33 research outputs found

    Quantitative Proteomic Profiling of Early and Late Responses to Salicylic Acid in Cucumber Leaves

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    <div><p>Salicylic acid (SA) is an important phytohormone that plays vital regulatory roles in plant growth, development, and stress responses. However, studies on the molecular mechanism of SA, especially during the early SA responses, are lagging behind. In this study, we initiated a comprehensive isobaric tag for relative and absolute quantitation (iTRAQ)-based proteomic analysis to explore the early and late SA-responsive proteins in leaves of cucumber (<i>Cucumis sativus</i> L.) seedlings. Upon SA application through the roots, endogenous SA accumulated in cucumber leaves. By assaying the changes in marker gene expression and photosynthetic rate, we collected samples at 12 h and 72 h post treatment (hpt) to profile the early and late SA responsiveness, respectively. The iTRAQ assay followed by tandem mass spectrometry revealed 135 differentially expressed proteins (DEPs) at 12 hpt and 301 DEPs at 72 hpt. The functional categories for these SA-responsive proteins included in a variety of biochemical processes, including photosynthesis, redox homeostasis, carbohydrate and energy metabolism, lipid metabolism, transport, protein folding and modification, proteolysis, cell wall organization, and the secondary phenylpropanoid pathway. Conclusively, based on the abundant changes of these DEPs, together with their putative functions, we proposed a possible SA-responsive protein network. It appears that SA could elicit reactive oxygen species (ROS) production via enhancing the photosynthetic electron transferring, and then confer some growth-promoting and stress-priming effects on cells during the late phase, including enhanced photosynthesis and ROS scavenging, altered carbon metabolic flux for the biosynthesis of amino acids and nucleotides, and cell wall reorganization. Overall, the present iTRAQ assay provides higher proteome coverage and deepened our understanding of the molecular basis of SA-responses.</p></div

    The Combination of Three Natural Compounds Effectively Prevented Lung Carcinogenesis by Optimal Wound Healing

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    <div><p>The tumor stroma has been described as “normal wound healing gone awry”. We explored whether the restoration of a wound healing-like microenvironment may facilitate tumor healing. Firstly, we screened three natural compounds (shikonin, notoginsenoside R1 and aconitine) from wound healing agents and evaluated the efficacies of wound healing microenvironment for limiting single agent-elicited carcinogenesis and two-stage carcinogenesis. The results showed that three compounds used alone could promote wound healing but had unfavorable efficacy to exert wound healing, and that the combination of three compounds made up treatment disadvantage of a single compound in wound healing and led to optimal wound healing. Although individual treatment with these agents may prevent cancer, they were not effective for the treatment of established tumors. However, combination treatment with these three compounds almost completely prevented urethane-induced lung carcinogenesis and reduced tumor burden. Different from previous studies, we found that urethane-induced lung carcinogenesis was associated with lung injury independent of pulmonary inflammation. LPS-induced pulmonary inflammation did not increase lung carcinogenesis, whereas decreased pulmonary inflammation by macrophage depletion promoted lung carcinogenesis. In addition, urethane damaged wound healing in skin excision wound model, reversed lung carcinogenic efficacy by the combination of three compounds was consistent with skin wound healing. Further, the combination of these three agents reduced the number of lung cancer stem cells (CSCs) by inducing cell differentiation, restoration of gap junction intercellular communication (GJIC) and blockade of the epithelial-to-mesenchymal transition (EMT). Our results suggest that restoration of a wound healing microenvironment represents an effective strategy for cancer prevention.</p></div

    The wound healing microenvironment restored GJIC but did not directly decrease tumor cells.

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    <p><b>(</b>A) The combination of three compounds reversed GJIC loss in urethane-treated BEAS-2B cells. Images were taken at ×10 magnification. (B and C) The combination of three compounds led to angiogenesis and optimal fibroblast proliferation but did not directly decrease A549 cancer cells. The results are presented as mean±SE (n = 5/group) **<i>p</i> < 0.01 <i>vs</i> control group.</p

    The effect of DCO-6 on cell viability of murine macrophages.

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    <p>(A) The chemical structure of DCO-6. (B) RAW264.7 cells or peritoneal macrophages from BALB/c mice were treated with various concentrations of DCO-6. After 24 h of incubation, LDH in the culture supernatant was tested. The absorbance value at 420 nm was measured by a microplate reader. The percentage of LDH released from the cells was determined using the formula: % release  =  LDH activity in supernatant/(LDH activity in supernatant + LDH activity in cell lysate). Data are shown as means ± S.D. of three independent experiments. *P<0.05 vs medium control.</p

    The conserved histidine boxes of the membrane-bound FAD proteins in cucumber.

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    <p>The conserved histidine boxes of the membrane-bound FAD proteins in cucumber.</p

    Inhibition of LPS-induced NO, IL-1β and IL-6 production by DCO-6 in murine macrophages.

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    <p>RAW264.7 cells or peritoneal macrophages from BALB/c mice were treated with various concentrations of DCO-6 in the absence or presence of LPS. (A) The levels of NO, IL-1β and IL-6 in the cell culture medium were determined 24 h after LPS stimulation as described in Methods. (B) The levels of iNOS, IL-1β and IL-6 mRNA were determined by real-time quantitative PCR 8 h after LPS stimulation. β-actin was used as an invariant control. Data are shown as means ± S.D. of three independent experiments. *P<0.05 vs LPS control.</p

    Protective effect of DCO-6 against LPS-induced septic shock in mice.

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    <p>BALB/c mice were administered LPS (10 mg/kg) and DCO-6 (10 or 20 mg/kg) or vehicle (olive oil) intraperitoneally. (A) Survival rate. (B) Serum cytokine levels of IL-1β and IL-6 were measured by ELISA 3 h after LPS injection. Data are shown as means ± S.D. of three independent experiments. n  = 10. *P<0.05 vs model control. (C) Peritoneal macrophages were isolated from the mice 3 h after LPS injection. Whole cell lysates were prepared, and the protein levels of total and phosphorylated p38 was detected by Western blotting analysis. Traces shown are representative of three independent experiments. Bands from (C) were analyzed by densitometry. Quantitative data are shown. *P<0.05 vs model control. (D) Peritoneal macrophages were isolated from the mice 3 h after LPS injection and incubated with DCFH-DA. DCF fluorescence distribution was detected by flow cytrometry. Data were analyzed by Cell Quest software. *P<0.05 vs model control.</p

    The screened three wound healing agents suppressed lung carcinogenesis and their combination led to optimal preventive efficacy.

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    <p>(A) Three wound healing agents were administered to animals following the first carcinogen treatment and were found to prevent lung carcinogenesis. (B) They were administered following the last carcinogen treatment but did not prevent lung carcinogenesis, whereas the combination of these three agents remained effective. (C) Summary data of lung tumor incidence (n = 20). (D) Three wound healing agents restored the lung barrier in urethane-induced lung carcinogenesis. The results are presented as mean±SE (n = 10/group). *<i>p</i> < 0.05, **<i>p</i> < 0.01, vs control group.</p

    tBlastn result using AtFAD family proteins as queries.

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    <p>tBlastn result using AtFAD family proteins as queries.</p

    The wound healing microenvironment prevented lung cell malignant transformation.

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    <p>(A) The combination of three compounds reduced the population expressing stem cell markers or EMT markers examined by flow cytometry and in A549 cells. (B) The combination of three compounds decreased A549 cell self-renew shown as the number of tumor spheres and soft agar colonies. (C) The combination of three compounds decreased the population expressing EMT markers examined by flow cytometry in urethane-treated BEAS-2B cells. The results are presented as mean±SE (n = 5/group) **<i>p</i> < 0.01 <i>vs</i> control group.</p
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