Hes1 deficiency causes hematopoietic stem cell exhaustion.

The transcriptional repressor Hairy Enhancer of Split 1 (HES1) plays an essential role in the development of many organs by promoting the maintenance of stem/progenitor cells, controlling the reversibility of cellular quiescence, and regulating both cell fate decisions. Deletion of Hes1 in mice results in severe defects in multiple organs and is lethal in late embryogenesis. Here we have investigated the role of HES1 in hematopoiesis using a hematopoietic lineage-specific Hes1 knockout mouse model. We found that while Hes1 is dispensable for steady-state hematopoiesis, Hes1-deficient hematopoietic stem cells (HSCs) undergo exhaustion under replicative stress. Loss of Hes1 upregulates the expression of genes involved in PPARγ signaling and fatty acid metabolism pathways, and augments fatty acid oxidation (FAO) in Hes1 f/f Vav1Cre HSCs and progenitors. Functionally, PPARγ targeting or FAO inhibition ameliorates the repopulating defects of Hes1 f/f Vav1Cre HSCs through improving quiescence in HSCs. Lastly, transcriptome analysis reveals that disruption of Hes1 in hematopoietic lineage alters expression of genes critical to HSC function, PPARγ signaling, and fatty acid metabolism. Together, our findings identify a novel role of HES1 in regulating stress hematopoiesis and provide mechanistic insight into the function of HES1 in HSC maintenance.

Overexpression of Hes1 is involved in sensitization of K562 cells to Imatinib.

Tyrosine kinase inhibitor (TKI)-based therapy has created promising results among much chronic myeloid leukemia (CML) patients. Imatinib as a relatively specific inhibitor of Bcr-Abl is at present one of the undisputed therapeutic agent for newlydiagnosed patients with CML. However, the occurrence of imatinib-resistance enlightens the urgent need to identify other therapeutic agents against CML. Juglone (5-hydroxy-2-methyl-1, 4-naphthoquinone) exerts cytotoxic effects against various human cancer cell lines. However, the mechanisms through which Juglone induces anticancer effects in CML especially in comparison with imatinib treatment remain unknown. Our results revealed that Juglone-inhibited K562 cells growth through inducing apoptosis. Based on our Western blot analyses, Juglone significantly reduced p-Akt levels and increased the expression level of Forkhead box O1 (FoxO1) and FoxO3a proteins. Moreover, hairy/enhancer of split-1 (Hes1) protein, overexpressed under the influence of Juglone, is apparently involved in Juglone-induced apoptosis among K562 cells. Conversely, treatment with imatinib attenuated Hes1 protein expression. Considering the different functional mechanism of Juglone compared with imatinib, it seems that Juglone treatment could be a useful alternative strategy for the treatment of patients with imatinib-resistance.

Baicalein suppresses the proliferation of human cervical cancer cells via Notch 1/Hes signaling pathway.

BACKGROUND: Baicalein is an active compound extracted from the roots of Scutellaria baicalensis georgi, which is widely and traditionally used in the anticancer therapy. Notch signaling pathway is usually abnormally activated in kinds of human cancers. The aim of the present study is to investigate the antitumor effects of baicalein in human cervical cancer and explore whether baicalein treatment affects notch signaling pathway in human cervical cancers. MATERIALS AND METHODS: Cervical cancer cells were treated with increasing concentrations of baicalein for 24, 48, and 72 h, respectively. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to determine cell viability of cervical cancer cells. The apoptosis rate was determined by FACS assay. Furthermore, the molecular mechanism was investigated. The expression levels of Notch 1, Notch 2, Notch 3, hairy enhancer of split-1 (Hes-1), and Hes-5 were determined by western blotting analysis. RESULTS: MTT assay results revealed that baicalein inhibited cell proliferation of HeLa cells and SiHa cells in a time- and dose-dependent manner. The data from FACS assay demonstrated that baicalein-induced cell apoptosis of cervical cancer cells at the final concentration of 100 µM for 24 h. Furthermore, baicalein treatment downregulated Notch 1/Hes-1, Hes-5 signaling pathway, and there was no obvious change on the expression of Notch 2 and Notch 3. CONCLUSION: Baicalein inhibited the proliferation of human cervical cancer cells via Notch 1/Hes signaling Pathway. The study would provide some new clues in the clinical therapy of human cervical cancers.

Fanconi anemia core complex-dependent HES1 mono-ubiquitination regulates its transcriptional activity.

OBJECTIVE: The Hairy Enhancer of Split 1 (HES1) is a transcriptional repressor that regulates cellular proliferation and differentiation during development. We previously found an interaction between HES1 and Fanconi anemia (FA) proteins. FA is a hematological and developmental disorder caused by mutations in more than 20 different genes. Eight FA gene products form a nuclear core complex containing E3 ligase activity required for mono-ubiquitination of FANCD2 and FANCI, both of which are FA proteins. Given that HES1 interacts with members of the FA core complex, the aim of this study was to determine whether HES1 is mono-ubiquitinated via the FA core complex. RESULTS: We show that HES1 is mono-ubiquitinated on a highly-conserved lysine residue that is located within a FA-like recognition motif. HES1 modification is dependent on a functional FA complex. Absence of HES1 mono-ubiquitination affects transcriptional repression of its own promoter. This study uncovers a novel post-translational modification of HES1 that regulates its transcriptional activity and suggests that ubiquitination of HES1 occurs in a FA core complex-dependent manner.

Hyperhomocysteinemia induces injury in olfactory bulb neurons by downregulating Hes1 and Hes5 expression.

Hyperhomocysteinemia has been shown to be associated with neurodegenerative diseases; however, lesions or histological changes and mechanisms underlying homocysteine-induced injury in olfactory bulb neurons remain unclear. In this study, hyperhomocysteinemia was induced in apolipoprotein E-deficient mice with 1.7% methionine. Pathological changes in the olfactory bulb were observed through hematoxylin-eosin and Pischingert staining. Cell apoptosis in the olfactory bulb was determined through terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) staining. Transmission electron microscopy revealed an abnormal ultrastructure of neurons. Furthermore, immunoreactivity and expression of the hairy enhancer of the split 1 (Hes1) and Hes5 were measured using immunohistochemistry, immunofluorescence, and western blot assay. Our results revealed no significant structural abnormality in the olfactory bulb of hyperhomocysteinemic mice. However, the number of TUNEL-positive cells increased in the olfactory bulb, lipofuscin and vacuolization were visible in mitochondria, and the expression of Hes1 and Hes5 decreased. These findings confirm that hyperhomocysteinemia induces injury in olfactory bulb neurons by downregulating Hes1 and Hes5 expression.

Increased Hes1 expression to regulate differentiation of liver epithelial progenitor cells into cholangiocytes / 西安交通大学学报(医学版)

【Objective】 To use hairy enhancer of split 1 (Hes1) to regulate the differentiation of liver epithelial progenitor cells (LEPCs) into cholangiocytes. 【Methods】 The vectors, pTet-on and pTRE2hyg-Hes1, were transfected into LEPCs. The expression of Hes1 was induced by doxycycline (DOX) with different concentrations (0, 0.1, 1, 5, 10, 50, 100 and 500 μg/mL). The expressions of Hes1, molecular markers of hepatocyte and cholangiocyte, glutathione synthetase (Gss), keratin 19 (Krt19) and hepatic nuclear factor 1β (HNF1β) in LEPCs were verified by Western blotting, RT-PCR, Real-time PCR, immunocytochemistry and immunofluorescence. 【Results】 The expression of Hes1 in LEPCs transfected by pTet-on/pTRE2hyg-Hes1 was increased by 11.21 fold when induced by DOX at 10 ug/mL, which drove the LEPCs to differentiate into biliary epithelial cells. With increasing expression of Hes1, cholangiocyte markers, Krt19 and HNF1β, were significantly upregulated, while the hepatocyte marker, Gss, was obviously downregulated. 【Conclusion】 DOX at 10 μg/mL may induce a suitably up-regulated expression of Hes1 in LEPCs double-transfected by pTet-on and pTRE2hyg-Hes1, and the suitable high-expression rather than over-expression of Hes1 can regulate LEPCs to differentiate into cholangiocytes.

Baicalein suppresses the proliferation of human cervical cancer cells via Notch 1/Hes signaling pathway

Background: Baicalein is an active compound extracted from the roots of Scutellaria baicalensis georgi, which is widely and traditionally used in the anticancer therapy. Notch signaling pathway is usually abnormally activated in kinds of human cancers. The aim of the present study is to investigate the antitumor effects of baicalein in human cervical cancer and explore whether baicalein treatment affects notch signaling pathway in human cervical cancers. Materials and Methods: Cervical cancer cells were treated with increasing concentrations of baicalein for 24, 48, and 72 h, respectively. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to determine cell viability of cervical cancer cells. The apoptosis rate was determined by FACS assay. Furthermore, the molecular mechanism was investigated. The expression levels of Notch 1, Notch 2, Notch 3, hairy enhancer of split-1 (Hes-1), and Hes-5 were determined by western blotting analysis. Results: MTT assay results revealed that baicalein inhibited cell proliferation of HeLa cells and SiHa cells in a time- and dose-dependent manner. The data from FACS assay demonstrated that baicalein-induced cell apoptosis of cervical cancer cells at the final concentration of 100 μM for 24 h. Furthermore, baicalein treatment downregulated Notch 1/Hes-1, Hes-5 signaling pathway, and there was no obvious change on the expression of Notch 2 and Notch 3. Conclusion: Baicalein inhibited the proliferation of human cervical cancer cells via Notch 1/Hes signaling Pathway. The study would provide some new clues in the clinical therapy of human cervical cancers

Protein hairy enhancer of split-1 expression during differentiation of muscle-derived stem cells into neuron-like cells.

Muscle-derived stem cells were isolated from the skeletal muscle of Sprague-Dawley neonatal rats aged 3 days old. Cells at passage 5 were incubated in Dulbecco's modified Eagle's medium supplemented with 10% (v/v) fetal bovine serum, 20 µg/L nerve growth factor, 20 µg/L basic fibroblast growth factor and 1% (v/v) penicillin for 6 days. Cells presented with long processes, similar to nerve cells. Connections were formed between cell processes. Immunocytochemical staining with neuron specific enolase verified that cells differentiated into neuron-like cells. Immunofluorescence cytochemistry and western blot results revealed that the expression of protein hairy enhancer of split-1 was significantly reduced. These results indicate that low expression of protein hairy enhancer of split-1 participates in the differentiation of muscle-derived stem cells into neuron-like cells.