Physiological responses of reared sea bream (Sparus aurata Linnaeus, 1758) to an Amyloodinium ocellatum outbreak

Amyloodiniosis represents a major bottleneck for semi-intensive aquaculture production in Southern Europe, causing extremely high mortalities. Amyloodinium ocellatum is a parasitic dinoflagellate that can infest almost all fish, crustacean and bivalves that live within its ecological range. Fish mortalities are usually attributed to anoxia, associated with serious gill hyperplasia, inflammation, haemorrhage and necrosis in heavy infestations; or with osmoregulatory impairment and secondary microbial infections due to severe epithelial damage in mild infestation. However, physiological information about the host responses to A.ocellatum infestation is scarce. In this work, we analysed the proteome of gilthead sea bream (Sparus aurata) plasma and relate it with haematological and immunological indicators, in order to enlighten the different physiological responses when exposed to an A.ocellatum outbreak. Using 2D-DIGE, immunological and haematological analysis and in response to the A.ocellatum contamination we have identified several proteins associated with acute-phase response, inflammation, lipid transport, homoeostasis, and osmoregulation, wound healing, neoplasia and iron transport. Overall, this preliminary study revealed that amyloodiniosis affects some fish functional pathways as revealed by the changes in the plasma proteome of S. aurata, and that the innate immunological system is not activated in the presence of the parasite.DIVERSIAQUA, Portugal [MAR2020]Fundacao para a Ciencia e Tecnologia [SFRH/BD/118601/2016]info:eu-repo/semantics/publishedVersio

Temperature controls on aquatic bacterial production and community dynamics in arctic lakes and streams

The impact of temperature on bacterial activity and community composition was investigated in arctic lakes and streams in northern Alaska. Aquatic bacterial communities incubated at different temperatures had different rates of production, as measured by 14 C-leucine uptake, indicating that populations within the communities had different temperature optima. Samples from Toolik Lake inlet and outlet were collected at water temperatures of 14.2°C and 15.9°C, respectively, and subsamples incubated at temperatures ranging from 6°C to 20°C. After 5 days, productivity rates varied from 0.5 to ∼13.7 µg C l −1 day −1 and two distinct activity optima appeared at 12°C and 20°C. At these optima, activity was 2- to 11-fold higher than at other incubation temperatures. The presence of two temperature optima indicates psychrophilic and psychrotolerant bacteria dominate under different conditions. Community fingerprinting via denaturant gradient gel electrophoresis (DGGE) of 16S rRNA genes showed strong shifts in the composition of communities driven more by temperature than by differences in dissolved organic matter source; e.g. four and seven unique operational taxonomic units (OTUs) were found only at 2°C and 25°C, respectively, and not found at other incubation temperatures after 5 days. The impact of temperature on bacteria is complex, influencing both bacterial productivity and community composition. Path analysis of measurements of 24 streams and lakes sampled across a catchment 12 times in 4 years indicates variable timing and strength of correlation between temperature and bacterial production, possibly due to bacterial community differences between sites. As indicated by both field and laboratory experiments, shifts in dominant community members can occur on ecologically relevant time scales (days), and have important implications for understanding the relationship of bacterial diversity and function.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/79247/1/j.1462-2920.2010.02176.x.pd

Dramatic Repositioning of c-Myb to Different Promoters during the Cell Cycle Observed by Combining Cell Sorting with Chromatin Immunoprecipitation

The c-Myb transcription factor is a critical regulator of proliferation and stem cell differentiation, and mutated alleles of c-Myb are oncogenic, but little is known about changes in c-Myb activity during the cell cycle. To map the association of c-Myb with specific target genes during the cell cycle, we developed a novel Fix-Sort-ChIP approach, in which asynchronously growing cells were fixed with formaldehyde, stained with Hoechst 33342 and separated into different cell cycle fractions by flow sorting, then processed for chromatin immunoprecipitation (ChIP) assays. We found that c-Myb actively repositions, binding to some genes only in specific cell cycle phases. In addition, the specificity of c-Myb is dramatically different in small subpopulations of cells, for example cells in the G2/M phase of the cell cycle, than in the bulk population. The repositioning of c-Myb during the cell cycle is not due to changes in its expression and also occurs with ectopically expressed, epitope-tagged versions of c-Myb. The repositioning occurs in established cell lines, in primary human CD34+ hematopoietic progenitors and in primary human acute myeloid leukemia cells. The combination of fixation, sorting and ChIP analysis sheds new light on the dynamic nature of gene regulation during the cell cycle and provides a new type of tool for the analysis of gene regulation in small subsets of cells, such as cells in a specific phase of the cell cycle

Antioxidant effects of insulin-like growth factor-I (IGF-I) in rats with advanced liver cirrhosis

BACKGROUND: The exogenous administration of Insulin-like Growth Factor-I (IGF-I) induces hepatoprotective and antifibrogenic actions in experimental liver cirrhosis. To better understand the possible pathways behind the beneficial effect of IGF-I, the aim of this work was to investigate severe parameters involved in oxidative damage in hepatic tissue from cirrhotic animals treated with IGF-I (2 μg. 100 g(-1). day(-1)). Iron and copper play an important role in oxidative mechanisms, producing the deleterious hydroxyl radical (*OH) that peroxides lipid membranes and damages DNA. Myeloperoxidase (MPO) and nitric oxide (NO) are known sources of free radicals and induce reduction of ferritin-Fe(3+ )into free Fe(2+), contributing to oxidative damage. METHODS: Liver cirrhosis was induced by CCl(4 )inhalation in Wistar male rats for 30 weeks. Healthy controls were studied in parallel (n = 10). Fe and Cu were assessed by atomic absoption spectrometry and iron content was also evaluated by Perls' staining. MPO was measured by ELISA and transferrin and ferritin by immunoturbidimetry. iNOS expression was studied by immuno-histochemistry. RESULTS: Liver cirrhosis was histologically proven and ascites was observed in all cirrhotic rats. Compared to controls untreated cirrhotic rats showed increased hepatic levels of iron, ferritin, transferrin (p < 0.01), copper, MPO and iNOS expression (p < 0.01). However, IGF-treatment induced a significant reduction of all these parameters (p < 0.05). CONCLUSION: the hepatoprotective and antifibrogenic effects of IGF-I in cirrhosis are associated with a diminution of the hepatic contents of several factors all of them involved in oxidative damage

A sensitive and specific ELISA detects methionine sulfoxide-containing apolipoprotein A-I in HDL*

Oxidized HDL has been proposed to play a key role in atherogenesis. A wide range of reactive intermediates oxidizes methionine residues to methionine sulfoxide (MetO) in apolipoprotein A-I (apoA-I), the major HDL protein. These reactive species include those produced by myeloperoxidase, an enzyme implicated in atherogenesis. The aim of the present study was to develop a sensitive and specific ELISA for detecting MetO residues in HDL. We therefore immunized mice with HPLC-purified human apoA-I containing MetO86 and MetO112 (termed apoA-I+32) to generate a monoclonal antibody termed MOA-I. An ELISA using MOA-I detected lipid-free apoA-I+32, apoA-I modified by 2e-oxidants (hydrogen peroxide, hypochlorous acid, peroxynitrite), and HDL oxidized by 1e- or 2e-oxidants and present in buffer or human plasma. Detection was concentration dependent, reproducible, and exhibited a linear response over a physiologically plausible range of concentrations of oxidized HDL. In contrast, MOA-I failed to recognize native apoA-I, native apoA-II, apoA-I modified by hydroxyl radical or metal ions, or LDL and methionine-containing proteins other than apoA-I modified by 2e-oxidants. Because the ELISA we have developed specifically detects apoA-I containing MetO in HDL and plasma, it should provide a useful tool for investigating the relationship between oxidized HDL and coronary artery disease

Stem cell-derived erythrocytes as upcoming players in blood transfusion

BACKGROUND: Blood transfusion is current standard-of-care for genetic forms of anemia that would be otherwise lethal and allows implementation of aggressive cytotoxic/surgical therapies developed for numerous types of cancer. In developed countries the blood supply is adequate and sporadically even in excess. However, difficulties exist in finding blood with rare phenotypes to treat alloimmunized patients and the progressive ageing of the human population predicts that blood will become scarce by 2050. These considerations establish the need for the development of techniques to generate cultured red blood cell (cRBCs) as transfusion products. MATERIALS AND METHODS: Recent progress in cell culture techniques is revolutionizing organ replacement therapies. Two new disciplines, cell therapy and tissue engineering, have been developed to generate in vitro therapeutic products for a variety of applications ranging from skin grafts to organ-function repairs. It is currently believed that these advances will eventually allow ex-vivo production of various cell types in numbers so great that, in the case of red cells, would be clinically adequate for transfusion. RESULTS: Proof-of-principle in animal models indicate that cRBCs generated from murine embryonic stem cells protect mice from lethal anemia. Conditions to generate small amounts of clinical grade cRBCs have been established and the first-in-man administration of autologous cRBCs perfomed. The results of this trial indicate that cRBCs survive in vivo at least as long as their natural counterpart. DISCUSSION: These ground-breaking reports have raised great excitement for clinical evaluation of cRBCs for transfusion. However, skepticism still persist that production of cRBCs in numbers sufficient for transfusion will ever be possible. This paper will discuss diagnostic and clinical goals pursuable with numbers of cRBCs that may be generated with current technology. CONCLUSION: We are confident that development of relevant clinical goals achievable with current technologies will not only improve clinical care in transfusion medicine but will also foster studies to overcome scientific and technical barriers that render transfusion with cRBCs of the general population impractical today