Bone marrow-derived dendritic cell vaccination of dogs with naturally occurring melanoma by using human gp100 antigen

Canine malignant melanoma (CMM) is a common and aggressive form of cancer in dogs. Established therapeutic approaches such as surgery, chemotherapy, and radiation therapy (RT) have not proven curative. As a coadjuvant of RT and to enhance the antimelanoma immune response, we characterized dendritic cells (DCs) from the bone marrow (BM) of dogs with CMM, ex vivo, for use in therapeutic vaccines. BM mononuclear cells from 3 dogs with melanoma and from 1 healthy dog were cultured for 12 days in media supplemented with recombinant human granulocyte-macrophage colony stimulating factor, stem cell factor, tumor necrosis factor, and Flt-3 ligand. On day 11, DCs were transduced with an adenovirus vector encoding a xenoantigen, human melanoma antigen gp100. Each dog received 3 subcutaneous vaccinations over a 4-month period. Phenotypic analysis of the expanded DC population demonstrated expression of CD11c/CD18 and major histocompatibility complex class II surface markers, and ultrastructural features characteristic of DCs were observed on electron microscopy. On functional analysis, these DCs were able to stimulate allo-reactivity and capture and express gp100. One dog demonstrated antigen-specific cytotoxic T lymphocyte (CTL) activity in peripheral blood lymphocytes. This dog has displayed no clinical signs, either locally or systemically, of recurrent melanoma 48 months after initial DC injection. However, another dog, which was CTL negative, relapsed 22 months after vaccination. Ex vivo DC expansion is feasible for immunotherapy of spontaneous cancers in outbred dogs.     

International regulatory considerations pertaining to the development of stem cell-based veterinary medicinal products

Stem cell-based products (SCPs) are an emerging field of veterinary medicine that focuses on the regeneration, repair, or replacement of damaged tissues or organs. However, there are some issues in applying the traditional regulatory guideline for the approval of SCPs as veterinary medicinal products. This article describes the positions of Korea, US, and EU regarding SCPs, and compares the regulatory guidelines of each country for their safety evaluation. Although there are some differences in the regulatory guidelines, similar considerations in identifying the quality of SCPs and their safety has adopted. Overall, these guidelines need to be harmonized among countries.     

Sulfo-Gambierones,Two New Analogs of Gambierone Produced by Gambierdiscus excentricus

Ciguatera poisoning is caused by the ingestion of fish or shellfish contaminated with ciguatoxins produced by dinoflagellate species belonging to the genera Gambierdiscus and Fukuyoa. Unlike in the Pacific region, the species producing ciguatoxins in the Atlantic Ocean have yet to be definitely identified, though some ciguatoxins responsible for ciguatera have been reported from fish. Previous studies investigating the ciguatoxin-like toxicity of Atlantic Gambierdiscus species using Neuro2a cell-based assay identified G. excentricus as a potential toxin producer. To more rigorously characterize the toxin profile produced by this species, a purified extract from 124 million cells was prepared and partial characterization by high-resolution mass spectrometry was performed. The analysis revealed two new analogs of the polyether gambierone: sulfo-gambierone and dihydro-sulfo-gambierone. Algal ciguatoxins were not identified. The very low ciguatoxin-like toxicity of the two new analogs obtained by the Neuro2a cell-based assay suggests they are not responsible for the relatively high toxicity previously observed when using fractionated G. excentricus extracts, and are unlikely the cause of ciguatera in the region. These compounds, however, can be useful as biomarkers of the presence of G. excentricus due to their sensitive detection by mass spectrometry.     

甜、糯玉米耐热性体细胞的筛选

以不同品种甜玉米和糯玉米幼胚为供试材料,研究了不同培养基对愈伤组织诱导、继代和分化的影响,并在高温条件下筛选玉米耐热性变异体。结果表明：采用含有低浓度2,4-D的N₆为玉米幼胚诱导培养基,可产生一定比例的变异,愈伤组织诱导率高,质量较好;MS培养基适于分化;温度渐变法可以保证幼胚能够在产生耐热变异的同时不至于死亡,并在最后进行了恢复培养,即温度恢复到适温,淘汰不稳定愈伤组织,得到相对稳定的耐热性变异体;通过测定愈伤组织游离脯氨酸浓度、再生植株叶片质膜相对透性和叶片相对失水率,确定玉米胚变异体的耐热程度。     

微生物细胞传感器在环境监测中的应用研究进展

微生物细胞传感器是以微生物细胞作为生物敏感元件,可以感应环境中的总毒性以及各类污染物,从而实现快速监测的目的。关于该领域的研究报道越来越多,随着研究的不断发展,该项技术日趋成熟,从开始的仅仅在环境总毒性监测方面的应用,发展到能够特异性地检测多种污染物,如重金属类(Hg、Cd、As、Pb、Cu、Ag、Ni等)、有机污染物类(苯系物质、PAHs、酚类物质等)、抗生素类(四环素、β-内酰胺等)以及N、P类营养元素等等。由于微生物细胞自身的一些特点,使得其在环境监测领域具有非常显著的特点和优势,特别是在检测污染物的生物有效性方面,同时该技术可以满足环境监测中原位、快速、低耗的需要,所以其在环境监测领域必将得到越来越广泛的应用。本文重点综述了微生物细胞传感器在环境监测中的一些应用研究进展,并展望了其未来的发展趋势。     

Alternative Methods for the Detection of Emerging Marine Toxins: Biosensors,Biochemical Assays and Cell-Based Assays

The emergence of marine toxins in water and seafood may have a considerable impact on public health. Although the tendency in Europe is to consolidate, when possible, official reference methods based on instrumental analysis, the development of alternative or complementary methods providing functional or toxicological information may provide advantages in terms of risk identification, but also low cost, simplicity, ease of use and high-throughput analysis. This article gives an overview of the immunoassays, cell-based assays, receptor-binding assays and biosensors that have been developed for the screening and quantification of emerging marine toxins: palytoxins, ciguatoxins, cyclic imines and tetrodotoxins. Their advantages and limitations are discussed, as well as their possible integration in research and monitoring programs.     

Comparative Study on the Performance of Three Detection Methods for the Quantification of Pacific Ciguatoxins in French Polynesian Strains of Gambierdiscus polynesiensis

Gambierdiscus and Fukuyoa dinoflagellates produce a suite of secondary metabolites, including ciguatoxins (CTXs), which bioaccumulate and are further biotransformed in fish and marine invertebrates, causing ciguatera poisoning when consumed by humans. This study is the first to compare the performance of the fluorescent receptor binding assay (fRBA), neuroblastoma cell-based assay (CBA-N2a), and liquid chromatography tandem mass spectrometry (LC-MS/MS) for the quantitative estimation of CTX contents in 30 samples, obtained from four French Polynesian strains of Gambierdiscus polynesiensis. fRBA was applied to Gambierdiscus matrix for the first time, and several parameters of the fRBA protocol were refined. Following liquid/liquid partitioning to separate CTXs from other algal compounds, the variability of CTX contents was estimated using these three methods in three independent experiments. All three assays were significantly correlated with each other, with the highest correlation coefficient (r² = 0.841) found between fRBA and LC-MS/MS. The CBA-N2a was more sensitive than LC-MS/MS and fRBA, with all assays showing good repeatability. The combined use of fRBA and/or CBA-N2a for screening purposes and LC-MS/MS for confirmation purposes allows for efficient CTX evaluation in Gambierdiscus. These findings, which support future collaborative studies for the inter-laboratory validation of CTX detection methods, will help improve ciguatera risk assessment and management.