Epigenetic Regulation of Neuregulin-1 Tunes White Adipose Stem Cell Differentiation

Expansion of subcutaneous adipose tissue by differentiation of new adipocytes has been linked to improvements in metabolic health. However, an expandability limit has been observed wherein new adipocytes cannot be produced, the existing adipocytes become enlarged (hypertrophic) and lipids spill over into ectopic sites. Inappropriate ectopic storage of these surplus lipids in liver, muscle, and visceral depots has been linked with metabolic dysfunction. Here we show that Neuregulin-1 (NRG1) serves as a regulator of adipogenic differentiation in subcutaneous primary human stem cells. We further demonstrate that DNA methylation modulates NRG1 expression in these cells, and a 3-day exposure of stem cells to a recombinant NRG1 peptide fragment is sufficient to reprogram adipogenic cellular differentiation to higher levels. These results define a novel molecular adipogenic rheostat with potential implications for the expansion of adipose tissue in vivo.

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Epigenetic regulation of Neuregulin-1 tunes white adipose stem cell differentiation

10.1101/2020.03.23.004275 ◽

2020 ◽

Author(s):

Alyssa D. Cordero ◽

Evan C. Callihan ◽

Rana Said ◽

Yasir Alowais ◽

Emily S. Paffhausen ◽

...

Keyword(s):

Stem Cells ◽

Adipose Tissue ◽

Adipogenic Differentiation ◽

Stem Cell Differentiation ◽

Metabolic Dysfunction ◽

Human Stem Cells ◽

Adipose Stem Cell ◽

Neuregulin 1 ◽

Subcutaneous Adipose

AbstractExpansion of subcutaneous adipose tissue by differentiation of new adipocytes has been linked to improvements in metabolic health. However, an expandability limit has been observed wherein new adipocytes cannot be produced, the existing adipocytes become enlarged (hypertrophic) and lipids spill over into ectopic sites. Inappropriate ectopic storage of these surplus lipids in liver, muscle, and visceral depots has been linked with metabolic dysfunction. Here we show that Neuregulin-1 (NRG1) serves as a regulator of adipogenic differentiation in subcutaneous primary human stem cells. We further demonstrate that DNA methylation modulates NRG1 expression in these cells, and a 3-day exposure of stem cells to a recombinant NRG1 peptide fragment is sufficient to reprogram adipogenic cellular differentiation to higher levels. These results define a novel molecular adipogenic rheostat with potential implications for the expansion of adipose tissue in vivo.

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Time-Dependent Reduction of Calcium Oscillations in Adipose-Derived Stem Cells Differentiating towards Adipogenic and Osteogenic Lineage

Biomolecules ◽

10.3390/biom11101400 ◽

2021 ◽

Vol 11 (10) ◽

pp. 1400

Author(s):

Enrico C. Torre ◽

Mesude Bicer ◽

Graeme S. Cottrell ◽

Darius Widera ◽

Francesco Tamagnini

Keyword(s):

Stem Cells ◽

Osteogenic Differentiation ◽

Adipogenic Differentiation ◽

Specific Treatment ◽

Stem Cell Differentiation ◽

Calcium Oscillations ◽

Multipotent Stem Cells ◽

Over Time

Adipose-derived mesenchymal stromal cells (ASCs) are multipotent stem cells which can differentiate into various cell types, including osteocytes and adipocytes. Due to their ease of harvesting, multipotency, and low tumorigenicity, they are a prime candidate for the development of novel interventional approaches in regenerative medicine. ASCs exhibit slow, spontaneous Ca2+ oscillations and the manipulation of Ca2+ signalling via electrical stimulation was proposed as a potential route for promoting their differentiation in vivo. However, the effects of differentiation-inducing treatments on spontaneous Ca2+ oscillations in ASCs are not yet fully characterised. In this study, we used 2-photon live Ca2+ imaging to assess the fraction of cells showing spontaneous oscillations and the frequency of the oscillation (measured as interpeak interval—IPI) in ASCs undergoing osteogenic or adipogenic differentiation, using undifferentiated ASCs as controls. The measurements were carried out at 7, 14, and 21 days in vitro (DIV) to assess the effect of time in culture on Ca2+ dynamics. We observed that both time and differentiation treatment are important factors associated with a reduced fraction of cells showing Ca2+ oscillations, paralleled by increased IPI times, in comparison with untreated ASCs. Both adipogenic and osteogenic differentiation resulted in a reduction in Ca2+ dynamics, such as the fraction of cells showing intracellular Ca2+ oscillations and their frequency. Adipogenic differentiation was associated with a more pronounced reduction of Ca2+ dynamics compared to cells differentiating towards the osteogenic fate. Changes in Ca2+ associated oscillations with a specific treatment had already occurred at 7 DIV. Finally, we observed a reduction in Ca2+ dynamics over time in untreated ASCs. These data suggest that adipogenic and osteogenic differentiation cell fates are associated with specific changes in spontaneous Ca2+ dynamics over time. While this observation is interesting and provides useful information to understand the functional correlates of stem cell differentiation, further studies are required to clarify the molecular and mechanistic correlates of these changes. This will allow us to better understand the causal relationship between Ca2+ dynamics and differentiation, potentially leading to the development of novel, more effective interventions for both bone regeneration and control of adipose growth.

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Insights into Differentiation of Melanocytes from Human Stem Cells and Their Relevance for Melanoma Treatment

Cancers ◽

10.3390/cancers12092508 ◽

2020 ◽

Vol 12 (9) ◽

pp. 2508

Author(s):

Madalina Mirea ◽

Stefan Eckensperger ◽

Markus Hengstschläger ◽

Mario Mikula

Keyword(s):

Stem Cells ◽

Cell Differentiation ◽

Developmental Process ◽

Epithelial Mesenchymal Transition ◽

Stem Cell Differentiation ◽

Human Stem Cells ◽

Mesenchymal Transition ◽

Melanocytic Differentiation

Malignant melanoma represents a highly aggressive form of skin cancer. The metastatic process itself is mostly governed by the so-called epithelial mesenchymal transition (EMT), which confers cancer cells migrative, invasive and resistance abilities. Since EMT represents a conserved developmental process, it is worthwhile further examining the nature of early developmental steps fundamental for melanocyte differentiation. This can be done either in vivo by analyzing the physiologic embryo development in different species or by in vitro studies of melanocytic differentiation originating from embryonic human stem cells. Most importantly, external cues drive progenitor cell differentiation, which can be divided in stages favoring neural crest specification or melanocytic differentiation and proliferation. In this review, we describe ectopic factors which drive human pluripotent stem cell differentiation to melanocytes in 2D, as well as in organoid models. Furthermore, we compare developmental mechanisms with processes described to occur during melanoma development. Finally, we suggest differentiation factors as potential co-treatment options for metastatic melanoma patients.

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282 ADIPOGENIC DIFFERENTIATION IN VITRO OF PORCINE ADULT MESENCHYMAL STEM CELLS

Reproduction Fertility and Development ◽

10.1071/rdv21n1ab282 ◽

2009 ◽

Vol 21 (1) ◽

pp. 238 ◽

Cited By ~ 1

Author(s):

E. Monaco ◽

A. Lima ◽

S. Wilson ◽

S. Lane ◽

M. Bionaz ◽

...

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Bone Marrow ◽

Adipose Tissue ◽

Subcutaneous Adipose Tissue ◽

Adipogenic Differentiation ◽

Expression Patterns ◽

Mrna Abundance ◽

Subcutaneous Adipose

The quantity and accessibility of subcutaneous adipose tissue in humans make it an attractive alternative to bone marrow as a source of adult stem cells for therapeutic purposes. However, before such a cell source substitution can be proposed, the properties of stem cells derived from adipose tissue (ADSC) and bone marrow (BMSC), and their differentiated progeny must be compared in an animal model, such as swine, that adequately simulates the structure and physiology of humans. The objective of this work was to induce adult porcine stem cells isolated from subcutaneous adipose tissue and bone marrow to differentiate in vitro along the adipogenic lineage and to compare their transcript profile properties. ADSC and BMSC were isolated from subcutaneous adipose tissue and femurs of adult pigs, respectively, and differentiated along the adipogenic lineage using specific inducing medium. Cells were incubated up to 4 weeks with medium replaced every 3 days. Histological staining with Oil Red O was performed at 0, 2, 4, 7, 14, 21, 28 days of differentiation (dd) to confirm the adipogenic differentiation. RNA was also extracted at these time points. qPCR was performed on PPARG, DBI, ACSL1, CD36, CEBPA, DGAT2, ADFP, ADIPOQ, SCD. The geometrical mean of GTF2H3, NUBP, and PPP2CB was used as an internal control. Gene expression was analyzed using a mixed model of SAS with repeated time. The adipogenic differentiation of both ADSC and BMSC was confirmed by the Oil Red O positive staining. The relative mRNA abundance of all the genes at dd0 was similar between the ADSC and BMSC. The relative mRNA abundance of most of the genes was also similar between ADSC and BMSC throughout the adipogenic differentiation. ACSL1 and ADIPOQ had analogous expression patterns among the cell types. ACSL1 had relatively large mRNA abundance before differentiation, but ADIPOQ was barely detectable. As a consequence of differentiation, ACSL1 increased in relative mRNA abundance about 10-fold, whereas ADIPOQ mRNA increased about 1000-fold. Temporal expression patterns of SCD, DGAT2, and ADFP were similar. The increase in gene expression was >800% for SCD, >500% for ADFP, and >50 000% for DGAT2 after 7dd. ADSC had significantly higher expression of those genes compared to BMSC at 14 and 28dd. Both ADIPOQ and DGAT2 were almost undetectable prior to differentiation. mRNA expression of CD36 and DBI was similar with a significantly larger increase in expression of ADSC compared with BMSC. Relative mRNA abundance of CEBPA and PPARG was also larger in ADSC compared with BMSC; however, BMSC had a remarkable increase in temporal expression of those genes throughout adipogenic differentiation. These results suggest both cell types can differentiate towards the adipogenic lineage but with quantitatively different gene expression patterns. More investigation is needed before the ADSC can be considered a practical alternative source for stem cells in future human clinical applications. This research was supported by the Illinois Regenerative Medicine Institute.

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Subcutaneous Adipose Stem Cells in Obesity: The Impact of Bariatric Surgery

10.5772/intechopen.95542 ◽

2021 ◽

Author(s):

Veronica Mocanu ◽

Daniel V. Timofte ◽

Ioana Hristov

Keyword(s):

Insulin Resistance ◽

Bariatric Surgery ◽

Stem Cells ◽

Weight Loss ◽

Adipose Tissue ◽

Adipogenic Differentiation ◽

Low Grade ◽

Potential Weight ◽

Subcutaneous Adipose ◽

The Impact

Adipocyte expansion, which involves adipose tissue-derived mesenchymal stem cells (ASCs), is a critical process with implications in the pathogenesis of metabolic syndrome and insulin resistance associated with obesity. Impaired subcutaneous adipogenesis leads to dysfunctional, hypertrophic adipocytes, chronic low-grade inflammation, and peripheric insulin resistance. Alternatively, it has also been proposed that the preservation of the functionality of subcutaneous adipocyte precursors could contribute to some obese individuals remaining metabolically healthy. Very few studies evaluated the changes in the adipogenic differentiation for human subcutaneous ASCs following bariatric surgery. Weight loss after bariatric surgery involves extensive remodeling of adipose tissue, comprising the hyperplasia-hypertrophy balance. Subcutaneous ASCs may be implicated in the variations of bariatric outcomes, through a different restoration in their proliferative and adipogenic potential. Weight loss induced by bariatric surgery correlates to the subcutaneous ASC functions and could explain the variability of metabolic improvement. Limited research data are available to the present and these data support the importance of diagnosis of subcutaneous ASCs functions as predictors of metabolic improvement after bariatric surgery.

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401 MIGRATION AND THERAPEUTIC POTENTIAL OF PORCINE ADULT ADIPOSE-DERIVED MESENCHYMAL STEM CELLS

Reproduction Fertility and Development ◽

10.1071/rdv22n1ab401 ◽

2010 ◽

Vol 22 (1) ◽

pp. 357 ◽

Cited By ~ 2

Author(s):

S. M. Wilson ◽

E. Monaco ◽

M. S. Goldwasser ◽

S. G. Clark ◽

W. L. Hurley ◽

...

Keyword(s):

Stem Cells ◽

Adipose Tissue ◽

Subcutaneous Adipose Tissue ◽

Adult Stem Cells ◽

Bone Defects ◽

Adipose Derived Stem Cells ◽

Subcutaneous Adipose ◽

Time Point

Bone marrow is one current source of adult stem cells for therapeutic purposes; however, the magnitude and accessibility of subcutaneous adipose tissue in humans make it an attractive alternative. Numerous in vitro studies have been conducted to determine how these cells act in vitro, but it is imperative to determine the vast abilities of these cells in vivo. The objective of this study was to evaluate in vivo migration and bone healing ability after transplanting adipose-derived stem cells (ADSC) in a swine model. Adipose-derived stem cells were isolated from subcutaneous adipose tissue of adult Yorkshire pigs and cultured in vitro. At 80 to 90% confluence/passage 3, the cells were trypsinized and labeled in suspension with carboxyfluorescein succinimidyl ester (CFDA-SE). This project included 20 pigs weighing between 63.5 and 81.7 kg. Bilateral mandibular osteoectomies with 10-mm defects were performed on each pig. Of the 20 pigs, half received a treatment of 2.5 million CFDA-SE labeled stem cells administered directly into each defect (DI), and the remaining half received a treatment of approximately 5 million CFDA-SE labeled stem cells through an ear vein injection via catheter (EVI). The time points were 1 h and 2 and 4 wk, with 2 pigs per time with the DI and EVI treatments. Pigs were slaughtered at each time, and spleen, liver, lung, kidney, ear vein, blood, and mandible tissues were collected. Blood samples were collected from the jugular vein with EDTA and processed via flow cytometry after collection. Tissues were fixed in 10% buffered formalin for histology. Fluorescent microscopy (CFDA-SE excitation/emission is 492/517 nm) has confirmed that transplanted ADSC do indeed migrate to a site of injury or trauma. Labeled cells were also present in blood collected from the 1-h time point group. Currently, we have not seen the presence of labeled ADSC in the other tissues (spleen, liver, lung, and kidney) after the 1-h time point. We did observe that ADSC administered by DI and EVI were able to significantly heal and regenerate bone defects within 4 wk post-surgery (P < 0.05, ANOVA with F-test), in contrast to bone defects in pigs that did not receive cell injections (control). Evidence of ADSC-related healing and bone regeneration was evident by gross visualization, dual-energy x-ray absorptiometry (DXA) and micro computer tomography (microCT) analysis. The clinical implications of these results are significant for treating many diseases in which inflammation or defects exist, such as cardiac disease, neurological disease, or traumatic injuries to both soft and hard tissue. If the adult stem cells can be harvested from fat, encouraged to produce bone or cartilage, and then reinserted into defects, treatment protocols for trauma victims could be developed that would reduce the need for alternate harvesting techniques for bone. This work was support by a grant from the Illinois Regenerative Medicine Institute (IDPH # 63080017).

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Differentiation of mesenchymal stem cells from humans and animals into insulin-producing cells: An overview in vitro induction forms

Current Stem Cell Research & Therapy ◽

10.2174/1574888x16666201229124429 ◽

2020 ◽

Vol 16 ◽

Author(s):

Bruna O. S. Câmara ◽

Bruno M. Bertassoli ◽

Natália M. Ocarino ◽

Rogéria Serakides

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Culture Medium ◽

Adipogenic Differentiation ◽

Medium Composition ◽

Cell Therapies ◽

Insulin Producing Cells ◽

Msc Differentiation

The use of stem cells in cell therapies has shown promising results in the treatment of several diseases, including diabetes mellitus, in both humans and animals. Mesenchymal stem cells (MSCs) can be isolated from various locations, including bone marrow, adipose tissues, synovia, muscles, dental pulp, umbilical cords, and the placenta. In vitro, by manipulating the composition of the culture medium or transfection, MSCs can differentiate into several cell lineages, including insulin-producing cells (IPCs). Unlike osteogenic, chondrogenic, and adipogenic differentiation, for which the culture medium and time are similar between studies, studies involving the induction of MSC differentiation in IPCs differ greatly. This divergence is usually evident in relation to the differentiation technique used, the composition of the culture medium, the cultivation time, which can vary from a few hours to several months, and the number of steps to complete differentiation. However, although there is no “gold standard” differentiation medium composition, most prominent studies mention the use of nicotinamide, exedin-4, ß-mercaptoethanol, fibroblast growth factor b (FGFb), and glucose in the culture medium to promote the differentiation of MSCs into IPCs. Therefore, the purpose of this review is to investigate the stages of MSC differentiation into IPCs both in vivo and in vitro, as well as address differentiation techniques and molecular actions and mechanisms by which some substances, such as nicotinamide, exedin-4, ßmercaptoethanol, FGFb, and glucose, participate in the differentiation process.

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Methionine and Arginine Supply Alters Abundance of Amino Acid, Insulin Signaling, and Glutathione Metabolism-Related Proteins in Bovine Subcutaneous Adipose Explants Challenged with N-Acetyl-d-sphingosine

Animals ◽

10.3390/ani11072114 ◽

2021 ◽

Vol 11 (7) ◽

pp. 2114

Author(s):

Yusheng Liang ◽

Nana Ma ◽

Danielle N. Coleman ◽

Fang Liu ◽

Yu Li ◽

...

Keyword(s):

Adipose Tissue ◽

Amino Acid ◽

Insulin Signaling ◽

Elongation Factor ◽

In Vivo Studies ◽

Glutathione Metabolism ◽

Protein Abundance ◽

Related Proteins ◽

Subcutaneous Adipose

The objective was to perform a proof-of-principle study to evaluate the effects of methionine (Met) and arginine (Arg) supply on protein abundance of amino acid, insulin signaling, and glutathione metabolism-related proteins in subcutaneous adipose tissue (SAT) explants under ceramide (Ce) challenge. SAT from four lactating Holstein cows was incubated with one of the following media: ideal profile of amino acid as the control (IPAA; Lys:Met 2.9:1, Lys:Arg 2:1), increased Met (incMet; Lys:Met 2.5:1), increased Arg (incArg; Lys:Arg 1:1), or incMet plus incArg (Lys:Met 2.5:1 Lys:Arg 1:1) with or without 100 μM exogenous cell-permeable Ce (N-Acetyl-d-sphingosine). Ceramide stimulation downregulated the overall abundance of phosphorylated (p) protein kinase B (AKT), p-mechanistic target of rapamycin (mTOR), and p-eukaryotic elongation factor 2 (eEF2). Without Ce stimulation, increased Met, Arg, or Met + Arg resulted in lower p-mTOR. Compared with control SAT stimulated with Ce, increased Met, Arg, or Met + Arg resulted in greater activation of mTOR (p-mTOR/total mTOR) and AKT (p-AKT/total AKT), with a more pronounced response due to Arg. The greatest protein abundance of glutathione S-transferase Mu 1 (GSTM1) was detected in response to increased Met supply during Ce stimulation. Ceramide stimulation decreased the overall protein abundance of the Na-coupled neutral amino acid transporter SLC38A1 and branched-chain alpha-ketoacid dehydrogenase kinase (BCKDK). However, compared with controls, increased Met or Arg supply attenuated the downregulation of BCKDK induced by Ce. Circulating ceramides might affect amino acid, insulin signaling, and glutathione metabolism in dairy cow adipose tissue. Further in vivo studies are needed to confirm the role of rumen-protected amino acids in regulating bovine adipose function.

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Regeneration of pulp-dentin complex using human stem cells of the apical papilla: in vivo interaction with two bioactive materials

Clinical Oral Investigations ◽

10.1007/s00784-021-03840-9 ◽

2021 ◽

Author(s):

Diana B. Sequeira ◽

Ana Rafaela Oliveira ◽

Catarina M. Seabra ◽

Paulo J. Palma ◽

Carlos Ramos ◽

...

Keyword(s):

Stem Cells ◽

Human Stem Cells ◽

Bioactive Materials ◽

Apical Papilla

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Comparative analysis of mouse bone marrow and adipose tissue mesenchymal stem cells for critical limb ischemia cell therapy

Stem Cell Research & Therapy ◽

10.1186/s13287-020-02110-x ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Pegah Nammian ◽

Seyedeh-Leili Asadi-Yousefabad ◽

Sajad Daneshi ◽

Mohammad Hasan Sheikhha ◽

Seyed Mohammad Bagher Tabei ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Adipose Tissue ◽

Cell Therapy ◽

Femoral Artery ◽

Critical Limb Ischemia ◽

Limb Ischemia

Abstract Introduction Critical limb ischemia (CLI) is the most advanced form of peripheral arterial disease (PAD) characterized by ischemic rest pain and non-healing ulcers. Currently, the standard therapy for CLI is the surgical reconstruction and endovascular therapy or limb amputation for patients with no treatment options. Neovasculogenesis induced by mesenchymal stem cells (MSCs) therapy is a promising approach to improve CLI. Owing to their angiogenic and immunomodulatory potential, MSCs are perfect candidates for the treatment of CLI. The purpose of this study was to determine and compare the in vitro and in vivo effects of allogeneic bone marrow mesenchymal stem cells (BM-MSCs) and adipose tissue mesenchymal stem cells (AT-MSCs) on CLI treatment. Methods For the first step, BM-MSCs and AT-MSCs were isolated and characterized for the characteristic MSC phenotypes. Then, femoral artery ligation and total excision of the femoral artery were performed on C57BL/6 mice to create a CLI model. The cells were evaluated for their in vitro and in vivo biological characteristics for CLI cell therapy. In order to determine these characteristics, the following tests were performed: morphology, flow cytometry, differentiation to osteocyte and adipocyte, wound healing assay, and behavioral tests including Tarlov, Ischemia, Modified ischemia, Function and the grade of limb necrosis scores, donor cell survival assay, and histological analysis. Results Our cellular and functional tests indicated that during 28 days after cell transplantation, BM-MSCs had a great effect on endothelial cell migration, muscle restructure, functional improvements, and neovascularization in ischemic tissues compared with AT-MSCs and control groups. Conclusions Allogeneic BM-MSC transplantation resulted in a more effective recovery from critical limb ischemia compared to AT-MSCs transplantation. In fact, BM-MSC transplantation could be considered as a promising therapy for diseases with insufficient angiogenesis including hindlimb ischemia.

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