黄皮素内酯Ⅱ在黄皮植物体的分布及对稗草生化代谢的影响

[目的]测定黄皮素内酯Ⅱ(2′,3′-epoxyanisolactone)在黄皮[Clausena lansium(Lour.)Skeels]植株的花、枝叶和果核中的含量,明确其对稗草生化代谢的影响。[方法]采用液相色谱法进行含量的测定,小杯法培养供试稗草。[结果]黄皮素内酯Ⅱ在花粗提物中的含量较少,仅为0.43%,在枝叶和果核中的含量分别为3.33%和3.21%。黄皮素内酯Ⅱ处理稗草植株,处理后稗草根中蛋白含量和氨基酸含量与未经处理存在明显的差异,在50μg/mL的浓度下,可以显著地降低稗草植株内的蛋白含量和增加植株内的氨基酸含量。当处理浓度为50、25、12.5、6.25、3.125μg/mL时,样品中的含氮量分别为1.402 8、1.317 2、1.250 2、1.128 7、0.903 2mg/g,比对照0.832 1mg/g均有所增加。黄皮素内酯Ⅱ处理后,稗草的蛋白含量与对照相比有所下降,处理浓度为50、25、12.5、6.25、3.125μg/mL时,蛋白含量比对照分别降低75.91%、66.43%、62.43%、58.57%和58.03%。[结论]黄皮素内酯Ⅱ对稗草代谢有一定影响。

臭假黄皮中的新大环内酰胺——黄皮素

从臭假黄皮(Clausena excavata)叶的乙酸乙酯提取部分分离到9个化合物,化合物Ⅰ为一新的大环内酰胺,经各种波谱分析,其化学结构鉴定为1,1′-二十二(碳)内二酰胺,命名为黄皮素(1)。化合物Ⅱ、Ⅲ、Ⅳ、Ⅴ及Ⅵ分别为11β-羟基黄柏酮(2)、5,7,3′,4′,5′-五羟基-3-0-鼠李糖黄酮甙(3)、阿魏酸(4)、7-羟基香豆素(5)及三乙胺氢碘酸盐。

Hydroxysafflor Yellow A Promotes Vascular Endothelial Cell Proliferation via VEGF/VEGF Receptor

Aim To study the proliferative effeet of hydroxysaftlor yellow A （HSYA） on cultured canine aortic endothelial cell （VEC） in normoxic （21% O2 ） or hypoxic （10% O2 ） culture and the underlying mechanism. Methods The endothelial cells were scratched from trypsined canine aorta endothelium. HSYA was added to the cells at final concentrations of 1 × 10^-3, 1 × 10^-4 and 1 × 10^-5 mol· L^-1, respectively. VEGF （2.6 × 10^-7 mol· L^-1 ）-treated cells were used as the positive control. The proliferative effect of HSYA on VEC was determined at 48, 72, 96, and 120 h in normoxic culture by MTI＂ assay. Similarly, the proliferation of VEC was determined at 12, 24, 48, and 72 h in hypoxic culture by MTF assay. The effects of HSYA on VEC proliferation and VEGF secretion were investigated by MTr and ELISA assays at the presence of the antibodies to VEGF and VEGF receptors. Results Pretreatment with HSYA at concentrations of 1 × 10^-3 and 1 × 10^-4 mol· L^-1 enhanced VEC proliferation in normoxic culture. The most significant enhancing effect of HSYA on VEC proliferation was achieved at 24, 48, and 72 h in hypoxic culture in concentration-dependent and time-dependent manner. HSYA at 1 × 10^-3 mol·L^-1 showed a potency similar to VEGF at 2.6 × 10^-7 mol·L^-1 . Pretreatment with the antibodies of Flt-1, KDR or VEGF blocked the proliferative effect of HSYA with similar potencies. Antibodies of Fit-1 or VEGF antagonized the promoting effect of HSYA on VEGF secretion. Conclusion HSYA promotes VEC proliferation either in normoxic or hypoxic culture, especially in the latter condition. This effect of HSYA is at least partly mediated by VEGF and VEGF receptor.

Curcumin suppresses PPARδ expression and related genes in HT-29 cells

AIM:To investigate the effects of curcumin on the expression of peroxisome proliferator-activated receptorδ(PPARδ)and related genes in HT-29 cells. METHODS:HT-29 cells were treated with curcumin (0-80μmol/L)for 24 h.The effects of curcumin on the morphology of HT-29 cells were studied by Hoechst 33342 staining.The activity of caspase-3 was determined using DEVD-p NA as substrate.The levels of peroxisome PPARδ,14-3-3εand vascular endothelial growth factor(VEGF)in HT-29 cells were determined by Western blotting analysis and their mRNA expression was determined by real-time quantitative RT-PCR. RESULTS:Treatment with 10-80μmol/L curcumin induced typical features of apoptosis and activated the caspase-3 in HT-29 cells.The expression of PPARδ, 14-3-3εand VEGF was reduced and the activity of β-catenin/Tcf-4 signaling was inhibited by curcumin treatment. CONCLUSION:Curcumin can induce apoptosis of HT-29 cells and down-regulate the expression of PPARδ,14-3-3εand VEGF in HT-29.

Quercetin,the key constituent of Astragali Radix,modulates ferroptosis in PASMCs and attenuates hypoxia pulmonary hypertension via the MAPK signaling pathway

This study delved into the mechanism by which the principal component of Astragali Radix regulated ferroptosis in the context of hypoxia-induced pulmonary hypertension,employing a combination of network pharmacology and experimental validation techniques.Active constituents of Astragali Radix and their corresponding targets were identified using the TCMSP database,while therapeutic targets associated with hypoxia-induced pulmonary hypertension were sourced from the GeneCards database.The Venn online tool facilitated the identification of overlapping targets between the active constituents of Astragali Radix and hypoxia-induced pulmonary hypertension.Interaction network diagrams depicting the relationship between Astragali Radix’s active constituents and their targets were constructed using Cytoscape software,with core targets and sub-networks identified using the CytoHubba plug-in.Gene ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analyses were conducted using the DAVID database.Additionally,the FerrDb database was consulted to analyze genes implicated in regulating ferroptosis.The investigation revealed 18 active constituents selected from Astragali Radix,with quercetin emerging as the key component.A total of 35 potential targets associated with Astragali Radix in regulating ferroptosis and addressing hypoxia-induced pulmonary hypertension were predicted.Experimental validation demonstrated that quercetin could inhibit the MAPK signaling pathway,resulting in reduced Fe2+and lipid peroxide levels,increased GPX4 expression,and the reversal of ferroptosis.In summary,this study elucidated the fundamental constituents and pivotal signaling pathways through which Astragali Radix modulated ferroptosis and mitigated hypoxia-induced pulmonary hypertension.Specifically,quercetin,a core constituent of Astragali Radix,was observed to inhibit ferroptosis in pulmonary arterial smooth muscle cells via the MAPK pathway and alleviate hypoxia-induced pulmonary hypertension.

Stability and degradation of hydroxysafflor yellow A and anhydrosafflor yellow B in the Safflower injection studied by HPLC-DAD-ESI-MS^n

Safflower is a popular Chinese medicinal plant and Safflower injection is extensively used for the clinical treatment of cerebrovascular and cardiovascular diseases. In this study, HPLC-DAD-ESI-MSn was utilized to study the stability and degradation of the two major but chemically unstable bioactive compounds hydroxysaffior yellow A and anhydrosaffior yellow B, in Safflower injection. The impact of light irradiation, temperature, and pH on the stability of these two compounds were studied. The results showed that hydroxysafflor yellow A and anhydrosafflor yellow B could degrade at high temperature （〉60 ℃） or extreme pHs （pH ≤ 3.0 or 〉7.0）, but not under light irradiation. The common degradation product was p-coumaric acid. Chemical structures of the other degradation products were characterized by LC-MS. Hypothetical degradation pathways were proposed. In addition, ADP-induced platelet aggregation tests showed that the degradation of anhydrosaffior yellow B could reduce the anticoagulation activities of Safflower injection. Our results suggest that temperature and pH are critically important for the preparation and storage of Safflower injection.

HORMONAL CONTROL OF THE VITELLOGENESIS IN THE JAPANESE OAK SILKWORM, ANTHERAEA YAMAMAI (LEPIDOPTERA : SATURNIIDAE)

Effects of ecdysteroid and juvenile hormone (JH) on vitellogenesis of the Japanese oak silkworm, Antheraea yamamai are reported in this article. After topical treatment with 20 hydroxyecdysone alone or JH analog (i.e. methoprene) alone and combined treatment with these two chemicals, vitellogenin (Vg) titers in the fat body and haemolymph at the pupal stage were mostly higher than those of the control, indicating that both ecdysteroid and JH exerted a promoting effect on the synthesis of Vg. In contrast, the Vg uptake was markedly inhibited by JH while stimulating effect of the ecdysteroid could be shown that vitellin (Vt) titer in the ovary was lower after methoprene treatments, but higher after 20 hydroxyecdyson treatments. Meanwhile, effects of these two hormones on Vg synthesis in the fat body were also tested with the incubation in vitro with Grace medium containing 3H leucine and the hormones. The results demonstrated that Vg synthesis was stimulated after treating with methoprene alone or 20 hydroxyecdysone alone and combined treating with these two chemicals, and particularly ecdysteroid had more marked positive effect. To comprehensively concluded our results, it could be regarded that ecdysteroid play the main role in the regulation of vitellogenesis for the Japanese oak silkworm.

Expression, subcellular localization and protein-protein interaction of four isoforms of EcR/USP in the common cutworm

Ecdysone receptor （EcR） and ultraspiracle （USP） form heterodimers to mediate ecdysteroid signaling during molting and metamorphosis. Various EcR/USP heterodimers have been reported. However, it is unclear what kind of EcR/USP combination is adopted by lepidopteran insects during the larval-pupal metamorphosis and whether the EcR/USP heterodimer varies among different tissues. To address these questions, two isoforms of each EcR and USP were cloned from the common cutworm, their messenger RNA expression patterns were examined by real-time quantitative polymerase chain reaction in different tissues during the larval-pupal metamorphosis and in the midgut in response to hormonal induction. Furthermore, their subcellular localization and protein-protein interaction were explored by transient expression and far-western blotting, respectively. All the four genes were significantly up-regulated in prepuae and/or pupae. The expression profiles of EcRB1 and USP1 were nearly identical to each other in the epidermis, fat body and midgut, and a similar situation also applied to EcRA and USP2. The three genes responded to 20-hydroxyecdysone （20E） induction except for USP2, and USP1 could be up-regulated by both 20E and juvenile hormone. The four proteins mainly localized in the nucleus and the nuclear localization was promoted by 20E. The protein-protein interaction between each EcR and USP was found in vitro. These results suggest that two types of EcR/USP heterodimer （EeRA/USP2 and EcRB 1/USP1） may exist simultaneously in the common cutworm, and the latter should play more important roles during the larval-pupal metamorphosis. In addition, the types of EcR/USP heterodimer do not vary in the tissues which undergo histolysis and regeneration during metamorphosis.

Curcumin co-treatment ameliorates resistance to gefitinib in drug-resistant NCI-H1975 lung cancer cells

OBJECTIVE: To examine whether a combinative treatment with curcumin enhances the effects of the epidermal growth factor receptor-tyrosine kinase inhibitor(EGFR-TKI) gefitinib on cell proliferation, clonogenic capacity and apoptosis in the drug-resistant lung cancer cell line NCI-H1975, and further investigate the molecular mechanisms involved.METHODS: NCI-H1975 cells were treated with curcumin and gefitinib alone or in combination, and cell proliferation, clonogenic capacity and apoptosis were examined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay, clone forming experiments, and flow cytometry, respectively, while p38, extracellular regulated protein kinase(ERK)1/2, and protein kinase B(AKT)phosphorylation were examined using Western blotting.RESULTS: Compared with the effects of either agent alone, the combination of curcumin and gefitinib had a stronger suppressive effect on proliferation and the clonogenic capacity(P < 0.05), and showed an increased ability to promote apoptosis(P < 0.05) and reduce p38, ERK1/2, and AKT phosphorylation(P < 0.05).CONCLUSION: Co-treatment of curcumin and gefitinib significantly improves the ability of gefitinib to inhibit cell proliferation, suppress the clonogenic capacity and enhance apoptosis in NCI-H1975 cells,and these effects are possibly mediated via a decrease in phosphorylation of proteins in downstream pathways of the epidermal growth factor receptor.