Expression of Hes-1 (Notch-1 Effector) during “In Vitro” Normal Erythroid Differentiation.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4287-4287
Author(s):  
Maria D. Cappellini ◽  
Ilaria V. Libani ◽  
Elisabetta Calzavara ◽  
Luisa Ronzoni ◽  
Raffaella Chiaramonte ◽  
...  
Keyword(s):  
Stem Cells ◽  
Notch Signaling ◽  
Cell Fate ◽  
Notch Pathway ◽  
Erythroid Differentiation ◽  
Notch Receptor ◽  
Erythroid Progenitors ◽  
Notch 1 ◽  
Multipotent Stem Cells

Abstract Hematopoiesis involves highly regulated proliferation and differentiation during which a small number of multipotent stem cells give rise to differentiated progenies. In several developmental systems stem cells fate is influenced by soluble molecules acting via cell-cell interaction, including those mediate by the Notch receptor family. Members of Notch family transmembrane receptors are found on primitive hematopoietic precursors, suggesting a role for Notch signaling in mammalian blood cells development. Notch signaling regulates cell fate controlling asymmetric cell division during stem/progenitor cell differentiation. A previous study on K562 cell line showed that Notch signaling inhibits erythroid/megakaryocytic development by suppressing GATA-1 activity. Furthermore there are evidences that Notch is expressed in early murine erythroid precursors. Probably Notch signaling in these uncommittted precursors may lead to enhanced survival, preserving multilineage potential. The role of Notch pathway during human adult erytropiesis has not been described. The aim of this study is to investigate the modulation of Notch activity during “in vitro” human erythropiesis. Human CD34+ from perpheral blood of normal adult subjects were differentiate “in vitro” for two weeks by the addiction of IL-3, SCF and Epo. This method of colture reproduces all stages of adult erythropoiesis. We analized the modulation of the expression of Notch-1, of its effector Hes-1 and of several erythoid specific genes, at different stages of differentiation using real time PCR. Our analysis shows that Hes-1 expression, which indicates the activation of Notch-1 pathway, is very high in the early steps of differentiation (BFU-E, CFU-E) while in the late stages rapidly decreases to undetectable levels. The Notch-1 gene expression doesn’t seem to be modulated way, but we didn’t invstigate the protein levels yet. These data suggest that Notch pathway is involved in the early stages of erythroid differentiation where it may enhance erythroid progenitors survival up to CFU-E, as hypothisized in mouse model, preventing them from apoptotic stimuli and promoting their proliferation. Involvement of Notch-1 signaling in preventing erythroid progenitors from apoptosis during erythroid differentiation could be important in some erythropoietic disorders such as b-Thalassemia syndromes or diserythropoietic anemias.

scholarly journals Notch Signaling Regulation in HCC: From Hepatitis Virus to Non-Coding RNAs

Cells ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 521
Author(s):  
Catia Giovannini ◽  
Francesca Fornari ◽  
Fabio Piscaglia ◽  
Laura Gramantieri
Keyword(s):  
Notch Signaling ◽  
Cell Fate ◽  
Hepatitis Virus ◽  
Notch Pathway ◽  
Notch Receptor ◽  
Gamma Secretase ◽  
Stem Cell Maintenance ◽  
Pathway Activation ◽  
Promising Therapeutic Approach ◽  
Conserved Genes

The Notch family includes evolutionary conserved genes that encode for single-pass transmembrane receptors involved in stem cell maintenance, development and cell fate determination of many cell lineages. Upon activation by different ligands, and depending on the cell type, Notch signaling plays pleomorphic roles in hepatocellular carcinoma (HCC) affecting neoplastic growth, invasion capability and stem like properties. A specific knowledge of the deregulated expression of each Notch receptor and ligand, coupled with resultant phenotypic changes, is still lacking in HCC. Therefore, while interfering with Notch signaling might represent a promising therapeutic approach, the complexity of Notch/ligands interactions and the variable consequences of their modulations raises concerns when performed in undefined molecular background. The gamma-secretase inhibitors (GSIs), representing the most utilized approach for Notch inhibition in clinical trials, are characterized by important adverse effects due to the non-specific nature of GSIs themselves and to the lack of molecular criteria guiding patient selection. In this review, we briefly summarize the mechanisms involved in Notch pathway activation in HCC supporting the development of alternatives to the γ-secretase pan-inhibitor for HCC therapy.

scholarly journals POS0042 NOTCH 1 INHIBITION INCREASES OSTEOCLAST PROGENITOR ACTIVITY IN THE MOUSE MODEL OF RHEUMATOID ARTHRITIS

2021 ◽  
Vol 80 (Suppl 1) ◽  
pp. 226.3-226
Author(s):  
M. Filipović ◽  
A. Šućur ◽  
D. Flegar ◽  
Z. Jajić ◽  
M. Ikić Matijašević ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Notch Signaling ◽  
Neutralizing Antibodies ◽  
Osteoclast Differentiation ◽  
Notch Receptor ◽  
Notch 1 ◽  
Collagen Induced Arthritis ◽  
Notch Receptors

Background:Osteoclasts mediate periarticular and systemic bone loss in rheumatoid arthritis (RA). Osteoclast progenitor cells (OCPs) derived from the myeloid lineage are susceptible to regulation through Notch signaling. Murine bone marrow and splenic OCPs, identified as CD45+Ly6G-CD3-B220-NK1.1-CD11blo/+CD115+CCR2+ cells, are specifically increased in arthritis. We previously identified an increased frequency of OCPs expressing Notch receptors in arthritic mice.Objectives:Several studies suggested that Notch signaling modulation affects the course of experimental arthritis. We aimed to determine the effects of Notch receptor signaling inhibition on OCP activity and arthritis severity in murine collagen-induced arthritis (CIA).Methods:Male C57/Bl6 and DBA mice were immunized with chicken type II collagen and treated with i.p. injections of anti-Notch 1 neutralizing antibodies (1mg/kg). Notch receptor 1 through 4 expression on OCPs was analyzed by flow cytometry in periarticular bone marrow (PBM) and spleen (SPL). Gene expression of Notch receptors, ligands and transcription targets as well as osteoclast differentiation genes RANK, cFos and cFms was determined by qPCR from tissues and sorted OCPs. FACS sorted OCPs were stimulated by osteoclastogenic factors (M-CSF and RANKL), in control, IgG, Jagged (Jag)1 or Delta-like (DLL)1 coated wells, with or without anti-Notch 1 antibodies. Research was approved by the Ethics Committee.Results:We confirmed the expression of Notch receptors on OCPs by flow cytometry with Notch 1 and 2 being most abundantly expressed (around 25% and 40% positive OCPs in PBM and 35% and 20% in SPL respectively), with a significant increase of Notch 2 expression in arthritis. Seeding OCPs on DLL1 coated wells significantly increased while seeding on Jag1 coated wells significantly decreased osteoclastogenesis as reflected on the number of TRAP+ osteoclasts and expression of osteoclast differentiation genes. The addition of anti-Notch 1 antibodies to ligand-stimulated OCPs resulted in an increased number of TRAP+ osteoclasts, partially reversing Jag1 inhibition. In vivo treatment with anti-Notch 1 antibodies did not affect total OCP frequency, but increased expression of Notch 4 both in PBM and SPL as seen by flow cytometry and qPCR. Additionally, anti-Notch 1 treatment stimulated Notch transcription factors HES and HEY. Both PBM and SPL cultured OCPs from anti-Notch 1 treated mice produced a higher number of large TRAP+ osteoclasts, doubling the area covered with osteoclasts in the latter compared to untreated mice. Increased osteoclastogenesis in vitro was further confirmed by an increased expression of osteoclast differentiation genes in the treated group.Conclusion:Our results confirm that Notch signaling may represent an important therapeutic target for the regulation of osteoclast activity in arthritis. Both in vitro and in vivo anti-Notch 1 neutralizing antibodies enhanced osteoclastogenesis in CIA model, implying an inhibitory role of Notch 1 signaling in osteoclast differentiation. As Notch 2 expression is increased on OCPs of arthritic mice, we next plan to determine the effects of Notch 2 neutralization on osteoclast activity and arthritis severity.References:[1]Ikić Matijašević M, Flegar D, Kovačić N, Katavić V, Kelava T, Šućur A, et al. Increased chemotaxis and activity of circulatory myeloid progenitor cells may contribute to enhanced osteoclastogenesis and bone loss in the C57BL/6 mouse model of collagen-induced arthritis. Clin Exp Immunol. 2016;186(3):321–35.[2]Šućur A, Filipović M, Flegar D, Kelava T, Šisl D, Lukač N, et al. Notch receptors and ligands in inflammatory arthritis – a systematic review. Immunology Letters 2020 Vol. 223, p. 106–14.Acknowledgements:The work has been supported by Croatian Science Foundation projects IP-2018-01-2414, UIP-2017-05-1965 and DOK-2018-09-4276.Disclosure of Interests:None declared.

scholarly journals Cancer Stem Cells, Quo Vadis? The Notch Signaling Pathway in Tumor Initiation and Progression

Cells ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1879 ◽  
Author(s):  
Christian T. Meisel ◽  
Cristina Porcheri ◽  
Thimios A. Mitsiadis
Keyword(s):  
Stem Cells ◽  
Cancer Stem Cells ◽  
Notch Signaling ◽  
Signaling Pathway ◽  
Cell Fate ◽  
Adult Life ◽  
Notch Pathway ◽  
Notch Signaling Pathway ◽  
Fine Tuning ◽  
Cell Fate Decisions

The Notch signaling pathway regulates cell proliferation, cytodifferentiation and cell fate decisions in both embryonic and adult life. Several aspects of stem cell maintenance are dependent from the functionality and fine tuning of the Notch pathway. In cancer, Notch is specifically involved in preserving self-renewal and amplification of cancer stem cells, supporting the formation, spread and recurrence of the tumor. As the function of Notch signaling is context dependent, we here provide an overview of its activity in a variety of tumors, focusing mostly on its role in the maintenance of the undifferentiated subset of cancer cells. Finally, we analyze the potential of molecules of the Notch pathway as diagnostic and therapeutic tools against the various cancers.

scholarly journals Delta-1 Activation of Notch-1 Signaling Results inHES-1 Transactivation

1998 ◽  
Vol 18 (12) ◽  
pp. 7423-7431 ◽  
Author(s):  
Sophie Jarriault ◽  
Odile Le Bail ◽  
Estelle Hirsinger ◽  
Olivier Pourquié ◽  
Frédérique Logeat ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Drosophila Melanogaster ◽  
Notch Signaling ◽  
Cell Fate ◽  
Notch Signaling Pathway ◽  
Notch Receptor ◽  
Cell Fate Determination ◽  
Notch 1 ◽  
Promoter Construct ◽  
Enhancer Of Split

ABSTRACT The Notch receptor is involved in many cell fate determination events in vertebrates and invertebrates. It has been shown inDrosophila melanogaster that Delta-dependent Notch signaling activates the transcription factor Suppressor of Hairless, leading to an increased expression of the Enhancer of Splitgenes. Genetic evidence has also implicated the kuzbaniangene, which encodes a disintegrin metalloprotease, in the Notch signaling pathway. By using a two-cell coculture assay, we show here that vertebrate Dl-1 activates the Notch-1 cascade. Consistent with previous data obtained with active forms of Notch-1 aHES-1-derived promoter construct is transactivated in cells expressing Notch-1 in response to Dl-1 stimulation. Impairing the proteolytic maturation of the full-length receptor leads to a decrease in HES-1 transactivation, further supporting the hypothesis that only mature processed Notch is expressed at the cell surface and activated by its ligand. Furthermore, we observed that Dl-1-inducedHES-1 transactivation was dependent both on Kuzbanian and RBP-J activities, consistent with the involvement of these two proteins in Notch signaling in Drosophila. We also observed that exposure of Notch-1-expressing cells to Dl-1 results in an increased level of endogenous HES-1 mRNA. Finally, coculture of Dl-1-expressing cells with myogenic C2 cells suppresses differentiation of C2 cells into myotubes, as previously demonstrated for Jagged-1 and Jagged-2, and also leads to an increased level of endogenousHES-1 mRNA. Thus, Dl-1 behaves as a functional ligand for Notch-1 and has the same ability to suppress cell differentiation as the Jagged proteins do.

scholarly journals Notch pathway activation targets AML-initiating cell homeostasis and differentiation

2013 ◽  
Vol 210 (2) ◽  
pp. 301-319 ◽  
Author(s):  
Camille Lobry ◽  
Panagiotis Ntziachristos ◽  
Delphine Ndiaye-Lobry ◽  
Philmo Oh ◽  
Luisa Cimmino ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Notch Signaling ◽  
Myeloproliferative Neoplasms ◽  
Notch Pathway ◽  
Notch Signaling Pathway ◽  
Notch Receptor ◽  
Pathway Activation ◽  
Function Models

Notch signaling pathway activation is known to contribute to the pathogenesis of a spectrum of human malignancies, including T cell leukemia. However, recent studies have implicated the Notch pathway as a tumor suppressor in myeloproliferative neoplasms and several solid tumors. Here we report a novel tumor suppressor role for Notch signaling in acute myeloid leukemia (AML) and demonstrate that Notch pathway activation could represent a therapeutic strategy in this disease. We show that Notch signaling is silenced in human AML samples, as well as in AML-initiating cells in an animal model of the disease. In vivo activation of Notch signaling using genetic Notch gain of function models or in vitro using synthetic Notch ligand induces rapid cell cycle arrest, differentiation, and apoptosis of AML-initiating cells. Moreover, we demonstrate that Notch inactivation cooperates in vivo with loss of the myeloid tumor suppressor Tet2 to induce AML-like disease. These data demonstrate a novel tumor suppressor role for Notch signaling in AML and elucidate the potential therapeutic use of Notch receptor agonists in the treatment of this devastating leukemia.

scholarly journals Influence of the endothelial cells on the erythroid differentiation of umbilical cord blood hematopoietic stem cells in vitro

2020 ◽  
Vol 17 (1) ◽  
pp. 78-86
Author(s):  
A. S. Voytehovich ◽  
E. V. Vasina ◽  
V. S. Kastsiunina ◽  
I. N. Seviaryn ◽  
N. V. Petyovka
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Erythroid Differentiation ◽  
Erythroid Progenitors ◽  
Hematopoietic Stem ◽  
Vascular Niche

The objective is to study the effect of umbilical cord blood endothelial cells on the hematopoietic cells growth and the maturation in the erythroid direction in co-culture, as well as the expression of adult and fetal hemoglobin genes during erythroid differentiation under the conditions of vascular niche modeling in vitro. We used the following research methods: cultural, flow cytometry, real-time PCR and morphological analysis. We have developed the method of hematopoietic cord blood stem cells erythroid differentiation in co-culture using cord blood endothelial cell progenitors. CD34+CD31+CD144+CD105+CD90–CD45– progenitors of endothelial cells stimulate the erythroid differentiation of hematopoietic CD34+ cord blood cells and the growth of erythroid progenitors in co-culture from the 4th to 11th day in the presence of the stem cell factor, the erythropoietin and the fibroblast growth factor-2. The in vitro modeling of the vascular niche increases the mature CD36–CD235a+ erythroid cells 2.5 times higher than those in the liquid culture. The microenvironment of endothelial cells does not affect the level and expression ratio of fetal and adult hemoglobin during the erythroid differentiation in vitro.

Notch Signaling Is Necessary and Sufficient for Runx1-Dependent Stem Cell Induction in the Embryonic Aorta-Gonad-Mesonephros (AGM) Region.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4161-4161
Author(s):  
Caroline Erter Burns ◽  
Leonard I. Zon
Keyword(s):  
Stem Cell ◽  
Notch Signaling ◽  
Cell Fate ◽  
Notch Pathway ◽  
Notch Receptor ◽  
Dorsal Aorta ◽  
Loss Of Function ◽  
Cardinal Vein ◽  
Hematopoietic Stem

Abstract Vertebrate hematopoiesis can be divided into two embryonic phases: a short primitive wave predominantly generating erythrocytes and a definitive (fetal/adult) wave producing long-term hematopoietic stem cells (HSCs). The definitive wave occurs in the embryonic aorta-gonad-mesonephros (AGM) region through the asymmetric induction of HSCs from the ventral, but not dorsal, aortic endothelial wall. Since Notch signaling is critical for orchestrating a variety of developmental cell fate choices from invertebrates to humans and has been implicated in affecting the differentiation of some hematopoietic lineages, we analyzed whether the Notch pathway regulates definitive HSC induction in vivo. The zebrafish mutant mindbomb harbors a mutation in an essential E3 ligase that ubiquitylates Delta, which in turn allows the Notch intercellular domain to be released and activate downstream target gene transcription. Thus, in the absence of Mindbomb function Notch signaling does not occur. We found that although mindbomb mutants show normal primitive hematopoiesis, definitive c-myb and runx1 HSC expression is lacking. Since embryos injected with synthetic morpholinos designed to inhibit proper splicing of runx1 RNA ( runx morphants) show the same hematopoietic phenotype as mindbomb mutants, we next addressed the epistatic relationship between notch and runx1 using classic gain-of-function and loss-of-function analyses. In runx1 morphants expression of a notch receptor, notch3, and a delta ligand, deltaC, in the developing dorsal aorta was normal. Moreover, injection of runx1 RNA rescued HSCs in the AGM of mindbomb mutants. Together, these results suggest that Runx1 functions downstream of Notch in promoting HSC fate. We next analyzed whether a constitutively activated form of Notch (NICD) is sufficient for HSC specification in the AGM using an inducible binary transgenic system. Zebrafish carrying the heat-shock promoter driving the activator gal4 were mated to animals carrying 6 gal4 -responsive tandem upstream activating sequences (UAS) driving NICD. At the 10 somite-stage the embryos were heat-shocked at 37°C for 1 hour to activate NICD throughout the double transgenic animals. Surprisingly, expression of both HSC markers, c-myb and runx1, were expanded from their normal restricted domain in the ventral endothelium to the entire circumference of the dorsal aorta. Most interestingly, the presence of ectopic c-myb and runx1 transcripts were observed in the developing post-cardinal vein, a vessel that normally does not produce HSCs. These data imply that activation of the Notch pathway generates increased numbers of HSCs in vivo. When runx1 RNA is injected into wild-type embryos a similar expansion of c-myb transcripts is seen throughout the entire dorsal aorta and post-cardinal vein, further indicating that Runx1 functions downstream of Notch in HSC induction. In summary, discovery of the molecular programs essential and sufficient for fetal/adult hematopoietic ontogeny will lead to a further understanding of the physiologic and pathologic processes regulating stem cell homeostasis and translate into more effective therapies for blood disorders.

scholarly journals Notch Signaling in Breast Cancer: A Role in Drug Resistance

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2204
Author(s):  
McKenna BeLow ◽  
Clodia Osipo
Keyword(s):  
Breast Cancer ◽  
Stem Cells ◽  
Cancer Stem Cells ◽  
Notch Signaling ◽  
Cell Fate ◽  
Targeted Therapies ◽  
Notch Pathway ◽  
Breast Cancer Stem Cells ◽  
Breast Cancers ◽  
Mesenchymal Transition

Breast cancer is a heterogeneous disease that can be subdivided into unique molecular subtypes based on protein expression of the Estrogen Receptor, Progesterone Receptor, and/or the Human Epidermal Growth Factor Receptor 2. Therapeutic approaches are designed to inhibit these overexpressed receptors either by endocrine therapy, targeted therapies, or combinations with cytotoxic chemotherapy. However, a significant percentage of breast cancers are inherently resistant or acquire resistance to therapies, and mechanisms that promote resistance remain poorly understood. Notch signaling is an evolutionarily conserved signaling pathway that regulates cell fate, including survival and self-renewal of stem cells, proliferation, or differentiation. Deregulation of Notch signaling promotes resistance to targeted or cytotoxic therapies by enriching of a small population of resistant cells, referred to as breast cancer stem cells, within the bulk tumor; enhancing stem-like features during the process of de-differentiation of tumor cells; or promoting epithelial to mesenchymal transition. Preclinical studies have shown that targeting the Notch pathway can prevent or reverse resistance through reduction or elimination of breast cancer stem cells. However, Notch inhibitors have yet to be clinically approved for the treatment of breast cancer, mainly due to dose-limiting gastrointestinal toxicity. In this review, we discuss potential mechanisms of Notch-mediated resistance in breast cancer cells and breast cancer stem cells, and various methods of targeting Notch through γ-secretase inhibitors, Notch signaling biologics, or transcriptional inhibitors. We also discuss future plans for identification of novel Notch-targeted therapies, in order to reduce toxicity and improve outcomes for women with resistant breast cancer.

P0019MICROVESICLES FROM MESENCHYMAL STEM CELLS OVEREXPRESSING MIR-34A INHIBIT TGF-BETA-1-INDUCED EPITHELIAL-MESENVHYMAL TRANSITION IN RENAL TUBULAR EPITHELIAL CELLS IN VITRO

2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
Juan He ◽  
Yali Jiang ◽  
Yan Wang ◽  
Xiujuan Tian ◽  
Shiren Sun
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Notch Signaling ◽  
Notch Signaling Pathway ◽  
Tubular Epithelial Cells ◽  
Renal Tubular Epithelial Cells ◽  
Notch 1 ◽  
Renal Tubular ◽  
Tgf Β1

Abstract Background and Aims The application of microRNAs (miRNAs) in the therapy of kidney diseases is hampered by difficulties in delivering miRNAs effectively. Nano-sized microvesicles (MVs) are known as natural carriers of small RNAs. Our prior work has demonstrated that MVs isolated from mesenchymal stem cells (MSCs) attenuated kidney injuries induced by unilateral ureteral obstruction and 5/6 subtotal nephrectomy in mice. The present work aimed at evaluating the effects of miR-34a-5p (miR-34a)-modified MSC-MVs on transforming growth factor (TGF)-β1 induced fibrosis and apoptosis in vitro. Method Bone marrow MSCs were further modified by lentiviruses overexpressing miR-34a, and MVs were collected from these MSCs to treat HK-2 renal tubular cells exposed to TGF-β1. Alterations in epithelial-mesenchymal transition (EMT) and cell survival were further determined. Results We first demonstrated that MVs generated by miR-34a-modified MSCs contained more miR-34a. By analyzing the expression levels of epithelial markers (E-cadherin and Tight Junction Protein 1 (TJP1)) and mesenchymal markers (α-SMA and fibronectin), we found that the pro-fibrotic TGF-β1 induced EMT was remarkably suppressed by miR-34a-enriched MSC-MVs. Notch-1 receptor and Jagged-1 ligand, two major molecules of Notch signaling pathway, are predicted targets of miR-34a. We further found that the elevation in Notch-1 and Jagged-1 induced by TGF-β1 was inhibited by miR-34a-enriched MSC-MVs. The inhibitory effects of miR-34a-enriched MSC-MVs on EMT and Notch signaling pathway were stronger than the control MSC-MVs. In addition, TGF-β1 exposure also induced apoptosis in HK-2 cells. Although miR-34amofidied MSC-MVs could inhibit TGF-β1-triggered apoptosis in HK-2 cells, the effects were less significant than the control MSC-MVs. This phenomenon may be resulted from the pro-apoptotic effects of miR-34a. Conclusion Our study demonstrates that miR-34a-overexpressing MSC-MVs inhibit EMT induced by pro-fibrotic TGF-β1 in renal tubular epithelial cells, possibly through inhibiting Jagged-1/Notch-1 pathway. Genetic modification of MSC-MVs with anti-fibrotic molecule may present a novel strategy for treatment of renal injuries.

scholarly journals DAPT inhibits the chondrogenesis of human umbilical cord mesenchymal stem cells

2015 ◽  
Vol 10 (1) ◽  
Author(s):  
Xiaoke Hu ◽  
Jiawen Zhang ◽  
Xinxin Shao ◽  
Ermei Luo ◽  
Li Yu
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Notch Signaling ◽  
Umbilical Cord ◽  
Quantitative Polymerase Chain Reaction ◽  
Butyl Ester ◽  
Notch Pathway ◽  
Human Umbilical Cord ◽  
Notch 1

AbstractNotch signaling plays a key role in cell proliferation and differentiation, and is important in several biological processes, but its role in the chondrogenesis of human umbilical cord mesenchymal stem cells (UC-MSCs) is still unknown. N-[N-(3,5- difluorophenacetyl-L-alanyl)]-(S)-phenylglycinet-butyl ester (DAPT) is the inhibitor of Notch pathway. The aim of this study is to investgate the influence of DAPT on the chondrogenesis of UC-MSCs. In our study, UC-MSCs were isolated from human umbilical cord and their characteristics were identified. The UC-MSCs were induced to differentiate into chondrocytes in vitro and treated with 5 μM DAPT. Glycosaminoglycan (GAG) and collagen type II (COL-2A1) were analyzed qualitatively and quantitatively. The gene expression of Notch-1, Hes-1, GAG and COL-2A1 were analyzed by quantitative polymerase chain reaction (qPCR). The UC-MSCs separated from human umbilical cord, followed the characteristics of Mesenchymal Stem Cells (MSCs). The gene expression of Notch-1 and Hes-1 decreased after chondrogenic induction but the percentage in G1 period and the content of GAG and COL-2A1 increased. The expression of all tested Notch signaling and proliferation genes declined when 5 μM DAPT was added, also the content of GAG and COL-2A1 also decreased. Our study revealed that Notch signaling exists in UC-MSCs and it may remain the proliferative activity of UC-MSCs. Once the chondrogenesis begins, Notch signaling strength decline evidently. DAPT inhibits the chondrogenesis of UC-MSCs.

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