抗肿瘤血管生成主动免疫治疗研究进展

肿瘤血管生成在肿瘤生长、侵袭、转移中具有十分重要的作用。调控血管生成的因子很多,包括血管内皮细胞生长因子（VEGF）、成纤维细胞生长因子（FGF）、表皮细胞生长因子（EGF）等。近年来抗血管生成的肿瘤治疗已经取得较大进展,特别是抗肿瘤血管形成主动免疫治疗,已经成为抗肿瘤研究的热点。因而,以异种同源分子和非异种同源分子为疫苗的研究很有意义。     

Inhibitory effect of Crocus sativus L. ethanol extract on adjuvant-induced arthritis

Saffron is a well-known spice produced from dried stigmas of Crocus sativus L. flowers. Apart from its wide use in food preparations, it also has a broad range of medical properties. We examined the potential anti-inflammatory effects of saffron ethanolic extract (SEE) using an animal model of arthritis. Adjuvant-induced arthritis was induced in Wistar rats by injection of Complete Freund's Adjuvant. The rats were then injected intraperitoneally every other day with 25–600 mg SEE/kg or dexamethasone (DEX, 2 mg/kg). Changes in body weight, paw oedema and arthritis indices were recorded over the subsequent 12 days of treatment. Results revealed that SEE particularly at the higher concentrations significantly reduced paw and tibiotarsal joint diameters and comparing with DEX caused no significant change in body weight. These observations suggest that SEE displays a considerable anti-inflammatory potency and could potentially be used as an anti-arthritic agent in control of inflammation in rheumatoid arthritis.     

Body surface potential mapping system equipped with a microprocessor for the dynamic observation of potential patterns

A body surface potential mapping system equipped with a microprocessor has been developed. It enables high-speed real-time and simultaneous observation of e.c.g. data of 96 lead points in a character display mode. The machine has functions not only to perform basic data processing such as baseline correction and linear interpolation for malfunctioned electrodes but also to generate an isopotential contour map, record data onto a floppy disc, make hard copies of displayed patterns and characters, etc. To increase the operation speed, all the assembler programs were put in one module. Thus, an operator can easily recognise the spatial motion pattern of the body surface potential map and, in addition, find malfunctioned channels by observing the pattern. A belt type electrode system allows quick and adaptable attachment. The practical usefulness of the system has been proved with the co-operation of cardiologists with over 120 cases of cardiac disease.     

Mosquito larvicidal activities of Solanum nigrum L. leaf extract against Culex quinquefasciatus Say

The present study was carried out to establish the larvicidal activities of crude and solvent extracts of Solanum nigrum L. leaves against Culex quinquefasciatus Say as target species. The results indicated that the mortality rates at 0.5% concentration were highest amongst all concentrations of the crude extracts tested against all the larval instars at 24, 48 and 72 h of exposure. Result of log probit analysis (at 95% confidence level) revealed that lethal concentration LC₅₀ and LC₉₀ values gradually decreased with the exposure periods in bioassay experiment with the crude plant extract. The results of regression analysis of crude extract of S. nigrum revealed that the mortality rate is positively correlated with the concentration of the extracts. Mature leaves of S. nigrum were also extracted with six different solvents [viz. petroleum ether, benzene, ethyl acetate, chloroform: methanol (1:1 v/v), acetone and absolute alcohol] to determine the best extractant for subsequent isolation and characterization of active ingredient. Mortality rate with ethyl acetate extract was significantly higher (p < 0.05) than other extracts when 50-ppm doses were used. The corresponding LC₅₀ value of acetone, absolute alcohol, petroleum ether, chloroform: methanol (1:1 v/v), benzene and ethyl acetate extracts were 72.91 ppm, 59.81 ppm, 54.11 ppm, 32.69 ppm, 27.95 ppm and 17.04 ppm, respectively, after 24 h of exposure period. Results of this study show that the ethyl acetate extract of S. nigrum may be considered as a potent source of a mosquito larvicidal agent.     

A potential role for the nucleolus in L1 retrotransposition

Determining the subcellular localization of the L1 ORF2 protein (ORF2p) has been impossible to date because of technical limitations in detecting either endogenous or overexpressed forms of the protein. Here we report visualization of the full-length ORF2p in cultured human cells following expression in a modified vaccinia virus/T7 RNA polymerase (MVA/T7RP) system. The MVA/T7RP system was used to ascertain subcellular localization of L1 ORF1p and ORF2p both as fusions with green fluorescent protein and by immunocytochemistry. Full-length ORF2p was predominantly cytoplasmic, while carboxy-terminal-deleted ORF2p localized additionally to the nucleolus. We mapped a functional nucleolar localization signal in ORF2p. ORF1p appeared in the cytoplasm with a speckled pattern and colocalized with ORF2p in nucleoli in a subset of cells. These findings help explain the presence of chimeras between L1s and small RNA gene sequences recently discovered in the human genome.     

Delta-like ligand 4-targeted nanomedicine for antiangiogenic cancer therapy

Tumor angiogenesis is a multistep process involved with multiple molecular events in cancer microenvironment. Several molecular-targeted agents aiming to suppress tumor angiogenesis have been successfully translated into cancer clinic. However, new strategies are still urgently desired to be excavated to overcome the poor response and resistance in some antiangiogenic therapies. Recently, Delta-like ligand 4 (Dll4) is identified to be specifically over-expressed on tumor vascular endothelial cells (EC), and the Dll4-Notch pathway serves as a critical regulator in the development and maintenance of tumor angiogenesis. The intensively up-regulated phenotype of Dll4 on the membrane of tumor vascular EC implies that Dll4 may act as a targetable address for drug delivery system (DDS) to achieve targeted antiangiogenic cancer therapy. Here, a nano-DDS, GD16 peptide (H₂N-GRCTNFHNFIYICFPD-CONH₂, containing a disulfide bond between Cys₃ and Cys₁₃) conjugated nanoparticles loading paclitaxel (GD16-PTX-NP), which can specifically target the angiogenic marker Dll4, was fabricated for the investigation of antiangiogenic therapeutic efficacy in human head and neck cancer FaDu (Dll4-negative) xenograft in nude mice. The results demonstrate that GD16-PTX-NP achieved controlled drug release and exhibited favorable in vivo long-circulating feature. GD16-PTX-NP exerted enhanced antiangiogenic activity in the inhibition of human umbilical vein endothelial cell (HUVEC) viability, motility, migration, and tube formation, and in the Matrigel plug model as well, which can be definitely ascribed to the active internalization mediated by the interaction of GD16 and the over-expressed Dll4 on EC. GD16-PTX-NP showed accurate in vivo tumor neovasculature targeting property in FaDu tumor, where the paclitaxel was specifically delivered into the tumor vascular EC, leading to significant apoptosis of tumor vascular EC and necrosis of tumor tissues. The antiangiogenic activity of GD16-PTX-NP significantly contributed to its in vivo anticancer efficacy in Fadu tumor; moreover, no overt toxicity to the mice was observed. Our research firstly presents the potency and significance of a Dll4-targeted nanomedicine in antiangiogenic cancer therapy.     

Effects of ethanol extract of Artemisia annua L. against monogenean parasites of Heterobranchus longifilis

Ethanol extract of Artemisia annua was effective in the dislodgement and mortality of monogenean parasites of juvenile Heterobranchus longifilis at concentrations ranging from 50 to 200 mg/l. Five hundred 1-week-old juvenile fish were stocked in hapa in earthen pond for 7 days to accumulate parasites. The approximate number of parasites per fish was confirmed by counting the number of parasites attached to body surfaces and the gills with a stereo-microscope before being exposed to the extract under in vivo conditions. The bioactivity of the extract was conducted in plastic Petri dishes with three replications and controls. The results obtained from A. annua extract were matched against those produced by pure artemisinin and artesunate powder, respectively, under similar experimental conditions. There was a faster effect of pure artemisinin crystals on the parasites as compared to A. annua extract and artesunate. Coagulation of parasite cells was observed with artemisinin treatment, whereas parasites were merely dislodged from their attachment organs and killed some hours later in the same concentration of A. annua. There were positive correlations between the number of parasites dislodged/killed and the concentration of A. annua extract, artemisinin, and artesunate powder, respectively, as well as the duration of exposure of affected fish to the substances. This led to the conclusion that A. annua contains substances that are effective against helminthes parasites of H. longifilis.     

The wound healing effect of aqueous extract from Piper amalago L. in diabetic patient

Diabetes patients present a complex healing process due to several factors directly linked to their pathology. The use of medicinal plants that aid in tissue repair can bring great benefits to such individuals. This case report describes how the topical application of the aqueous extract produced from the leaves of Piper amalago L. was used to aid the healing of a lacerated wound in the left thumb of a patient with type 2 diabetes mellitus.The aqueous extract of the leaves of Piper amalago L. was prepared in boiling water. During the boiling process the dried leaves were submerged in the boiling water and left for five min. The injured thumb was submerged in the solution and the leaves were placed on the injury. The action of the aqueous extract obtained from the leaves of P. amalago was shown to be promising in the healing of a wound in a patient with type 2 diabetes mellitus. The topical application of the aqueous extract produced from the leaves of P. amalago assisted in the healing of a lacerated wound in the left thumb of a patient with type 2 diabetes mellitus over a period of 15 days.     

Spirodela polyrhiza (L.) Sch. ethanolic extract inhibits LPS-induced inflammation in RAW264.7 cells

Objective: Spirodela polyrhiza (L.) Sch. is widely used in Korean traditional medicine. No previous work has investigated in detail the anti-inflammatory activities of S. polyrhiza or assessed in vitro their potential underlying mechanism(s). We assessed the effects of S. polyrhiza ethanolic extract (SPEE) on the production of inflammatory mediators in lipopolysaccharide (LPS)-stimulated RAW264.7 cells and investigated some potential underlying mechanisms. Additionally, we performed simultaneous determination of seven flavonoids in S. polyrhiza by high-performance liquid chromatography (HPLC)-photodiode array (PDA).

Materials and methods: RAW264.7 cells were subjected to 5, 10, 20, and 50 μg/mL of SPEE for 1 h then treated with LPS for 24 h. Production of namely nitric oxide (NO), prostaglandin E₂ and cytokine levels were measured by the Griess reagent and ELISA, respectively. To investigate the underlying mechanisms of the anti-inflammatory activities of SPEE, expression of NO synthase (iNOS), cyclooxygenase-2 (COX-2), and nuclear factor-kappa B (NF-κB) proteins were evaluated by western blot analysis. HPLC analysis was performed using a Gemini C₁₈ column at 40°C and PDA detection at 340 nm.

Results: SPEE treatment significantly inhibited the LPS-induced production of NO, prostaglandin E₂, interleukin-6, and tumor necrosis factor-α and inhibited the expression of iNOS and COX-2 via attenuation of NF-κB p65 expression. The contents of the seven flavonoids in S. polyrhiza range from 0.25 to 8.77 mg/g.

Conclusions: These results indicate that the anti-inflammatory activity of SPEE may be NF-κB p65 signaling. Also, the method will help to improve quality control of S. polyrhiza.     

Spirodela polyrhiza (L.) Sch. ethanolic extract inhibits LPS-induced inflammation in RAW264.7 cells

Objective: Spirodela polyrhiza (L.) Sch. is widely used in Korean traditional medicine. No previous work has investigated in detail the anti-inflammatory activities of S. polyrhiza or assessed in vitro their potential underlying mechanism(s). We assessed the effects of S. polyrhiza ethanolic extract (SPEE) on the production of inflammatory mediators in lipopolysaccharide (LPS)-stimulated RAW264.7 cells and investigated some potential underlying mechanisms. Additionally, we performed simultaneous determination of seven flavonoids in S. polyrhiza by high-performance liquid chromatography (HPLC)-photodiode array (PDA). Materials and methods: RAW264.7 cells were subjected to 5, 10, 20, and 50 μg/mL of SPEE for 1 h then treated with LPS for 24 h. Production of namely nitric oxide (NO), prostaglandin E(2) and cytokine levels were measured by the Griess reagent and ELISA, respectively. To investigate the underlying mechanisms of the anti-inflammatory activities of SPEE, expression of NO synthase (iNOS), cyclooxygenase-2 (COX-2), and nuclear factor-kappa B (NF-κB) proteins were evaluated by western blot analysis. HPLC analysis was performed using a Gemini C(18) column at 40°C and PDA detection at 340 nm. Results: SPEE treatment significantly inhibited the LPS-induced production of NO, prostaglandin E(2), interleukin-6, and tumor necrosis factor-α and inhibited the expression of iNOS and COX-2 via attenuation of NF-κB p65 expression. The contents of the seven flavonoids in S. polyrhiza range from 0.25 to 8.77 mg/g. Conclusions: These results indicate that the anti-inflammatory activity of SPEE may be NF-κB p65 signaling. Also, the method will help to improve quality control of S. polyrhiza.     

Petiveria alliacea L. extract protects mice against Listeria monocytogenes infection--effects on bone marrow progenitor cells

In this study we have investigated the effects of Petiveria alliacea on the hematopoietic response of mice infected with Listeria monocytogenes. Our results demonstrate a protective effect of the crude extract of P. alliacea since the survival of the treated/infected was higher than that in the infected group. Moreover, the number of granulocyte/macrophage colonies (CFU-GM) and the serum colony stimulating activity levels were increased in the treated/infected mice in relation to the infected group. These results suggest an immunomodulation of Petiveria alliacea extract on hematopoiesis, which may be responsible, at least in part, for the increased resistance of mice to Listeria monocytogenes infection.     

Evaluation and characterization of the immunomodulatory activity of the protein extract from Citrullus colocynthis L.

Citrullus colocynthis L. is a plant widely used in Moroccan traditional medicine as a strong laxative and has many others uses. The aim of the present work is to evaluate the immunomodulatory activity of the protein extract (PE) from seeds of C. colocynthis. The results of this study showed that the PE has an immunosuppressive activity in vitro. The activity was assessed through the evaluation of the proliferation of splenocytes induced by the mitogen concanavalin-A factor. The PE showed no agglutination activity against rabbit erythrocytes, and was separated into two fractions F1 and F2 with molecular weight of 160 and 79 kDa, respectively. Fraction F1 showed the highest immunosuppressive activity (44.62%) in dose-dependent manner (IC₅₀ 60 µg/mL) and is characterized into three sub-units of 53.65 kDa. The sub-units of F1 were determined by the polyacrylamide gel electrophoresis. The fraction F1 of the PE could be a promising immunosuppressing agent extracellular.     

Preclinical safety evaluation of rhuMAbVEGF, an antiangiogenic humanized monoclonal antibody.

Recombinant humanized antivascular endothelial growth factor (rhuMAbVEGF) is a monoclonal IgG1 antibody that is being developed as an antiangiogenic agent for use in treating a variety of solid tumors. Preclinical safety studies included an immunohistochemical tissue cross-reactivity study, in vitro hemolytic potential and blood compatibility studies, and multiple dose toxicity studies. Toxicity studies were conducted in cynomolgus monkey because rhuMAbVEGF is pharmacologically active in this species and does not bind rat or mouse vascular endothelial growth factor (VEGF). Following twice weekly administration of rhuMAbVEGF for 4 or 13 wk, young adult cynomolgus monkeys exhibited physeal dysplasia characterized by a dose-related increase in hypertrophied chondrocytes, subchondral bony plate formation, and inhibition of vascular invasion of the growth plate. In addition, decreased ovarian and uterine weights and an absence of corpora lutea were observed in females receiving 10 and 50 mg/kg/dose in the 13-wk study. Both the physeal and ovarian changes were reversible with cessation of treatment. No other treatment-related effects were observed following rhuMAbVEGF administration at doses up to 50 mg/kg. These findings indicate that VEGF is required for longitudinal bone growth and corpora lutea formation and that rhuMAbVEGF can reversibly inhibit physiologic neovascularization at these sites.     

Activity of anthocyanins from fruit extract of Ribes nigrum L. against influenza A and B viruses

Earlier, we have detected antiviral activity in an extract from Ribes nigrum L. fruits ("Kurokarin", name of the one species of black currant in Japanese) against influenza A and B viruses, and herpes simplex virus 1 (Knox et al., Food Processing 33, 21-23, 1998). In the present study, the antiviral activity of constituents of a Kurokarin extract and the mechanism of its antiviral action were examined. Kurokarin extracts were separated to fractions A to D by column chromatography. The major constituents of the fraction D were estimated as anthocyanins. The fraction D was further fractionated by thin-layer chromatography (TLC) to fractions A' to G'. The fraction E' consisted of 3-O-alpha-L-rhamnopyranosyl-beta-D-glucopyranosyl-cyanidin and 3-O-beta-D-glucopyranosyl-cyanidin, and the fraction F' consisted of 3-O-alpha-L-rhamnopyranosyl-beta-D-glucopyranosyl-delphinidin and 3-O-beta-D-glucopyranosyl-delphinidin, identified by high performance liquid chromatography (HPLC) with standards and by high resolution mass spectrometry. The fractions D' to G' showed potent antiviral activity against influenza viruses A and B. The additive antiviral effect of a combination of the fractions E' and F' was assessed. Anthocyanins in the fraction F' did not directly inactivate influenza viruses A and B, but they inhibited virus adsorption to cells and also virus release from infected cells.     

Anti-inflammatory efficacy of methanolic extract of Muntingia calabura L. leaves in Carrageenan induced paw edema model

The plant Muntingia calabura L. is a well-known herb which gained attention due to its pharmacological value. The necessity of this plant in human ailments is illustrious in old medical practices. Muntingia calabura L. leaves were therapeutically used for ulcer, fever, headache etc. The study was designed to assess the acute toxicity and anti-inflammatory potential of methanolic extract of Muntingia calabura L. (MEMC) in in vivo models. Two different doses (550, 2000 mg/kg body weight) of MEMC were taken to evaluate the acute toxicity response. The drugs were given orally to wistar rats and were monitored for behavioral changes and mortality for 14 days period. The blood parameter analysis, serum analysis of liver and kidney injury markers and histopathological evaluation of kidney, heart and liver were carried out. The Carrageenan induced paw edema model was performed to inspect the anti-inflammatory response of MEMC. The level of CRP in serum and the histological alterations in the paw tissue were evaluated. There were no evident symptoms of toxicity observed in animals treated with MEMC at the dose of 2000 mg/kg body weight. The Carrageenan induced paw edema model study established the anti-inflammatory potential of MEMC. The MEMC, which is innoxious, can act as a potential anti-inflammatory drug.