Search PubMed⌕ Search

Biomedical subjects

A Wiseman

Publications and source records attributed to A Wiseman.

At least 19 recordsLinked to original sources
PubMed2006

Initial results of screening of nondiabetic organ donors for expression of islet autoantibodies.

CONTEXT: Type 1A diabetes is characterized by a long prodromal phase during which autoantibodies to islet antigens are present. Nevertheless, we lack data on the pancreatic pathology of subjects who are positive for islet autoantibodies (to islet autoantigens GAD65, insulin, and ICA512). OBJECTIVE: In this manuscript, we describe a novel strategy in obtaining pancreata and pancreatic lymph nodes from islet autoantibody-positive organ donors that involves careful coordination among the laboratory and the organ donor provider organization. DESIGN: We developed a rapid screening protocol for islet autoantibodies measurement of organ donors to allow identification of positive subjects before organ harvesting. In this way we were able to obtain pancreata and pancreatic lymph nodes from subjects with and without islet autoimmunity. SETTING: The organ donors used in this study were obtained from the general community. SUBJECTS: The population studied consisted of 112 organ donors (age range 1 month to 86 yr, mean age 39 yr). MAIN OUTCOME MEASURE: The main outcome measure of this study consisted of evaluating the pancreatic histology and identify T cells autoreactive for islet antigens in the pancreatic lymph nodes. RESULTS: To date we have identified three positive subjects and obtained the pancreas for histological evaluation from one of the autoantibody-positive donors who expressed ICA512 autoantibodies. Although this subject did not exhibit insulitis, lymphocytes derived from pancreatic lymph nodes reacted to the islet antigen phogrin. CONCLUSION: In summary, these results indicate that it is possible to screen organ donors in real time for antiislet antibodies, characterize pancreatic histology, and obtain viable T cells for immunological studies.

Adolescent↗
PubMed2005

Avoidance of oxidative-stress perturbation in yeast bioprocesses by proteomic and genomic biostrategies?

AIMS: Bioprocess oxidative stress caused by many reactive oxygen species (ROS) can lead to largely irreversible perturbation of yeast bioprocesses. These include the production of proteins derived from recombinant DNA yeast technology (aerobically grown Saccharomyces cerevisiae). These proteins include rennin, amyloglucosidases (glucamylases), interferons, interleukins, insulin, monoclonal antibodies, tissue plasminogen activators (t-PA), sexually transmitted disease antigens, and measles, mumps and rubella antigens, growth hormones, somatotropin, blood clotting factors VIII and XIII. In addition, there may be a demand for severe acute respiratory syndrome-coronavirus antigens, hepatitis A, B and C viral-selected antigens, HIV retroviral antigens, influenza antigens, trypanosomal antigens, and foot and mouth disease antigens. Prevention of oxidative stress has been achieved by application of antioxidant redox metalloenzymes such as superoxide dismutases (containing Cu/Zn cytosolic, Mn mitochondrial and Fe bacterial) glutathione peroxidases (and other Se-containing proteins and enzymes such as the thioredoxins), catalases (Fe-containing), cytochrome c peroxidases (Fe-containing), ceruloplasmins (Cu-containing), metallothionines (these cysteine thiol-rich proteins bind ions of cadmium and mercury) and tyrosinases(Cu-containing). METHODS AND RESULTS: ROS are generated inadvertently by single metal valency couples such as FeII/FeIII and by FeIII/FeV present in 2700 (including 57 human) isoforms in cytochromes P450 mixed-function oxidases (EC 1.14.14.1; O2 : mono-oxygenase NADPH/NADH requiring). In addition, mixed-metal couples such as valency unmatched forms in CuI/FeII and FeIII/MnIV can recycle electrons. Moreover, proteins/protein chaperone couples can recycle electrons, often where futile-recycling systems have been instigated. Furthermore, oxidized membrane phospholipids (R) can form ROOH (lipid hydroperoxides) and ROH (lipid alkoxides) that can generate ROS through Fenton chemistry (iron-catalysed) chain reactions. Utilization of chain-breaking antioxidants such as vitamin E (alpha-tocopherol) in the lipid phase and vitamin C (ascorbate) in the aqueous phase can terminate these ROS-producing reactions. CONCLUSIONS: The main significance of the study is that proteomic strategies of relief from bioprocess perturbation by ROS of yeast fermentations (used to manufacture proteins required in the food and therapeutic bioindustries) may become possible through addition of selected proteins (including metalloenzymes). The main impact of the study is that the utilization of genetically modified (GM) yeast produced by recombinant DNA technology genomic strategies could circumvent the bioprocessing problems that otherwise result from the bioprocess perturbations: this is as a result of oxidative stress caused by ROS, which is avoidable by deployment of appropriate antioxidants such as vitamins E, C and D (and antioxidant proteins and enzymes often of microbial origin via recombinant DNA technology).

Antioxidants↗
PubMed2004

Duration of immunity of a quadrivalent vaccine against respiratory diseases caused by BHV-1, PI3V, BVDV, and BRSV in experimentally infected calves.

Several laboratory studies assessed the duration of immunity of a quadrivalent vaccine (Rispoval 4, Pfizer Animal Health) against bovine respiratory diseases (BRD) caused by bovine herpes-virus type-1 (BHV-1), parainfluenza type-3 virus (PI3V), bovine viral-diarrhoea virus type 1 (BVDV), or bovine respiratory syncytial virus (BRSV). Calves between 7 weeks and 6 months of age were allocated to treatment and then were injected with two doses of either the vaccine or the placebo 3 weeks apart. Six to 12 months after the second injection, animals were challenged with BHV-1 (n=16), PI3V (n=31), BVDV (n=16), or BRSV (n=20) and the course of viral infection was monitored by serological, haematological (in the BVDV study only), clinical, and virological means for > or =2 weeks. Infection induced mild clinical signs of respiratory disease and elevated rectal temperature in both vaccinated and control animals and was followed by a dramatic rise in neutralising antibodies in all treatment groups. Titres reached higher levels in vaccinated calves than in control calves after challenge with BHV-1, BVDV, or BRSV. On day 3 after PI3V challenge, virus shedding was reduced from 3.64 log10TCID50 in control animals to 2.59 log10TCID50 in vaccinated animals. On days 6 and 8 after BRSV challenge, there were fewer vaccinated animals (n=2/10 and 0/10, respectively) shedding the virus than control animals (n=8/10 and 3/10, respectively). Moreover, after challenge, the mean duration of virus shedding was reduced from 3.8 days in control animals to 1 day in vaccinated animals in the BVDV study and from 3.4 days in control animals to 1.2 days in vaccinated animals in the BRSV study. The duration of immunity of >or =6 months for PI3V, BHV-1 and BVDV, and 12 months for BRSV, after vaccination with Rispoval 4, was associated mainly with enhanced post-challenge antibody response to all four viruses and reduction of the amount or duration of virus shedding or both.

Animals↗
PubMed2003

Novel cytochromes P450 applications arising from the directed-evolution of recombinant micro-organisms.

AIMS: Directed (forced) evolution of cytochromes P450 (overall 2700 CYP isoforms in non-recombinant biota) is a method that has been investigated in yeasts (and other micro-organisms) by aerobically growing brewers' yeast Saccharomyces cerevisiae in very high glucose (20%) media. METHODS AND RESULTS: Mitochondrial repression subverts cytochrome oxidase biosynthesis into manifest cytochromes P450 accumulation in brewers' yeast. A similar phenomenon is observed with the acridine-induced petit mutant. Cytochromes P450 EC 1.14.14.1 (and mimics) display a range of redox iron-mediated bioconversions in food processing, with mixed function oxidase (O2:mono-oxygenase) intervention results. Unfortunately these enzymes generate reactive oxygen species (ROS) through redox electron recycling, whilst isoform CYP 1A1 can activate precarcinogens such as benzo(a)pyrene to the ultimate (proximate) carcinogen that binds to nuclear DNA. CONCLUSIONS: In conclusion, another 5000 CYP isoforms, for example, might be identified in micro-organisms and many more made to order through recombinant DNA technology and utilized both in vitro and in vivo for aimed bioconversions in industry and in the environment, as part of the impact of greener-approach supporting strategies to minimize global pollution.

Bacteria↗
PubMed2002

Protection against oronasal challenge with virulent feline leukaemia virus lasts for at least 12 months following a primary course of immunisation with Leukocell 2 vaccine.

The duration of immunity provided by a feline leukemia virus (FeLV) vaccine, Leukocell 2, was determined. Kittens were vaccinated when 9 and 12 weeks of age and were challenged 12 months later with FeLV-A/Glasgow-1. An oronasal challenge protocol without corticosteroid enhancement was developed in order to induce a persistent viraemia in a high proportion of adult cats. Fourteen of 18 (80%) of the vaccinated cats challenged in this way remained non-viraemic while 9/15 (60%) of age-matched controls became persistently infected, a preventable fraction of 63%. This difference was statistically significant (P=0.038). For comparison, 10 of 12 (83%) 15-17-week-old kittens challenged in the same way became persistently infected, confirming the relative resistance of adult animals to FeLV. Tests for virus neutralising and anti-feline oncornavirus-associated cell membrane antigen (FOCMA) antibodies suggested that the former were more important than the latter in protection. Thus, Leukocell 2 protected a significant proportion of cats from FeLV challenge 1 year after primary vaccination as kittens.

Animals↗
PubMed2000

CD4 T cell-mediated cardiac allograft rejection requires donor but not host MHC class II.

Numerous studies indicate that CD4 T cells are required for acute cardiac allograft rejection. However, the precise role for CD4 T cells in this response has remained ambiguous owing to the multipotential properties of this T-cell subpopulation. In the current study, we demonstrate the capacity of CD4 T cells to serve as direct effector cells of cardiac allograft rejection. We show that CD4 T cells are both necessary and sufficient for acute graft rejection, as indicated by adoptive transfer experiments in immune-deficient SCID and rag1(-/-) recipients. We have analyzed the contribution of direct (donor MHC class II restricted) and indirect (host MHC class II restricted) antigen recognition in CD4-mediated rejection. Acute CD4 T cell-mediated rejection required MHC class II expression by the allograft, indicating the importance of direct graft recognition. In contrast, reciprocal experiments indicate that CD4 T cells can acutely reject allogeneic cardiac allografts established in rag1(-/-) hosts that were also MHC class II deficient. This latter result indicates that indirect presentation of donor antigens by host MHC class II is not required for acute CD4-mediated rejection. Taken together, these results indicate that CD4 T cells can serve as effector cells for primary acute cardiac allograft rejection, predominantly via direct donor antigen recognition and independent of indirect reactivity.

Acute Disease↗
PubMed1999

Molecular modelling of lanosterol 14 alpha-demethylase (CYP51) from Saccharomyces cerevisiae via homology with CYP102, a unique bacterial cytochrome P450 isoform: quantitative structure-activity relationships (QSARs) within two related series of antifungal azole derivatives.

The construction of a three-dimensional molecular model of the fungal form of cytochrome P450 (CYP51) from Saccharomyces cerevisiae, based on homology with the haemoprotein domain of CYP102 from Bacillus megaterium (a unique bacterial P450 of known crystal structure) is described. It is found that the endogenous substrate, lanosterol, can readily occupy the putative active site of the CYP51 model such that the known mono-oxygenation reaction, leading to C14-demethylation of lanosterol, is the preferred route of metabolism for this particular substrate. Key amino acid contacts within the CYP51 active site appear to orientate lanosterol for oxidative attack at the C14-methyl group, and the position of the substrate relative to the haem moiety is consistent with the phenyl-iron complexation studies reported by Tuck et al. [J. Biol. Chem., 267, 13175-13179 (1992)]. Typical azole inhibitors, such as ketoconazole, are able to fit the putative active site of CYP51 by a combination of haem ligation, hydrogen bonding, pi-pi stacking and hydrophobic interactions within the enzyme's haem environment. The mode of action of azole antifungals, as described by the modelling studies, is supported by quantitative structure-activity relationship (QSAR) analyses on two groups of structurally related fungal inhibitors. Moreover, the results of molecular electrostatic isopotential (EIP) energy calculations are compatible with the proposed mode of binding between azole antifungal agents and the putative active site of CYP51, although membrane interactions may also have a role in the antifungal activity of azole derivatives.

Amino Acid Sequence↗