[Show abstract][Hide abstract] ABSTRACT: Resveratrol exerts inhibitory effects on ovarian cancer cells, while its underlying mechanism and critical molecular target(s) have been lesser known. Activations of Wnt, Notch and STAT3 signaling are frequent in ovarian cancers/OCs and supposed to be important for OC formation and progression, while the impacts of resveratrol on these signaling pathways in OC cells remain obscure.
In this study, two human ovarian cancer cell lines, OVCAR-3 and CAOV-3, were treated by 120 μM resveratrol and their responses to the treatment and the statuses of Wnt, Notch and STAT3 signaling in them were analyzed by multiple experimental approaches. Selective inhibitors of Wnt, Notch or STAT3 signaling were employed to treat OVCAR-3 and CAOV-3 cells to elucidate the significance of individual signaling pathways for ovarian cancers.
The results demonstrated distinct inhibitory effects of resveratrol on human ovarian cancer cells in terms of remarkable G1 phase accumulation, increased apoptosis fraction and concurrent suppression of Wnt, Notch and STAT3 signaling as well as their downstream cancer-related gene expression. Treatments with Wnt, Notch or STAT3 selective inhibitor revealed that only AG490, a JAK-specific inhibitor, inhibits OVCAR-3 and CAOV-3 cells in the extent as similar as that of resveratrol.
Our results suggest the significance of STAT3 activation in the maintenance and survival of ovarian cancer cells. The activated STAT3 signaling is the critical molecular target of resveratrol. Resveratrol would be a promising candidate in the management of ovarian cancers, especially the ones with resistance to conventional therapeutic agents.
Journal of Ovarian Research 04/2015; 8(1):25. DOI:10.1186/s13048-015-0152-4 · 2.43 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glioblastomas respond differently to all-trans retinoic acid (RA) for unknown reasons. Because CRABP-II and FABP5 mediate RA intracellular signaling respectively and lead to distinct biological consequences, their expression patterns in different grades of astrocytomas and the glioblastoma cells lines LN18, LN428 and U251 were examined to identify potential correlations with RA sensitivities. The response of glioblastoma cells to RA, decitabine or the FABP5 competitive inhibitor, BMS309403, was analyzed. CRABP-II and FABP5 were expressed to varying degrees by the 84-astrocytoma cases examined. Treatment of LN428, U251 and LN18 cells with RA failed to suppress their growth; however, U251 proliferation was inhibited by decitabine. The combination of decitabine and RA suppressed the growth of all three cell lines and induced significant apoptosis of LN428 and U251 cells. Both CRABP-II and FABP5 were transcribed in the three cell lines but FABP5 proteins were undetectable in U251 cells. The ratio of CRABP-II to FABP5 was not altered after RA, decitabine or RA and decitabine treatment and the resistance of cells to RA was not reversed by BMS309403 treatment. In conclusion, CRABP-II and FABP5 expression patterns are neither related to the tumor grades nor correlated with RA sensitivity. Additional molecular factors may be present that determines the sensitivity of glioblastoma cells to RA. Dicitabine may improve the sensitivity of glioblastoma cells to RA, however, its underlying mechanism and its in vivo feasibility need to be investigated.
[Show abstract][Hide abstract] ABSTRACT: Resveratrol possesses anti-tumor activities against central nervous system (CNS) tumors in vitro but has not yet been used clinically due to its low bioavailability, particularly in the CNS. This study thus aimed to elucidate brain bioavailability of trans-resveratrol by monitoring brain concentrations and dwell times following administration of resveratrol through intragastric, intraperitoneal, external carotid artery/ECA and intrathecal routes. In parallel, we evaluated the biological responses of rat RG2 glioblastoma cells as well as RG2-formed rat intracranial glioblastomas treated with resveratrol via intrathecal administration. The results revealed that resveratrol was detected in rat brains except when administered systemically. Intrathecal administration of reseveratrol led to abundant apoptotic foci and increased staining of the autophagy proteins, LC-3 and Beclin-1 and shrinkage of the intracranial tumors. In conclusion, the BBB penetrability of resveratrol is remarkably increased by intracthecal administration. Regular short-term resveratrol treatments suppress growth and enhance autophagic and apoptotic activities of rat RG2 glioblastoma cells in vitro and in vivo. Therefore, intrathecal administration of resveratrol could be an optimal intervention approach in the adjuvant management of brain malignancies.
Journal of the American Society for Experimental NeuroTherapeutics 01/2015; 12(2). DOI:10.1007/s13311-014-0334-6 · 5.05 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Maggot extracts promote wound healing but their bioactive part(s) and molecular effects on the regenerating tissues/cells remain largely unclear. These issues are addressed here by treating rat skin wounds, human keratinocyte line/HaCat and fibroblasts with maggot secretion/excretion and the extracts of maggots without and with secretion/excretion. The wound closure rates, cell proliferation activities and statuses of wound healing-related signaling pathways (STAT3, Notch1, Wnt2, NF-κB and TGF-beta/Smad3) and their downstream gene expression (c-Myc, cyclin D1 and VEGF) are evaluated by multiple approaches. The results reveal that the maggot extracts, especially the one from the maggots without secretion/excretion shows the best wound healing promoting effects in terms of quicker wound closure rates and more rapid growth of keratinocytes and fibroblasts. Of the 5 signaling pathways checked, the ones mediated by TGF-beta/Smad3 and STAT3 are activated in the untreated wounds and become further enhanced by the maggot extracts, accompanied with c-Myc, VEGF and cyclin D1 up-regulation. Our results thus demonstrate 1) that both body extract and secretion/excretion of maggots contain favorable wound healing elements and 2) that the enhancement of TGF-beta/Smad3 and STAT3 signaling activities may be the main molecular effects of maggot extracts on the wound tissues.
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[Show abstract][Hide abstract] ABSTRACT: S100A4 promotes cancer metastasis but is frequently silenced in human cutaneous squamous cell carcinomas/c-SCCs due to DNA methylation, which may explain the less metastasized property of c-SCCs.
Cancer biomarkers: section A of Disease markers 08/2014; 14(5):325-33. DOI:10.3233/CBM-140414 · 1.72 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cervical cancers/CCs are one of the commonest malignancies and the second leading cause of cancer-related death in women. Resveratrol inhibits CC cell growth but its molecular target(s) remains unclear. Since the signaling pathways mediated by STAT3, Notch1 and Wnt2 play beneficial roles in CC formation and progression, the effects of resveratrol on them in cervical adenocarcinoma (HeLa) and squamous cell carcinoma (SiHa) cells were analyzed. The biological significances of the above signaling for HeLa and SiHa cells were evaluated by treating the cells with STAT3, Wnt or Notch selective inhibitors. The frequencies of STAT3, Notch and Wnt activations in 68 cases of CC specimens and 38 non-cancerous cervical epithelia were examined by tissue microarray-based immunohistochemical staining. The results revealed that HeLa and SiHa cells treated by 100μM resveratrol showed extensive apoptosis, accompanied with suppression of STAT3, Notch and Wnt activations. Growth inhibition and apoptosis were found in HeLa and SiHa populations treated by AG490, a STAT3/JAK3 inhibitor but not the ones treated by Notch inhibitor L-685,458 or by Wnt inhibitor XAV-939. Immunohistochemical staining performed on the tissue microarrays showed that the frequencies of Notch1, Notch2, Hes1, Wnt2, Wnt5a and p-STAT3 detection as well as β-catenin nuclear translocation in CC samples were significantly higher than that of noncancerous group (p<0.01), while the expression rate of PIAS3 was remarkably low in cancer samples (p<0.01). Our results thus demonstrate that STAT3, Wnt and Notch signaling are frequently co-activated in human CC cells and specimens and resveratrol can concurrently inhibit those signaling activations and meanwhile lead cervical squamous cell carcinoma and adenocarcinoma cells to growth arrest and apoptosis. STAT3 signaling is more critical for CC cells and is the major target of resveratrol because selective inhibition of STAT3 rather than Wnt or Notch activation commits SiHa and HeLa cells to apoptosis.
[Show abstract][Hide abstract] ABSTRACT: Conventional adjuvant chemotherapies for bladder transitional cell carcinomas (TCCs) may cause strong systemic toxicity and local irritation. Non-toxic resveratrol inhibits TCC cell growth but its feasibility in clinical management of TCCs remains obscure. This study aimed to evaluate the safety and anti-TCC efficacy of resveratrol, using the experimental models closer to the clinical treatment condition. Human TCC EJ cells were exposed to 100 µM, 150 µM and 200 µM resveratrol respectively for 1 hour and 2 hours to mimic intravesical drug instillation and the cell responses were analyzed by multiple experimental approaches. An orthotopic TCC nude mouse model was established by injecting EJ cells into the sub-urothelial layer and used for short-term intravesical resveratrol instillation. The safety of resveratrol instillation was evaluated and compared with that of MCC. The results revealed that 2 h 150 µM or 200 µM resveratrol treatment leaded to remarkable S phase arrest and apoptosis at 72 h time-point, accompanied with attenuated phosphorylation, nuclear translocation and transcription of STAT3, down-regulation of STAT3 downstream genes (survivin, cyclinD1, c-Myc and VEGF) and nuclear translocations of Sirt1 and p53. The importance of STAT3 signaling in cell growth was confirmed by treating EJ cells with JAK2 inhibitor tyrphostin AG490. The efficacy and safety of resveratrol instillation were proved by the findings from nude mouse orthotopic xenograft models, because this treatment caused growth suppression, distinctive apoptosis and STAT3 inactivation of the transplanted tumors without affecting normal urothelium. Our results thus suggest for the first time the practical values of resveratrol as a safe and effective agent in the post-operative treatment of TCCs.
PLoS ONE 02/2014; 9(2):e89806. DOI:10.1371/journal.pone.0089806 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Wound measurement is an objective and direct way to trace the course of wound healing and to evaluate therapeutic efficacy. Nevertheless, the accuracy and efficiency of the current measurement methods need to be improved. Taking the advantages of reliability of transparency tracing and the accuracy of computer-aided digital imaging, a transparency-based digital imaging approach is established, by which data from 340 wound tracing were collected from 6 experimental groups (8 rats/group) at 8 experimental time points (Day 1, 3, 5, 7, 10, 12, 14 and 16) and orderly archived onto a transparency model sheet. This sheet was scanned and its image was saved in JPG form. Since a set of standard area units from 1 mm(2) to 1 cm(2) was integrated into the sheet, the tracing areas in JPG image were measured directly, using the "Magnetic lasso tool" in Adobe Photoshop program. The pixel values/PVs of individual outlined regions were obtained and recorded in an average speed of 27 second/region. All PV data were saved in an excel form and their corresponding areas were calculated simultaneously by the formula of Y (PV of the outlined region)/X (PV of standard area unit) × Z (area of standard unit). It took a researcher less than 3 hours to finish area calculation of 340 regions. In contrast, over 3 hours were expended by three skillful researchers to accomplish the above work with traditional transparency-based method. Moreover, unlike the results obtained traditionally, little variation was found among the data calculated by different persons and the standard area units in different sizes and shapes. Given its accurate, reproductive and efficient properties, this transparency-based digital imaging approach would be of significant values in basic wound healing research and clinical practice.
PLoS ONE 05/2012; 7(5):e38069. DOI:10.1371/journal.pone.0038069 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glioblastoma multiforme (GBM) cells show different responses to resveratrol, for unknown reasons. Our data from human medulloblastoma cells and primary cultures of rat brain cells revealed an inverse correlation of sulfonation activity with resveratrol sensitivities, providing a clue to the underlying mechanisms of the variable sensitivities of GBM cells to resveratrol. In this study, we found that U251 cells were sensitive and LN229 cells were insensitive to resveratrol. Thus, these two cell lines were taken as comparable models for elucidating the influence of sulfonation activities on resveratrol sensitivity. HPLC showed identical resveratrol metabolic patterns in both cell lines. LC/MS and high-resolution mass MS analyses further demonstrated that resveratrol monosulfate generated by sulfotransferases (SULTs) was the major metabolite of human GBM cells. The levels of brain-associated SULT (SULT1A1, SULT1C2, and SULT4A1) expression in U251 cells were lower than those in LN229 cells, suggesting the inverse relationship of SULT-mediated sulfonation activity with high intracellular resveratrol bioavailability and resveratrol sensitivity of human GBM cells. Furthermore, immunohistochemical staining revealed reductions in expression of the three brain-associated SULTs in 72.8%, 47.5% and 66.3% of astrocytomas, respectively. Therefore, the levels of brain-associated SULTs and sulfonation activity mediated by them could be important parameters for evaluating the potential response of human GBM cells to resveratrol, and may have value in the personalized treatment of GBMs with resveratrol.
Database Nucleotide sequence data for SULT1A1, SULT1C2 and SULT4A1 are available in the GenBank database under the accession numbers BC110887, BC005353, and BC028171.
[Show abstract][Hide abstract] ABSTRACT: Medulloblastoma is a primitive neuroectodermal tumor, which originates in the cerebellum, presumably due to the alterations of some neurogenetic elements. Sirtuin 1 (SIRT1), a class III histone deacetylase (HDAC), regulates differentiation of neuronal stem cells but its status in medulloblastomas remains largely unknown. The current study aimed to address this issue by checking SIRT1 expression in noncancerous cerebellar tissues, medulloblastoma tissues and established cell lines. The roles of SIRT1 in proliferation and survival of UW228-3 medulloblastoma cells were analyzed by SIRT1 small interfering RNA (siRNA) transfection and SIRT1 inhibitor nicotinamide treatment. The results revealed that the frequency of SIRT1 expression in medulloblastoma tissues was 64.17% (77/120), while only one out of seven tumor-surrounding noncancerous cerebellar tissues showed restricted SIRT1 expression in the cells within the granule layer. Of the three morphological subtypes, the rates of SIRT1 detection in the large cell/anaplastic cell (79.07%; 34/43) and the classic medulloblastomas (60.29%; 41/68) are higher than that (22.22%; 2/9) in nodular/desmoplastic medulloblastomas (P < 0.01 and P < 0.05, respectively). Heterogeneous SIRT1 expression was commonly observed in classic medulloblastoma. Inhibition of SIRT1 expression by siRNA arrested 64.96% of UW228-3 medulloblastoma cells in the gap 1 (G1) phase and induced 14.53% of cells to apoptosis at the 48-h time point. Similarly, inhibition of SIRT1 enzymatic activity with nicotinamide brought about G1 arrest and apoptosis in a dose-related fashion. Our data thus indicate: (i) that SIRT1 may act as a G1-phase promoter and a survival factor in medulloblastoma cells; and (ii) that SIRT1 expression is correlated with the formation and prognosis of human medulloblastomas. In this context, SIRT1 would be a potential therapeutic target of medulloblastomas.
[Show abstract][Hide abstract] ABSTRACT: To identify differentially expressed lumcian in scleral tissue of eyes with primary open-angle glaucoma (POAG) and high mopia (HM).
Total RNA was isolated from scleral tissue of cadaveric eyes derived from normal donors, patient's eyes with diagnosed glaucoma and high mopia who accepted trabeculectomy. RNA was amplified, RT-PCR was used to measure the levels of mRNA. The ratio of the electrophoresis strips'gray scale values of the β-actin over the lumican gene was obtained for ANOVA analysis.
β-actin/LUM of normal eye was significantly higher than that of POAG and POAG + HM eyes (P < 0.01), but there was no significant difference between POAG and POAG + HM eyes (P > 0.05).
Differentially expressed lumican between POAG and control groups identified in this study have not been previously investigated for their role in the pathogenesis of POAG and thus are novel factors for further study of the mechanism of the disease and for their possible use as diagnostic markers.
Zhonghua yi xue za zhi 03/2012; 92(9):608-11. DOI:10.3760/cma.j.issn.0376-2491.2012.09.010
[Show abstract][Hide abstract] ABSTRACT: Medulloblastoma cells exhibit varied responses to therapy by all-trans retinoic acid (RA). The underlying mechanism for such diverse effects however remains largely unclear. In this study, we attempted to elucidate the molecular basis of RA resistance through the study of RA signaling components in both RA-sensitive (Med-3) and RA-resistant (UW228-2 and UW228-3) medulloblastoma cells. The results revealed that RARα/β/γ and RXRα/β/γ were found in the three cell lines. Expression of CRABP-I and CRABP-II was seen in Med-3 cells, up-regulated when treated with RA, but was absent in UW228-2 and UW228-3 cells regardless of RA treatment. Bisulfite sequencing revealed 8 methylated CG sites at the promoter region of CRABP-II in UW228-2 and UW228-3 but not in Med-3 cells. Demethylation by 5-aza-2'-deoxycytidine recovered CRABP-II expression. Upon restoration of CRABP-II expression, both UW228-2 and UW228-3 cells responded to RA treatment by forming neuronal-like differentiation, synaptophysin expression, β-III tubulin upregulation, and apoptosis. Furthermore, CRABP-II specific siRNA reduced RA sensitivity in Med-3 cells. Tissue microarray-based immunohistochemical staining showed variable CRABP-II expression patterns among 104 medulloblastoma cases, ranging from negative (42.3%), partly positive (14.4%) to positive (43.3%). CRABP-II expression was positively correlated with synaptophysin (rs = 0.317; p = 0.001) but not with CRABP-I expression (p > 0.05). In conclusion, aberrant methylation in CRABP-II reduces the expression of CRABP-II that in turn confers RA resistance in medulloblastoma cells. Determination of CRABP-II expression or methylation status may enable a personalized RA therapy in patients with medulloblastomas and other types of cancers.
[Show abstract][Hide abstract] ABSTRACT: Trans-resveratrol rather than its biotransformed monosulfate metabolite exerts anti-medulloblastoma effects by suppressing STAT3 activation. Nevertheless, its effects on human glioblastoma cells are variable due to certain unknown reason(s).
Citing resveratrol-sensitive UW228-3 medulloblastoma cell line and primarily cultured rat brain cells/PBCs as controls, the effect of resveratrol on LN-18 human glioblastoma cells and its relevance with metabolic pattern(s), brain-associated sulfotransferase/SULT expression and the statuses of STAT3 signaling and protein inhibitor of activated STAT3 (PIAS3) were elucidated by multiple experimental approaches. Meanwhile, the expression patterns of three SULTs (SULT1A1, 1C2 and 4A1) in human glioblastoma tumors were profiled immunohistochemically. The results revealed that 100 µM resveratrol-treated LN-18 generated the same metabolites as UW228-3 cells, while additional metabolite in molecular weight of 403.0992 in negative ion mode was found in PBCs. Neither growth arrest nor apoptosis was found in resveratrol-treated LN-18 and PBC cells. Upon resveratrol treatment, the levels of SULT1A1, 1C2 and 4A1 expression in LN-18 cells were more up-regulated than that expressed in UW228-3 cells and close to the levels in PBCs. Immunohistochemical staining showed that 42.0%, 27.1% and 19.6% of 149 glioblastoma cases produced similar SULT1A1, 1C2 and 4A1 levels as that of tumor-surrounding tissues. Unlike the situation in UW228-3 cells, STAT3 signaling remained activated and its protein inhibitor PIAS3 was restricted in the cytosol of resveratrol-treated LN-18 cells. No nuclear translocation of STAT3 and PIAS3 was observed in resveratrol-treated PBCs. Treatment with STAT3 chemical inhibitor, AG490, committed majority of LN-18 and UW228-3 cells but not PBCs to apoptosis within 48 hours.
LN-18 glioblastoma cells are insensitive to resveratrol due to the more inducible brain-associated SULT expression, insufficiency of resveratrol to suppress activated STAT3 signaling and the lack of PIAS3 nuclear translocation. The findings from PBCs suggest that an effective anticancer dose of resveratrol exerts little side effect on normal brain cells.
PLoS ONE 11/2011; 6(11):e27484. DOI:10.1371/journal.pone.0027484 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The effect of all-trans retinoic acid (ATRA) on cutaneous squamous cell carcinomas (c-SCC) has been poorly described. Because the imbalance of CRABP-II-mediated anticancer signalling and FABP5-mediated growth-promoting signalling was supposed to be related with ATRA sensitivities of cancer cells, COLO16 human c-SCC cell line was selected to check underlying mechanism leading to ATRA resistance by multiple experimental approaches. The results revealed that COLO 16 cells were resistant to 15 μm ATRA treatment. FABP5 as well as the elements related with CRABP-II signalling (CYP26A1, CYP26B1, CRABP-I, RARα/β/γ and RXRα/β/γ) and with FABP5 signalling (PPARβ/δ) were expressed, but CRABP-II was undetectable in COLO 16 cells. 5-Aza treatment enhanced CRABP-II expression but further bisulfite sequencing PCR-DNA sequencing revealed no methylation in CRABP-II promoter region. Transfection of CRABP-II-expressing plasmids or FABP5 siRNA or both successfully manipulated the level(s) of target gene expression but failed to overcome ATRA resistance in the transfectants. In conclusion, CRABP-II and FABP5 expression were imbalanced in ATRA-resistant COLO 16 cells. 5-Aza-enhanced CRABP-II expression and unmethylation in CRABP-II promoter region suggest the methylation of certain CRABP-II regulatory gene(s) in COLO 16 cells. As neither restoration of CRABP-II expression nor the increased CRABP-II versus FABP5 ratio can overcome ATRA resistance of COLO 16 cells, additional ATRA-resistant mechanism(s) may present in human c-SCCs and COLO 16 cells would be of value in addressing this issue.
[Show abstract][Hide abstract] ABSTRACT: Resveratrol promotes differentiation and apoptosis of medulloblastoma cells by suppressing STAT3 signaling and a range of cancer-associated gene expression. However, Bcl-2, a common target of STAT3 and NF-κB signaling, is distinctly up-regulated in resveratrol-treated medulloblastoma cells, indicating potential effects of NF-κB in Bcl-2 expression and anti-medulloblastoma efficiency of resveratrol. To clarify this point, the status of NF-κB signaling and the consequence of NF-κB inhibition in UW228-2 and UW228-3 medulloblastoma cells without and with resveratrol treatment were evaluated by several experimental approaches. The results revealed that resveratrol activated NF-κB signaling in both cell lines at the 4-h treatment point, and the treated cells sequentially exhibited Bcl-2 up-regulation, neuronal-like phenotype with synaptophisin expression, and, eventually, apoptosis. Pyrrolidine dithiocarbamate (PDTC) treatment inhibited NF-κB activation and Bcl-2 expression and committed resveratrol-treated cells to apoptosis at the 8-h time point without the step of neuron-oriented differentiation. On the other hand, a single 50 μg/ml lipopolysaccharide (LPS) treatment activated NF-κB signaling accompanied with sustained proliferation and neuron-like differentiation. Tissue microarray-based immunohistochemical staining showed significantly different (P < 0.001) p65 nuclear translocation between the neurons of tumor-surrounding cerebella (10/10; 100%) and medulloblastoma tissues (20/117; 17.09%). Additionally, synaptophysin production was found in 83.64% of p65-positive and in 40.35% of p65-negative medulloblastoma cases. Our in-vitro and in-vivo results thus demonstrate the dual effects of NF-κB signaling on medulloblastoma cells by delaying resveratrol-induced apoptosis by up-regulating Bcl-2 expression or by involvement in neuronal-like differentiation in the absence of resveratrol. Therefore, appropriate inhibition of NF-κB activation may enhance the anti-medulloblastoma efficacy of resveratrol.
Journal of Neuro-Oncology 12/2010; 104(1):169-77. DOI:10.1007/s11060-010-0496-y · 3.07 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cancer preventive reagent trans-resveratrol is intracellularly biotransformed to different metabolites. However, it is still unclear whether trans-resveratrol exerts its biological effects directly or through its metabolite(s). This issue was addressed here by identifying the metabolic pattern and the bioactive form of resveratrol in a resveratrol-sensitive human medulloblastoma cell line, UW228-3. The cell lysates and condition media of UW228-3 cells with or without 100 microM resveratrol treatment were analyzed by HPLC and LC/MS which revealed (1) that resveratrol was chemically unstable and the spontaneous generation of cis-resveratrol reduced resveratrol's anti-medulloblastoma efficacy and (2) that resveratrol monosulfate was the major metabolite of the cells. To identify the bioactive form of resveratrol, a mixture-containing approximately half fraction of resveratrol monosulfate was prepared by incubating trans-resveratrol with freshly prepared rat brain lysates. Medulloblastoma cells treated by 100 microM of this mixture showed attenuated cell crisis. The overall levels of the three brain-associated sulfotransferases (SULT1A1, 1C2 and 4A1) were low in medulloblastoma cells in vivo and in vitro in comparison with that in human noncancerous and rat normal cerebella; resveratrol could more or less up-regulate the production of these enzymes in UW228-3 cells but their overall level was still lower than that in normal cerebellum tissue. Our study thus demonstrated for the first time that trans-resveratrol is the bioactive form in medulloblastoma cells in which the expression of brain-associated SULTs was down-regulated, resulting in the increased intracellular bioavailability and anti-medulloblastoma efficacy of trans-resveratrol.
[Show abstract][Hide abstract] ABSTRACT: S100A4 appears important for cancer metastasis and its overexpression is common in a variety of human malignancies, but its status in epidermal cancers remains lesser known. Likewise, E-cadherin downregulation and Wingless (Wnt) activation are frequent cancer-associated alterations, whereas their potential correlations with S100A4 expression in skin lesions have not been characterized. These issues were addressed in the present study using tissue microarray-based immunohistochemical staining, reverse transcriptase polymerase chain reaction and western blotting. Meanwhile, the underlying epigenetic mechanism leading to the altered S100A4 expression in epidermal tumors was elucidated. Immunohistochemistry revealed that S100A4 expression frequencies were 100% (8/8) in normal epidermis, 80.6% (25/31) in tumor-surrounding non-cancerous epidermis, 66.7% (10/15) in premalignant diseases, 8.3% (1/11) in Bowen's disease and 7.7-26.3% in different cancer tissues. The incidence of S100A4 detection in the normal and non-cancerous epidermis was significantly different from that of epidermal cancers (P = 0.000). Accordingly, human immortalized keratinocyte line HaCat but not skin squamous cell carcinoma (SCC) line colo16 was positive in S100A4 expression. S100A4 downregulation, E-cadherin reduction and Wnt activation coexisted in most of epidermal cancers but unnecessarily overlapped. Methylation DNA sequencing revealed methylation of four critical (cytosine and guanine separated by a phosphate or -C-phosphate-G-) CpG sites within S100A4 intron first in S100A4-negative colo16 cells and skin SCCs, and demethylator/5-aza-2'-deoxycytidine treatment efficiently recovered S100A4 expression in colo16 cells. Our findings demonstrate that S100A4 downregulation, as the consequence of DNA methylation, is closely correlated with skin tumor formation. Wnt activation and E-cadherin reduction and S100A4 down-regulation are paralleled molecular events in skin tumors, which may serve as the biomarkers for predicting epidermal cancer risk.
[Show abstract][Hide abstract] ABSTRACT: S100A4 promotes cancer metastasis, but its overall status and splicing manner during gastrocarcinogenesis remains less known. We therefore examined S100A4 frequencies, splicing pattern(s) and the underlying reason(s) for S100A4 expression in gastric cancers. Immunohistochemistry revealed frequent S100A4 expression in intestinal gastric cancers (37/45; 82%) and diffuse gastric cancers (12/20; 60%), but uncommon in noncancerous epithelia (0/12), chronic gastritis (2/24; 8%), and intestinal metaplasia (3/15; 20%). Of 65 primary tumors, 18 were found with focal S100A4 expression, while their LN metastases showed homogenous distribution. S100A4-oriented reverse transcription-polymerase chain reaction yielded a transcript containing exons 1, 3, and 4 (AS1) in 20% of noncancerous, 84% premalignant, and 92% tumor tissues and a transcript harboring exons 1 to 4 (AS2) in 65% of gastric cancers (GCs), 26% premalignant but none in noncancerous tissues. Further analyses found AS1 expression in stromal but not epithelial cells of premalignant tissues, absence of AS2 in endoscopic inflammatory mucosa, and the coexistence of AS1/AS2 in the cultured fibroblasts. Methylation DNA sequencing revealed hypermethylation of four critical CpG sites within S100A4 intron first among S100A4-negative gastric tissues and hypomethylation in S100A4-expressing GC tissues/cell lines. E-cadherin reduction and Wnt activation were common in gastric cancers, which were closely correlated but unnecessarily overlapped with S100A4 expression. Our findings suggest that S100A4 expression is closely related with GC formation, which, as a hypomethylation event, is accompanied with E-cadherin reduction and Wnt activation. The preferential S100A4 AS2 expression in GC cells would have potential values in GC surveillance and prognostic assessment.
[Show abstract][Hide abstract] ABSTRACT: In this study, the potential influence of resveratrol (3,5,4'-trihydroxy-trans-stilbene) in signal transducer and activator of transcription 3 (STAT3) signaling of medulloblastoma cells was evaluated by checking the status of STAT3 sig- naling and its downstream gene expression in two medulloblastoma cell lines (UW228-2 and UW228-3) with and without resveratrol treatment. The results revealed that resveratrol induced neuronal differentiation of medulloblas- toma cells. Signal transducer and activator of transcription 3 expression and phosphorylation were detected in normally cultured UW228-2 and UW228-3 cells that were apparently attenuated after resveratrol treatment. The expression of STAT3 downstream genes, survivin, cyclin D1, Cox-2 ,a ndc-Myc, was suppressed but Bcl-2 was enhanced by resveratrol. Meanwhile, the production and secretion of leukemia inhibitory factor, a STAT3 activator, became active in resveratrol-treated cells. To further ascertain the significance of STAT3 signaling for medulloblas- toma cells, AG490, a selective inhibitor of STAT3 phosphorylation, was used to treat UW228-3 cells. Phosphory- lation of STAT3 was inhibited by AG490 accompanied with growth suppression, differentiation-like changes, and down-regulation of survivin, cyclin D1, Cox-2, and c-Myc. Our data thus suggest the importance of STAT3 signaling in maintenance and survival of medulloblastoma cells. This signaling may be the major target of resveratrol. En- hanced leukemia inhibitory factor and Bcl-2 expressions in resveratrol-treated cells might reflect a compensatory response to the loss of STAT3 function.
Neoplasia (New York, N.Y.) 08/2008; 10(7). DOI:10.1593/neo.08304 · 4.25 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Altered Notch signaling seems linked with medulloblastoma (MB) formation and resveratrol exhibits anti-medulloblastoma effects. However, the influence of resveratrol in Notch signaling of MB cells has not been described. This issue was addressed here by checking Notch1 and Notch2 statuses in three MB cell lines with and without resveratrol treatment. Notch1 and Notch2 were detected in the cytoplasm of three cell lines under normal condition, which were up-regulated by resveratrol along with differentiation, apoptosis and enhanced Hes1 nuclear translocation. Nevertheless, blockage of Notch enzymatic cleavage with gamma-seacretase inhibitors, DAPT and L-685,458, neither interrupted resveratrol-caused cellular events nor affected MB cell growth. These results demonstrate that Notch signaling has little relevance with resveratrol-induced differentiation and apoptosis and may not be a universal critical factor of MB cells.