A Naturally Occurring Canine Model of Autosomal Recessive Congenital Stationary Night Blindness

Congenital stationary night blindness (CSNB) is a non-progressive, clinically and genetically heterogeneous disease of impaired night vision. We report a naturally-occurring, stationary, autosomal recessive phenotype in beagle dogs with normal daylight vision but absent night vision. Affected dogs had normal retinas on clinical examination, but showed no detectable rod responses. They had “negative-type” mixed rod and cone responses in full-field ERGs. Their photopic long-flash ERGs had normal OFF-responses associated with severely reduced ON-responses. The phenotype is similar to the Schubert-Bornschein form of complete CSNB in humans. Homozygosity mapping ruled out most known CSNB candidates as well as CACNA2D4 and GNB3. Three remaining genes were excluded based on sequencing the open reading frame and intron-exon boundaries (RHO, NYX), causal to a different form of CSNB (RHO) or X-chromosome (NYX, CACNA1F) location. Among the genes expressed in the photoreceptors and their synaptic terminals, and mGluR6 cascade and modulators, reduced expression of GNAT1, CACNA2D4 and NYX was observed by qRT-PCR in both carrier (n = 2) and affected (n = 2) retinas whereas CACNA1F was down-regulated only in the affecteds. Retinal morphology revealed normal cellular layers and structure, and electron microscopy showed normal rod spherules and synaptic ribbons. No difference from normal was observed by immunohistochemistry (IHC) for antibodies labeling rods, cones and their presynaptic terminals. None of the retinas showed any sign of stress. Selected proteins of mGluR6 cascade and its modulators were examined by IHC and showed that PKCα weakly labeled the rod bipolar somata in the affected, but intensely labeled axonal terminals that appeared thickened and irregular. Dendritic terminals of ON-bipolar cells showed increased Goα labeling. Both PKCα and Goα labeled the more prominent bipolar dendrites that extended into the OPL in affected but not normal retinas. Interestingly, RGS11 showed no labeling in the affected retina. Our results indicate involvement of a yet unknown gene in this canine model of complete CSNB.     

Serum Bile Acids in Repaired Tetralogy of Fallot: A Marker for Liver and Heart?

Background and Aims

Patients with repaired tetralogy of Fallot may develop chronic right ventricular dysfunction and hepatic congestion over time. We hypothesized that bile acid metabolism is altered in repaired tetralogy of Fallot patients and therefore sought to correlate right ventricular indices with serum bile acid levels.

Methods

Indexed right ventricular end diastolic volume, as assessed by cardiac magnetic-resonance imaging, was classified as <100ml/m² (Group 1, n = 5), 100–150ml/m² (Group 2, n = 18), and >150ml/m² (Group 3, n = 6) in 29 patients with repaired tetralogy of Fallot. Pulmonary regurgitation fraction and right ventricular ejection fraction were calculated. The serum bile acid profile, including 15 species, in these patients was determined by liquid chromatography coupled with mass spectrometry.

Results

Serum bile acid levels increased from Group 1 to Group 3 (2.5 ± 0.7; 4.1 ± 2.5; 6.0 ± 2.8 μmol/l, respectively) with significantly increased bile acid values in Group 3 compared to Group 1 (p≤0.05). In Group 3, but not in Group 1 and 2, a significant increase in glycine-conjugated bile acids was observed. Pulmonary regurgitation fraction increased (12 ± 1; 28 ± 16; 43 ± 3%, Groups 1–3, respectively) and right ventricular ejection fraction decreased (48.4 ± 6.4; 48.5 ± 6.5; 42.1 ± 5.3%, Groups 1–3, respectively) with rising indexed right ventricular end diastolic volume.

Conclusions

These preliminary results suggest that serum bile acid levels are positively correlated with indexed right ventricular end-diastolic volume in patients with repaired tetralogy of Fallot; however, this needs to be confirmed in a larger patient cohort.     

Bacterial communities of the coronal sulcus and distal urethra of adolescent males

Lactobacillus-dominated vaginal microbiotas are associated with reproductive health and STI resistance in women, whereas altered microbiotas are associated with bacterial vaginosis (BV), STI risk and poor reproductive outcomes. Putative vaginal taxa have been observed in male first-catch urine, urethral swab and coronal sulcus (CS) specimens but the significance of these observations is unclear. We used 16 S rRNA sequencing to characterize the microbiota of the CS and urine collected from 18 adolescent men over three consecutive months. CS microbiotas of most participants were more stable than their urine microbiotas and the composition of CS microbiotas were strongly influenced by circumcision. BV-associated taxa, including Atopobium, Megasphaera, Mobiluncus, Prevotella and Gemella, were detected in CS specimens from sexually experienced and inexperienced participants. In contrast, urine primarily contained taxa that were not abundant in CS specimens. Lactobacilllus and Streptococcus were major urine taxa but their abundance was inversely correlated. In contrast, Sneathia, Mycoplasma and Ureaplasma were only found in urine from sexually active participants. Thus, the CS and urine support stable and distinct bacterial communities. Finally, our results suggest that the penis and the urethra can be colonized by a variety of BV-associated taxa and that some of these colonizations result from partnered sexual activity.     

Abortive phage resistance mechanism AbiZ speeds the lysis clock to cause premature lysis of phage-infected Lactococcus lactis

The conjugative plasmid pTR2030 has been used extensively to confer phage resistance in commercial Lactococcus starter cultures. The plasmid harbors a 16-kb region, flanked by insertion sequence (IS) elements, that encodes the restriction/modification system LlaI and carries an abortive infection gene, abiA. The AbiA system inhibits both prolate and small isometric phages by interfering with the early stages of phage DNA replication. However, abiA alone does not account for the full abortive activity reported for pTR2030. In this study, a 7.5-kb region positioned within the IS elements and downstream of abiA was sequenced to reveal seven additional open reading frames (ORFs). A single ORF, designated abiZ, was found to be responsible for a significant reduction in plaque size and an efficiency of plaquing (EOP) of 10(-6), without affecting phage adsorption. AbiZ causes phage phi31-infected Lactococcus lactis NCK203 to lyse 15 min early, reducing the burst size of phi31 100-fold. Thirteen of 14 phages of the P335 group were sensitive to AbiZ, through reduction in either plaque size, EOP, or both. The predicted AbiZ protein contains two predicted transmembrane helices but shows no significant DNA homologies. When the phage phi31 lysin and holin genes were cloned into the nisin-inducible shuttle vector pMSP3545, nisin induction of holin and lysin caused partial lysis of NCK203. In the presence of AbiZ, lysis occurred 30 min earlier. In holin-induced cells, membrane permeability as measured using propidium iodide was greater in the presence of AbiZ. These results suggest that AbiZ may interact cooperatively with holin to cause premature lysis.     

Regulation of hepcidin in HepG2 and RINm5F cells

Despite the high impact of the antimicrobial peptide hepcidin in iron homeostasis, the regulation of this hormone is still not completely understood. Studies concerning hepcidin regulation are performed at the mRNA level. For the first time we analyzed the regulation of hepcidin not only at mRNA, but also at protein level in a hepatoma and a pancreatic beta cell line using quantitative RT-PCR and immunoblot analysis. Our data show, that hepcidin is present in HepG2 and RINm5F cells. A significant up-regulation of hepcidin was observed in both cell lines by the inflammatory cytokine interleukin-6, lipopolysaccharide, and a slight upregulation by deferoxamine. A down-regulation was detected after stimulation with erythropoietin. Hepcidin was regulated by iron in a dose dependent manner: low doses up to 3 microM increased hepcidin expression, high doses of iron (65 microM) revealed a switch-over to down-regulation of hepcidin expression. Regulation of hepcidin in HepG2 and RINm5F cells at mRNA and protein level by these substances indicates its involvement in inflammation and iron metabolism.     

Innate recognition network driving herpes simplex virus-induced corneal immunopathology: role of the toll pathway in early inflammatory events in stromal keratitis

Ocular infection with herpes simplex virus (HSV) sets off an array of events that succeed in clearing virus from the cornea but leaves the tissue with a CD4(+) T-cell-orchestrated chronic inflammatory lesion that impairs vision. We demonstrate that Toll-like receptor (TLR) signaling forms a part of the recognition system that induces the syndrome that eventually culminates in immunopathology. Accordingly, in a comparison of the outcomes of infection in wild-type (WT) mice and those lacking TLR function, it was apparent that the absence of TLR2 and, to a lesser extent, TLR9 resulted in significantly diminished lesions. Similarly, mice lacking the adapter molecule MyD88 were resistant to lesion development, but such animals were also unable to control infection, with most succumbing to lethal encephalitis. The susceptibility of TLR4(-/-) animals was also evaluated. These animals developed lesions, which were more severe, more rapidly than did WT animals. We discuss the possible mechanisms by which early recognition of HSV constituents impacts the subsequent development of immunopathological lesions.     

Synthesis and evaluation of alkenyl indazoles as selective Aurora kinase inhibitors

A series of alkenyl indazoles were synthesized and evaluated in Aurora kinase enzyme assays. Several promising leads were optimized for selectivity towards Aurora B. Excellent binding affinity and good selectivity were achieved with optimized compounds in isolated Aurora subfamily assays.     

How can malaria rapid diagnostic tests achieve their potential? A qualitative study of a trial at health facilities in Ghana

Malaria is still a major public health problem in Brazil, with approximately 306 000 registered cases in 2009, but it is estimated that in the early 1940s, around six million cases of malaria occurred each year. As a result of the fight against the disease, the number of malaria cases decreased over the years and the smallest numbers of cases to-date were recorded in the 1960s. From the mid-1960s onwards, Brazil underwent a rapid and disorganized settlement process in the Amazon and this migratory movement led to a progressive increase in the number of reported cases. Although the main mosquito vector (Anopheles darlingi) is present in about 80% of the country, currently the incidence of malaria in Brazil is almost exclusively (99,8% of the cases) restricted to the region of the Amazon Basin, where a number of combined factors favors disease transmission and impair the use of standard control procedures. Plasmodium vivax accounts for 83,7% of registered cases, while Plasmodium falciparum is responsible for 16,3% and Plasmodium malariae is seldom observed. Although vivax malaria is thought to cause little mortality, compared to falciparum malaria, it accounts for much of the morbidity and for huge burdens on the prosperity of endemic communities. However, in the last few years a pattern of unusual clinical complications with fatal cases associated with P. vivax have been reported in Brazil and this is a matter of concern for Brazilian malariologists. In addition, the emergence of P. vivax strains resistant to chloroquine in some reports needs to be further investigated. In contrast, asymptomatic infection by P. falciparum and P. vivax has been detected in epidemiological studies in the states of Rondonia and Amazonas, indicating probably a pattern of clinical immunity in both autochthonous and migrant populations. Seropidemiological studies investigating the type of immune responses elicited in naturally-exposed populations to several malaria vaccine candidates in Brazilian populations have also been providing important information on whether immune responses specific to these antigens are generated in natural infections and their immunogenic potential as vaccine candidates. The present difficulties in reducing economic and social risk factors that determine the incidence of malaria in the Amazon Region render impracticable its elimination in the region. As a result, a malaria-integrated control effort - as a joint action on the part of the government and the population - directed towards the elimination or reduction of the risks of death or illness, is the direction adopted by the Brazilian government in the fight against the disease.