: Recent publications suggested that monocytes might be an attractive cell type to transdifferentiate into various cellular phenotypes. Aim was, therefore, to evaluate the potential of blood monocytes to transdifferentiate into osteoblasts. Monocytes isolated from peripheral blood were subjected to two previously published treatments to obtain unique, multipotent cell fractions, named programmable cells of monocytic origin (PCMOs) and monocyte-derived mesenchymal progenitor cells (MOMPs). Subsequently, MOMPs and PCMOs were treated with osteogenic differentiation medium (including either vitamin D or dexamethasone) for 14 days. Regarding a variety of surface markers, no differences between MOMPs, PCMOs, and primary monocytes could be detected. The treatment with osteogenic medium neither resulted in loss of hematopoietic markers nor in adoption of mesenchymal phenotype in all cell types. No significant effect was observed regarding the expression of osteogenic transcription factors, bone-related genes, or production of mineralized matrix. Osteogenic medium resulted in activation of monocytes and appearance of osteoclasts. In conclusion, none of the investigated monocyte cell types showed any transdifferentiation characteristics under the tested circumstances. Based on our data, we rather see an activation and maturation of monocytes towards macrophages and osteoclasts.
    Full-text · Article · Apr 2012 · The Scientific World Journal
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