Glycosyl conjugates of biotinylated diaminopyridine applied for study of carbohydrate-to-carbohydrate interaction

Previous studies by us and others established that cell-cell adhesion is mediated by specific carbohydrate-to-carbohydrate interaction (CCI). Those previous studies were based on various biochemical and biophysical approaches, including the use of labeled glycosyl epitopes with fluorescent tag. However, these methods ideally require that the glycosyl epitope must be fixed to a solid phase molecule, preferably with multivalency. The purpose of the present study is to establish a CCI process using specific glycosyl residues conjugated to biotinylated diaminopyridine (BAP), and to observe: (i) clear occurrence of homotypic CCI between “Os Fr.B” having 5–6 GlcNAc termini, vs. absence of such homotypic CCI between “Os Fr.1” having 2 GlcNAc termini; (ii) occurrence of heterotypic CCI between GM3 ganglioside and Os Fr.B, vs. absence of such heterotypic CCI between GM3 and Os Fr.1. Interaction between Os Fr.B-BAP conjugate and Os Fr.B-ceramide mimetic (Os Fr.B-mCer) was demonstrated based on two experiments: (i) dose-dependent binding of Os Fr.B-BAP conjugate to polystyrene plates coated with Os Fr.B-mCer was observed in the presence of bivalent cation, a prerequisite for all CCI processes, and such binding was abolished by EDTA; (ii) binding between equal nanomolar Os Fr.B-BAP and Os Fr.B-mCer was inhibited by mM concentration Os Fr.B without conjugate, in dose-dependent manner. Thus, cell adhesion processes based on homotypic CCI between N-linked glycans having multiple GlcNAc termini, and heterotypic CCI between GM3 and such glycans, were clearly observed using BAP conjugates of glycosyl epitopes.     

The extraction of a tissue collagenase associated with ovulation in the rat

A method has been developed to assay collagenase in ovarian extracts in the presence of tissue inhibitors. Rat ovarian tissue is first extracted with Triton X-100 and then heated to 60 degrees C in 50 mM Tris buffer containing 100 mM CaCl2. This extract contains collagenase activity and putative inhibitor(s). The inhibitory activity is removed by reduction with dithiothreitol and alkylation with iodoacetamide. Collagenase is then activated with aminophenylmercuric acetate and assayed using 3H-acetylated collagen from which the telopeptides have been removed. Identification of this activity as collagenase was performed by using the metalloprotease inhibitors EDTA and o-phenanthroline and by demonstration of the typical collagen cleavage fragments on sodium dodecyl sulfate-gel electrophoresis. To investigate the changes in collagenase activity associated with ovulation, immature rats received 20 IU of pregnant mare's serum gonadotropin and 52 h later 10 IU of human chorionic gonadotropin (hCG). After hCG administration, ovaries were removed at intervals from 0 to 20 h. Collagenase activity rose from 4.9 +/- 1.4% digestion of the 3H-collagen at 0 time to a maximum of 24.7 +/- 1.5% digestion at 8 h after hCG and remained high at 12 h (time of ovulation) and up to 20 h (18.7 +/- 1.9% and 16.1 +/- 1.6% digestion, respectively). These findings support a role of collagenase in the rupture of the follicle and they suggest a further role for this enzyme in the events following ovulation.     

Type I Antifreeze Proteins Enhance Ice Nucleation above Certain Concentrations

In this study, we examined the effects that antifreeze proteins have on the supercooling and ice-nucleating abilities of aqueous solutions. Very little information on such nucleation currently exists. Using an automated lag time apparatus and a new analysis, we show several dilution series of Type I antifreeze proteins. Our results indicate that, above a concentration of ∼8 mg/ml, ice nucleation is enhanced rather than hindered. We discuss this unexpected result and present a new hypothesis outlining three components of polar fish blood that we believe affect its solution properties in certain situations.     

Contractile properties of reinnervated skeletal muscle and their relation to the degree of nerve damage

Experiments with 22 rats have shown that the anterior tibial muscle in the stage of incomplete reinnervation is marked by decreased force and retardation of the semi-relaxation of an isometric contraction. In completely reinnervated muscles, the changes in the contractility are determined by the degree of nerve damage. The group of animals with the sciatic nerve injury demonstrated the contractility characteristic of a slower muscle, in contrast to the group with the fibular nerve damage.     

Competitive interactions between corticosterone and 11-dehydrocorticosterone in binding to receptors in fetal mouse tissues

The binding of ³H-corticosterone and ³H-11-dehydrocorticosterone to receptors in cytosol and nucleus was examined in fetal mouse brain and placenta using Sephadex gel filtration or charcoal to separate bound and unbound steroid. In the cytosol, competitive displacement of each steroid by the other was observed. The binding was unaffected by RNase, DNase, dithiothreitol or N-ethyl maleimide but was diminished by Pronase. Nuclei were isolated by hypotonic shock using dilute MgCl₂ and the steroid receptor-complexes of both steroids were obtained from the nuclear sap. Receptor-complexes of both steroids were observed in brain and placental tissues. Competitive displacement of each steroid by the other was also observed in nuclear binding. Both 11-dehydrocorticosterone and 11-deoxycorticosterone bound to a chromatin fraction as did the hormone corticosterone. Identity of the steroids was established by using chromatography and co-crystallization techniques. This work raises the possibility that in the fetal mouse, 11-dehydrocorticosterone, previously considered biologically inactive and an abundant metabolite in fetal mouse tissues, may in fact play a more positive role in regulation.     

Using diffusion distances for flexible molecular shape comparison

Background  

Many molecules are flexible and undergo significant shape deformation as part of their function, and yet most existing molecular shape comparison (MSC) methods treat them as rigid bodies, which may lead to incorrect shape recognition.     

Effect of interlok, remantadine and biotop on normal monocyte functional activity and in acute viral hepatitis

The influence of interlok, biotop and remantadine on the functional activity of monocytes was determined with the use of the NBT test permitting the quantitative evaluation of this characteristic. In this study interlok was found to give promising results in the treatment of acute viral hepatitis, due both to its antiviral action and to the stimulation of the functional activity of monocytes. biotop proved to render a sharply pronounced stimulating effect on the functional activity of monocytes, thus enhancing antiviral resistance. Remantadine suppressed the functional activity of monocytes and increased their lesion by the virus, thus creating favorable conditions for virus persistence in monocytes.     

Monoclonal antibodies to antigens found on the neutrophils, activated T-lymphocytes and blast cells in human acute leukemia

Two hybridomas secreting mouse cytotoxic monoclonal antibodies to in vivo and in vitro activated human T-lymphocyte and neutrophil surface membrane antigenic determinants have been produced. One of these monoclonal antibodies (Ta/H-2) appeared to be also specifically reactive to blast cells in the majority of non-T-non-B and T acute lymphoblastic leukemia patients.     

Factors associated with whole carcass condemnation rates in provincially-inspected abattoirs in Ontario 2001-2007: implications for food animal syndromic surveillance

Background  

Ontario provincial abattoirs have the potential to be important sources of syndromic surveillance data for emerging diseases of concern to animal health, public health and food safety. The objectives of this study were to: (1) describe provincially inspected abattoirs processing cattle in Ontario in terms of the number of abattoirs, the number of weeks abattoirs process cattle, geographical distribution, types of whole carcass condemnations reported, and the distance animals are shipped for slaughter; and (2) identify various seasonal, secular, disease and non-disease factors that might bias the results of quantitative methods, such as cluster detection methods, used for food animal syndromic surveillance.     

Estimation of transport distance of fine particulate organic matter in relation to channel morphology in tailwaters of the Lake Biwa and reservoir dams

Knowledge on transport and deposition of fine particulate organic matter (FPOM) from reservoir dams is increasingly required for habitat management and restoration of dam tailwater ecosystems. Variations in the transport distance of FPOM, however, have never been studied well, particularly in relation to channel morphology, due to channel size restrictions of artificial tracers such as corn pollen when applied to larger river channels. This study aims to show the relations between FPOM retention efficiency and channel morphology in dam tailwaters using lentic plankters as tracers. We estimated the mean transport distance, S _p, by calculating downstream reduction ratios of lentic tracer plankters and calculated the deposition velocity, v _dep. Suspended FPOM samples were collected in tailwaters of two river channels below reservoir dams and two artificial canals below Lake Biwa in the Yodo River system. The longest S _p (19.2 km) and the shortest one (2.2 km) were recorded in the deep canal and shallow canal, respectively, showing a positive correlation with channel hydraulic radius. The values of v _dep were 4.7–6.4 times higher in river channels than in artificial canals. These results indicate that increasing complexity of bed morphology can minimize S _p, whereas bed degradation and armored bed materials may lead to increased S _p. Advantages of lentic plankters as tracers for estimating distance ranges of reservoir dam impact on river ecosystems are also discussed.