Water-Soluble Dinitrosyl Iron Complex (DNIC): a Nitric Oxide Vehicle Triggering Cancer Cell Death via Apoptosis

Nitric oxide (NO) is an important cellular signaling molecule that modulates various physiological activities. Angiogenesis-promoting activities of NO-donor drugs have been explored in both experimental and clinical studies. In this study, a structurally well characterized and water-soluble neutral {Fe­(NO)₂}⁹ DNIC [(S­(CH₂)₂OH)­(S­(CH₂)₂NH₃)­Fe­(NO)₂] (DNIC <b>2</b>) was synthesized to serve as a NO-donor species. The antitumor activity of DNIC <b>2</b> was determined by MTT assay, confocal imaging, and Annexin-V/PI staining. The IC₅₀ values of DNIC <b>2</b> were 18.8, 42.9, and 38.6 μM for PC-3, SKBR-3, and CRL5866 tumor cells, respectively. Moreover, DNIC <b>2</b> promoted apoptotic cell death via activation of apoptosis-associated proteins and inhibition of survival associated proteins. In particular, DNIC <b>2</b> treatment suppressed PC-3 tumor growth by 2.34- and 19.3-fold at 7 and 21 days, in comparison with the control group. These results indicate that water-soluble DNIC <b>2</b> may serve as a promising drug for cancer therapy

Decreased autophagy and increased apoptosis in CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells after CLP.

<p>Spenocytes were obtained at 4h, 9h, 18h and 24h after CLP. Gating strategy for CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells by flow cytometry analysis (A) Cells were initially gated on a forward- and side-sactter lymphocytes gate to exclude dead cells, monocytes and granulocytes. The gated lymphocytes were then selected for either CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells. Cyto-ID Green (B), acridine orange (C) and Annexin-V (D) staining were further analyzed by flow cytometry. Representative histograms were gated on CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells. Values were shown as mean fluorescence intensities (MFI) for Cyto-ID Green/acridine orange and percentage for Annexin-V staining. In the histogram of Annexin-V staining, light gray histogram represents a staining control without adding Annexin-V. Results obtained from 5–6 animals in each group are shown as mean ± SEM in the bar graph. Data are compared by one-way analysis of variance and Student-Newman Keul's test. CLP: cecal ligation and puncture. *<i>P</i><0.05 vs. sham-operated mice.</p

Impaired number of CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells in CLP-induced Atg7^f/fCD4-Cre mice.

<p>Splenocytes obtained at 18h after surgery were stained for surface markers (CD4 and CD8) and analyzed by flow cytometry. Viable lymphocytes were gated by using forward scatter versus side scatter (excluding cell debris). The gated lymphocytes were further gated on the CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells. Representative histograms were shown percentage of lymphocytes (A). The absolute numbers of CD4⁺CD8⁻ and CD4⁻CD8⁺ T-cell subsets in the spleen were enumerated (B). Results obtained from 3 animals in each group are shown as mean ± SEM in the bar graph. Data are compared by one-way analysis of variance and Student-Newman Keul's test. CLP: cecal ligation and puncture. *<i>P</i><0.05 vs. respective sham mice. #<i>P</i><0.05 vs. CLP-induced Atg7^f/f mice.</p

Suppression of LC3-II and ATG7 protein levels in CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells after CLP.

<p>Splenic CD4⁺ and CD8⁺ T cell extracts were used for LC3-II and ATG7 protein expressions by Western blot analysis. CD4⁺ and CD8⁺ T cells obtained at 24h after CLP were isolated using CD4 and CD8 microbeads, respectively. A, Representative immunoblots of LC3-II and ATG7. B, Densitometric values of LC3-II. C, Densitometric values of ATG7. Actin was used as a loading control. Data are shown as mean ± SEM of 3 animals in each group and compared by two-tailed Student t-test. *<i>P</i><0.05 vs. sham-operated mice.</p

MOESM1 of Inhibitory effect of trans-ferulic acid on proliferation and migration of human lung cancer cells accompanied with increased endogenous reactive oxygen species and β-catenin instability

Additional file 1. DPPH radical-scavenging capacity of vitamin C as a positive control. (A) Vitamin C as a positive control in DPPH assay. (B) Quantificative analysis of (A). The radical-scavenging capacity of Vitamin C at indicated concentrations was quantified as the percentage decrease in absorbance at 492 nm against the blank control. *P < 0.05 and **P < 0.001 for trans-FA treatments against vehicle respectively

Decreased CD4⁺ cell cytokine production, macrophage phagocytosis and bacterial clearance in CLP-induced Atg7^f/fCD4-Cre mice.

<p>Splenocytes obtained at 18h after surgery were isolated with CD4 MicroBeads, and stimulated by anti-CD3/CD28 for 24h for cytokine production (A). In vitro phagocytosis, splenocytes obtained at 18h after CLP and cultured with <i>E. coli</i> BioParticles for 1h (B). Cells were then stained with surface marker (F4/80) and analyzed by flow cytometry. The light gray histogram represents a staining control without adding <i>E. coli</i> BioParticles. Blood and spleen tissues were collected at 18h after CLP and analyzed for bacteria loads (C). For bacterial loads, results were expressed as CFU per milliliter of blood and CFU per spleen tissue. Values of cytokine production and bacterial loads are shown as mean ± SEM of 6–8 animals in each group. Values of phagocytosis are shown as mean ± SEM of 3 animals in each group. Data are compared by one-way analysis of variance and Student-Newman Keul's test for cytokine production and phagocytosis. Data are compared by two-tailed Student t-test for bacterial loads. CLP: cecal ligation and puncture. †<i>P</i><0.05 vs. sham-operated Atg7^f/f mice. *<i>P</i><0.05 vs. respective sham mice. #<i>P</i><0.05 vs. CLP-induced Atg7^f/f mice.</p

Efficacy of autophagy inhibition in Atg7^f/fCD4-Cre mice after CLP.

<p>Atg7^floxp/floxp mice (Atg7^f/f) were crossed with CD4-Cre transgenic mice to generate doubly transgenic mice (Atg7^f/fCD4-Cre) in which <i>Atg7</i> gene was specifically deleted in T cells. ATG7 protein levels in Atg7^f/f and Atg7^f/fCD4-Cre mice were detected by Western blot analysis. Splenic CD4⁺ and CD8⁺ T cells were isolated using CD4 and CD8 MicroBeads, respectively. The efficacy of protein deletion of ATG7 from CD4⁺ and CD8⁺ cells was determined in normal Atg7^f/f and Atg7^f/f/CD4-Cre mice (A). ATG7 protein levels were further detected in sham and CLP-induced Atg7^f/fCD4-Cre and Atg7^f/f mice (cells obtained at 18h after CLP) (B). Actin was used as a loading control. Autophagy was further evaluated by Cyto-ID Green (C) and acridine orange (D) staining and followed by flow cytometry analysis. Cells were initially gated on a forward- and side-sactter lymphocytes gate to exclude dead cells, monocytes and granulocytes. T cells were divided into CD4⁺CD8⁻ and CD4⁻CD8⁺. Representative histograms were gated on CD4⁺CD8⁻ and CD4⁻CD8⁺ T cells, and shown as mean fluorescence intensities (MFI) of Cyto-ID Green and acridine orange staining. Data are shown as mean ± SEM of 3 animals in each group and compared by two-tailed Student t-test. CLP: cecal ligation and puncture. †<i>P</i><0.05 vs. sham-operated Atg7^f/f mice. *<i>P</i><0.05 vs. respective sham mice. #<i>P</i><0.05 vs. CLP-induced Atg7^f/f mice.</p

Ultrastructural features of autophagic vacuoles after CLP.

<p>Autophagy was mophologically characterized by transmission electron microscopy (A). Splenic tissues were harvested at 24h after CLP. In sham mice (a and b), splenocytes were normal in appearance with proper mitochondria distribution. Sham mice revealed autophagic vacuoles (arrowheads) in the cytosol with double- or single-membrane structures containing digested cytoplasmic components. CLP mice (c and d) displayed less autophagic vacuolization. A representative cell showed apoptosis in CLP mice with cell shrinkage, nuclear condensation and cellular disorganization (d, arrow). Quantification of autophagic vacuoles in sham and CLP mice (B). The number of autophagic vacuoles was counted under the microscope at 7,500× from 30 non-repeating micrographs for each mouse. Data are shown as mean ± SEM of 3 animals in each group and compared by two-tailed Student t-test. *<i>P</i><0.05 vs. sham-operated mice. CLP: cecal ligation and puncture.</p

Bioactive 6<i>S</i>‑Styryllactone Constituents of <i>Polyalthia parviflora</i>

Parvistones A–E (<b>1</b>–<b>5</b>), five new styryllactones possessing a rare α,β-lactone moiety and a 6<i>S</i> configuration, were isolated from a methanolic extract of <i>Polyalthia parviflora</i> leaves. The structures and the absolute configuration of the isolates were elucidated using NMR spectroscopy, specific rotation, circular dichroism, and X-ray single-crystal analysis. Compounds <b>8</b>, <b>9</b>, <b>11</b>, and <b>12</b> were isolated for the first time. The results were supported by comparing the data measured to those of 6<i>R</i>-styryllactones. Moreover, a plausible biogenetic pathway of the isolated compounds was proposed. The structure–activity relationship of the compounds in an in vitro anti-inflammatory assay revealed the 6<i>S</i>-styryllactones to be more potent than the 6<i>R</i> derivatives. However, the effect was opposite regarding their cytotoxic activity. In addition, 6<i>S</i>-styrylpyrones isolated showed more potent anti-inflammatory and cytotoxic activity when compared to the 1<i>S</i>-phenylpyranopyrones obtained

Bioactive 6<i>S</i>‑Styryllactone Constituents of <i>Polyalthia parviflora</i>

Parvistones A–E (<b>1</b>–<b>5</b>), five new styryllactones possessing a rare α,β-lactone moiety and a 6<i>S</i> configuration, were isolated from a methanolic extract of <i>Polyalthia parviflora</i> leaves. The structures and the absolute configuration of the isolates were elucidated using NMR spectroscopy, specific rotation, circular dichroism, and X-ray single-crystal analysis. Compounds <b>8</b>, <b>9</b>, <b>11</b>, and <b>12</b> were isolated for the first time. The results were supported by comparing the data measured to those of 6<i>R</i>-styryllactones. Moreover, a plausible biogenetic pathway of the isolated compounds was proposed. The structure–activity relationship of the compounds in an in vitro anti-inflammatory assay revealed the 6<i>S</i>-styryllactones to be more potent than the 6<i>R</i> derivatives. However, the effect was opposite regarding their cytotoxic activity. In addition, 6<i>S</i>-styrylpyrones isolated showed more potent anti-inflammatory and cytotoxic activity when compared to the 1<i>S</i>-phenylpyranopyrones obtained