Alteration of the glutamate and GABA transporters in the hippocampus of the Niemann-Pick disease, type C mouse using proteomic analysis

Niemann-Pick disease type C (NPC) is a fatal autosomal recessive cholesterol disorder characterized by severe progressive neurodegeneration. To unveil the mechanism of neurodegeneration, proteomic and morphological approaches were applied to the hippocampus in NPC -/- mouse. Two-DE was utilized to resolve the hippocampal protein expression profiles of 4- and 8-week-old NPC +/+ and -/- mice. Differentially expressed protein spots were identified by MALDI-TOF MS and database searching. At 4 weeks of age, there was no significant difference in protein profiles between NPC +/+ and -/- mice. However, at the age of 8 weeks, NPC +/+ and -/- mice showed marked difference in protein expressions. Among these, glutamate receptor 2 precursor was identified. The immunohistochemical study on neurotransporters showed that glial GABA transporter (GAT-3) increased in both 4- and 8-week-old NPC -/- mouse and glutamic acid decarboxylase (GAD-6) increased in 8-week-old NPC -/- mouse. Glial glutamate transporter, excitatory amino acids carrier-1 (EAAC1), decreased in 8-week-old NPC -/- mouse. In conclusion, our data may provide insight into the understanding of the basic mechanism through perturbation of protein networks and neurotransporter systems in a single gene knockout model of NPC disease.     

Biosynthetic origin of [R-(Z)]-4-amino-3-chloro-2-pentenedioic acid in Streptomyces viridogenes

The biosynthesis of the chlorinated amino acid [R-(Z)]-4-amino-3-chloro-2-pentenedioic acid (ACPA) was investigated. Feeding studies with Streptomyces viridogenes were conducted in resting cells. Substantial incorporation from [(15)N]- and [(13)C]-enriched glutamate and proline indicated that the biosynthetic origin of ACPA is one of these amino acids. Experiments with deuterated glutamate and proline imply that chlorination does not occur via a radical mechanism, but rather suggest that a FADH(2)-dependent halogenase is involved.     

The Effects of Gibberellin4+7 on the Vase Life and Flower Quality of Alstroemeria Cut Flowers

Leaf yellowing is a major problem in Alstroemeria and absence of leaf senescence symptoms is an important quality attribute. Two Alstroemeria cultivars ‘Yellow King’ and ‘Marina’ were sourced from a commercial farm and harvested when sepals began to reflex. Stems were re-cut under water and kept in vase solutions of gibberellin A₄₊₇ (0, 2.5, 5.0, 7.5, 10.0, 12.5 or 15.0 mg l⁻¹ [Provide^r]). Treatments and cultivars were combined in a factorial fashion and arranged in a completely randomised design. Application of GA₄₊₇ in the holding solution at 2.5–10.0 mg l⁻¹ significantly delayed the onset of leaf senescence by around 7 days and significantly increased days to 50% petal fall by ca. 2 days. Additionally, these GA₄₊₇ concentrations resulted in higher retention of leaf nitrogen, leaf chlorophyll and also increased leaf water content, while reducing leaf dry weight, all relative to untreated controls. Cultivar ‘Yellow King’ had significantly longer vase life and a better retention of leaf quality than ‘Marina’. Our results suggest that a concentration of 10 mg l⁻¹ GA₄₊₇ can be used to prolong vase life, delay leaf senescence and enhance post-harvest quality of Alstroemeria cut flowers during their transport to market.     

Endothelin antagonism prevents early EGFR transactivation but not increased matrix metalloproteinase activity in diabetes

Although past studies have demonstrated decreased renal matrix metalloproteinase (MMP) activity in type 1 diabetes and in mesangial cells grown under high glucose conditions, renal MMP expression and activity in type 2 diabetes and the regulation of MMPs by profibrotic factors involved in diabetic renal complications such as endothelin-1 (ET-1) remained unknown. The renal expression and activity of MMPs in type 2 diabetic Goto-Kakizaki (GK) rats treated with vehicle or ET(A) receptor selective antagonist ABT-627 for 4 wk were assessed by gelatin zymography, fluorogenic gelatinase assay, and immunoblotting. In addition, expression and phosphorylation of epidermal growth factor receptor (EGFR) and connective tissue growth factor were evaluated by immunoblotting. Renal sections stained with Masson trichrome were used to investigate kidney structure. MMP-2 activity and protein levels were significantly increased in both cortical and medullary regions in the GK rats. Membrane-bound MMP (MT1-MMP), MMP-9, and fibronectin levels were also increased, and ABT-627 treatment did not have an effect on MMP activity and expression. Histological analysis of kidneys did not reveal any structural changes. Phosphorylation of EGFR was significantly increased in the diabetic animals, and ABT-627 treatment prevented this increase, suggesting ET-1-mediated transactivation of EGFR. These results suggest that there is early upregulation of renal MMPs in the absence of any kidney damage. Although the ET(A) receptor subtype is not involved in the early activation of MMPs in type 2 diabetes, ET-1 contributes to transactivation of growth-promoting and profibrotic EGFR.     

Shear stress analysis of mammalian cell suspensions for prediction of industrial centrifugation and its verification

This article describes the use of ultra scale-down studies requiring milliliter quantities of process material to study the clarification of mammalian cell culture broths using industrial-scale continuous centrifuges during the manufacture of a monoclonal antibody for therapeutic use. Samples were pretreated in a small high-speed rotating-disc device in order to mimic the effect on the cells of shear stresses in the feed zone of the industrial scale centrifuges. The use of this feed mimic was shown to predict a reduction of the clarification efficiency by significantly reducing the particle size distribution of the mammalian cells. The combined use of the rotating-disc device and a laboratory-scale test tube centrifuge successfully predicted the separation characteristics of industrial-scale, disc stack centrifuges operating with different feed zones. A 70% reduction in flow rate in the industrial-scale centrifuge was shown to arise from shear effects. A predicted 2.5-fold increase in throughput for the same clarification performance, achieved by the change to a centrifuge using a feed zone designed to give gentler acceleration of the bioprocess fluid, was also verified at large-scale.     

Sex differences in the rate of fatigue development and recovery

Background

Many musculoskeltal injuries in the workplace have been attributed to the repetitive loading of muscle and soft tissues. It is not disputed that muscular fatigue is a risk factor for musculoskeltal injury, however the disparity between gender with respect to muscular fatigability and rate of recovery is not well understood. Current health and safety guidelines do not account for sex differences in fatiguability and may be predisposing one gender to greater risk. The purpose of this study was to quantify the sex differences in fatigue development and recovery rate of lower and upper body musculature after repeated bouts of sustained isometric contractions.

Methods

Twenty-seven healthy males (n = 12) and females (n = 15) underwent bilateral localized fatigue of either the knee extensors (male: n = 8; female: n = 8), elbow flexors (male: n = 8; female: n = 10), or both muscle groups. The fatigue protocol consisted of ten 30-second sub-maximal isometric contractions. The changes in maximum voluntary contraction (MVC), electrically evoked twitches, and motor unit activation (MUA) were assessed along with the ability to control the sustained contractions (SLP) during the fatigue protocol using a mixed four-factor repeated measures ANOVA (gender × side × muscle × time) design with significance set at p < 0.05.

Results

There was a significant loss of MVC, MUA, and evoked twitch amplitude from pre- to post-fatigue in both the arms and legs. Males had greater relative loss of isometric force, a higher rate of fatigue development, and were less capable of maintaining the fatiguing contractions in the legs when compared to the females.

Conclusion

The nature of the induced fatigue was a combination of central and peripheral fatigue that did not fully recover over a 45-minute period. The results appear to reflect sex differences that are peripheral, and partially support the muscle mass hypothesis for explaining differences in muscular fatigue.

     

Nonpeptide GPIIB/IIIA receptor antagonists. Part 21: C-6 flexibility and amide bond orientation are important factors in determining the affinity of compounds for activated or resting platelet receptors

Compound affinity for activated and resting GPIIb/IIIa receptors may differ, and comparison of those differences determines selectivity. Structural features that influence selectivity are discussed.     

Exposure and Exclusion: Disenfranchised Biological Citizenship among the First-Generation Korean Americans

Based on fieldwork with a highly uninsured and underinsured Korean American population, this article maps how the current healthcare system in the United States disenfranchises those of marginal insurance status. The vulnerability of these disenfranchised biological citizens is multiplied through exposure to disproportional health risks compounded by exclusion from essential healthcare. The first-generation Korean Americans, who commonly work in small businesses, face the double burden of increased health risks from long, stress-laden work hours and lack of access to healthcare due to the prohibitive costs of health insurance for small business owners. Even as their health needs become critical, their insurance status and costly medical bills discourage them from visiting healthcare institutions, leaving Korean Americans outside the ??political economy of hope?? (Good, Cult Med Psychiatry 52:61?C69, 2001). Through an ethnographic examination of the daily practice of doing-without-health among a marginalized sub-group in American society, this paper articulates how disenfranchised biological citizenship goes beyond creating institutional barriers to healthcare to shaping subjectivities of the disenfranchised.