Postoperative resveratrol administration improves prognosis of rat orthotopic glioblastomas.

BACKGROUND: Although our previous study revealed lumbar punctured resveratrol could remarkably prolong the survival of rats bearing orthotopic glioblastomas, it also suggested the administration did not completely suppress rapid tumour growth. These evidences led us to consider that the prognosis of tumour-bearing rats may be further improved if this treatment is used in combination with neurosurgery. Therefore, we investigated the effectiveness of the combined treatment on rat orthotopic glioblastomas. METHODS: Rat RG2 glioblastoma cells were inoculated into the brains of 36 rats. The rats were subjected to partial tumour removal after they showed symptoms of intracranial hypertension. There were 28 rats that survived the surgery, and these animals were randomly and equally divided into the control group without postoperative treatment and the LP group treated with 100 µl of 300 µM resveratrol via the LP route. Resveratrol was administered 24 h after tumour resection in 3-day intervals, and the animals received 7 treatments. The intracranial tumour sizes, average life span, cell apoptosis and STAT3 signalling were evaluated by multiple experimental approaches in the tumour tissues harvested from both groups. RESULTS: The results showed that 5 of the 14 (35.7%) rats in the LP group remained alive over 60 days without any sign of recurrence. The remaining nine animals had a longer mean postoperative survival time (11.0 ± 2.9 days) than that of the (7.3 + 1.3 days; p < 0.05) control group. The resveratrol-treated tumour tissues showed less Ki67 labelling, widely distributed apoptotic regions, upregulated PIAS3 expression and reduced p-STAT3 nuclear translocation. CONCLUSIONS: This study demonstrates that postoperative resveratrol administration efficiently improves the prognosis of rat advanced orthotopic glioblastoma via inhibition of growth, induction of apoptosis and inactivation of STAT3 signalling. Therefore, this therapeutic approach could be of potential practical value in the management of glioblastomas.

Inhibition of NF-κB signaling commits resveratrol-treated medulloblastoma cells to apoptosis without neuronal differentiation.

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.

A novel threefold-interpenetrating primitive cubic network based on a dinuclear Zn2 node.

In the mixed-ligand metal-organic polymeric compound poly[[mu(2)-1,4-bis(imidazol-1-yl)benzene](mu(2)-terephthalato)dizinc(II)], [Zn(2)(C(8)H(4)O(4))(2)(C(12)H(10)N(4))](n) or [Zn(2)(bdc)(2)(bib)](n) [H(2)bdc is terephthalic acid and bib is 1,4-bis(imidazol-1-yl)benzene], the asymmetric unit contains one Zn(II) ion, with two half bdc anions and one half bib molecule lying around inversion centers. The Zn(II) ion is in a slightly distorted tetrahedral environment, coordinated by three carboxylate O atoms from three different bdc anions and by one bib N atom. The crystal structure is constructed from the secondary building unit (SBU) [Zn(2)(CO(2))(2)N(2)O(2)], in which the two metal centers are held together by two bdc linkers with bis(syn,syn-bridging bidentate) bonding modes. The SBU is connected by bdc bridges to form a two-dimensional grid-like (4,4)-layer, which is further pillared by the bib ligand. Topologically, the dinuclear SBU can be considered to be a six-connected node, and the extended structure exhibits an elongated primitive approximately cubic framework. The three-dimensional framework possesses a large cavity with dimensions of approximately 10 x 13 x 17 A in cross-section. The potential porosity is filled with mutual interpenetration of two identical equivalent frameworks, generating a novel threefold interpenetrating network with an alpha-polonium topology [Abrahams, Hoskins, Robson & Slizys (2002). CrystEngComm, 4, 478-482].

(E)-4-Bromo-N'-(2-chloro-benzyl-idene)benzohydrazide.

In the title compound, C(14)H(10)BrClN(2)O, the dihedral angle between the two benzene rings is 11.4 (2)°. In the crystal structure, mol-ecules are connected via inter-molecular N-H⋯O hydrogen bonds into one-dimensional chains running parallel to the c axis.

A combined plasma photolysis (CPP) method for removal of CS2 from gas streams at atmospheric pressure.

A combined plasma photolysis (CPP) reactor that utilizes a dielectric barrier discharge (DBD) plasma and 207 nm UV radiation from discharge-driven KrBr* excimers was designed and constructed. Gas streams containing CS2 were treated with stand-alone DBD and CPP at atmospheric pressure. In comparison to DBD, CPP greatly enhanced the removal efficiency at the same applied voltage, waste gas concentration and gas residence time. Thus the applied voltage could be reduced to a certain extent in the plasma processing of industrial wastes. Influences of the KrBr* radiation, inlet CS2 concentration and gas residence time on CS2 removal by CPP were also studied. In addition, the likely reaction mechanisms for the removal of CS2 by CPP are suggested based on the byproducts analysis. The enhanced removal efficiency and reaction mechanisms implied that the CPP process would probably be suitable for the removal of a large number of gaseous pollutants.

[Relationship between sensitivity of tumor cells to chemotherapeutic agent in vivo and in vitro: experiment with mouse lymphoma cells].

OBJECTIVE: To study the relationship of the sensitivity of tumor cells to chemotherapeutic agent between in vivo and in vitro. METHODS: Mouse lymphoma cells of the line E14 were cultured and melphalan resistant EL4 cell line (EL4/melphalan) was established by culturing EL4 cells with continuous low-concentration and intermittent gradually-increasing-concentration of melphalan in vitro. MTT assay was used to evaluate the drug sensitivity and the resistance index of the EL4/melphalan cells to melphalan was calculated. EL4/melphalan and EL4 cells of the concentration of 5 x 10(8)/L were inoculated separately into 20 C57BL/6 mice subcutaneously. 12 days later, the EL4 and EL4/melphalan tumor-bearing mice were randomly divided into 2 groups respectively, 5 mice in each group. Treatment groups were given 7.5 mg/kg melphalan intraperitoneally, and control groups were given the same volume of normal saline. The tumor size was observed every other day. RESULTS: Compared with the EL4 cells, the EL4/melphalan cells had no obvious changes morphologically. They could grow in RPMI 1640 medium containing 5 mg/ml melphalan. The resistance index was 2.87 against melphalan. After the treatment of melphalan of the dose 7.5 mg/kg, the tumor sizes of the treatment groups and control groups inoculated with both EL4 cells and the EL4/melphalan cells gradually decreased at the similar speed, and about one week later all tumors disappeared. However, the tumors of the control groups grew progressively and all the mice died at last. CONCLUSION: The chemotherapeutic effects of tumors in vivo have nothing to do with the effects of the chemotherapeutic agents on tumor cells in vitro. The tumor cells resistant to melphalan in vitro remain sensitive to the drug in vivo.

The anxiolytic- and antidepressant-like effects of ATPM-ET, a novel κ agonist and µ partial agonist, in mice.

RATIONALE: Opioid receptors are implicated in the regulation of motivation and emotion. However, animal studies show that activation of κ opioid receptor produces contrasting mood-altering effects in models of anxiety-like and depressive-like behaviors, and consequently, the role of κ receptor in mood control remains unsettled. The effect of κ/µ opioid combination in emotion regulation was unexplored. OBJECTIVES: The aim of the study was to investigate the effects of (-)-3-N-ethylaminothiazolo [5,4-b]-N-cyclopropylmethylmorphinan hydrochloride (ATPM-ET), a novel κ agonist and µ partial agonist, in regulating emotional responses. METHODS: The emotional responses of ATPM-ET were detected in the elevated plus maze (EPM), open field test (OFT), forced swim test (FST), and tail suspension test (TST). Selective κ antagonist nor-binaltorphimine (nor-BNI) and µ antagonist ß-funaltrexamine (ß-FNA) were applied to determine the type of receptor involved. The conditioned place aversion model was used to evaluate the effects on aversive emotion. RESULTS: In the EPM and OFT, ATPM-ET (1 and 2 mg/kg, s.c.) significantly increased the time spent in the open arm and in the central area, respectively. In the FST and TST, ATPM-ET (0.5 and 1 mg/kg, s.c.) significantly reduced the duration of immobility. These effects were prevented by nor-BNI (10 mg/kg, i.p., -24 h), but not by ß-FNA (10 and20 mg/kg, i.p., -24 h) pretreatment. At the dose of 2 mg/kg, ATPM-ET did not induce conditioned place aversion. CONCLUSIONS: ATPM-ET, at doses from 0.5 to 2 mg/kg, produced anxiolytic- and antidepressant-like effects without inducing aversive emotion. These effects were more closely mediated by activation of κ receptor than µ receptor.

[Toxicity attenuation and efficacy potentiation effects of sijunzi decoction on bladder carcinoma treated by chemotherapy in mice].

OBJECTIVE: To investigate the toxicity attenuation and efficacy potentiation effects of Sijunzi decoction (SJZD) on bladder carcinoma treated by chemotherapy in mice. METHODS: T739 mice were randomly divided into 8 groups after subcutaneous inoculation of bladder carcinoma cells, the control group (A); two mitomycin C (MMC) group, treated with MMC of routine dosage (B) and low-dosage (C) respectively; three SJZD groups, treated with SJZD of high (D), medium (E) and low-dosage (F) respectively; and two combined treatment groups, treated with SJZD of high-dosage + MMC of routine dosage(G) and SJZD of high-dosage + MMC of low-dosage(H). The medication was begun at 24 hrs after inoculation. The tumor inhibitory rate, activity of peritoneal macrophages after 14 days of treatment and change of peripheral white blood cells after 7 days of treatment were determined and the survival time of mice was observed. RESULTS: The survival time of mice in Group D was significantly higher than that in Group A (P < 0.05), while those in Group E and F showed insignificant difference as compared with those in Group A (P > 0.05). The highest tumor inhibitory rate was shown in Group B, but the survival time in that group showed no significant difference as compared to those in Group A (P > 0.05). The longest survival time (32.7 +/- 1.3 days) was shown in Group H, which was obviously different to that in other groups (P < 0.05). And the leukocyte counts and macrophage activity in Group H were better than those in Group B, C and G (P < 0.05), except that the tumor inhibitory rate was significantly lower than that in Group B, C and G (P < 0.05). CONCLUSION: Combined chemotherapy of SJZD with low dosage MMC has definite effect in inhibiting tumor growth in mice with bladder carcinoma, displaying special effects of toxicity attenuation and efficacy potentiation.

Antagonism of κ opioid receptor in the nucleus accumbens prevents the depressive-like behaviors following prolonged morphine abstinence.

The association between morphine withdrawal and depressive-like symptoms is well documented, however, the role of dynorphin/κ opioid receptor system and the underlying neural substrates have not been fully understood. In the present study, we found that four weeks morphine abstinence after a chronic escalating morphine regimen significantly induced depressive-like behaviors in mice. Prodynorphin mRNA and protein levels were increased in the nucleus accumbens (NAc) after four weeks of morphine withdrawal. Local injection of κ opioid receptor antagonist nor-Binaltorphimine (norBNI) in the NAc significantly blocked the expression of depressive-like behaviors without influencing general locomotor activity. Thus, the present study extends previous findings by showing that prolonged morphine withdrawal-induced depressive-like behaviors are regulated by dynorphin/κ opioid receptor system, and shed light on the κ opioid receptor antagonists as potential therapeutic agents for the treatment of depressive-like behaviors induced by opiate withdrawal.

Diffusion Efficiency and Bioavailability of Resveratrol Administered to Rat Brain by Different Routes: Therapeutic Implications.

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.

Short-term resveratrol exposure causes in vitro and in vivo growth inhibition and apoptosis of bladder cancer cells.

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.

Evaluation of resveratrol sensitivities and metabolic patterns in human and rat glioblastoma cells.

PURPOSE: To further elucidate the correlation of resveratrol sensitivities with biotransformation activities of human and rat glioblastoma cells for personalized anti-glioblastoma therapy. METHODS: Resveratrol sensitivity of human U251 and rat RG2 and C6 glioblastoma cells was evaluated by 3-[4,5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide/MTT, flow cytometry, and TUNEL assays. The metabolic patterns of those cell lines were analyzed by high-performance liquid chromatography/HPLC coupled with tandem mass spectrum/MS/MS, and high-resolution mass spectrometry/HRMS. Immunocytochemical staining and Western blotting were employed to check resveratrol metabolic enzyme expression. RESULTS: Both rat RG2 and C6 and human U251 glioblastoma cells are sensitive to 100 µM resveratrol in terms of growth arrest and increased apoptotic fraction. The main resveratrol metabolite in U251 cells is monosulfate biotransformed by sulfotransferases/SULTs and in RG2 and C6 cells is monoglucuronide generated by UDP-glucuronosyltransferase/UGT. Both metabolites show lesser therapeutic efficacy. Although brain-associated UGTs (UGT1A6, 2B7, and 8) and SULTs (SULT1A1, 1C2, and 4A1) are expressed in rat and human glioma cells, the overall level of UGTs is predominant in the rat and SULTs in human glioblastoma cells. In similar to SULT expression pattern, UGT1A6, 2B7, and 8 are frequently downregulated (84.6 %, 82/97; 90.7 %, 88/97; 80.4 %, 78/97) in human glioblastoma tissues. CONCLUSION: Our results suggest (1) the decreased resveratrol biotransforming activity in rat and human resveratrol-sensitive glioblastoma cells; (2) the discrepant resveratrol metabolic patterns between human and rat glioblastoma cells; (3) the more powerful anti-glioblastoma efficacy of trans-resveratrol rather than resveratrol monoglucuronide or monosulfate; and (4) the value of RG2 and C6 cells in establishing resveratrol-based rat in vivo therapeutic model.

Distinct sulfonation activities in resveratrol-sensitive and resveratrol-insensitive human glioblastoma cells.

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.

CRABP-II methylation: a critical determinant of retinoic acid resistance of medulloblastoma cells.

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/ß/γ and RXRα/ß/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, ß-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.

Microbial transformation of deoxyandrographolide by Alternaria alternata AS 3.4578.

Biotransformation of deoxyandrographolide (1) by Alternaria alternata AS 3.4578 gave five derivatives identified by spectral methods including 2D NMR as the known dehydroandrographolide (2) and 9beta-hydroxy-dehydroandrographolide (3) and the new compounds 9beta-hydroxy-deoxyandrographolide (4), 3alpha,17,19-trihydroxy-8,13-ent-labdadien-15,16-olide (5) and 3-oxo-9beta-hydroxy-deoxyandrographolide (6).

Metabolic patterns and biotransformation activities of resveratrol in human glioblastoma cells: relevance with therapeutic efficacies.

BACKGROUND: 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). METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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.

Antitumor effects of recombinant human interleukin-6 on mouse bladder carcinoma through Fas-mediated apoptosis.

PURPOSE: An apoptosis-inducing therapy is gradually becoming a new strategy for cancer treatment. The aim of this study was to investigate the mechanism of growth-inhibitory effects of recombinant human interleukin-6 (rhIL-6) on bladder tumor-bearing T739 mice in vivo. METHODS: Murine bladder transitional carcinoma cells (BTT739) were inoculated subcutaneously into T739 mice as a tumor model for evaluating the antitumor effects of rhIL-6. Then the mice were divided randomly into 5 groups: A, B, C, D and E. Different doses (0, 2, 4, 8 x 10(6) IU/kg body weight) of rhIL-6 were injected intraperitoneally twice per day and administered for 14 days, and 1 mg/kg/d mitomycin-C(MMC) was used as control. Tumor size was measured and determined as the mean of the largest diameter and the diameter at right angle. Animals were killed by CO(2) inhalation on the 15th day after tumor cell inoculation. Then, tumors were removed, weighed and collected. The tumor growth inhibition rate of rhIL-6 was calculated. The morphological characteristic changes of tumor cells were observed under electron microscope, and cell cycle analysis was determined by flow cytometry. The expressions of Fas, FasL and Bcl-2 protein on tumor cells were qualitatively detected by immunofluorescence cell staining, and their relative contents (rate of positive cells, RPC) were quantitatively determined with flow cytometry. RESULTS: rhIL-6 could inhibit bladder tumor growth in a dose-dependent manner in vivo. The tumor growth-inhibitory rates of 2, 4, 8 x 10(6) IU/kg rhIL-6 and 1 mg/kg MMC were 11.8, 39.5, 39.7 and 68.8%, respectively. Flow cytometry results showed that a hypodiploid peak before G1 phase could be found in tumor cells treated with rhIL-6. Moreover, the cells treated with rhIL-6 displayed disappearance of nucleoli, chromatin gathering under the nuclear membrane in mass or ring-shape under transmission electron microscopy. The rates of Fas, FasL protein-positive cells estimated by flow cytometry in rhIL-6-treated mice were (12.57 +/- 0.83) and (20.1 +/- 0.87) %, respectively, significantly higher than that (4.66 +/- 0.17) and (14.1 +/- 0.83) % in control mice (P < 0.01). There was no significant difference in the rate of Bcl-2 protein-positive cells between the mice in these two groups (P > 0.05). CONCLUSIONS: rhIL-6 had obvious antitumor effects on mouse bladder carcinoma in vivo, and the Fas signaling pathway might play an important role in rhIL-6-induced bladder carcinoma cell apoptosis.

Identification of metabolic pattern and bioactive form of resveratrol in human medulloblastoma cells.

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.

[Changes and significance of peripheral blood count in tumor rejection induced by a low dose of melphalan in C57BL/6 mice].

AIM: To investigate the relationship between changes of peripheral blood counts and tumor rejection induced by a low dose of melphalan in C57BL/6 mice. METHODS: Mouse lymphoma EL4 cells were inoculated subcutaneously into wild type C57BL/6 mice. Twelve days later, 7.5 mg/kg melphalan were administered intraperitoneally and the same volume of Normal Saline as control. Tumor sizes were observed and recorded subsequently. Blood samples were obtained from orbital venous sinus on different days before and after melphalan treatment, and then complete blood counts were performed and the relationship between the alterations of blood counts and tumor shrinkage after melphalan treatment was analyzed. RESULTS: Tumor sizes decreased and tumors disappeared after 7.5 mg/kg melphalan treatment; while tumors grow continuously in control mice. The number of WBC was increased a little (10.6 + or - 2.3) x 10(9)/L 6 h after melphalan treatment, but there was no significant difference with mice before melphalan injection (9.8 + or - 0.32) x 10(9)/L (P>0.05); The number of WBC decreased significantly at 4(th) day after melphalan treatment (P<0.01); Later it increased a little, but at 28(th); day after melphalan it still obviously lower than that of the normal (P<0.01). Hemoglobin (Hb) concentration decreased from (132 + or - 7) g/L before melphalan treatment to (110 + or - 14) g/L at 6 h after melphalan treatment (P<0.05). Later, the amount of Hb was decreasing and at 7th day it got to its lowest point (96 + or - 5) g/L. It increased gradually back to normal in 2 weeks after melphalan treatment. The platelet count increased to (1502 + or - 142) x 10(9)/L 6 h after melphalan treatment, significantly higher that that (914 + or - 322) x 10(9)/L before melphalan injection (P<0.01). It maintained at a high level for one week and it recovered back to normal level at 28(th) day after melphalan treatment. CONCLUSION: Tumor shrinkage after melphalan treatment was not related to the decreased number of WBC or RBC, but correlated with the increased number of platelet in 10 days after melphalan treatment.

[The significance and function of IFN-gamma on the changes of peripheral blood platelet count during tumor-rejection induced by a low dose of melphalan in C57BL/6 mice].

AIM: To investigate the significance and function of IFN-gamma on the changes of peripheral blood platelet count during tumor-rejection induced by a low dose of melphalan in C57BL/6 mice. METHODS: Mouse tumor rejection model induced by a single dose of melphalan was used in this experiment. Different gene-type tumor-bearing mice (IFN-gamma(+/-) and IFN-gamma(-/-)), which had the same genetic background of C57BL/6, were treated intraperitoneally with melphalan (7.5 mg/kg). Tumor size was observed and recorded every one to three days in these different gene-type mice subsequently. Blood samples were obtained from orbital venous sinus on different days before and after melphalan treatment, and then complete blood counts were performed. The function of IFN-gamma on the efficacy of chemotherapy and the changes of blood platelet count in IFN-gamma(+/-) and IFN-gamma(-/-) mice after melphalan treatment was analyzed. RESULTS: There was no significant difference in tumor sizes and blood platelet count between IFN-gamma(-/-) and IFN-gamma(+/-) mice (P>0.05). On the first day after melphalan (7.5 mg/kg) treatment, there were no significant changes in tumor sizes between mice in these two groups (P>0.05). Tumors shrank a little in IFN-gamma(-/-) mice and then grew gradually. Tumors relapsed in 2 w after melphalan injection in all IFN-gamma(-/-) mice, while tumor volumes decreased progressively and tumor cured at last in IFN-gamma(+/-) mice. The number of blood PLT in IFN-gamma(+/-) mice increased to (1935+/-378) x 10(9)/L 6 h after melphalan treatment, significantly higher than before (P<0.01); While in IFN-gamma(-/-) mice it was (1183+/-186) x 10(9)/L 6 h after melphalan treatment, no obvious increase than before. There was significant difference in blood PLT 6 h after melphalan treatment between IFN-gamma(+/-) and IFN-gamma(-/-) mice (P<0.01). Later, the numbers of blood PLT in IFN-gamma(+/-) mice decreased gradually and it dropped to normal (1158+/-270) x 10(9)/L on 11th day after melphalan treatment (P>0.05); While it sustained in normal range in IFN-gamma(-/-) mice. There was no significant difference in blood platelet count between IFN-gamma(-/-) and IFN-gamma(+/-) mice. CONCLUSION: Peripheral blood platelet count increased on the first day after melphalan treatment and tumors cured in IFN-gamma(+/-) mice; While tumors relapsed and there is no increase in blood platelet count on the first day after melphalan treatment in IFN-gamma(-/-) mice. These data indicated that the increase of blood PLT count was related to the function of IFN-gamma in tumor-bearing mice in vivo during tumor rejection induced by a low dose of melphalan.