Knowledge,Attitudes, and Practices regarding Diarrhea and Cholera following an Oral Cholera Vaccination Campaign in the Solomon Islands

BackgroundIn response to a 2011 cholera outbreak in Papua New Guinea, the Government of the Solomon Islands initiated a cholera prevention program which included cholera disease prevention and treatment messaging, community meetings, and a pre-emptive cholera vaccination campaign targeting 11,000 children aged 1–15 years in selected communities in Choiseul and Western Provinces.ConclusionsThis pre-emptive OCV campaign in a cholera-naïve community provided a unique opportunity to assess household-level knowledge, attitudes, and practices regarding diarrhea, cholera, and water, sanitation, and hygiene (WASH). Our findings suggest that education provided during the vaccination campaign may have reinforced earlier mass messaging about cholera and diarrheal disease in vaccinated communities.     

A neural-specific F-box protein Fbs1 functions as a chaperone suppressing glycoprotein aggregation

Fbs1 is an F-box protein present abundantly in the nervous system. Similar to the ubiquitously expressed Fbs2, Fbs1 recognizes N-glycans at the innermost position as a signal for unfolded glycoproteins, probably in the endoplasmic reticulum-associated degradation pathway. Here, we show that the in vivo majority of Fbs1 is present as Fbs1-Skp1 heterodimers or Fbs1 monomers but not SCF(Fbs1) complex. The inefficient SCF complex formation of Fbs1 and the restricted presence of SCF(Fbs1) bound on the endoplasmic reticulum membrane were due to the short linker sequence between the F-box domain and the sugar-binding domain. In vitro, Fbs1 prevented the aggregation of the glycoprotein through the N-terminal unique sequence of Fbs1. Our results suggest that Fbs1 assists clearance of aberrant glycoproteins in neuronal cells by suppressing aggregates formation, independent of ubiquitin ligase activity, and thus functions as a unique chaperone for those proteins.     

Induction of steady-state glomeruloid sphere by self-assembly from human embryonic kidney cells

The glomerulus is a network of capillaries known as a tuft, located at the beginning of a nephron in the kidney. Here we describe a novel method for the induction of a macroscopically visible three-dimensional glomerulus-like sphere (GLS). This procedure did not require any additional cytokines and completed the formation of spheres within 24?h. After the formation was complete, GLS maintained a steady state for at least five days without proliferation and without a decrease in viability. Therefore, this procedure assists various assays for a prolong period of time. Overall, our protocol allows for a very simple mixing of cells from different sources to obtain fine-grained and highly dispersed GLSs. The kidney filtration barrier is a unique structure characterized by a complex three-dimensional framework of podocytes and endothelial cells. GLS exhibited the induction of many podocyte-specific gene profiles similar to those in adult human kidneys, suggesting that the sphere formation process is important for the maturation of podocytes. Focal segmental glomerulosclerosis (FSGS) is one of the major causes of steroid-resistant nephrotic syndrome, and some circulating permeability factors in the patient's serum FSGS have been implicated in the pathogenesis of the disease. Serum from patients with FSGS induced the collapse of GLS, which imitates the appearance of glomerulosclerosis in patients. In conclusion, the investigation and use of GLS may provide a novel method to elucidate the molecular mechanisms underlying complicated and unexplained events in glomeruli in a similar condition in adult kidneys.     

Rat liver serine dehydratase. Bacterial expression and two folding domains as revealed by limited proteolysis

A pCW vector harboring rat liver serine dehydratase cDNA was expressed in Escherichia coli. The expressed level was about 5-fold higher in E. coli BL21 than in JM109 cell extract; the former lacked two kinds of proteases. Immunoblot analysis revealed the occurrence of a derivative other than serine dehydratase in the JM109 cell extract. The recombinant enzyme was purified to homogeneity. Staphylococcus aureus V8 protease and trypsin cleaved the enzyme at Glu-206 and Lys-220, respectively, with a concomitant loss of enzyme activity. Spectrophotometrically, the nicked enzyme showed a approximately 50% reduced capacity for binding of the coenzyme pyridoxal phosphate and no spectral change of circular dichroism in the region at 300-480 nm, whereas circular dichroism spectra of both enzymes in the far-UV region were similar, suggesting that proteolysis impairs the coenzyme binding without an accompanying gross change of the secondary structure. Whereas the nicked enzyme behaved like the intact enzyme on Sephadex G-75 column chromatography, it was dissociated into two fragments on the column containing 6 M urea. Upon the removal of urea, both fragments spontaneously refolded. These results suggest that serine dehydratase consists of two folding domains connected by a region that is very susceptible to proteases.     

Somatostatin suppresses ghrelin secretion from the rat stomach

Ghrelin is an acylated peptide that stimulates food intake and the secretion of growth hormone. While ghrelin is predominantly synthesized in a subset of endocrine cells in the oxyntic gland of the human and rat stomach, the mechanism regulating ghrelin secretion remains unknown. Somatostatin, a peptide produced in the gastric oxyntic mucosa, is known to suppress secretion of several gastrointestinal peptides in a paracrine fashion. By double immunohistochemistry, we demonstrated that somatostatin-immunoreactive cells contact ghrelin-immunoreactive cells. A single intravenous injection of somatostatin reduced the systemic plasma concentration of ghrelin in rats. Continuous infusion of somatostatin into the gastric artery of the vascularly perfused rat stomach suppressed ghrelin secretion in both dose- and time-dependent manner. These findings indicate that ghrelin secretion from the stomach is regulated by gastric somatostatin.     

Evolutionary history and mechanism of the Drosophila Cecropin gene family

 Upon bacterial infection, insects secrete a set of synthesized antibacterial proteins into the hemolymph and initiate synergistic destruction of invaders. Cecropin is one such antibacterial protein which is also found in vertebrates. To study the evolutionary history and mechanism of the Cecropin gene family, we determined DNA sequences of one isogenic In(3R)C and six isofemale lines of Drosophila melanogaster as well as one line of D. simulans and of D. yakuba. The phylogenetic analysis of these sequences together with those published for D. virilis and Sarcophaga peregrina reveals frequent gene re-organization. It was also found that silent nucleotide differences within D. melanogaster are quite heterogeneous across the gene region of approximately 3 kilobases and the extent of polymorphism is unusually usually high. These data suggest that the Cecropin gene region of D. melanogaster underwent intragenic recombination as well as introgression from a closely related sibling species, D. simulans. Received: 31 July 1997 / Revised: 24 October 1997     

Intra-Arterial Transplantation of Allogeneic Mesenchymal Stem Cells Mounts Neuroprotective Effects in a Transient Ischemic Stroke Model in Rats: Analyses of Therapeutic Time Window and Its Mechanisms

Objective

Intra-arterial stem cell transplantation exerts neuroprotective effects for ischemic stroke. However, the optimal therapeutic time window and mechanisms have not been completely understood. In this study, we investigated the relationship between the timing of intra-arterial transplantation of allogeneic mesenchymal stem cells (MSCs) in ischemic stroke model in rats and its efficacy in acute phase.

Methods

Adult male Wistar rats weighing 200 to 250g received right middle cerebral artery occlusion (MCAO) for 90 minutes. MSCs (1×10⁶cells/ 1ml PBS) were intra-arterially injected at either 1, 6, 24, or 48 hours (1, 6, 24, 48h group) after MCAO. PBS (1ml) was intra-arterially injected to control rats at 1 hour after MCAO. Behavioral test was performed immediately after reperfusion, and at 3, 7 days after MCAO using the Modified Neurological Severity Score (mNSS). Rats were euthanized at 7 days after MCAO for evaluation of infarct volumes and the migration of MSCs. In order to explore potential mechanisms of action, the upregulation of neurotrophic factor and chemotactic cytokine (bFGF, SDF-1α) induced by cell transplantation was examined in another cohort of rats that received intra-arterial transplantation at 24 hours after recanalization then euthanized at 7 days after MCAO for protein assays.

Results

Behavioral test at 3 and 7 days after transplantation revealed that stroke rats in 24h group displayed the most robust significant improvements in mNSS compared to stroke rats in all other groups (p’s<0.05). Similarly, the infarct volumes of stroke rats in 24h group were much significantly decreased compared to those in all other groups (p’s<0.05). These observed behavioral and histological effects were accompanied by MSC survival and migration, with the highest number of integrated MSCs detected in the 24h group. Moreover, bFGF and SDF-1α levels of the infarcted cortex were highly elevated in the 24h group compared to control group (p’s<0.05).

Conclusions

These results suggest that intra-arterial allogeneic transplantation of MSCs provides post-stroke functional recovery and reduction of infarct volumes in ischemic stroke model of rats. The upregulation of bFGF and SDF-1α likely played a key mechanistic role in enabling MSC to afford functional effects in stroke. MSC transplantation at 24 hours after recanalization appears to be the optimal timing for ischemic stroke model, which should guide the design of clinical trials of cell transplantation for stroke patients.     

Stress responses in Salix gracilistyla cuttings subjected to repetitive alternate flooding and drought

To determine the tolerance of Salix gracilistyla to repetitive alternate flooding and drought, we measured leaf stomatal conductance, pre-dawn water potential, osmotic adjustment, and biomass production under greenhouse conditions. We used a control and nine crossed treatments (F1-D1–F3-D3) in which we combined 1-, 2-, or 3-week floodings (F) and droughts (D). Leaf stomatal conductance was lowest in 3 weeks of flooding or drought when the preceding event (flood or drought) was also of a 3-week duration. Leaf pre-dawn water potential was reduced in 3 weeks of drought when preceded by 2 or 3 weeks of flooding. Cuttings had slight osmotic adjustments in repetitions of long floodings and droughts. During longer durations of drought in crossed experiments, plants had low root and shoot mass, few hypertrophic lenticels, and reduced leaf mass; when flooding duration increased in crossed experiments, root mass was reduced, there were more hypertrophic lenticels, and the leaf area was reduced. Cuttings achieved stress tolerance by inhibition of transpiration, osmotic adjustment, reduction of transpiration area, and development of hypertrophic lenticels. Stress tolerance was weak when repetitive 2- or 3-week floodings were combined with 3-week droughts. The duration of flooding and drought periods under which S. gracilistyla achieves stress tolerance may be critical in determining distributions along riverbanks.     

Sensitive determination of carnosine in urine by high-performance liquid chromatography using 4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride as a fluorescent labeling reagent

A simple and highly sensitive high-performance liquid chromatography procedure was developed for the determination of carnosine in urine. Carnosine was derivatized with 4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride at 70 °C for 15 min in borate buffer (20 mmol l^?1, pH 9.0) to produce fluorescent sulfonamides. After hydrolysis of the reaction mixture with formic acid at 100 °C for 15 min, the fluorescent derivative of carnosine was separated on a reversed-phase column with a linear gradient elution using solvents of (A) acetate buffer (0.1 mmol l^?1, pH 7.0) and (B) acetonitrile at a flow-rate of 1.0 ml/min and was detected at excitation and emission wavelengths of 318 and 400 nm, respectively. The detection limit of carnosine was 4 fmol at a signal-to-noise ratio of 3. The within-day and day-to-day relative standard deviations were 2.7–4.6% and 0.4–5.2%, respectively. The concentration of carnosine in normal human urine was found to be 4.6–125 nmol (mg creatinine)^?1 (mean ± SD: 21.6 ± 26.6 nmol (mg creatinine)^?1, n = 20).