Direct evidence for the role of haem doming as the primary event in the cooperative transition of haemoglobin

The study of cooperative ligand binding among the four subunits of haemoglobin has played a central role in the understanding of allosteric transitions in a large number of enzymes. Haem iron out-of-plane motion has been suggested to be the trigger for the cooperative transition of haemoglobin. To function as a trigger in a dynamic sense, haem-iron doming must be the first conformational change to occur following ligand dissociation. Here we present the first direct demonstration that haem-iron doming occurs on the same time scale as the breaking of the iron-ligand bond, thus establishing haem-iron doming as the primary event which lead to the R-->T transition in haemoglobin.     

Use of a multileaf collimator for the production of intensity-modulated beams

The continuous release of nitric oxide (NO) from the constitutive, endothelial isoform of nitric oxide synthase (e-NOS) serves mainly to keep the vasculature in a continuous state of active vasodilation. Although it has been suggested that NO production from e-NOS might also be affected by hemorrhagic shock (HS), this relationship is still controversial. Therefore, the roles of NO in the pathophysiology in hemorrhagic shock were reviewed. According to the previous reports, NO might play an important role in the pathophysioliogy of HS. In the early phase of HS, it may be possible that NO delivered from e-NOS serves a cytoprotective function in preventing shock-induced organ injury. This opinion suggests that endothelial NO production has a significant modulatory effect on vascular tone during hemorrhage, and that inhibition of NO production permits greater vasconstrictor influences leading to organ injury. NO production in the late phases of HS has an adverse effect on survival rate in the HS model. Moreover, the findings from an animal study of prolonged periods of HS suggest that excessive NO formation, including those produced from i-NOS, induces vascular hypoactivity and they have suggested that NOS inhibitors may improve the therapeutic outcome for patients suffering from HS. Therefore, it may be suggested that NO might play a biphasic role, cytoprotective during the early phase and cytotoxic late in HS.     

Mechanism of Rab geranylgeranylation: formation of the catalytic ternary complex

Rab proteins are geranylgeranylated on one or two C-terminal cysteines by Rab geranylgeranyl transferase (RabGGTase). The reaction is dependent on a Rab-binding protein, termed Rab escort protein (REP). Here, we studied the role of REP in the geranylgeranylation reaction. We first characterized the interaction between REP and ungeranylgeranylated Rab using analytical ultracentrifugation and a fluorescence-based assay. We measured an equilibrium dissociation constant of 0.2 microM for the formation of a 1:1 REP-Rab complex and showed that this interaction relies mostly on ionic bonds and does not involve the two C-terminal cysteine residues. Second, we show that REP is required for recognition of Rab by RabGGTase and therefore that the REP-Rab complex is the true substrate for RabGGTase. Third, we show that free REP inhibits the geranylgeranylation reaction, suggesting that the complex is recognized by RabGGTase primarily via a REP-binding site. Our data suggest a model whereby REP behaves kinetically as an essential activator of the reaction.     

Effects of growth hormone, insulin-like growth factor-I, and cortisol on periparturient antibody response profiles of dairy cattle

The objectives of this study were to determine hormone and antibody response profiles from the prepartum period to peak lactation, and evaluate potential immunomodulatory effects of the classic endocrine hormones, growth hormone (GH), insulin-like growth factor-I (IGF-I) and cortisol. Specifically, 33 Holstein cows were immunized with ovalbumin (OVA) and Escherichia coli J5 at weeks -8 and -3 prior to parturition. At parturition (week 0), cows received an additional immunization of OVA. Blood was collected at weeks -8, -3, 0, 3 and 6 relative to parturition and various samples were used to determine plasma hormone concentration, serum immunoglobulin (Ig), and specific antibody response to OVA and E. coli. Colostrum and milk samples were also collected post-parturition to monitor local immunoglobulin and antibody responses. Results indicated that not all periparturient cows exhibited depressed immune response, and that antibody response to OVA could be used to partition cows into 3 groups recognizing animals with sustained measurable antibody response before and after parturition (Group 1), animals which responded poorly to immunization at parturition (Group 2), and animals which did not respond to immunizations at week -3 or parturition (Group 3). Cows with the highest antibody response to OVA (Group 1) also tended (P < or = 0.10) to have the highest response to E. coli J5 at parturition and had the lowest incidence of disease, particularly mastitis. Antibody response to OVA measured in milk tended to be higher in Group 1 cows, particularly at week 0 (P < or = 0.06) compared to cows of Group 3. IGF-I was higher (P < or = 0.05) in cows of Group 1 than Group 3 at peak lactation (week 6).     

Bacterial flora of processed broiler chicken skin after successive washings in mixtures of potassium hydroxide and lauric acid

Changes in the size of populations of different groups of bacteria composing the normal flora of processed broiler skin were examined after each of five consecutive washings in mixtures of potassium hydroxide (KOH) and lauric acid (LA). Portions of skin from commercially processed broiler carcasses were washed in distilled water (control) or in mixtures of 0.25% KOH-0.5% LA or 0.5% KOH-1% LA by using a stomacher laboratory blender to agitate the skin in the solutions. After each wash, skin was transferred to fresh solutions, and washing was repeated to provide samples washed one to five times in each solution. Bacteria in rinsates of the washed skin were enumerated on plate count (PC) agar, Staphylococcus (STA) agar, Levine eosin methylene blue (EMB) agar, lactic acid bacteria (LAB) agar, and Perfringens (PER) agar with TSC supplement. Selected isolates recovered on each medium were identified. Overall, no significant differences were observed in numbers of bacteria recovered on PC, STA, or EMB agars from skin after repeated washing in water, but there were significant reductions in the number of bacteria recovered on LAB and PER agars. Repeated washing of skin in 0.25% KOH-0.5% LA or 0.5% KOH-1% LA generally produced significant reductions in the number of bacteria recovered on all media. Furthermore, no bacteria were recovered on PER agar from skin washed five times in 0.25% KOH-0.5% LA. Likewise, no bacteria were recovered on EMB or LAB agars from skin washed three or more times in 0.5% KOH-1% LA or on PER agar from skin washed four or five times in this solution. Staphylococcus spp. were identified as the skin isolates with the highest degree of resistance to the bactericidal activity of KOH-LA. Findings indicate that although bacteria may be continually shed from poultry skin after repeated washings, bactericidal surfactants can be used to remove and kill several types of bacteria found on the surface of the skin of processed broilers.     

Molecular and immunophenotypical characterization of a feline immunodeficiency virus (FIV)-associated lymphoma: a direct role for FIV in B-lymphocyte transformation?

We describe the histopathological, immunohistochemical, and molecular characterization of a lymphoma arising in a 7-year-old cat following experimental infection with feline immunodeficiency virus (FIV). The tumor was high grade and of B-cell lineage. The transformed cell had an immature phenotype (CD79a+, CD79b-, CD21-, immunoglobulin heavy and light chain negative), confirmed by antigen receptor gene analysis, which showed germ line configuration. Single-copy, clonally integrated FIV provirus was detected in tumor genomic DNA. FIV p24 antigen was not detected in tumor cells by immunostaining. This study provides the first evidence that the feline lentivirus may play a direct role in cell transformation under certain circumstances.