Community-based use of the larvivorous fish Poecilia reticulata to control the dengue vector Aedes aegypti in domestic water storage containers in rural Cambodia

A community-based study of the distribution of larvivorous fish, Poecilia reticulata (common name: guppy), in water storage containers for dengue control was undertaken in 14 villages and approximately 1,000 households in Cambodia. Community volunteers reared guppies and distributed them in water jars and tanks in households for which they were responsible. A nearby control area received no intervention. One year after project commencement, 56.9% of eligible containers contained guppies and there was a 79.0% reduction in Aedes infestation in the intervention community compared with the control. Smaller or discarded containers unsuitable for guppy distribution in the intervention area also had 51% less infestation than those in the control area, suggesting a "community-wide" protective effect. In addition, there was less infestation in villages with higher rates of fish uptake, suggesting that the presence of fish was responsible for a reduction in Aedes infestation. This applied vector control model was well-accepted, effective, efficient, and shows promise as a sustainable community-based, non-insecticidal intervention for dengue vector control in large domestic water storage containers in rural Cambodia and elsewhere.     

Biochemical isolation of myonuclei employed to define changes to the myonuclear proteome that occur with aging

Skeletal muscle aging is accompanied by loss of muscle mass and strength. Examining changes in myonuclear proteins with age would provide insight into molecular processes which regulate these profound changes in muscle physiology. However, muscle tissue is highly adapted for contraction and thus comprised largely of contractile proteins making the nuclear proteins difficult to identify from whole muscle samples. By developing a method to purify myonuclei from whole skeletal muscle, we were able to collect myonuclei for analysis by flow cytometry, biochemistry, and mass spectrometry. Nuclear purification dramatically increased the number and intensity of nuclear proteins detected by mass spectrometry compared to whole tissue. We exploited this increased proteomic depth to investigate age‐related changes to the myonuclear proteome. Nuclear levels of 54 of 779 identified proteins (7%) changed significantly with age; these proteins were primarily involved in chromatin maintenance and RNA processing. To determine whether the changes we detected were specific to myonuclei or were common to nuclei of excitatory tissues, we compared aging in myonuclei to aging in brain nuclei. Although several of the same processes were affected by aging in both brain and muscle nuclei, the specific proteins involved in these alterations differed between the two tissues. Isolating myonuclei allowed a deeper view into the myonuclear proteome than previously possible facilitating identification of novel age‐related changes in skeletal muscle. Our technique will enable future studies into a heretofore underrepresented compartment of skeletal muscle.     

Access to artemisinin combination therapy for malaria in remote areas of Cambodia

Background

Within the context of increasing antimalarial costs and or decreasing malaria transmission, the importance of limiting antimalarial treatment to only those confirmed as having malaria parasites becomes paramount. This motivates for this assessment of the cost-effectiveness of routine use of rapid diagnostic tests (RDTs) as an integral part of deploying artemisinin-based combination therapies (ACTs).

Methods

The costs and cost-effectiveness of using RDTs to limit the use of ACTs to those who actually have Plasmodium falciparum parasitaemia in two districts in southern Mozambique were assessed. To evaluate the potential impact of introducing definitive diagnosis using RDTs (costing $0.95), five scenarios were considered, assuming that the use of definitive diagnosis would find that between 25% and 75% of the clinically diagnosed malaria patients are confirmed to be parasitaemic. The base analysis compared two ACTs, artesunate plus sulfadoxine/pyrimethamine (AS+SP) costing $1.77 per adult treatment and artemether-lumefantrine (AL) costing $2.40 per adult treatment, as well as the option of restricting RDT use to only those older than six years. Sensitivity analyses considered lower cost ACTs and RDTs and different population age distributions.

Results

Compared to treating patients on the basis of clinical diagnosis, the use of RDTs in all clinically diagnosed malaria cases results in cost savings only when 29% and 52% or less of all suspected malaria cases test positive for malaria and are treated with AS+SP and AL, respectively. These cut-off points increase to 41.5% (for AS+SP) and to 74% (for AL) when the use of RDTs is restricted to only those older than six years of age. When 25% of clinically diagnosed patients are RDT positive and treated using AL, there are cost savings per malaria positive patient treated of up to $2.12. When more than 29% of clinically diagnosed cases are malaria test positive, the incremental cost per malaria positive patient treated is less than US$ 1. When relatively less expensive ACTs are introduced (e.g. current WHO preferential price for AL of $1.44 per adult treatment), the RDT price to the healthcare provider should be $0.65 or lower for RDTs to be cost saving in populations with between 30 and 52% of clinically diagnosed malaria cases being malaria test positive.

Conclusion

While the use of RDTs in all suspected cases has been shown to be cost-saving when parasite prevalence among clinically diagnosed malaria cases is low to moderate, findings show that targeting RDTs at the group older than six years and treating children less than six years on the basis of clinical diagnosis is even more cost-saving. In semi-immune populations, young children carry the highest risk of severe malaria and many healthcare providers would find it harder to deny antimalarials to those who test negative in this age group.     

Increased production of apolipoprotein B-containing lipoproteins in the absence of hyperlipidemia in transgenic mice expressing cholesterol 7alpha-hydroxylase

The finding that expression of a cholesterol 7alpha-hydroxylase (CYP7A1) transgene in cultured rat hepatoma cells caused a coordinate increase in lipogenesis and secretion of apoB-containing lipoproteins led to the hypothesis that hepatic production of apoB-containing lipoproteins may be linked to the expression of CYP7A1 (Wang, S.-L., Du, E., Martin, T. D., and Davis, R. A. (1997) J. Biol. Chem. 272, 19351-19358). To examine this hypothesis in vivo, a transgene encoding CYP7A1 driven by the constitutive liver-specific enhancer of the human apoE gene was expressed in C56BL/6 mice. The expression of CYP7A1 mRNA (20-fold), protein ( approximately 10-fold), and enzyme activity (5-fold) was markedly increased in transgenic mice compared with non-transgenic littermates. The bile acid pool of CYP7A1 transgenic mice was doubled mainly due to increased hydrophobic dihydroxy bile acids. In CYP7A1 transgenic mice, livers contained approximately 3-fold more sterol response element-binding protein-2 mRNA. Hepatic expression of mRNAs encoding lipogenic enzymes (i.e. fatty-acid synthase, acetyl-CoA carboxylase, stearoyl-CoA desaturase, squalene synthase, farnesyl-pyrophosphate synthase, 3-hydroxy-3-methylglutaryl-CoA reductase, and low density lipoprotein receptor) as well as microsomal triglyceride transfer protein were elevated approximately 3-5-fold in transgenic mice. CYP7A1 transgenic mice also displayed a >2-fold increase in hepatic production and secretion of triglyceride-rich apoB-containing lipoproteins. Despite the increased hepatic secretion of apoB-containing lipoproteins in CYP7A1 mice, plasma levels of triglycerides and cholesterol were not significantly increased. These data suggest that the 5-fold increased expression of the low density lipoprotein receptor displayed by the livers of CYP7A1 transgenic mice was sufficient to compensate for the 2-fold increase production of apoB-containing lipoproteins. These findings emphasize the important homeostatic role that CYP7A1 plays in balancing the anabolic lipoprotein assembly/secretion pathway with the cholesterol catabolic bile acid synthetic pathway.     

SNAREs contribute to the specificity of membrane fusion

Intracellular membrane fusion is mediated by the formation of a four-helix bundle comprised of SNARE proteins. Every cell expresses a large number of SNARE proteins that are localized to particular membrane compartments, suggesting that the fidelity of vesicle trafficking might in part be determined by specific SNARE pairing. However, the promiscuity of SNARE pairing in vitro suggests that the information for membrane compartment organization is not encoded in the inherent ability of SNAREs to form complexes. Here, we show that exocytosis of norepinephrine from PC12 cells is only inhibited or rescued by specific SNAREs. The data suggest that SNARE pairing does underlie vesicle trafficking fidelity, and that specific SNARE interactions with other proteins may facilitate the correct pairing.     

SNARE complex formation is triggered by Ca2+ and drives membrane fusion

Neurotransmitter exocytosis, a process mediated by a core complex of syntaxin, SNAP-25, and VAMP (SNAREs), is inhibited by SNARE-cleaving neurotoxins. Botulinum neurotoxin E inhibition of norepinephrine release in permeabilized PC12 cells can be rescued by adding a 65 aa C-terminal fragment of SNAP-25 (S25-C). Mutations along the hydrophobic face of the S25-C helix result in SNARE complexes with different thermostabilities, and these mutants rescue exocytosis to different extents. Rescue depends on the continued presence of both S25-C and Ca2+ and correlates with complex formation. The data suggest that Ca2+ triggers S25-C binding to a low-affinity site, initiating trans-complex formation. Pairing of SNARE proteins on apposing membranes leads to bilayer fusion and results in a high-affinity cis-SNARE complex.     

Degradation of Spectrin via Calpains in the Ventral Horn after Transient Spinal Cord Ischemia in Rabbits

In the present study, we investigated chronological changes of μ-calpain, m-calpain and cleaved spectrin αII immunoreactivity in the ventral horn after transient spinal cord ischemia to investigate relationship between calpains and vulnerability to ischemia using abdominal aorta occlusion model in rabbits. Spinal cord sections at the level of L₇ were immunostained with calpains and cleaved spectrin αII monoclonal antibodies. μ-Calpain and m-calpain immunoreactivity was significantly increased in the ischemic ventral horn at 30 min and 1 h after ischemia/reperfusion, respectively. Thereafter, they were decreased with time after ischemia/reperfusion: at 48 h after ischemia, their immunoreactivities nearly disappeared in the ischemic ventral horn. Cleaved spectrin αII immunoreactivity was significantly increased in the ventral horn of spinal cord at 12 h after ischemia/reperfusion, and thereafter, its immunoreactivity was decreased with time after ischemia/reperfusion. In addition, spectrin αII protein level (280 kDa) was decreased from 12 h after ischemia/reperfusion; in contrast, cleaved spectrin αII protein level (150 kDa) was significantly increased at 12 h after ischemia/reperfusion. In conclusion, our observations in this study indicate that calpain is associated with neuronal degeneration in the ventral horn at early time after transient spinal cord ischemia via the proteolysis of spectrin αII.Jae-Chul Lee and In Koo Hwang equally contribute to this article.     

大鼠肝脏细胞核糖皮质激素受体的测定

低浓度25nM以下的[~3H]皮质酮和大鼠肝脏细胞核的特异结台位点具有低容量、高亲和力,和甾体激素结合的特异性等特点,其特异结合量和由胞浆转位到核内的糖皮质激素——受体复合物的量平行,证明该结合位点是具有生理意义的糖皮质激素核受体。实验表明,用20nM的[~3H]皮质酮作一点分析是测定大鼠肝脏细胞核糖皮质激素受体的简便方法。用此法测定了13只成年雌性大鼠肝脏细胞核内的糖皮质激素受体,结果为34.5±4.5fmol/mg DNA。