前体饲喂对细胞培养生产紫杉烷的促进作用

研究了前体化合物对中国红豆杉细胞培养生产紫杉烷的影响.0.5 mmol/L 丙酮酸钠使紫杉醇和1β-羟基巴卡亭Ⅰ的产率分别增加236 %和225.8 %.2 mmol/L 3-甲基 -2-丁烯 -1-醇和0.4 mmol/L苯甲酸钠使2α,5α,10β,14β-四乙酰氧基 -紫杉 -4(20),11-二烯产率分别提高218.6 %和245.2 %.1 mmol/L乙酸钠和0.5 mmol/L 丙酮酸钠使14β-2-甲基丁酰氧基 -2α,5α,10β-三乙酰氧基-紫杉 -4(20),11-二烯的产率分别提高193 %和210.4 %.结果表明添加适当浓度的前体有利于紫杉烷的合成.     

Identification of a cell-wall peptidase (NlpC/P60) from Nocardia seriolae which induces apoptosis in fathead minnow cells

Nocardia seriolae, a Gram-positive bacterium, is the main pathogen of fish nocardiosis. Protein NlpC/P60 is a cell-wall peptidase and a potential virulence factor of N. seriolae. Subcellular localization research revealed that both NlpC/P60-GFP and NlpC/P60Δsig-GFP fusion proteins were evenly distributed in the whole cell of fathead minnow (FHM) cells. Furthermore, typical apoptotic features, such as nuclear pyrosis and apoptotic bodies, were observed in the transfected FHM cells and grouper spleen cells by the overexpression of protein NlpC/P60. Then, quantitative assays of mitochondrial membrane potential (ΔΨm) value, caspase-3 activity and apoptosis-related gene (Bax, BNIP3, TNF1 and TNF6) mRNA expression were conducted. The results showed that ΔΨm was decreased, caspase-3 was significantly activated, and the mRNA expression of pro-apoptotic genes (Bax and BNIP3) and tumour necrosis factors (TNF1 and TNF6) was up-regulated in NlpC/P60-overexpressed cells. Taken together, the results indicated that the protein NlpC/P60 of N. seriolae might involve in apoptosis regulation. This study may lay the foundation for further study on the function of N. seriolae NlpC/P60 and promote the understanding of the virulence factors and pathogenic mechanism of N. seriolae.     

鱼类生殖细胞移植的研究进展及应用前景

生殖细胞移植是指将供体的生殖细胞移植到同种或异种受体体内,供体生殖细胞嵌合到受体性腺,经过增殖、分化并最终发育为功能性配子的过程。作为辅助生殖技术,它不仅为珍稀濒危动物的繁育和保护提供了新途径,同时也为生殖干细胞的功能研究提供了有效手段。鱼类生殖细胞移植研究首先在模式鱼类斑马鱼中开展,经过十多年的发展,取得了一系列突破性的进展:主要包括先后建立了以胚胎、仔鱼和成鱼为受体的生殖细胞移植体系,精原和卵原干细胞的发现拓宽了供体生殖细胞的选择,受体的选择与制备方法的完善。该技术在缩短鱼类性成熟周期、性控育种、珍稀濒危鱼类保护等方面具有巨大的应用前景,已成功在多种淡水和海水鱼类中开展了研究和应用。本文结合作者的研究实践和经验,系统地梳理和总结了鱼类生殖细胞移植的研究进展,指出了该技术实践应用的关键问题,并探讨了其应用前景。     

Partial replacement of fishmeal with Clostridium autoethanogenum single‐cell protein in the diet for juvenile black sea bream (Acanthopagrus schlegelii)

Clostridium autoethanogenum protein (CAP) is a single‐cell protein derived from ethanol production and may have potential to become a substitute for fishmeal in aquafeeds. A 70‐day feeding trial was conducted with black sea bream (Acanthopagrus schlegelii) (mean initial weight 6.03 ± 0.09 g) to evaluate partial replacement of fishmeal with CAP in diets. Seven isonitrogenous and isoenergetic diets were formulated with graded levels of CAP (0, 4.85, 9.70, 14.55, 19.40, 38.80 and 58.20%) to replace fishmeal. The inclusion of CAP at all dietary levels tested did not significantly affect the growth performance (p > .05). Fish fed the CAP58.20% diet showed a significantly lower feeding rate, with significantly higher protein efficiency ratio and feed efficiency ratio compared with fish fed the other diets (p < .05). No statistical differences were found in dorsal muscle and whole‐body compositions. Total superoxide dismutase in serum of fish fed CAP58.20% diet was significantly lower compared with that of the control. Malondialdehyde, catalase, total antioxidant capacity and digestive enzyme activities revealed no significant differences among dietary treatments. Phosphorus retention efficiency significantly increased, and phosphorus discharge showed a downward trend with increasing CAP inclusion levels. In conclusion, the results indicated that CAP is a safe and effective alternative protein source, which can replace fishmeal in the diet of black sea bream up to 58.20%, without adverse effects on growth performance, antioxidation and digestive enzyme activity. This study has shown the potential of converting industrial waste into a high protein feed ingredient for aquafeeds.     

Fishmeal alternative from renewable CO2 for rainbow trout feed

The rapid global growth in fish farming and limited supply of fish meal (FM) has consequently reduced FM inclusion levels in compound feeds leading to a higher reliance on alternative protein sources. Sasya is a single cell protein (SCP) product that has a similar amino acid profile as FM. The aim of this study was to evaluate the effects of replacing FM with SCP on in vivo digestibility, growth, feed efficiency, whole‐body proximate/amino acid composition and gene expression levels of various hepatic enzymes in rainbow trout (Oncorhynchus mykiss). Three isonitrogenous (470 g/kg crude protein) and isolipidic (18 g/kg crude lipid) diets were formulated as follows: Diet 1: control (30% FM); Diet 2:24% FM + 6% SCP and Diet 3:18% FM + 12% SCP. Each diet was hand‐fed to triplicate tanks containing 30 rainbow trout fingerlings (4.99 ± 0.20 g) for 9 weeks. Apparent digestibility coefficients of SCP for dry matter, crude protein, lipid and energy were 60, 80, 93 and 74% respectively. Growth performance (final weight: 69–71 g), feed conversion ratios (0.91–0.94) as well as whole‐body protein and amino acid composition were unaffected by diets. However, Diet 3 significantly increased whole‐body crude fat and energy. Fish fed the SCP‐based diets had significantly higher expression for carnitine palmitoyltransferase‐1b (CPT1b), fatty acid delta 6 desaturase (FADS6) and fatty acid elongase 5 compared to the control. Overall, the quality of the SCP was similar as FM. Therefore, this product could enlarge the portfolio of alternative protein sources that can be used in fish diets and thus open a new market opportunity for use of a new feed resource in the feed industry.     

Identification of a mitochondrial‐targeting secretory protein from Nocardia seriolae which induces apoptosis in fathead minnow cells

Nocardia seriolae is the main pathogen responsible for fish nocardiosis. A mitochondrial‐targeting secretory protein (MTSP) 3141 with an N‐terminal transit peptide (TP) from N. seriolae was predicted by bioinformatic analysis based on the genomic sequence of the N. seriolae strain ZJ0503. However, the function of the MTSP3141 and its homologs remains totally unknown. In this study, mass spectrometry analysis of the extracellular products from N. seriolae proved that MTSP3141 was a secretory protein, subcellular localization research showed the MTSP3141‐GFP fusion protein co‐localized with mitochondria in fathead minnow (FHM) cells, the TP played an important role in mitochondria targeting, and only the TP located at N‐terminus but not C‐terminus can lead to mitochondria directing. Moreover, quantitative assays of mitochondrial membrane potential (ΔΨm) value, caspase‐3 activity and apoptosis‐related gene (Bcl‐2, Bax, Bad, Bid and p53) mRNA expression suggested that cell apoptosis was induced in FHM cells by the overexpression of both MTSP3141 and MTSP3141ΔTP (with the N‐terminal TP deleted) proteins. Taken together, the results of this study indicated that the MTSP3141 of N. seriolae was a secretory protein, might target mitochondria, induce apoptosis in host cells and function as a virulence factor.     

Reestablishment of transzonal projections and growth of bovine oocytes in vitro

Transzonal projections (TZPs) that maintain bidirectional communication between oocytes and granulosa cells or cumulus cells are important structures for oocyte growth. However, whether TZPs develop between TZP-free oocytes and granulosa cells, and whether reestablished TZPs support oocyte growth, is unknown. We first examined changes in TZPs after denudation of bovine oocytes collected from early antral follicles (0.5–0.7 mm). Twenty-four hours after denudation, almost all the TZPs disappeared. We also examined the reestablishment of TZPs by coculturing TZP-free denuded oocytes (DOs) with mural granulosa cells (MGCs) collected from early antral follicles. In addition, to confirm if the reestablished TZPs were functional, the reconstructed complexes (DO+MGCs) were subjected to in vitro growth culture and found that the MGCs adhered to TZP-free DOs and TZPs were reestablished. During in vitro growth culture, DO+MGCs developed and formed antrum-like structures. After culture, the number of TZPs in DO+MGCs increased, and the oocytes grew fully and acquired meiotic competence. These results suggest that reestablished TZPs are able to support oocyte growth.     

Development of a monoclonal antibody for the detection of anti-canine CD20 chimeric antigen receptor expression on canine CD20 chimeric antigen receptor-transduced T cells

Chimeric antigen receptor (CAR) CAR-T cell therapy targeting CD20 can be a novel adoptive cell therapy for canine patients with B-cell malignancy. After injection of the CAR-T cells in vivo, monitoring circulating CAR-T cells is essential to prove in vivo persistence of CAR-T cells. In this study, we developed a novel monoclonal antibody against canine CD20 CAR, whose single-chain variable fragment was derived from the our previously reported anti-canine CD20 therapeutic antibody. Furthermore, we proved that this monoclonal antibody can detect therapeutic anti-canine CD20 chimeric antibody in the serum from healthy beagle dogs injected with the therapeutic antibody for safety study. This monoclonal antibody is a useful tool for monitoring both canine CD20-CAR-T cells and anti-canine CD20 therapeutic antibody for canine lymphoma.     

砷在铜藻中的亚细胞分布及细胞壁吸附作用研究

为揭示铜藻(Sargassum horneri)细胞壁在砷(As)生物吸附中的作用,以铜藻幼苗为研究对象,测定了不同As胁迫浓度(50、100、150、200、250μmol·L^-1)下细胞亚组分中的As含量变化,并对细胞壁的吸附动力学、细胞壁多糖As吸附效果以及细胞壁多糖傅里叶红外光谱进行了分析研究。结果表明：在低浓度(50μmol·L^-1)As胁迫时,50.0%的As分布于细胞壁,明显高于其他组分;而在高浓度(≥150μmol·L^-1)As胁迫时,As优势分布由细胞壁转向细胞可溶组分,细胞壁中的As比例下降至36.4%～37.3%。随着As胁迫浓度升高,细胞壁As含量呈上升趋势,当As胁迫浓度超过100μmol·L^-1时,细胞壁As含量上升趋于平缓。细胞壁吸附动力学实验表明酯化改性和甲基化改性可降低细胞壁As吸附的平衡容量,其降幅分别为68.5%和42.9%,同时细胞壁多糖红外光谱分析结果也表明羟基、氨基、羧基、硫酸盐基等官能团为细胞壁As吸附提供了结合位点。细胞壁多糖体外吸附实验表明随着多糖浓...