OBJECTIVE: To observe the effects of a traditional Chinese medicine (TCM) capsule for replenishing qi, nourishing yin and activating blood on Notch/Hes1 signaling pathway in the renal tissue and vascular endothelial CD34 and CD144 expressions in a rat model of diabetic nephropathy. METHODS: Rat models of early-stage diabetic nephropathy were established by left nephrectomy and high- fat and high- sugar feeding combined with intraperitoneal injection of STZ. The rats were randomized into model group, benazepril group, and high-, moderate-, and low-dose TCM capsule groups for corresponding treatments, with 6 normal rats as the control group. After 8 weeks of drug treatment, blood glucose and 24-h urinary albumin of the rats were measured, and the renal histopathology was observed with HE staining; Hes1 expression in the renal tissue was detected with immunohistochemical staining, and the renal expressions of CD34 and CD144 were detected using Western blotting. RESULTS: Compared with the normal control group, the rat models of diabetic nephropathy showed obvious abnormalities in 24- h urinary albumin and expressions of Hes1, CD34 and CD144d. The TCM capsule at both the high and moderate doses significantly reduced 24-h urinary albumin in the rats; the renal expressions of Hes1 and CD34 was significantly reduced in all the dose groups, and the expression of CD144 was significantly reduced in the high- dose group. Compared with benazepril group, the TCM capsule obviously reduced CD34 expression at all the 3 doses and lowered CD144 expression at the low dose. Histopathologically, the rats in the model group showed glomerular hypertrophy, increased mesenteric matrix, thickening and widening of the mesenteric membrane, and nodular hyperplasia. These pathologies were obviously alleviated by treatment with the TCM capsule at the high and moderate doses. CONCLUSIONS The Traditional Chinese medicine (TCM) capsule for replenishing qi, nourishing yin and activating blood can reduce Hes1, CD34 and CD144 in kidney tissue of model rats, play a protective role on kidney function and delay the development of DN.
Animals ; Diabetic Nephropathies ; Drugs, Chinese Herbal ; Medicine, Chinese Traditional ; Qi ; Rats ; Signal Transduction ; Transcription Factor HES-1

OBJECTIVETo elucidate the impact of Hes1 on the proliferation and apoptosis of acute myeloid leukemia (AML) cells.

METHODSThe expression levels of Hes1 and p21 in AML patient samples and myeloid leukemia cell lines were analyzed by real-time PCR. Hes1 was up-regulated by retrovirus transfection in AML cell lines and the proliferation capacity were assayed by MTT, cell cycle by Hoechst/PY, apoptosis by AnnexinV.

RESULTSThe expression of Hes1 in primary AML cells and HL-60, U937, KG1a cell lines were 0.67 ± 0.24, 0.59 ± 0.43, 0.42 ± 0.03, and 0.32 ± 0.26, respectively, and p21 were 0.54 ± 0.01, 0.44 ± 0.12, 0.36 ± 0.12, and 0.59 ± 0.43, respectively. Hes1 expression levels after transduction in HL-60, U937, KG1a were 4.9 ± 0.2, 5.2 ± 0.4, 5.8 ± 0.5, respectively. Induced activation of Hes1 led to AML cells growth arrest and apoptosis, which was associated with an enhanced p21 expression. Besides, activated Hes1 led to AML cells growth inhibition in vivo.

CONCLUSIONHes1 could mediate growth arrest and apoptosis in AML cells, which may be a novel target for AML.

Apoptosis ; Basic Helix-Loop-Helix Transcription Factors ; Cell Cycle ; Cell Line, Tumor ; Homeodomain Proteins ; Humans ; Leukemia, Myeloid, Acute ; Transcription Factor HES-1 ; Up-Regulation

Mesenchymal stem cell (MSC)-based therapy has emerged as a promising treatment for spinal cord injury (SCI), but improving the neurogenic potential of MSCs remains a challenge. Mixed lineage leukemia 1 (MLL1), an H3K4me3 methyltransferases, plays a critical role in regulating lineage-specific gene expression and influences neurogenesis. In this study, we investigated the role and mechanism of MLL1 in the neurogenesis of stem cells from apical papilla (SCAPs). We examined the expression of neural markers, and the nerve repair and regeneration ability of SCAPs using dynamic changes in neuron-like cells, immunofluorescence staining, and a SCI model. We employed a coimmunoprecipitation (Co-IP) assay, real-time RT-PCR, microarray analysis, and chromatin immunoprecipitation (ChIP) assay to investigate the molecular mechanism. The results showed that MLL1 knock-down increased the expression of neural markers, including neurogenic differentiation factor (NeuroD), neural cell adhesion molecule (NCAM), tyrosine hydroxylase (TH), βIII-tubulin and Nestin, and promoted neuron-like cell formation in SCAPs. In vivo, a transplantation experiment showed that depletion of MLL 1 in SCAPs can restore motor function in a rat SCI model. MLL1 can combine with WD repeat domain 5 (WDR5) and WDR5 inhibit the expression of neural markers in SCAPs. MLL1 regulates Hairy and enhancer of split 1 (HES1) expression by directly binds to HES1 promoters via regulating H3K4me3 methylation by interacting with WDR5. Additionally, HES1 enhances the expression of neural markers in SCAPs. Our findings demonstrate that MLL1 inhibits the neurogenic potential of SCAPs by interacting with WDR5 and repressing HES1. These results provide a potential therapeutic target for promoting the recovery of motor function in SCI patients.
Animals ; Humans ; Rats ; Cell Differentiation ; Intracellular Signaling Peptides and Proteins/therapeutic use* ; Leukemia/metabolism* ; Mesenchymal Stem Cells ; Neurogenesis ; Stem Cells ; Transcription Factor HES-1/metabolism*

OBJECTIVE: To analyze and investigate the expression levels of HES1, C-MYC and NF-kB in peripheral blood of patients with T cell acute lymphoblastic leukemia (T-ALL) and their significance. METHODS: Sixty patients with T-ALL and 60 patients with acute myelogenous leukemia (AML) diagnosed in our hospital from June 2012 to March 2015 were enrolled in T-ALL group and AML group, respectively. Another 30 healthy people were enrolled in the control group. Peripheral blood was collected to detect the expression levels of HES1, C-MYC and NF-kB by RT-PCR. The general data and the expression of HES1, C-MYC and NF-kB in peripheral blood were compared among the patients with different type of leukemia, cytogenetical types and different prognosis. RESULTS: There was no significant difference in baseline data, such as age and sex among the 3 groups (P＞0.05). The Hb level, WBC and Plt count, BM blast cell ratio in T-ALL and AML groups all were significantly higher than those in control group (P＜0.01), but there were no statistical difference in above-mentioned indicators between T-ALL and AML groups (P＞0.05). The expression levels of HES1, C-MYC and NF-kB in peripheral blood among 3 groups were significantly differenct (P＜0.01), the expressions levels of HES1, C-MYC and NF-kB in T-ALL and AML groups were significantly higher than those in control were significantly group (P＜0.01), moreover, the expression levels of above-mentional indicators in T-ALL groups were significantly higher than than those in AML group (P＜0.01). The expression levels of HES1, C-MYC and NF-kB iin T-ALL patients with poor prognosis were significantly higher than those in T-ALL patients with favorable prognosis (P＜0.01); the expression levels of HES1, C-MYC and NF-kB in peripheral blood of patients with different theraptic efficacy were follow: complete remission group＜partial remission group＜no remission group (P＜0.01). CONCLUSION The HES1, C-MYC and NF-kB are highly expressed in peripheral blood of the patients with T-ALL, moreover, the expression levels maybe different, because of the cytogenetic, and theraptic efficacy.
Humans ; Leukemia, Myeloid, Acute ; NF-kappa B ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma ; Remission Induction ; T-Lymphocytes ; Transcription Factor HES-1

OBJECTIVETo observe the effect of activation of Notch1 signaling pathway by Notch intracellular domain (NICD) plasmid transfection on pancreatic cancer cell proliferation and explore the underlying mechanism.

METHODSThe transfection rates were observed under microscope with fluorescence stimulation, and mRNA expression levels of Hes1 were detected by real-time PCR. Cell proliferation changes were evaluated by CCK-8 after NICD and control plasmid transfection in pancreatic cancer cells. Caspase 3 activity was examined using a caspase 3 detection kit.

RESULTSThe transfection rates of NICD plasmid were up to 80% by fluorescence stimulation observation. Hes1 expression was significantly increased after NICD plasmid transfection, suggesting the activation of Notch1 signaling pathway. NICD plasmid transfection significantly promoted cancer cell proliferation compared to control plasmid transfeciton. The activities of caspase 3 were obviously decreased after NICD plasmid transfection in 3 pancreatic cancer cell lines.

CONCLUSIONActivation of Notch1 signaling pathway by NICD plasmid transfection can promote the proliferation of pancreatic cancer cells by inhibiting the apoptosis pathway.

Apoptosis ; Basic Helix-Loop-Helix Transcription Factors ; metabolism ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Homeodomain Proteins ; metabolism ; Humans ; Pancreatic Neoplasms ; metabolism ; pathology ; Plasmids ; Receptor, Notch1 ; genetics ; metabolism ; Signal Transduction ; Transcription Factor HES-1 ; Transfection

The aim of this study was to explore the effect of DAPT (N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycinet-butyl ester) on proliferation in vitro of human multiple myeloma cell line RPMI8226 and its underlying mechanism. The proliferation of RPMI8226 cells was detected by CCK-8 method; flow cytometry was employed to assay the cell apoptosis rate;the expressions of Notch1 and Hes1 proteins were detected by Western blot. The results indicated that the proliferation of human RPMI8226 cells significantly decreased after treatment with DAPT 0.5 - 5.0 µmol/L for 24 - 72 h (P < 0.05) in a concentration- and time-dependent manner. DAPT significantly induced apoptosis of RPMI8226 cells (P < 0.05). The expressions of Notch1 and Hes1 proteins were gradually downregulated with the increase of DAPT concentration. It is concluded that the DAPT can inhibit the proliferation of RPMI8226 cells, which may be related with the down-regulation of the protein expression of Notchl and Hes1.
Apoptosis ; drug effects ; Basic Helix-Loop-Helix Transcription Factors ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dipeptides ; pharmacology ; Homeodomain Proteins ; metabolism ; Humans ; Multiple Myeloma ; metabolism ; pathology ; Receptor, Notch1 ; metabolism ; Transcription Factor HES-1

OBJECTIVETo study the changes of Hes-1, the target gene of Notch signaling pathway, and its relationship with airway inflammation and remodeling in a rat model of asthma.

METHODSForty-eight rats were randomly divided into an asthma group and a control group. The rats in the asthma group were sensitized and challenged by ovalbumin (OVA), and normal saline was used in the control group. Two groups were further divided into 3 subgroups according to time points after challenging, i.e. 4 weeks, 8 weeks and 12 weeks (n=8 rats each). Pathological changes of lungs were observed by light microscopy and the thickness of bronchial smooth muscle layer (Wam) was measured. The levels of IL-4 and INF-γ in rat serum and bronchoalveolar lavage fluids (BALF) were measured using ELISA. Expression levels of Hes-1 protein and mRNA were determined by immunohistochemistry and quantitative real-time PCR respectively.

RESULTSTogether with the extension of challenging, the Wam of rats in the asthma group increased, a decrease of INF-γ level and an increase of IL-4 level in serum and BALF were also observed, and the differences were statistically significant compared with those in the corresponding control group (P<0.05). Hes-1 protein and mRNA levels also increased gradually after OVA challenging and were higher than those in the control group (P<0.05). The levels of Hes-1 protein and mRNA were positively correlated with Wam and IL-4 in serum and BALF, but were inversely correlated with INF-γ in serum and BALF (P<0.05).

CONCLUSIONSLevels of Hes-1 protein and mRNA increased, which were closely related with the levels of airway inflammatory factors and remodeling of airway smooth muscle. Hes-1 may play an important role in the pathogenesis of asthma.

Airway Remodeling ; Animals ; Asthma ; etiology ; Basic Helix-Loop-Helix Transcription Factors ; analysis ; genetics ; physiology ; Disease Models, Animal ; Homeodomain Proteins ; analysis ; genetics ; physiology ; Interferon-gamma ; analysis ; Interleukin-4 ; analysis ; Male ; Rats ; Rats, Sprague-Dawley ; Transcription Factor HES-1

OBJECTIVETo detect the expression of Math1, Hes1 and Hes5 in greater epithelial ridge (GER) cells of rat cochlear and explore their influence on hair cell differentiation.

METHODSPostnatal day 0 (P0), day 1 (P1) , day 3 (P3) day 4 (P4) and day 5 (P5) rat cochlear were dissected respectively and then pure GER cells were separated by a combinatorial approach of attachment and mechanical separation. The total RNA of GER cells was extracted by Trizol one step method and the expression of Math1, Hes1 and Hes5 in GER cells was detected with reverse transcription polymerase chain reaction.

RESULTSMath1 was expressed in P0 - P5 rat GER cells and Hes1 was expressed only in PO - P3 rat GER cells, while there was no expression of Hes5 in P0 - P5 rat GER cells.

CONCLUSIONSProbably only when the expression of Math1 reaches a certain level can it induce GER cells to differentiate into hair cells. Meanwhile this process might controlled by Hes1 to some extent.

Animals ; Basic Helix-Loop-Helix Transcription Factors ; genetics ; metabolism ; Cell Differentiation ; Cochlea ; cytology ; metabolism ; Epithelial Cells ; metabolism ; Gene Expression Regulation, Developmental ; Hair Cells, Auditory ; cytology ; metabolism ; Homeodomain Proteins ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Transcription Factor HES-1

OBJECTIVE: To explore the relationship between the expression levels of JARID1B,Hes1 and MMP-9 genes and the stages of chronic myelogenous leukemia(CML) and the curative effect of imatinib mesylate (IM). METHODS: Peripheral blood samples of 15 cases of CML in chronic phase and 10 cases of CML in progressive phase were collected from the Hematology Department of Taihe Hospital affiliated to Hubei University of Medicine and 15 cases of healthy people in the Physical Examination Center. CML patients were divided into effective group and ineffective group based on the efficacy after treatment with IM, then real-time PCR was used to detect the expression levels of JARID1B, Hes1 and MMP-9 mRNA, finally, the differences in the level of gene expression and their correlations with CML stages and IM curative efficacy were analysed. RESULTS: The expression levels of Hes1 and MMP-9 in initially diagnosed patients in chronic and progressive phase without IM treatment were significantly higher than those of health people（P＜0.05）. There was no significant difference in the expression level of JARID1B between chronic phase patients and health people（P＞0.05）, but the expression level of JARID1B in the progressive phase patients was higher than that of health people （P＜0.05）. The expression levels of JARID1B and Hes1 in the IM-effective group were not significantly different from those in the IM-ineffective group (P=0.85,P=0.82）, while the expression level of MMP-9 in the IM-effective group [JP2]was significantly lower than that in the IM-ineffective group（P＜0.05）. CONCLUSION The expression levels of JARID1B Hes1 and MMP-9 relate with the different phase of CML; The expression levels of JARID1B and Hes1 have not significant relationship with IM curative efficacy, the MMP-9 gene expression level relates with IM curative efficacy.
Antineoplastic Agents ; therapeutic use ; Humans ; Imatinib Mesylate ; therapeutic use ; Jumonji Domain-Containing Histone Demethylases ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; drug therapy ; Matrix Metalloproteinase 9 ; Nuclear Proteins ; Repressor Proteins ; Transcription Factor HES-1

The Notch signaling pathway has been implicated in the regulation of cell-fate decisions such as differentiation of embryo stem cells and neural stem cells into neurons. We cultured human mesenchymal stem cells (hMSCs) in vitro and induced hMSCs to differentiate into neural cells by beta-mercaptoethanol (beta-ME), DMSO and 3-tert-butyl-4-hydroxyanisole (BHA). Immunocytochemistry was utilized to detect neuron-specific enolase (NSE) and Nissl body, and flow cytometry was used to determine cell growth phases. The expressions of signal molecules involved in the Notch pathway such as Notch1, Jagged 1 (JAG1), presenilin 1 (PS1) and hairy and enhancer of split 1(HES1) were observed by RT-PCR and immunofluorescent techniques. The results were as follows: (1) Before induction, the percentage of hMSCs at G(0)/G(1) was 58.5%, and the percentage at S+G(2)/M was 41.5%. After induction, the percentage of hMSCs at G(0)/G(1) increased to 73.1%, 76.2% and 78.1%, respectively on days 2, 4 and 6, and the percentage at S+G(2)/M decreased to 26.8%, 24.8% and 21.9%, respectively; The percentage of NSE-positive cells reached (77+/-0.35) %; Nisslos staining was positive in cytoplasm. (2) Notch1 and JAG1 were both expressed in hMSCs before and after induction, but the mRNA expressions of both Notch1 and JAG1, detected by RT-PCR, decreased obviously after induction(P<0.05). Notch1 mRNA/beta-actin was 1.157, 0.815, 0.756 and 0.570, and JAG1 mRNA/beta-actin was 0.437, 0.350, 0.314 and 0.362, respectively, on days 0, 2, 4 and 6 after induction. The Notch pathway activation participant PS1 mRNA and Notch pathway target gene HES1 mRNA also decreased apparently after induction (P<0.05), and their mRNA/beta-actin was 0.990, 0.449, 0.441, 0.454 and 0.370, 0.256, 0.266, 0.240 on days 0, 2, 4 and 6, respectively. These observations indicate that the expressions of Notch signal molecules were suppressed when hMSCs were induced to differentiate into neural cells. Based on these findings, we propose that low level of Notch signaling activation may contribute to neural cell differentiation.
Basic Helix-Loop-Helix Transcription Factors ; genetics ; Calcium-Binding Proteins ; genetics ; Cell Cycle ; Cell Differentiation ; Flow Cytometry ; Homeodomain Proteins ; genetics ; Humans ; Intercellular Signaling Peptides and Proteins ; genetics ; Jagged-1 Protein ; Membrane Proteins ; genetics ; Mesenchymal Stromal Cells ; cytology ; Neurons ; cytology ; Receptor, Notch1 ; genetics ; Receptors, Notch ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Serrate-Jagged Proteins ; Signal Transduction ; Transcription Factor HES-1