Adiponectin-Mediated Analgesia and Anti-Inflammatory Effects in Rat

The adipose tissue-derived protein, adiponectin, has significant anti-inflammatory properties in a variety of disease conditions. Recent evidence that adiponectin and its receptors (AdipoR1 and AdipoR2) are expressed in central nervous system, suggests that it may also have a central modulatory role in pain and inflammation. This study set out to investigate the effects of exogenously applied recombinant adiponectin (via intrathecal and intraplantar routes; 10–5000 ng) on the development of peripheral inflammation (paw oedema) and pain hypersensitivity in the rat carrageenan model of inflammation. Expression of adiponectin, AdipoR1 and AdipoR2 mRNA and protein was characterised in dorsal spinal cord using real-time polymerase chain reaction (PCR) and Western blotting. AdipoR1 and AdipoR2 mRNA and protein were found to be constitutively expressed in dorsal spinal cord, but no change in mRNA expression levels was detected in response to carrageenan-induced inflammation. Adiponectin mRNA, but not protein, was detected in dorsal spinal cord, although levels were very low. Intrathecal administration of adiponectin, both pre- and 3 hours post-carrageenan, significantly attenuated thermal hyperalgesia and mechanical hypersensitivity. Intrathecal administration of adiponectin post-carrageenan also reduced peripheral inflammation. Intraplantar administration of adiponectin pre-carrageenan dose-dependently reduced thermal hyperalgesia but had no effect on mechanical hypersensitivity and peripheral inflammation. These results show that adiponectin functions both peripherally and centrally at the spinal cord level, likely through activation of AdipoRs to modulate pain and peripheral inflammation. These data suggest that adiponectin receptors may be a novel therapeutic target for pain modulation.     

A multi-faceted knowledge translation approach to support persons with stroke and cognitive impairment: evaluation protocol

Background

Patients with cognitive impairments following a stroke are often denied access to inpatient rehabilitation. The few patients with cognitive impairment admitted to rehabilitation generally receive services based on outdated impairment-reduction models, rather than recommended function-based approaches. Both reduced access to rehabilitation and the knowledge-to-practice gap stem from a reported lack of skills and knowledge regarding cognitive rehabilitation on the part of inpatient rehabilitation team members. To address these issues, a multi-faceted knowledge translation (KT) initiative will be implemented and evaluated. It will be targeted specifically at the inter-professional application of the cognitive orientation to daily occupational performance (CO-OP). CO-OP training combined with KT support is called CO-OP KT. The long-term objective of CO-OP KT is to optimize functional outcomes for individuals with stroke and cognitive impairments. Three research questions are posed:

1. 1.
   Is the implementation of CO-OP KT associated with a change in the proportion of patients with cognitive impairment following a stroke accepted to inpatient rehabilitation?
    
2. 2.
   Is the implementation of CO-OP KT associated with a change in rehabilitation clinicians’ practice, knowledge, and self-efficacy related to implementing the CO-OP approach, immediately following and 1 year later?
    
3. 3.
   Is CO-OP KT associated with changes in activity, participation, and self-efficacy to perform daily activities in patients with cognitive impairment following stroke at discharge from inpatient rehabilitation and at 1-, 3-, and 6-month follow-ups?
    

Methods/Design

Three interrelated studies will be conducted. Study 1 will be a quasi-experimental, interrupted time series design measuring monthly summaries of stroke unit level data. Study 2, which relates to changes in health care professional practice and self-efficacy, will be a single group pre-post evaluation design incorporating chart audits and a self-report survey. Study 3 will assess patient functional outcomes using a non-randomized design with historical controls. Assessments will occur during admission and discharge from rehabilitation and at 1, 3, and 6 months following discharge from rehabilitation.

Discussion

This project will advance knowledge about the degree to which the implementation of a supported KT initiative can sustainably change health system, knowledge, and patient outcomes.

     

Loss of Cell Cycle Checkpoint Control in Drosophila Rfc4 Mutants

Two alleles of the Drosophila melanogaster Rfc4 (DmRfc4) gene, which encodes subunit 4 of the replication factor C (RFC) complex, cause striking defects in mitotic chromosome cohesion and condensation. These mutations produce larval phenotypes consistent with a role in DNA replication but also result in mitotic chromosomal defects appearing either as premature chromosome condensation-like or precocious sister chromatid separation figures. Though the DmRFC4 protein localizes to all replicating nuclei, it is dispersed from chromatin in mitosis. Thus the mitotic defects appear not to be the result of a direct role for RFC4 in chromosome structure. We also show that the mitotic defects in these two DmRfc4 alleles are the result of aberrant checkpoint control in response to DNA replication inhibition or damage to chromosomes. Not all surveillance function is compromised in these mutants, as the kinetochore attachment checkpoint is operative. Intriguingly, metaphase delay is frequently observed with the more severe of the two alleles, indicating that subsequent chromosome segregation may be inhibited. This is the first demonstration that subunit 4 of RFC functions in checkpoint control in any organism, and our findings additionally emphasize the conserved nature of RFC's involvement in checkpoint control in multicellular eukaryotes.     

Endocytosed cation-independent mannose 6-phosphate receptor traffics via the endocytic recycling compartment en route to the trans-Golgi network and a subpopulation of late endosomes

Although the distribution of the cation-independent mannose 6-phosphate receptor (CI-MPR) has been well studied, its intracellular itinerary and trafficking kinetics remain uncertain. In this report, we describe the endocytic trafficking and steady-state localization of a chimeric form of the CI-MPR containing the ecto-domain of the bovine CI-MPR and the murine transmembrane and cytoplasmic domains expressed in a CHO cell line. Detailed confocal microscopy analysis revealed that internalized chimeric CI-MPR overlaps almost completely with the endogenous CI-MPR but only partially with individual markers for the trans-Golgi network or other endosomal compartments. After endocytosis, the chimeric receptor first enters sorting endosomes, and it then accumulates in the endocytic recycling compartment. A large fraction of the receptors return to the plasma membrane, but some are delivered to the trans-Golgi network and/or late endosomes. Over the course of an hour, the endocytosed receptors achieve their steady-state distribution. Importantly, the receptor does not start to colocalize with late endosomal markers until after it has passed through the endocytic recycling compartment. In CHO cells, only a small fraction of the receptor is ever detected in endosomes bearing substrates destined for lysosomes (kinetically defined late endosomes). These data demonstrate that CI-MPR takes a complex route that involves multiple sorting steps in both early and late endosomes.     

An Assessment of Routine Chromosomal Staining Procedures in Radioautography

Unlabeled human chromosome preparations were treated with commonly employed chromosome stains as follows: (I) they were stained, destained, coated with liquid emulsion, developed, fixed, and restained; (II) stained and coated directly; or (III) coated and then stained. Of the stains tested, the methylene blue-eosin type (Giemsa, MacNeal's, Wright's) was useful for application after coating, although a similar stain (eosin-Stevenel's blue) caused formation of a heavy precipitate in the emulsion when so used. None of these stains could be employed before coating, however, even though they were removed with acid alcohol prior to dipping, because they caused chemographic grain formation in the emulsion. Aceto-orcein and Feulgen could not be employed after coating because the procedures removed the emulsion from the slides. Safranin was also found to be ineffective for staining coated preparations due to chemical changes caused by the photographic processing. The only stain which did not cause chemography, and hence can be used before coating slides, is aceto-orcein. Since this stain fades during radioautographic processing and cannot be employed after coating, we recommend secondary use of one of the methylene blue-eosin type stains for revisualization of the chromosome spread.     

A model system for the study of food container leakage

A model system to study food container leakage was developed. The model system allows the independent investigation of the effect of physical factors such as vacuum and contents viscosity, and microbial factors on the leakage process. The design, construction and operation of the container leakage model system is described.     

Effect of pH,temperature, and growth conditions on the adhesion of a gliding bacterium and three nongliding bacteria to polystyrene

The effect of growth rate, growth phase, pH, and temperature on the permanent adhesion of a glidingFlexibacter sp. and three nongliding bacteria,Pseudomonas fluorescens, Enterobacter cloacae, andChromobacterium sp., to polystyrene substrata was investigated. The permanent adhesion of the flexibacter appeared to be related to growth, as levels of adhesion increased with increased growth rate in continuous culture and declined rapidly with death phase in batch culture. With the three nongliding bacteria, there was no relationship between growth rate and levels of permanent adhesion. The permanent adhesion of the nongliding bacteria was maximum between pH 5.5 and pH 7 and between 20 and 30°C, whereas the adhesion of the flexibacter progressively decreased with increasing temperature and pH. The effect of different nutrient conditions on the gliding motility of the flexibacter across agar was also investigated. Gliding motility was inhibited by increased nutrient concentration and was affected by carbon source. Inhibition appeared to be related to the accumulation of a viscous exopolymer. It is proposed that the differences in the permanent adhesion of the gliding and nongliding bacteria may be related to their adaptation to different ecological niches.     

The effect of physical and microbiological factors on food container leakage

The effect of physical and microbiological factors on food container leakage was investigated in a container leakage model system (CLMS). The leakage of Acineto-bacter calcoaceticus, Staphylococcus sp., Pseudomonas sp., Bacillus sp., a coryneform, Staph. aureus , and two biotest organisms ( Enterobacter cloacae NC1B 8151 and Ent. aerogenes MB31) was studied. The rate of bacterial leakage (log₁₀ cells/channel/s) was greater in the presence of a partial vacuum of 51–305 mm Hg than at atmospheric pressure. Fluid flow (ml) through leakage channels was increased by the application of vacuum. Leakage varied with vacuum, bacterial morphology, cell concentration, leakage channel size (0.78–120 μm²) and channel shape (straight or convoluted). The number of leaked cells was not proportional to vacuum or channel size. The effect of channel shape varied with bacterial species. Increased container medium viscosity decreased bacterial leakage. Fluid flow through leakage channels was generally reduced by the most viscous solution. Cells from biofilms and mono-layers of Ac. calcoaceticus or Staph. aureus attached to nylon (Hyfax) or stainless steel surfaces underwent leakage. Mixed bacterial populations had characteristic leakage rates against vacuum different from the leakage pattern of individual species in the population. The composition of the leaked population was different from the original inoculum. The results indicated that container leakage is a complex process involving a range of interdependent factors.     

A model system for the study of food container leakage

A model system to study food container leakage was developed. The model system allows the independent investigation of the effect of physical factors such as vacuum and contents viscosity, and microbial factors on the leakage process. The design, construction and operation of the container leakage model system is described.     

A comparative study of the effect of modification of the surface of human platelets on the receptors for aggregated immunoglobulins and for ristocetin-von Willebrand factor

The receptors for aggregated immunoglobulin G (IgG) (an Fc receptor) and for ristocetin-von Willebrand factor on human platelets were studied by means of various modifications of the platelet surface. The expression of these receptors was measured by the agglutination of platelets to ristocetin in the presence of von Willebrand factor, which is part of the factor VIII complex, and by the binding of aggregated IgG coupled to ³H-labelled diazobenzen. Treatment of platelets with chymotrypsin, trypsin, papain and pronase which removed protein and glycoprotein from the platelet under conditions where the release reaction was inhibited caused loss of the expression of the receptor for ristocetin-von Willebrand factor and an enhancement of that for aggregated IgG. Induction of membrane changes with ADP and of the release reaction with the ionophore A23187 abolished agglutination to ristocetin-von Willebrand factor but did not alter the receptor for aggregated IgG. Possible contributions of unspecific membrane changes, produced by protease treatment of platelets, to the modification of receptor expression were eliminated by the use of formaldehyde-treated platelets. Trypsin, papain and pronase destroyed the ability of these platelets to agglutinate to ristocetin-von Willebrand factor but produced no change in the binding of aggregated IgG. Therefore, the receptor for ristocetin-von Willebrand factor is truly sensitive to proteolysis while the Fc receptor is not, but is partially masked by protease-sensitive material.