Histone deacetylase (HDAC) has been highlighted as one of key players in tumorigenesis and angiogenesis. Recently, several derivatives of psammaplin (Psams) from a marine sponge have been known to inhibit the HDAC activity, but the molecular mechanism for the inhibition has not fully understood. Here, we explored the mode of action of Psams for the inhibition of HDAC activity in the molecular and cellular level. Among the derivatives, psammaplin A (Psam A) showed the potent inhibitory activity in enzyme assay and anti-proliferation assay with IC50 value of 0.003 and 1 microM, respectively. Psam A selectively induced hyperacetylation of histones in the cells, resulting in the upregulation of gelsolin, a well-known HDAC target gene, in a transcriptional level. In addition, reduced Psam A showed a stronger inhibitory activity than that of non-reduced one. Notably, glutathione-depleted cells were not sensitive to Psam A, implying that cellular reduction of the compound is responsible for the HDAC inhibition of Psam A after uptake into the cells. Together, these data demonstrate that Psam A could exhibit its activity under the reduced condition in the cells and be a new natural prodrug targeting HDAC.
Tyrosine/*analogs & derivatives/chemistry/pharmacology ; Prodrugs/chemistry/*pharmacology ; Oxidation-Reduction ; Molecular Structure ; Humans ; Histones/metabolism ; Histone Deacetylases/*antagonists & inhibitors/*classification/genetics/metabolism ; Hela Cells ; Enzyme Inhibitors/chemistry/*pharmacology ; Disulfides/chemistry/*pharmacology ; Cell Proliferation ; Biological Products/chemistry/*pharmacology ; Acetylation

OBJECTIVETo investigate the molecular mechanisms of Z-ajoene mitosis blocking and telomerase inhibitory effects on HL-60 cells.

METHODSProliferation inhibition of HL-60 cell line was evaluated by MTT assay. Z-ajoene-induced mitotic blocking effect was investigated by flow cytometry. Immunoblotting analysis was used to determine cell cycle regulatory proteins. The telomerase activity of HL-60 cells was detected by TRAP-silver stain assay. Telomerase hTRT and TP1 mRNA level were determined by RT-PCR.

RESULTSZ-ajoene displayed great proliferation inhibiting effect on HL-60 cells. Progressive increase in the percentage of mitotic block at G(2)/M phase was observed from 4 h to 12 h after treatment with 10 micromol/L Z-ajoene, with a peak at 10 h, which was 1.95 times higher than that in control. Z-ajoene also caused an increase in cyclin B1 accumulation and a decrease of p34(cdc2) expression. But Z-ajoene did not change the level of cyclin A. After treating with 10 micromol/L Z-ajoene for 24 h, the telomerase activity of HL-60 cells was also decreased in a dose-independent manner. Furthermore, telomerase hTRT and TP1 mRNA levels decreased after 10 micromol/L Z-ajoene treatment for 24 h.

CONCLUSIONThe results suggest that Z-ajoene has potent anti-cancer activity, and that its inhibitory effect on telomerase activity and on cell growth might be the result of G(2)/M phase blocking.

Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Cycle ; drug effects ; Disulfides ; pharmacology ; Garlic ; chemistry ; HL-60 Cells ; Humans ; Mitosis ; drug effects ; Telomerase ; metabolism

OBJECTIVETo investigate the effects of garlicin on fibroblasts proliferation and type I collagen synthesis and explore its anti-fibrosis mechanism.

METHODSGarlicin was added into the culture fluid of NIH3T3 cell, taking Radix Salviae miltiorrhizae as the control medicine. The spiking of H3-thymidine DNA was detected, also the hydroxyproline (HOP) concentration in the culture fluid by alkali digestion method and the protein expression of type I collagen in NIH3T3 cells by immunofluorescent staining.

RESULTSThe NIH3T3 cell growth and proliferation rate were obviously reduced after garlicin treatment concentration-dependently in range of 0.2 - 5 microg/mL; HOP level and protein expression of type I collagen also lowered.

CONCLUSIONGarlicin could inhibit NIH3T3 cell proliferation, reduce the synthesis and protein expression of type I collagen so as to exert the anti-fibrosis effect.

Allyl Compounds ; pharmacology ; Animals ; Cell Proliferation ; drug effects ; Collagen Type I ; biosynthesis ; Disulfides ; pharmacology ; Dose-Response Relationship, Drug ; Garlic ; chemistry ; Hydroxyproline ; analysis ; Mice ; NIH 3T3 Cells

To reveal the effects of disulfide bridges in rubella virus glycoprotein E1 on the membrane fusion activity, the recombinant plasmid pBSK-SPE2E1 and site-directed mutagenesis to mutate 11 cysteines individually in the ectodomain of E1 to remove a disulfide bridge from the wild-type E1 were constructed. All mutants and the wild-type plasmid were expressed on BHK-21 cell. Giemsa Staining was used to show the polykaryon formed in the transfected BHK-21 cells. The cell surface expression efficiency of the plasmids was assayed with fluorescence-activated cell sorter (FACS). Hemadsorption was performed to detect the receptor recognition activity of the recombinant plasmids. The results showed that all the 10 disulfide bridges in the ectodomain of E1 played an important role in the process of the membrane fusion. The removal of any disulfide bridge resulted in the loss of the fusion activity. The disulfide formed by the 5th and the 8th cysteine might be critical for the interaction of E1 and E2. While the disulfide bridges formed by the 3rd, the 4th, and the 13th might influence the membrane fusion activity of E1 directly.
Cell Membrane ; drug effects ; Cysteine ; chemistry ; Disulfides ; chemistry ; pharmacology ; Flow Cytometry ; Membrane Fusion ; drug effects ; Mutagenesis, Site-Directed ; Rubella virus ; chemistry ; Viral Envelope Proteins ; chemistry ; Viral Fusion Proteins ; Virus Internalization ; drug effects

OBJECTIVETo observe the effect of garlicin on adhension molecules CD11a and deformability of peripheral neutrophil in patients with acute cerebral infarction (ACI).

METHODSNeutrophils were separated from peripheral blood of healthy subjects and ACI patients, and incubated in 37 degrees C in vitro. The CD11a expression was detected by antibody fluorescence labeling method and the time of neutrophils passing millipore membrane were measured for calculation of the filter index.

RESULTSCD11a expression rate in healthy subjects was 34.64 +/- 25.34%, while in patients was 55.35 +/- 30.54%, difference between them was significant (P < 0.05). After garlicin treatment, it lowered to 49.16 +/- 31.68%, as compared with untreated group, P < 0.05. The neutrophil filter index in healthy group, untreated group, garlicin treated group and Nimodipine treated group was 0.87 +/- 0.46, 6.42 +/- 6.40, 3.47 +/- 3.67 and 5.03 +/- 3.72 respectively, comparison between that in the garlicin treated group and in untreated group showed significant difference (P < 0.05).

CONCLUSIONGarlicin could effectively inhibit the CD11a expression in peripheral blood neutrophils and improve the deformability of the neutrophils in ACI patients.

Aged ; Allyl Compounds ; pharmacology ; CD11a Antigen ; biosynthesis ; Cell Adhesion ; drug effects ; Cell Separation ; Cerebral Infarction ; blood ; Disulfides ; pharmacology ; Erythrocyte Deformability ; drug effects ; Female ; Garlic ; chemistry ; Humans ; Male ; Middle Aged ; Neutrophils ; physiology

The characteristics of uptake, transepithelial transport and efflux of Z- and E-ajoenes isolated from the bulbs of Allium sativum were studied. A human colon cell model Caco-2 cell monolayers in vitro cultured had been applied to study the characteristics of uptake, transepithelial transport and efflux of Z- and E-ajoenes. The quantitative determination of Z- and E-ajoenes was performed by high-performance liquid chromatography. Z- and E-Ajoenes can be detected only in the apical side and can be metabolized, but both compounds can not be transported from apical-to-basolateral and basolateral-to-apical directions in cultured Caco-2 cell monolayers. The metabolism of Z- and E-ajoenes in Caco-2 cell monolayers can be partially inhibited by vitamin C as an anti-oxidant, metyrapone as an inhibitor to subtype CYP3A of cytochrome P450 drug metabolism enzymes, and sodium azide as an inhibitor to ATP production. It is shown that neither Z-ajoene nor E-ajoene can pass through Caco-2 cell monolayers, and that they can be metabolized by the cells. The metabolism might be in correlation with cytochrome P450 drugs metabolism enzymes in Caco-2 cell monolayers.
Antioxidants ; pharmacology ; Ascorbic Acid ; pharmacology ; Biological Transport ; drug effects ; Caco-2 Cells ; Cell Membrane ; drug effects ; metabolism ; Cytochrome P-450 CYP3A ; metabolism ; Cytochrome P-450 CYP3A Inhibitors ; Disulfides ; chemistry ; isolation & purification ; pharmacokinetics ; Enzyme Inhibitors ; pharmacology ; Garlic ; chemistry ; Humans ; Metyrapone ; pharmacology ; Plants, Medicinal ; chemistry ; Sodium Azide ; pharmacology ; Stereoisomerism

The constrained alpha-helical structure of a C-peptide is useful for enhancing anti-HIV-1 activity. The i and i+3 positions in an alpha-helical structure are located close together, therefore D-Cys (dC) and L-Cys (C) were introduced at the positions, respectively, to make a dC-C disulfide bond in 28mer C-peptides. Accordingly, this study tested whether a dC-C disulfide bond would increase the alpha-helicity and anti-HIV-1 activity of peptides. A C-peptide can be divided into three domains, the N-terminal hydrophobic domain (HPD), middle interface domain (IFD), and C-terminal hydrogen domain (HGD), based on the binding property with an N-peptide. In general, the dC-C modifications in HPD enhanced the anti-HIV-1 activity, while those in IFD and HGD resulted in no or much less activity. The modified peptides with no activity clearly showed much less alpha-helicity than the native peptides, while those with higher activity showed an almost similar or slightly increased alpha-helicity. Therefore, the present results suggest that the introduction of a dC-C bridge in the N-terminal hydrophobic domain of a C-peptide may be useful for enhancing the anti-HIV-1 activity.
Amino Acid Sequence ; Anti-HIV Agents/chemical synthesis/*chemistry/isolation & purification/*pharmacology ; Cell Line ; Circular Dichroism ; Cysteine/chemistry ; Disulfides/chemistry ; HIV Envelope Protein gp41/*chemistry ; HIV-1/*drug effects/growth & development ; Humans ; Inhibitory Concentration 50 ; Models, Molecular ; Molecular Sequence Data ; Peptides/chemical synthesis/*chemistry/isolation & purification/*pharmacology ; Protein Structure, Secondary ; Protein Structure, Tertiary ; Research Support, Non-U.S. Gov't ; Structure-Activity Relationship