Decreased CD44 Expression Research Articles

MS364 COSTS AND CLINICAL OUTCOMES IN THE PATIENTS WITH OFF-PUMP VS. ON-PUMP CORONARY ARTERY BYPASS SURGERY

Malignant pleural mesothelioma (MPM) is a highly aggressive malignancy in which the mitogen-activated protein kinase pathway plays a critical role in the regulation of tumorigenesis. Hyaluronic acid (HA) is a major component of the extracellular matrix, and elevated HA levels with a concurrent increase in malignant properties are associated with MPM.We evaluated the effects of trametinib, a mitogen-activated protein kinase (MEK) inhibitor, and 4-methylumbelliferone (4-MU), an HA synthesis inhibitor, alone and in combination on MPM cells in vitro and in vivo. We studied the effects of trametinib, 4-MU, and their combination on MPM cells by using cell viability assays, Western blot analysis, and a mouse xenograft model.Trametinib and 4-MU exhibited antiproliferative activity in MPM cells. Trametinib blocked MEK-dependent extracellular signal-regulated kinase (ERK) phosphorylation and decreased CD44 expression in a concentration-dependent manner. Trametinib inhibited the expression of Fra-1 (the activator protein 1 [AP1] component), inhibited ERK phosphorylation, and decreased CD44 expression. 4-MU inhibited ERK phosphorylation but not CD44 expression. In a mouse xenograft model, trametinib and 4-MU alone suppressed tumor growth compared with a control. The combination had a greater inhibitory effect than either monotherapy. Immunohistochemical analysis showed that trametinib treatment alone significantly reduced expression of programmed cell death 1 ligand 1. Furthermore, the combination of trametinib and 4-MU resulted in higher expression of programmed cell death 1 and programmed cell death 1 ligand 1 than did the 4-MU treatment alone.Our results suggest that trametinib and 4-MU are promising therapeutic agents in MPM and that further study of the combination is warranted.

Y-Box Binding Protein-1 Induces the Expression of CD44 and CD49f Leading to Enhanced Self-Renewal, Mammosphere Growth, and Drug Resistance

Y-box binding protein-1 (YB-1) is an oncogenic transcription/translation factor expressed in >40% of breast cancers, where it is associated with poor prognosis, disease recurrence, and drug resistance. We questioned whether this may be linked to the ability of YB-1 to induce the expression of genes linked to cancer stem cells such as CD44 and CD49f. Herein, we report that YB-1 binds the CD44 and CD49f promoters to transcriptionally upregulate their expressions. The introduction of wild-type (WT) YB-1 or activated P-YB-1(S102) stimulated the production of CD44 and CD49f in MDA-MB-231 and SUM 149 breast cancer cell lines. YB-1-transfected cells also bound to the CD44 ligand hyaluronan more than the control cells. Similarly, YB-1 was induced in immortalized breast epithelial cells and upregulated CD44. Conversely, silencing YB-1 decreased CD44 expression as well as reporter activity in SUM 149 cells. In mice, expression of YB-1 in the mammary gland induces CD44 and CD49f with associated hyperplasia. Further, activated mutant YB-1(S102D) enhances self-renewal, primary and secondary mammosphere growth, and soft-agar colony growth, which were reversible via loss of CD44 or CD49f. We next addressed the consequence of this system on therapeutic responsiveness. Here, we show that paclitaxel induces P-YB-1(S102) expression, nuclear localization of activated YB-1, and CD44 expression. The overexpression of WT YB-1 promotes mammosphere growth in the presence of paclitaxel. Importantly, targeting YB-1 sensitized the CD44(High)/CD24(Low) cells to paclitaxel. In conclusion, YB-1 promotes cancer cell growth and drug resistance through its induction of CD44 and CD49f.

Expression of adhesion molecules in first trimester spontaneous abortions and their role in abortion pathogenesis

Early placental development is associated with complex regulatory mechanisms, and molecular communication problems that arise during the developmental process are dangerous for continuation of the pregnancy. As studies on the process of invasion and migration of trophoblast cells have shown the importance of cell-cell and cell-matrix interactions, we examined the effects of adhesion molecules on the mechanism(s) of spontaneous abortions and compared them to elective abortion materials using histopathological and immunohistochemical methods. To the best of our knowledge, this is the first study to investigate adhesion molecules in spontaneous abortions. Curettage materials from abortions were examined retrospectively in the Department of Pathology, Zonguldak Karaelmas University School of Medicine, Zonguldak, Turkey. CD31/PECAM-1 (endothelial cell marker), CD44v (variant 3), E-cadherin, CD54/ICAM-1, and CD106/VCAM-1 expression profiles were evaluated by immunohistochemistry, and cellular localization was determined under light microscopy. The results of spontaneous abortions were compared to those of elective abortions. The staining percentages of CD31, CD44, CD106, and E-cadherin decreased in cases of spontaneous abortion, but CD54 (ICAM-1) expression increased. Statistically significant differences were detected between spontaneous and elective abortion materials with regard to cytotrophoblasts (CTs), syncytiotrophoblasts (STs), and extravillous trophoblasts (EVTs) with the anti-CD31 antibody (p=0.0001). In addition, CD54 (p=0.007 and p=0.002) and E-cadherin (p=0.002 and p=0.02) expression in CTs and STs, respectively, were significantly different. Furthermore, CD44 expression (p=0.003) in decidual (D) cells and CD106 (p=0.0001) expression in vessels of endometrial (E) and villous tissues were also significantly different. Decreased CD31 expression in CTs that invade the spiral arterioles and mimic E cells in spontaneous abortion cases suggests that CD31/PECAM-1 is an important molecule in uteroplacental adequacy. Moreover, diminished expression of CD44 in D cells caused impaired stroma-villous connections. Enhancement of ICAM-1 in placental and invading STs may be useful as a diagnostic marker for patients who may have a tendency to have spontaneous abortions. A down-regulation of E-cadherin was observed, which may be responsible for impaired CT differentiation and loss of the pregnancy. Furthermore, decreased VCAM-1 expression in spontaneous abortions may be consistent with the importance of VCAM-1 in trophoblast-endothelial cell interactions. Many adhesion molecules are known to be effective in the normal development of a pregnancy, and the analysis of adhesion molecules in spontaneous abortions will provide useful information for clarifying the physiopathology of spontaneous abortions.

Effects of Age and Nutrition on Expression of CD25, CD44, and l-Selectin (CD62L) on T-cells from Neonatal Calves

Effects of the plane of nutrition and age on the proliferation and activation of lymphocyte subsets from milk replacer-fed calves were investigated in vitro. Holstein calves were fed a standard (0.45kg/d of a 20% crude protein, 20% fat milk replacer, n=4) or intensified (1.14kg/d of a 28% crude protein, 20% fat milk replacer, n=4) diet from 1 to 8wk of age. Average daily weight gain of intensified-diet (0.66kg/d) calves was greater than that of standard-diet (0.27kg/d) calves. Relative to the pokeweed mitogen-induced responses of CD4+ cells from steers (5 to 6 mo of age), CD4+ cells from 1-wk-old calves showed decreased proliferative activity, delayed increase in CD25 expression, and no demonstrable increase in CD44 expression or decrease in CD62L expression. Calf CD8+ and γδT-cell receptor+ cells, unlike T-cells from the older animals, did not demonstrate decreased expression of CD62L after stimulation with mitogen. The increased expression of CD44 by mitogen-stimulated γδT-cell receptor+ cells from older animals was not seen in γδT-cell receptor+ cells from 1-wk-old calves. At wk 8 of age, mitogen-induced proliferation and expression of activation antigens by T-cells from standard-fed calves were similar to responses of T-cells from steers indicating rapid maturation of T-cell function during the neonatal period. Feeding calves an intensified milk replacer was associated with decreased proliferation of mitogen-stimulated CD4+, CD8+, and γδT-cell receptor+ cells; decreased CD25 expression by mitogen-stimulated CD4+ and CD8+ cells; and decreased CD44 expression by mitogen-stimulated CD8+ cells. These results indicate that the functional capacity of the calf's T-cell population becomes more adult-like during the first weeks of life and suggest that nutrition modulates T-cell function during this period of immune maturation.

Wisteria floribunda gall extract inhibits cell migration in mouse B16F1 melanoma cells by regulating CD44 expression and GTP-RhoA activity

Extracts from galls grown on Wisteria floribunda are used as an anti-tumoral preparation in oriental traditional medicine. Here, we investigated the molecular mechanism of this anti-tumoral effect by first examining whether the extract inhibited cell migration in a B16 cell-based wound healing assay. The gall extract delayed wound healing in a dose- and time-dependent manner, indicating that one or more components of the fraction inhibited cell migration. Examination of two molecules known to be involved in metastasis, CD44, and RhoA-GTP, revealed that the gall extract decreased CD44 expression in a concentration-dependent manner, and also increased RhoA-GTP activity in comparison to untreated controls. Taken together, these results suggest that the Wisteria gall extract may inhibit cancer cell migration via inhibition of CD44 mRNA expression and activation of the GTP-RhoA protein.

Autocrine/Paracrine Prostaglandin E2 Production by Non-small Cell Lung Cancer Cells Regulates Matrix Metalloproteinase-2 and CD44 in Cyclooxygenase-2-dependent Invasion

Tumor cyclooxygenase-2 (COX-2) expression is known to be associated with enhanced tumor invasiveness. In the present study, we evaluated the importance of the COX-2 product prostaglandin E2 (PGE2) and its signaling through the EP4 receptor in mediating non-small cell lung cancer (NSCLC) invasiveness. Genetic inhibition of tumor COX-2 led to diminished matrix metalloproteinase (MMP)-2, CD44, and EP4 receptor expression and invasion. Treatment of NSCLC cells with exogenous 16,16-dimethylprostaglandin E2 significantly increased EP4 receptor, CD44, and MMP-2 expression and matrigel invasion. In contrast, anti-PGE2 decreased EP4 receptor, CD44, and MMP-2 expression in NSCLC cells. EP4 receptor signaling was found to be central to this process, because antisense oligonucleotide-mediated inhibition of tumor cell EP4 receptors significantly decreased CD44 expression. In addition, agents that increased intracellular cAMP, as is typical of EP4 receptor signaling, markedly increased CD44 expression. Moreover, MMP-2-AS treatment decreased PGE2-mediated CD44 expression, and CD44-AS treatment decreased MMP-2 expression. Thus, PGE2-mediated effects through EP4 required the parallel induction of both CD44 and MMP-2 expression because genetic inhibition of either MMP-2 or CD44 expression effectively blocked PGE2-mediated invasion in NSCLC. These findings indicate that PGE2 regulates COX-2-dependent, CD44- and MMP-2-mediated invasion in NSCLC in an autocrine/paracrine manner via EP receptor signaling. Thus, blocking PGE2 production or activity by genetic or pharmacological interventions may prove to be beneficial in chemoprevention or treatment of NSCLC.

Expression of cadherins and CD44 isoforms in human endometrium and peritoneal endometriosis.

To evaluate (a) cadherins and CD44 expression in normal endometrium and in peritoneal endometriosis, and (b) to correlate, their expressions with clinicopathological parameters. E- and N-cadherin, CD44 isoforms, CD44v3 and CD44v6 expressions were evaluated: (a) by immunoblotting in endometrium (n = 6) and in peritoneal endometriotic samples (n = 7) and, (b) by immunohistochemistry in endometrium (n = 15) and in peritoneal endometriotic samples (n = 23). By immunoblotting, endometrium expressed E- and N-cadherin, CD44 isoforms, CD44v3 and CD44v6. Similar results were observed in peritoneal endometriosis. By immunohistochemistry, in endometrium, E-cadherin was restricted to epithelial cells. Its expression remained constant throughout the menstrual cycle. N-cadherin was detected in both epithelial and stromal cells in the proliferative phase but was restricted to epithelial cells in the secretory phase. CD44 immunostaining was detected in the secretory but not in the proliferative phase. Decreased expression of E-cadherin (p < 0.01) and CD44 (p < 0.01) in epithelial cells was found in peritoneal endometriosis as compared with normal endometrium. In endometriotic stromal cells, decreased CD44 expression was found. In peritoneal endometriosis, we observed a decreased expression of E-cadherin in advanced stages of the disease (p < 0.01). Our results suggest that E-cadherin and CD44 proteins could be involved in the development of endometriotic lesions. Investigation of the mechanisms of altered adhesion molecule expression may contribute to the understanding of the behavior of endometriotic cells.

Expression of cadherins and CD44 isoforms in human endometrium and peritoneal endometriosis

Background. To evaluate (a) cadherins and CD44 expression in normal endometrium and in peritoneal endometriosis, and (b) to correlate, their expressions with clinicopathological parameters. Methods. E- and N-cadherin, CD44 isoforms, CD44v3 and CD44v6 expressions were evaluated: (a) by immunoblotting in endometrium (n = 6) and in peritoneal endometriotic samples (n = 7) and, (b) by immunohistochemistry in endometrium (n = 15) and in peritoneal endometriotic samples (n = 23). Results. By immunoblotting, endometrium expressed E- and N-cadherin, CD44 isoforms, CD44v3 and CD44v6. Similar results were observed in peritoneal endometriosis. By immunohistochemistry, in endometrium, E-cadherin was restricted to epithelial cells. Its expression remained constant throughout the menstrual cycle. N-cadherin was detected in both epithelial and stromal cells in the proliferative phase but was restricted to epithelial cells in the secretory phase. CD44 immunostaining was detected in the secretory but not in the proliferative phase. Decreased expression of E-cadherin (p < 0.01) and CD44 (p < 0.01) in epithelial cells was found in peritoneal endometriosis as compared with normal endometrium. In endometriotic stromal cells, decreased CD44 expression was found. In peritoneal endometriosis, we observed a decreased expression of E-cadherin in advanced stages of the disease (p < 0.01). Conclusion. Our results suggest that E-cadherin and CD44 proteins could be involved in the development of endometriotic lesions. Investigation of the mechanisms of altered adhesion molecule expression may contribute to the understanding of the behavior of endometriotic cells.

Correlation between nicotine-induced inhibition of hematopoiesis and decreased CD44 expression on bone marrow stromal cells

This study demonstrates that in vivo exposure to cigarette smoke (CS) and in vitro treatment of long-term bone marrow cultures (LTBMCs) with nicotine, a major constituent of CS, result in inhibition of hematopoiesis. Nicotine treatment significantly delayed the onset of hematopoietic foci and reduced their size. Furthermore, the number of long-term culture-initiating cells (LTC-ICs) within an adherent layer of LTBMCs was significantly reduced in cultures treated with nicotine. Although the production of nonadherent mature cells and their progenitors in nicotine-treated LTBMCs was inhibited, this treatment failed to influence the proliferation of committed hematopoietic progenitors when added into methylcellulose cultures. Bone marrow stromal cells are an integral component of the hematopoietic microenvironment and play a critical role in the regulation of hematopoietic stem cell proliferation and self-renewal. Exposure to nicotine decreased CD44 surface expression on primary bone marrow-derived fibroblastlike stromal cells and MS-5 stromal cell line, but not on hematopoietic cells. In addition, mainstream CS altered the trafficking of hematopoietic stem/progenitor cells (HSPC) in vivo. Exposure of mice to CS resulted in the inhibition of HSPC homing into bone marrow. Nicotine and cotinine treatment resulted in reduction of CD44 surface expression on lung microvascular endothelial cell line (LEISVO) and bone marrow-derived (STR-12) endothelial cell line. Nicotine treatment increased E-selectin expression on LEISVO cells, but not on STR-12 cells. These findings demonstrate that nicotine can modulate hematopoiesis by affecting the functions of the hematopoiesis-supportive stromal microenvironment, resulting in the inhibition of bone marrow seeding by LTC-ICs and interfering with stem cell homing by targeting microvascular endothelial cells.

CD31 Expression on Leukocytes Is Downregulated in vivo during Hemodialysis

Background/Aim: CD31 on leukocytes is the adhesion molecule involved in the leukocyte extravasation in inflammatory conditions. During hemodialysis with cellulosic membranes, it is considered that activated leukocytes adhere to endothelium, but do not show extravasation. However, it is not elucidated why activated leukocytes do not show endothelial transmigration during hemodialysis with cellulosic membranes. Methods: In the present study, changes in the expressions of Mac-1 and CD31 on granulocytes and monocytes were analyzed by flow cytometry during hemodialysis in 7 patients treated with regenerated-cellulose (RC) membranes and next with polysulfone (PS) membranes. Results: During dialysis with RC, Mac-1 expressions on granulocytes and monocytes both significantly increased as compared with predialysis values and across the dialyzer. During dialysis with RC, the CD31 expression on granulocytes and monocytes significantly decreased as compared with predialysis values. During dialysis with PS, changes in Mac-1 and CD31 expressions on granulocytes and monocytes were smaller than those during dialysis with RC. Conclusions: Decreased CD31 expression on leukocytes may affect leukocyte function more in patients chronically hemodialyzed with RC than in those hemodialyzed with PS, since CD31 is important in leukocyte transendothelial migration in inflammatory conditions.

Immunohistochemical Expression of Integrins and CD44 in Ameloblastomas.

To clarify the role of cell adhesion molecules in epithelial odontogenic tumors, expression of integrin α2, α3 and β4 subunits and the standard form of CD44 was immunohistochemically examined in 22 ameloblastomas as well as in 1 clear cell odontogenic tumor (CCOT). Integrin α2 and α3 subunits were expressed on the cell membrane of peripheral columnar or cuboidal cells in ameloblastomas. Most tumor cells in basal cell ameloblastomas and the CCOT were positive for these subunits. These features suggest that CCOT possesses characteristics similar to those of peripheral cells in main type ameloblastoma or neoplastic cells in basal cell ameloblastoma. Integrin β4 subunit was found along the basement membrane in ameloblastomas and the CCOT. The expression of these representative epithelial integrins at the epithelialmesenchymal interfaces suggests that they might mediate parenchymal-stromal cell interactions in epithelial odontogenic tumors. CD44 was detected in most tumor cells of ameloblastomas and the CCOT, but keratinizing areas in acanthomatous ameloblastomas showed decreased CD44 expression. These features suggest that CD44 is involved not only in cell adhesion but also in cellular differentiation in epithelial odontogenic tumors.

Expression of cell surface glycoprotein CD44 and integrins in breast cancers among Indian women.

Parsis, the sole surviving group of followers of Zoroaster who are settled in Bombay, have a fourfold higher incidence of breast cancer than the general population of Greater Bombay. CD44 expression was studied by immunohistochemistry in breast cancers of 50 non-Parsi and 35 Parsi women, 10 normal breast tissues, 10 proliferative lesions and 49 tissues adjoining a tumor mass. Alpha2 and beta1 integrins could be studied in only 42 malignant cases and five normal tissues. The immunohistochemistry results were correlated with other parameters including tumor grade and size, estrogen and progesterone receptor status, lymph node involvement and mitotic index. CD44 was not expressed in normal areas. Benign areas and tissues adjacent to tumor masses showed increased staining. Both Parsi and non-Parsi women showed significantly high CD44 expression. All Parsi ILCs were strongly positive for CD44. In both groups ER negativity was associated with strong CD44 positivity, while mitotic counts correlated with decreased CD44 expression in Parsis but not in non-Parsis. Alpha2 and beta1 integrins were strongly expressed on the basolateral surface of normal epithelium. However, they were downregulated in tumors. In general breast tumor tissues from Parsi and non-Parsi patients did not differ significantly with respect to most parameters. However, among Parsis lymph node involvement and CD44 correlated weakly whereas the mitotic index was inversely correlated with CD44. The reverse was true for non-Parsis. The deviation from the general pattern needs further study based on a large number of samples and appropriate use of splice variants.

Function and Expression of CD44 during Spreading, Migration, and Invasion of Murine Carcinoma Cells

The cell surface glycoprotein CD44 is proposed as a main participant in cell adhesion and migration. We studied the function, expression, and distribution of CD44 in the invasive and metastatic F3II murine carcinoma cell line during adhesion, spreading, migration, and invasion. A mAb anti-CD44 (KM 201) dramatically blocked F3II cell adhesion on both plastic and hyaluronic acid coatings, as well as spreading on uncoated plastic surfaces (P< 0.01). KM201 mAb significantly inhibited F3II cell migration and invasion in Transwell chambers. Immunocytochemistry of spreading cells revealed that CD44 distributed in bands on the cell surface, particularly in the tip of leading edges and in the perinuclear zones of the cell membrane. CD44 antigen was never detected in filopodia or lamellipodia nor in focal adhesion-like structures, but was also detectable as strong interlamellar bands. Fully spread cells showed a decreased CD44 signal compared to cells in early stages of spreading. This decrease correlated with a reduced expression of CD44 as detected by Western blot. We also investigated the signals that may regulate CD44 expression in F3II cells. Treatment of F3II cells, with phorbol myristate acetate (PMA) or phosphatidic acid (PA, the product of PLD-dependent hydrolysis of phosphatidylcholine), significantly enhanced CD44 expression. Conversely, the treatment of F3II cells with H7, a specific PKC inhibitor, or propranolol, which blocks PA conversion to DAG, significantly decreased CD44 expression levels. These results suggest the involvement of PKC and PLD pathways in CD44 expression. These results demonstrate that CD44 plays an important role during F3II cells adhesion, spreading, migration, and invasion. In addition we provide information linking the PLD- and PKC-dependent pathways with the regulation of CD44 expression.

Bovine chimeric offspring produced by transgenic embryonic stem cells generated from somatic cell nuclear transfer embryos

Thyroid cancer stem cells (CSCs) with ALDH and CD44 markers contribute to tumor growth and aggressiveness. We hypothesized that novel HSP90 inhibitors (KU711, WGA-TA) and 17-AAG can effectively target the function of thyroid CSCs in vitro and prevent migration and invasion.Validated papillary (TPC1), follicular (FTC238,WRO), and anaplastic (ACT1) human thyroid cancer cell lines were treated with 3 HSP90 inhibitors. CSCs were quantified for aldehyde dehydrogenase by flow cytometry, CD44 expression by Western blot, and thyrosphere formation assay. Cellular pathway proteins were analyzed by Western blot and migration/invasion by Boyden-chambers.WGA-TA and 17-AAG induced HSP70 compensation (not observed with KU711) on Western blot in all cell lines (>1,000 fold vs controls). Only WGA-TA degraded HSP90-Cdc37 complexing by 60–70% versus controls. Expression of HSP90 clients β-catenin, BRAF, Akt, and phospho-Akt were significantly inhibited by WGA-TA treatment (50–80%, 50–90%, >80%, and >90%) compared with controls, KU711, and 17-AAG treatment. KU711 and WGA-TA decreased CD44 expression in all cell lines (25–60% vs controls/17-AAG), decreased ALDEFLOR activity by 69–98% (P < .005), and decreased sphere formation by 64–99% (P < .05 each). Finally, cell migration was decreased by 31–98%, 100%, and 30–38%, and invasion by 75–100%, 100%, and 47% by KU711,WGA-TA, and 17-AAG treatment (P < .05) each, respectively.KU711 and WGA-TA are novel HSP90 inhibitors targeting CSC function and inhibiting cell migration/invasion in differentiated and anaplastic thyroid cancers, warranting further translational evaluation in vivo.