Comparative studies on calotropins DI and DII from the latex of Calotropis gigantea

Autodigestion of two cysteine proteinases, calotropins DI and DII isolated from the latex of Calotropis gigantea, has been studied at pH 7.5 and 37 degrees C in the presence of an activating agent. Calotropin DI is more susceptible to autodigestion than calotropin DII. During autodigestion no interconversion of one calotropin to another has occurred, as verified by polyacrylamide gel electrophoresis in the presence and absence of sodium dodecyl sulfate. Immunologically, both calotropins are closely related, but they differ from papain and ficin. Both calotropins have blocked N-terminal amino acid residues. Their C-terminal amino acid sequences, determined by treatment with carboxypeptidase Y, are -(Pro, Ala)-Ala-Val-Tyr for calotropin DI and -(Ala, Val)-Ala-Pro-Tyr for calotropin DII. The tryptic peptide maps of their reduced and S-carboxymethylated derivatives suggest that both calotropins share a high proportion of common regions in their amino acid sequences. Calotropins DI and DII are two distinct proteinases, and they do not appear to be produced by autodigestion of a single precursor. Although they are inert to the common synthetic substrates of papain and ficin, their specificities toward oxidized insulin B chain are comparable to those of papain and ficin.     

Phytochemical and cytotoxic studies on the roots of Euphorbia fischeriana

With the aim of supporting the folk applications of Euphorbia fischeriana, a phytochemical study was performed, which led to the discovery of 9 compounds, including three new ones (1–3) and six known ones (4–9). Their structures were determined by 1D, 2D NMR, and HRESIMS analysis. In the cytotoxic assays on Hep-3B cell line, 2 showed stronger inhibitory effects (IC₅₀ 8.1 μmol/L) than that of positive control, and 1, 8 and 9 also gave inhibitory effects in a certain degree with IC₅₀ values of 12.5, 12.0 and 18.7 μmol/L, respectively. While on A549, the cytotoxic activities of 1 (IC₅₀ 11.9 μmol/L) and 8 (IC₅₀ 9.4 μmol/L) were superior to that of 5-Fu, and those of 4 and 9 were moderate with IC₅₀ values of 28.2 and 29.8 μmol/L, respectively. In addition, both petroleum ether and dichloromethane extracts showed cytotoxic activities with different degree, while n-butanol extracts had no effect. The results clarified that the low-polarity fractions of E. fischeriana, including triterpenoids, abietane and tigliane-type diterpenoids might be the potential bioactive ingredients which will exert strong antitumor effects.     

Five new quassinoids and cytotoxic constituents from the roots of Eurycoma longifolia

Eurycoma longifolia has been widely used for various traditional medicinal purposes in South-East Asia. In this study, five new quassinoids, eurylactone E (1), eurylactone F (2), eurylactone G (3), eurycomalide D (4), and eurycomalide E (5), along with ten known quassinoids (6–15) were isolated from the roots of E. longifolia. Their structures were determined by extensive spectroscopic methods, including 1D and 2D NMR, and MS spectra data. Among the isolated compounds, 13β-methyl,21-dihydroeurycomanone (6) has been reported as a synthetic derivative. However, it was isolated from the natural product for the first time in this study. The cytotoxic activities of fifteen compounds were evaluated against human lung cancer cell line, A549 and human cervical cancer cell line, HeLa.     

白扁豆多糖对人胃癌细胞凋亡的作用及其机制

目的：探讨白扁豆多糖对人胃癌细胞凋亡的作用及其相关机制。方法：胃癌细胞HGC-27和SGC-7901经终浓度为16、8、4、2、1和0 μg/ml的白扁豆多糖作用24、48和72h,各设3个复孔。MTS法检测其增殖活性;分别取经4、0 μg/ml白扁豆多糖作用24h的HGC-27和SGC-7901细胞（各3个复孔）,JC-1染色观察线粒体膜电位,流式细胞仪分析细胞周期和凋亡率,QPCR法探讨Bcl-2、caspase-3和Bax基因的mRNA转录水平。结果：白扁豆多糖作用后,HGC-27和SGC-7901细胞线粒体膜电位降低;细胞增殖显著受抑制（P<0.01）,且效果与药物作用浓度和时间有关;细胞凋亡率分别为53.15%和38.77%,均较PBS处理组（8.07%和6.03%）明显增加（P<0.01）,而细胞周期无显著变化;同时,细胞内Bcl-2基因的转录水平明显受抑制,Bax和caspase-3基因的转录明显上调（P<0.01）。结论：白扁豆多糖可通过调节Bax-Bcl-2-caspase3通路,诱导胃癌细胞HGC-27和SGC-7901凋亡。     

布地奈德对人气道上皮9HTEo-细胞增殖的影响及机制

探究布地奈德(budesonide,BUD)对人气道上皮9HTEo-细胞增殖的影响及机制,利用CCK-8法检测BUD对9HTEo-细胞增殖的影响,荧光显微镜观察经BUD处理后9HTEo-细胞形态的改变,流式细胞术检测细胞周期分布变化,Western blot检测p27kip1蛋白表达.CCK-8法结果显示3～10 μmol/L BUD对细胞具有显著的抑制作用(P＜0.01);荧光显微镜下,1～10 μmol/L BUD处理后细胞可见明显的增殖受抑、凋亡形态;流式细胞术检测G0/G1期细胞随着BUD浓度的增加而增多,S期细胞逐渐减少;Western blot检测结果提示BUD能使p27kip1表达增加.由此可见,BUD对人气道上皮细胞株9HTEo-具有显著增殖抑制作用,其作用与提高p27kip1蛋白的表达,将细胞阻滞在G0/G1期有关.     

Glycosylation of serum ribonuclease 1 indicates a major endothelial origin and reveals an increase in core fucosylation in pancreatic cancer

Human pancreatic ribonuclease 1 (RNase 1) is a glycoprotein expressed mainly by the pancreas and also found in endothelial cells. The diagnosis of pancreatic cancer (PaC) remains difficult and therefore the search for sensitive and specific markers is required. Previous studies showed that RNase 1 from human healthy pancreas contained only neutral glycans, whereas RNase 1 from PaC cell lines contained sialylated structures. To determine whether these glycan tumor cell-associated changes were also characteristic of serum RNase 1 and could be used as a marker of PaC, we have analyzed the glycosylation of serum RNase 1. The origin of serum RNase 1 was also investigated. Serum RNase 1 from two PaC patients and two controls was purified and the glycans analyzed by high-performance liquid chromatography (HPLC)-based sequencing and mass spectrometry. Although normal and tumor serum RNase 1 contained the same glycan structures, there was an increase of 40% in core fucosylation in the main sialylated biantennary glycans in the PaC serum RNase 1. This change in proportion would be indicative of a subset of tumor-associated glycoforms of RNase 1, which may provide a biomarker for PaC. Two-dimensional electrophoresis of the RNase 1 from several endothelial cell lines, EA.hy926, human umbilical vein endothelial cells (HUVEC), human mammary microvessel endothelial cells (HuMMEC), and human lung microvessel endothelial cells (HuLEC), showed basically the same pattern and was also very similar to that of serum RNase 1. RNase 1 from EA.hy926 was then purified and presented a glycosylation profile very similar to that from serum RNase 1, suggesting that endothelial cells are the main source of this enzyme.     

Sensitivity of human pancreatic carcinoma cell line (MIA PaCa-2) to Bisantrene and Theprubicin in vitro

Summary We tested the effect of Bisantrene (BS) and Theprubicin (THP-ADR) on cell growth of a human pancreatic carcinoma cell line (MIA PaCa-2). After 1 h exposure ID₅₀ of BS or THP-ADR was 3×10⁻⁷ and 5×10⁻⁸ M, respectively. Increasing the exposure time from 1 h to continuous exposure for 5d resulted in 11-fold decrease in ID₅₀ for BS and a 6-fold decrease for THP-ADR. Both drugs inhibited [¹⁴C]thymidine incorporation to the same extent and caused an accumulation of cells into G₂+M phase of the cell cycle. This work is supported by a gift, from Mr. Issam Fares and U.S. Public Health Service Grants AM 07114 and CA 19182.     

Inhibitory effect of 8-substituted adenosine derivatives on Ca++ and modulator protein-dependent phosphodiesterase activity.

8-Substituted adenosine and cyclic AMP derivatives exhibited some negative Cotton effects in circular Dichroism at B_2u band in pH 7.5 solution, suggesting that these derivatives take syn conformation. The adenosine derivatives, as well as cyclic AMP derivatives, competitively inhibited the cyclic AMP hydrolyzing activity in Ca⁺⁺ and modulator protein-dependent phosphodiesterase preparation from hog brain cortex. The inhibitory potential of an adenosine derivative was lower than that of the cyclic AMP derivative having the same substituent by the lack of the phosphate moiety for which affinity was 0.5 kcal / mol. These results may suggest that the cyclic AMP hydrolyzing site on the enzyme requires the syn conformation of purine riboside.     

GATA1通过上调NANOG促进胰腺癌肿瘤干细胞形成

目的:探究GATA1在胰腺癌肿瘤干细胞形成中的功能和作用机制。方法:通过流式细胞术检测GATA1对胰腺癌肿瘤干细胞形成的影响;通过实时荧光定量PCR和Western印迹筛选和验证GATA1下游的干性基因;通过双萤光素酶报告基因实验和染色质免疫共沉淀明确GATA1的调控机制。结果:GATA1过表达细胞株中肿瘤干细胞含量增加;GATA1上调NANOG的mRNA和蛋白表达水平;GATA1可以增强NANOG启动子的活性;GATA1结合在NANOG启动子-527^-524bp处的GATA序列。结论:GATA1可以通过结合在NANOG启动子上激活其转录,促进胰腺癌肿瘤干细胞的形成。     

Phytochemical and chemotaxonomic studies on the stems and leaves of Schisandra chinensis (Turcz.) Baill

A comprehensive phytochemical investigation of the stems and leaves of Schisandra chinensis (Turcz.) Baill. resulted in isolation of seventeen compounds, including five lignans: meso-dihydroguaiaretic acid (1), licarin-A (2), pregomisin (3), gomisin A (4), acutissimanide (5), three phenylpropanoids: 2-[4-(3-hydroxypropyl)-2-methoxyphenoxy]-propane-1,3-diol (6), 2-methoxy-4-(2-propenyl) phenyl β-D-glucopyranoside (7), erigeside 2 (8), six sesquiterpenoids: 7′-hydroxy-abscisic acid (9), burmannic acid (10), (3S,5R,6R,7E)-3,5,6-trihydroxy-7-megastigmen-9-one (11), 3-Cyclohexene-1,2-diol, 4-(3-hydroxybutyl)- 3, 5, 5-trimethyl- (12), (−)-loliolide (13), (3Z,5R,8R,11R)-Caryophyll-3-ene-5,8,15-triol (14), one monoterpenoid: (6R,3Z)-6,7-dihydroxy-3,7-dimethyl-2-octenoic acid (15) and two other compounds: methyl shikimate (16), 4-Hydroxydodec-2-enedioic acid (17). Their chemical structures were confirmed through NMR, HRESIMS and comparison with the data in the literature. This is the first report of compounds 5, 6, 8–15, 17 from the family Schisandraceae and compounds 2, 16 from the genus Schisandra. Furthermore, we performed a chemotaxonomic study of the separated compounds.     

单叶蔓荆果实中多甲氧基黄酮类成分的分离、鉴定及细胞毒活性分析

Six polymethoxylated flavonoids were isolated from ethyl acetate fraction of ethanol extracts from Vitex rotundifolia Linn. f. fruit, and their structures were identified by NMR and MS methods, and cyt...     

Phytochemical and chemotaxonomic study on the leaves of Rhododendron amesiae

The phytochemical study of the leaves of Rhododendron amesiae (Ericaceae) led to the isolation and identification of 19 compounds, including six diterpenoids (1–6), six triterpenoids (7–12) and seven flavonoids (13–19). The chemical structures of these compounds were identified by spectroscopic data, as well as by comparison with previously reported data in literature. This is the first systematic study on the chemical constituents of Rhododendron amesiae. All the compounds were isolated from this plant for the first time. Compounds 12, 14 and 15 were first isolated and reported from the genus Rhododendron and the family Ericaceae. Furthermore, the chemotaxonomic significance of these compounds was discussed.     

Paclitaxel administration and its effects on clinically relevant human cancer and non cancer cell lines

Comparisons of the effects of clinically relevant concentrations of the anticancer agent paclitaxel on growth, viability, and apoptosis were determined using in vitro human cell cultures. Growth of the cervical cancer cell line, HeLa-S3, was significantly reduced, and apoptotic index was significantly increased, after 24 h in cultures treated with 12 nM paclitaxel. In contrast, hepatic carcinoma (HEpG2) cells capable of detoxifying paclitaxel were only affected at paclitaxel concentrations ge120 nM. The previously uncharacterized non-cancerous human microvessel endothelial cell line HMEC-1, was more sensitive to paclitaxel treatment than both HeLa-S3 and HEpG2 cells, demonstrating decreased growth and increased apoptosis with 1.2 nM paclitaxel. These results are significant in the design of in vitro cell culture systems to study drug metabolism and toxicity.     

Comparison of cytotoxic T lymphocyte responses against pancreatic cancer induced by dendritic cells transfected with total tumor RNA and fusion hybrided with tumor cell

Pancreatic cancer (PC) is a deadly human malignancy. Dendritic cell (DC)-based immunotherapy with whole tumor antigens demonstrates potential efficiency in cancer treatment. Tumor RNA and tumor fusion hybrid cells are sources of whole tumor antigens for preparing DC tumor vaccines. However, the efficacy of these sources in eliciting immune responses against PC has not yet to be directly compared. In the present study, patient-derived PC cells and DCs were fused (DC–tumor hybrids) and primary cultured PC cell-derived total RNA was electroporated into autologous DCs (DC–tumor RNA). The antitumor immune responses induced by DC–tumor hybrids and DC–tumor RNA were compared directly. The results showed that both RNA and hybrid methodologies could induce tumor-specific cytotoxic T lymphocyte (CTL) responses, but pulsing DCs with total tumor RNA could induce a higher frequency of activated CTLs and T-helper cells than fusing DCs with autologous tumor cells. In addition, DC–tumor RNA triggered stronger autologous tumor cell lysis than DC–tumor hybrids. It could be concluded that DCs pulsed with whole tumor RNA are superior to those fused with tumor cells in priming anti-PC CTL responses. Electroporation with total tumor RNA may be more suitable for DC-based PC vaccination.     

Modification of blood group A antigen expression in a pancreatic cancer cell line (PC-1) by inhibitors of N-glycan processing.

Pancreatic adenocarcinomas induced in Syrian hamsters by treatment with N-nitrosobis(2-oxopropyl) amine express blood group A antigen, which is absent in normal pancreatic cells. On membrane glycoproteins purified from tumors, blood group A antigen has been found to be expressed on multiantennary Asn-linked complex glycans. In this study, we investigated the effect of inhibitors of Asn-glycan processing on blood group A antigen bearing glycan structures in a cell line (PC-1) established from a primary induced pancreatic cancer. Expression of blood group A antigen on cells and in membrane preparations was blocked by treatment with 1-deoxymannojirimycin, an inhibitor of mannosidase I, but was retained after treatment with swainsonine, an inhibitor of mannosidase II. However, swainsonine treatment altered the glycan structure associated with blood group A antigen from an endoglycosidase H resistant type to a sensitive type, indicating that the blood group A structure might shift from a complex type to a hybrid type glycan by this treatment. These results demonstrate that Asn-linked glycans carry the major blood group A antigens in PC-1 cells.     

CSN5 promotes the invasion and metastasis of pancreatic cancer by stabilization of FOXM1

COP9 signalosome subunit 5 (CSN5) has been involved in the progression of diverse human cancers. MMP2 plays an important role in the metastasis of cancer cells. However, the roles and relationship of in pancreatic cancer (PC) is still unknown. Here, our data shown that both CSN5 and MMP2 were significantly upregulated in PC compared with the corresponding adjacent tissues, where a positive correlation in their expression and associated malignant characteristics were found. Further, silencing of CSN5 expression markedly inhibited PC invasion and metastasis in vitro and in vivo, accompanied by decreased MMP2 expression. Moreover, the anti-metastasis role of CSN5 silence was reversed by MMP2 overexpression, whereas knockdown of MMP2 decreased PC metastasis driven by upregulation of CSN5. Further investigation revealed that CSN5 regulated MMP2 expression via activation of FOXM1 in PC cells. Mechanistically, CSN5 directly bound FOXM1 and decreased its ubiquitination to enhance the protein stability of FOXM1. Taken together, the results indicate that CSN5 can contribute to PC invasion and metastasis through activation of FOXM1/MMP2 axis.