Cinnamaldehyde inhibits pancreatic cancer cell proliferation and sternness via modulation of CD44s/STAT3 signaling pathway / 中国药理学通报

Aim To explore the effects of cinnamaldehyde on the proliferation and stemness of pancreatic cancer PANC-1 cells, and its possible mechanism of action.Methods CCK8 assay was used to detect the effect of cinnamaldehyde treatment on cell viability at different concentrations(0, 5, 15, 20, 30, 50, 70, 100, 150 μmol·L-1)and different intervention time(24, 48, 72 h).CFSE proliferation assay was used to detect the inhibitory effects of cinnamaldehyde on PANC-1 cells.Colony formation assay was employed to determine the colony-forming ability of PANC-1 cells after cinnamaldehyde treatment.The sphere formation assay was employed to detect the effects of cinnamaldehyde on sphere-forming ability in PANC-1 cells.Western blot analysis and qRT-PCR analysis were applied to determine the expression levels of Nanog, Sox-2 and Oct-4.Flow cytometry was used to detect the percentage of CD44+CD24+ cells and ALDH+ cells in cinnamaldehyde treated and untreated PANC-1 cells.Western blot analysis was used to detect the effects of cinnamaldehyde on the expression of CD44s, p-STAT3 and STAT3 in PANC-1 cells.Results Cinnamaldehyde suppressed cell viability in a dose- and time-dependent manner, and inhibited tumor-cell proliferation and colony forming ability significantly in a dose-dependent manner in PANC-1 cells.Sphere-forming assay showed that cinnamaldehyde could significantly inhibit sphere-forming ability in suspension culture of PANC-1 cells.The mRNA and protein expression levels of three stemness-related genes were down-regulated after cinnamaldehyde treatment.In addition, cinnamaldehyde treatment significantly decreased the proportion of CD44+CD24+ cells and ALDH+ cells.Western blotting showed that cinnamaldehyde inhibited the expression of CD44s and p-STAT3, while it had no effect on the expression of STAT3.With the addition of STAT3 activator(Colivelin TFA), the inhibition of cinnamaldehyde on proliferation and tumor-cell stemness in PANC-1 cells was partially rescued.Conclusions Cinnamaldehyde significantly inhibits the proliferation and tumor-cell stemness of pancreatic cancer PANC-1 cells, and the mechanism could be related to the modulation of CD44s/STAT3 signaling pathway.

Overexpression of CD44 Standard Isoform Upregulates HIF-1α Signaling in Hypoxic Breast Cancer Cells

Cluster of differentiation 44 (CD44), a cell surface receptor for hyaluronic acid (HA), is involved in aggressive cancer phenotypes. Herein, we investigated the role of the CD44 standard isoform (CD44s) in hypoxia-inducible factor-1α (HIF-1α) regulation using MCF7 overexpressing CD44s (pCD44s-MCF7). When pCD44s-MCF7 was incubated under hypoxia, levels of HIF-1α, vascular endothelial growth factor, and the HIF-1α response element-derived luciferase activity were significantly increased compared to those in the control MCF7. Incubation of pCD44s-MCF7 cells with HA further increased HIF-1α accumulation, and the silencing of CD44s attenuated HIF-1α elevation, which verifies the role of CD44s in HIF-1α regulation. In addition, the levels of phosphorylated extracellular signal-regulated kinase (ERK) was higher in hypoxic pCD44s-MCF7 cells, and HIF-1α accumulation was diminished by the pharmacological inhibitors of ERK. CD44s-mediated HIF-1α augmentation resulted in two functional outcomes. First, pCD44s-MCF7 cells showed facilitated cell motility under hypoxia via the upregulation of proteins associated with epithelial-mesenchymal transition, such as SNAIL1 and ZEB1. Second, pCD44s-MCF7 cells exhibited higher levels of glycolytic proteins, such as glucose transporter-1, and produced higher levels of lactate under hypoxa. As a consequence of the enhanced glycolytic adaptation to hypoxia, pCD44s-MCF7 cells exhibited a higher rate of cell survival under hypoxia than that of the control MCF7, and glucose deprivation abolished these differential responses of the two cell lines. Taken together, these results suggest that CD44s activates hypoxia-inducible HIF-1α signaling via ERK pathway, and the CD44s-ERK-HIF-1α pathway is involved in facilitated cancer cell viability and motility under hypoxic conditions.

Expression of tumor stem cell markers CD133-2, CD24 and CD44S in head and neck squamous cell carcinoma tissues and their clinical significances / 肿瘤研究与临床

Objective To explore the expressions and significances of the tumor stem cell markers CD133-2,CD24 and CD44s in head and neck squamous cell carcinoma (HNSCC) tissues and their association with the clinical pathologic characteristics.Methods Expressions of CD133-2,CD24 and CD44s were analyzed by immunohistochemistry (SP) in 83 cases of primary HNSCC tissues and 46 cases of normal epithelia.Clinicopathological indexes were assessed.Results In primary HNSCC tissues and normal epithelia,the expression rates of CD133-2 and CD24 were 9.64 ％ (8/83),21.74 ％ (10/46) and 90.36 ％ (75/83),46.67 ％ (21/46)respectively,which had statistically significances (x2 =15.040,5.818,P ＜ 0.05).CD44s expression was detected in primary HNSCC tissues and normal epithelia,but their staining scores had statistical significance (Z =-4.262,P ＜ 0.05).In primary HNSCC tissues,the expression of CD133-2 had negative correlation with differentiation degree (x2 =7.246,P ＜ 0.05),but CD24 and CD44s had positive correlation with differentiation degree (x2 =9.005,44.765,P ＜ 0.05).In addition,the expression of CD44s in primary HNSCC tissues had negative correlation with T classification (x2 =4.650,P ＜ 0.05).Conclusion The expressions of CD24 and CD44s in primary HNSCC tissues are highly up-regulated with tumor cells differentiation,and further research needs to be performed to discover whether or not CD24 and CD44s could be the markers of tumor stem cells of HNSCC.The expression of CD133-2 in primary HNSCC tissues is highly down-regulated with tumor cell differentiation.As one of the tumor stem cell markers of HNSCC,CD133-2 may play an important role in the development and clinical outcomes of tumor.

CD44s and CD44v6 Are Predominantly Expressed in the Non-germinal Center B-Cell-like Type of Diffuse Large B-Cell Lymphomas

BACKGROUND: CD44 protein is known as a homing cellular adhesion molecule that is linked to diverse cellular functions such as adhesion, migration and invasion, which are all important in cancer progression and metastasis. The expression of CD44 standard and variant isoforms (CD44 standard isoform [CD44s] and CD44 splice variants containing exon v6 [CD44v6], respectively) is associated with an unfavorable clinical outcome in various neoplasms. METHODS: Forty patients who were diagnosed with diffuse large B-cell lymphoma (DLBCL) through biopsy at Hanyang University Hospital between 1996 and 2003 were included in this study. CD44 proteins expression was analyzed by immunohistochemical staining on a tissue microarray and the correlation of CD44 with the types of DLBCL and clinical parameters, including the factors defined by the International Prognostic Index, was evaluated. RESULTS: A high CD44s and intermediate to strong CD44v6 expression, including cytoplasmic membranous staining patterns, was present in 35% (14/40) and 25% (10/40) of DLBCL patients, respectively. High CD44s expression was correlated significantly with non-germinal center B-cell-like types (non-GCB, p=0.004) and patients with old age (p=0.041). CONCLUSIONS: High CD44s expression may be significantly associated with the non-GCB type compared to the GCB type and may be essential to the prediction of disease outcome in tumor stage III in DLBCL patients.

Expression of the Transmembrane Glycoprotein CD44s Is Potentially an Independent Predictor of Recurrence in Hepatocellular Carcinoma

BACKGROUND/AIMS: Cluster differentiation 44 standard isoform (CD44s) is a transmembrane glycoprotein. CD44s is a known prognostic factor in various cancers, due to its involvement in tumor cell growth, invasion and metastasis. Its prognostic role, however, is debated because it can be a positive or negative prognostic factor depending on tumor type and is still an ambiguous prognostic indicator in other cancers, especially hepatocellular carcinoma (HCC). We investigated the relationship between CD44s expression and survival in HCC patients. METHODS: A total of 260 HCC samples were collected to generate a tissue microarray. Staining of the arrays with a primary mouse CD44s monoclonal antibody was followed by evaluation of the relationship between CD44s expression and tumor differentiation. The effect of CD44s expression on patient survival was analyzed. RESULTS: CD44s protein expression correlated with histological grade (most and worst Edmondson grade) of the HCC (p=0.029 and p=0.039, respectively) and adversely affected the disease free survival period based on univariate and multivariate analyses (p=0.038 and p=0.077, respectively). CONCLUSIONS: High CD44s protein expression correlates with shorter disease free survival and poorly differentiated HCC. CD44s-targeted therapy may be efficacious for HCC treatment in the future.

The Effect and Significance of Cinobufagin on the Expression of CD44s in Ovarian Carcinoma 3AO Cells / 国际中医中药杂志

Objegtive To investigate the effect and significance of cinobufagin on the expression of CD44s in ovarian carcinoma 3AO cells cultured in vitro.Methods Ovarian carcinoma 3AO cells were cultured in vitro;after the intervention of tinobufagin with different concentration,the expression of CD44s was measured by RT-PCR,and the cellular apoptosis was tested by flow cytometry.Results Ovarian carcinoma 3AO cells expressed CD44S mRNA of 1.3±0.1 and the cellular apoptofic rate was(2.31±0.98)%:0.25 mg/ml cinobufagin did not affect CD44s mRNA and cellular apoptofic rate(P ＞0.05):when 2.5 mg/ml cinobufagin was intervened,the expression of CD44s mRNA was 1.0±0.1 and the cellular apoptotic rate Was(28.69±4.16)%,showing significant difference comparing with the control group of ovarian carcinoma 3AO ceils(P＜0.05):the intervention of 25 mg/ml cinobufagin was similar to that of 0.25 mg/ml cinobuhgin(P＞0.05).3AO cells.

Studies of CD44s and CD44v6 Expressions and Correlation with Clinicopathologic Parameters in Gastric Carcinoma

BACKGROUND: The aim of this study is to elucidate the relationship between the CD44s and CD44v6 expression level and the biological characteristics of a gastric carcinoma. METHODS: CD44s and CD44v6 expression was investigated in 56 gastric carcinomas, 18 dysplasias, and 22 normal mucosae by immunohischemical staining. RESULTS: The CD44s and CD44v6 expression rates in gastric carcinomas, dysplasia, and normal mucosae were 80.3% and 83.9%, 72.2% and 77.8%, and 13.6% and 4.5%, respectively. Statistical analysis showed significant difference after comparing a gastric carcinomas and dysplasia to the normal mucosae (p<0.001). The CD44s and CD44v6 expression rates in the cases with invasion to the muscle proper and serosa were 60.7% and 57.1%, and 82.4% and 88.2%, respectively. Both showed a significant statistical difference compared to the expression rates in the cases with invasion to the mucosae and submucosae. The CD44s and CD44v6 expression rates in gastric carcinomas with a lymph node metastasis showed a statistically significant difference compared to those without a lymph node metastasis (p<0.001 and p<0.01, respectively). CD44s and CD44v6 were also expressed in the normal basal cells around gastric carcinomas. CONCLUSIONS: The CD44s and CD44v6 expression showed a significant relationship with gastric carcinogenesis, toward an aggressive biologic behavior.

Expression of CD44S in benign, borderline, and malignant serous epithelial ovarian tumors / 대한산부인과학회잡지

OBJECTIVE: To determine the CD44S expression in the development and progression of serous ovarian tumor. METHODS: An immunohistochemical stain was undertaken of a series of 12 cases of benign, 15 cases of borderline, 30 cases of malignant serous ovarian tumors. Then, we analyzed the expression of CD44S to see whether expression of this adhesion molecule by tumor cells correlated with clinicopathological factors. RESULTS: In twelve benign tumors, CD44S was not expressed. Four of fifteen (27%) borderline tumors were positive for CD44S. Twelve of thirty (40%) malignant tumors were positive for CD44S. CD44S expression rate was significantly associated with non-benign tumors (P=0.003). But, there was no significant correlation between CD44S expression and stage, grade, peritoneal seeding or lymph node metastasis. CONCLUSION: Since benign ovarian tumors show negative CD44S, positive CD44S can provide partial diagnostic aid in ovarian malignant tumor.

The Expression of MUC1 and CD44s in Non-small Cell Lung Cancer / 결핵

BACKGROUND: MUC1 mucin is a heavily glycosylated large glycoprotein and is expressed aberrantly in carcinoma. CD44 is polymorphic family of cell surface glycoproteins participating in cell-cell adhesion and modulat ion of the cell-matrix interaction. MUC1 mucin and CD44 expression have been implicated in a tumor invasion and metastasis in certain malignancies. In this study, the expression of MUC1 and the standard form of CD44(CD44s) was examined in non-small cell lung cancer (NSCLC). METHODS: Immunohistochemical staining using monoclonal antibodies including MUC1 glycoprotein and CD44s was performed on 80 NSCLC surgical specimens. The association between MUC1 and CD44s expression and the histological type and tumor stage was investigated. RESULTS: Depolarized MUC1 expression in more than 10% of cancer cells was found in 12 (27.9%) out of 43 squamous cell carcinomas (SCCs) and 12 (32.4%) out of 37 adenocarcinomas (ACs). It was not associated with the tumor histological type and the TNM-stage in SCCs. Depolarized MUC1 expression correlated with the N-stage in ACs (p=0.036). CD44s was expressed in 36 (83.7%) out of 43 SCCs and 14(37.8%) out of 37ACs. Reduced CD44s expression correlated with the N-stage (p=0.031) and the TNM-stage (p=0.0006) in SCCs. CONCLUSIONS: Depolarized MUC1 expression was related to the nodal stage in NSCLC adenocarcinoma. Reduced CD44s expression was related to nodal involvement and the TNM-stage in squamous cell carcinoma. This suggests that MUC1 and CD44s expression in NSCLC might play important roles in tumor progression and can be used as prognostic variables.

Investigation on the relative association of CD44 and fibronectin expression with endometriosis / 中国免疫学杂志

Objective:To study the expression of CD44s and fibronectin in endometriosis.Methods:Expression of CD44s and fibronectin were evaluated by immunohistochemistry method in entopic from 22 patients and in ectopic from 45 patients with endometriosis and the endometrial tissues of 20 non-endometriosis.Results:CD44s is restricted to epithelial cells;fibronectin is restricted to stroma,their expression remained constants thoughout the menstrual cycle.Decreased expression of CD44s and fibronectin in epithelial and stroma in endometriosis was found in eutopic endometrium(P

Immunohistochemical Expression of CD44 Isoforms in Eccrine Gland Tumors / 대한피부과학회지

BACKGROUND: CD44 is the principal cell surface receptor for hyaluronate and exists as multiple isoforms generated by the alternative splicing of up to 10 variant exons. Although certain isofroms may play a role in tumor progression and metastasis formation, the precise function and expression of the variant isoforms are less clear. Since on normal eccrine glands CD44 standard isoform(CD44s) is expressed only in eccrine coil secretory cells, it can be considered as a possible marker of this type of differentiation. However little is known about the expression of CD44 variant isofroms(CD44v) in eccrine gland tumors. OBJECTIVE: The purpose of this study was to investigate the immunohistochemical expression of different CD44 isoforms(CD44s, CD44v4, CD44v6) in the eccrine gland tumors. METHODS: Formalin-fixed and paraffin-embedded tissues from 2 cases of eccrine hidrocystoma, 5 cases of syringoma, 2 cases of eccrine poroma, 2 cases of syringofibroadenoma, 2 cases of nodular hidradenoma were immunolabelled with monoclonal antibody directed CD44s, CD44v4, and CD44v6. RESULTS: Except for syringofibroadenoma, the most tumors cells with eccrine ductal differentiation showed negative staining for CD44s, and positive staining for CD44v4 and CD44v6. Syringofibroadenoma exhibited positive staining for CD44s and CD44v4, but negative staining for CD44v6. Eccrine poroma showed negative staining for CD44s, positive staining for CD44v4, and variable intensity of staining for CD44v6 in different areas of the tumors. In case of nodular hidradenoma, small tubular lumina and clear cells were positive for CD44s. CONCLUSION: Our results suggest that CD44 isoforms can not be a useful marker for an eccrine gland tumor with specific differentiation, but its characteristic pattern of distribution might reflect the variety of functional roles of CD44 isoforms in tumorigenesis of eccrine gland tumors.

Expression of CD44 Standard, Variant 6 and Relationship to the Lymph Node Metastasis in Gastric Adenocarcinoma / 대한암학회지

PURPOSE: The transmembrane glycoprotein CD44 exists in a variety of isoforms generated by alternative splicing of the pre-mRNA. We studied the role of CD44-standard (CD44s) and CD44-variant6 (CD44v6) in gastric adenocarcinoma. MATERIALS AND METHODS: Immunohistochemical staining was performed in 101 patients with gastric adenocarcinoma who underwent radical gastrectomy at KGCC, Seoul Paik Hospital. The relationship of CD44s, CD44v6 expressions to the clinicopathologic parameters, p53 and Ki-67 were evaluated. RESULTS: CD44s and CD44-v6 expressions were found in 56.4% and 48.5%, respectively. CD44s expression was significantly correlated with lymph node metastasis, lymphatic invasion, and Borr mann type. CD44v6 expression was significantly correlated with sex, lymph node metastasis, lymphatic invasion, and perineural invasion and had a tendency toward p53 expression. In inte stinal type adenocarcinoma, CD44s expression had correlations with lymph node metastasis and CD44v6 had correlations with lymph node metastasis, lymphatic invasion. However, in diffuse type adenocarcinoma, CD44s and CD44v6 expressions had correlations with only Borrmann type. In multivariate analysis, lymph node metastasis was the most significant risk factor for CD44s and CD44v6 expressions in total cases and intestinal type adenocarcinoma. CONCLUSION: These data suggest that expression of CD44 v6 may play an important role in the regulation of lymph node metastasis in intestinal type adenocarcinoma of stomach.