A new cytolytic protein from the sea anemone Urticina crassicornis that binds to cholesterol- and sphingomyelin-rich membranes

A new pore-forming cytolytic protein was isolated from the Northern red sea anemone, Urticina crassicornis. Its biochemical properties were characterized and partial N-terminal amino acid sequence was determined. The cytolysin, named UcI, has a molecular mass of around 30 kDa and lacks phospholipase A₂ activity. UcI lyses bovine erythrocytes at nanomolar concentrations. Hemolysis is a result of a colloid-osmotic shock caused by the opening of toxin-induced ionic pores and can be prevented by osmotic protectants of size >600 Da. The functional radius of an average pore was estimated to be about 0.66 nm. A more detailed study of the cytolytic activity of UcI was performed with lipid vesicles and monolayers. The toxin binds to monolayers and efficiently permeabilizes small lipid vesicles composed of sphingomyelin and cholesterol. However, the cytolytic activity is not prevented by preincubation with either pure cholesterol or sphingomyelin dispersions. We conclude that the presence of both sphingomyelin and cholesterol, key components of lipid rafts, greatly enhances toxin binding to membranes and probably facilitates pore formation. Alignment of the toxin partial amino acid sequence with sequences of cytolysins belonging to the actinoporin family reveals no sequence homology. We conclude that partial sequence of UcI resembles only the N-terminal part of UpI, a cytolytic protein isolated from a related sea anemone species, Urticina piscivora. The two proteins most probably belong to a separate family of sea anemone cytolysins that are worthy of further characterization.     

Plasma erythropoietin level and iron reserves in haemodialysis patients with and without acquired cystic kidney disease

Concentration of erythropoietin (Epo) and iron reserves (IR) belong to the essential factors determining erythropoiesis in haemodialysis patients. Patients on dialysis with acquired kidney disease (ACKD+) can control anaemia better than patients without acquired kidney disease (ACKD-). Therefore we decided to check if plasma Epo levels and IR differ significantly in both groups of patients. Forty chronically haemodialyzed patients after ultrasound diagnosis were divided into 18 patients (45%) with ACKD+ and 22 (55%) without ACKD-. In both groups of patients we compared their plasma levels of Epo and IR. Plasma erythropoietin and ferritin levels were measured by enzymatic immunoassay. Iron reserves were estimated by the formula: IR=400×[ln(ferritin)-ln(50)]. In the ACKD+ group 72% of patients and in the ACKD-group 32% of patients did not require rHu Epo therapy. Plasma levels of erythropoietin and iron reserves did not differ significantly between ACKD+ and ACKD—patients. There must be also other factors than erythropoietin levels and iron reserves regulating erythropoiesis in these patients.     

Electrophysiological testing of visual function after mirror telescope implantation: a case report

Purpose

The implantation of an intraocular telescope increases life quality in patients with end-stage age-related macular degeneration (AMD). The present study monitored changes in electrophysiological markers of visual processing before and during seventeen months after a novel mirror telescope implantation in two patients (OV—male 90 years, MZ—female 70 years) with the final-stage form of AMD.

Methods

Visual evoked potentials were recorded to high-contrast pattern-reversal (PR-VEP for check size 40′ and 10′), low-contrast motion-onset stimuli (in visual periphery M-VEP M20°, and in central part M-VEP C8°), and event-related potentials (ERPs) in the oddball visual paradigm.

Results

MZ’s more systematic responses showed attenuation and prolongation of the M-VEP M20° and the PR-VEP 40′ immediately after the telescope implantation with a slow amplitude recovery with unchanged prolonged latency. The implantation completely eradicated the M-VEP C8° without any restoration. The PR-VEP 10′ were not readable. Only a part of OV’s PR-VEP 40′ and M-VEP M20′ were of a repeatable and expected morphology. These OV’s VEPs were consistent with MZ’s findings. The ERPs did not show any effect of implantation in both patients. Post-implantation visual acuity and reaction time overcame the pre-implantation levels.

Conclusions

The mirror telescope preserved peripheral vision in contrast to classic telescopes; however, the telescope concurrently reduced the luminance of the magnified retinal image, which was likely responsible for the prolongation of the VEP latencies.

     

Lithium-induced changes in the brain levels of free amino acids in stress-exposed rats

The effects of stress and lithium on brain free amino acid levels in rats were investigated. Stress caused a significant decrease in the brain levels of alanine, ammonia, arginine, isoleucine, lysine and phenylalanine. Lithium by itself induced a significant increase in the brain levels of arginine and threonine, and a significant decrease in those of alanine, ammonia, aspartic acid, glutamic acid, glycine, lysine, phenylalanine and tyrosine. Lithium and stress together significantly decreased brain levels of alanine, arginine, aspartic acid, glycine, isoleucine, leucine, lysine, phenylalanine and tyrosine, and significantly increased that of serine.     

SORB-GEL method of analysis of 99mTc-labelled compounds

A SORB-GEL method has been worked out for analysing ^99mTc-labelled compounds which (in a few minutes) enables the pertechnetate content to be determined in a preparation. The method is based upon a different tupe of behaviour of ^99mTc-labelled compound, pertechnetate in the columns packed with Sephadex G-10, and with alumina during elution with saline.Polythene syringes 4.7 cm long and 1 cm in diameter were used as chromotographic columns. A syringe packed with Sephadex G-10 transferred 0.1 ml of ^99mTc-labelled compound into the column, and the activity of the column was then measured in the ionisation chamber. Using a syringe, 20 ml of saline was foreced through this column. The eluate was then removed. Using an injection needle, a second column, packed with alumina, was connected with the first. Similarly, a third column, packed with Sephadex G-10, was connected to the second. Through all of them 40 ml of saline was forced. The third column containing Sephadex G-10 was then disconnected and its activity was measured in the ionisation chamber. The pertechnetate content in the preparation was then calculated from the measured values.The method is suitable for determinating the free pertechnetate content in strong and weak chelate compounds and in particle preparations.     

Serum Vitamin D Levels in Children with Vernal Keratoconjunctivitis

Purpose: To evaluate serum 25-hydroxyvitamin D [25(OH)D3] levels of vernal keratoconjunctivitis (VKC) children.

Methods: A total of 62 non-atopic healthy children (64.5% male, mean age 10.79 ± 3.3 years) and 29 VKC children (75.9%, mean age 12.17 ± 2.7 years) were included in the study. Serum 25(OH)D3 levels measured by HPLC were compared between the two groups and a p value of <0.05 was considered as statistically significant.

Results: The mean serum 25(OH)D3 level of VKC group was significantly lower than in the control group (11.02 ± 5.16 ng/mL and 15.99 ± 7.36 ng/mL, respectively) (p = 0.002). Severe vitamin D deficiency (<10 ng/mL) was detected in 48.3% of VKC children and 22.6% of the controls (p = 0.017). Time spent outdoors during daylight was higher in the control group (229.5 ± 101.2 min) compared with the VKC group (160.7 ± 65.9 min) (p = 0.008), and showed a significant correlation with serum 25(OH)D3 levels (Spearman rho = 0.812) (p<0.001).

Conclusions: Children with VKC should be evaluated for vitamin D deficiency, which might occur secondary to sun avoidance.     

Molecular prevalence and ecoregion distribution of select tick‐borne pathogens in Texas dogs

Tick‐borne diseases (TBD), caused by borrelial, rickettsial and babesial pathogens, are common across the United States and can cause severe clinical disease in susceptible hosts, such as domestic dogs. However, there are limited TBD molecular epidemiological reports for dogs in Texas, and none for the non‐Lyme borrelial pathogen responsible for causing tick‐borne relapsing fever (TBRF). Therefore, data to support the prevalence of TBRF in the canine population is inadequate. This study aimed to characterize the molecular prevalence of 11 causative agents responsible for three TBD groups within domestic dogs with an emphasis on pathogen distribution within Texas ecoregions. A total representative number of 1,171 whole‐blood samples were collected opportunistically from two Texas veterinary diagnostic laboratories. A layerplex real‐time PCR assay was utilized to screen the dog samples for all 11 pathogens simultaneously. The overall molecular infection prevalence of disease was 0.68% borrelial, 1.8% rickettsial and 0.43% babesial pathogens, for a TBD total of 2.73% across Texas. Higher molecular prevalence was observed when analysed by ecoregion distinction, including 5.78% rickettsial infections by Ehrlichia canis and Anaplasma platys in the Rolling Plains ecoregion, and an average of 1.1% Borrelia turicatae and 1.0% Babesia gibsoni across detected ecoregions. To our knowledge, our findings indicate the first molecular detection of A. platys in Texas, and the first report of coinfections with E. canis and A. platys in dogs of Texas. The zoonotic concerns associated with TBDs, in conjunction with dogs’ implication as an effective sentinel for human disease, highlight the importance of characterizing and monitoring regions associated with active infections in dogs. Surveillance data obtained from this study may aid public health agencies in updating maps depicting high‐risk areas of disease and developing preventative measures for the affected areas.