Improved Framework for Tractography Reconstruction of the Optic Radiation

The optic radiation (OR) is one of the major components of the visual system and a key structure at risk in white matter diseases such as multiple sclerosis (MS). However, it is challenging to perform track reconstruction of the OR using diffusion MRI due to a sharp change of direction in the Meyer’s loop and the presence of kissing and crossing fibers along the pathway. As such, we aimed to provide a highly precise and reproducible framework for tracking the OR from thalamic and visual cortex masks. The framework combined the generation of probabilistic streamlines by high order fiber orientation distributions estimated with constrained spherical deconvolution and an automatic post-processing based on anatomical exclusion criteria (AEC) to compensate for the presence of anatomically implausible streamlines. Specifically, those ending in the contralateral hemisphere, cerebrospinal fluid or grey matter outside the visual cortex were automatically excluded. We applied the framework to two distinct high angular resolution diffusion-weighted imaging (HARDI) acquisition protocols on one cohort, comprised of ten healthy volunteers and five MS patients. The OR was successfully delineated in both HARDI acquisitions in the healthy volunteers and MS patients. Quantitative evaluation of the OR position was done by comparing the results with histological reference data. Compared with histological mask, the OR reconstruction into a template (OR-TCT) was highly precise (percentage of voxels within the OR-TCT correctly defined as OR), ranging from 0.71 to 0.83. The sensitivity (percentage of voxels in histological reference mask correctly defined as OR in OR-TCT) ranged from 0.65 to 0.81 and the accuracy (measured by F1 score) was 0.73 to 0.77 in healthy volunteers. When AEC was not applied the precision and accuracy decreased. The absolute agreement between both HARDI datasets measured by the intraclass correlation coefficient was 0.73. This improved framework allowed us to reconstruct the OR with high reliability and accuracy independently of the acquisition parameters. Moreover, the reconstruction was possible even in the presence of tissue damage due to MS. This framework could also be applied to other tracts with complex configuration.     

Beyond Bar and Line Graphs: Time for a New Data Presentation Paradigm

Figures in scientific publications are critically important because they often show the data supporting key findings. Our systematic review of research articles published in top physiology journals (n = 703) suggests that, as scientists, we urgently need to change our practices for presenting continuous data in small sample size studies. Papers rarely included scatterplots, box plots, and histograms that allow readers to critically evaluate continuous data. Most papers presented continuous data in bar and line graphs. This is problematic, as many different data distributions can lead to the same bar or line graph. The full data may suggest different conclusions from the summary statistics. We recommend training investigators in data presentation, encouraging a more complete presentation of data, and changing journal editorial policies. Investigators can quickly make univariate scatterplots for small sample size studies using our Excel templates.     

Delivery of a Chlamydial Adhesin N-PmpC Subunit Vaccine to the Ocular Mucosa Using Particulate Carriers

Trachoma, caused by the intracellular bacterium Chlamydia trachomatis (Ct), remains the world’s leading preventable infectious cause of blindness. Recent attempts to develop effective vaccines rely on modified chlamydial antigen delivery platforms. As the mechanisms engaged in the pathology of the disease are not fully understood, designing a subunit vaccine specific to chlamydial antigens could improve safety for human use. We propose the delivery of chlamydia-specific antigens to the ocular mucosa using particulate carriers, bacterial ghosts (BGs). We therefore characterized humoral and cellular immune responses after conjunctival and subcutaneous immunization with a N-terminal portion (amino acid 1–893) of the chlamydial polymorphic membrane protein C (PmpC) of Ct serovar B, expressed in probiotic Escherichia coli Nissle 1917 bacterial ghosts (EcN BGs) in BALB/c mice. Three immunizations were performed at two-week intervals, and the immune responses were evaluated two weeks after the final immunization in mice. In a guinea pig model of ocular infection animals were immunized in the same manner as the mice, and protection against challenge was assessed two weeks after the last immunization. N-PmpC was successfully expressed within BGs and delivery to the ocular mucosa was well tolerated without signs of inflammation. N-PmpC-specific mucosal IgA levels in tears yielded significantly increased levels in the group immunized via the conjunctiva compared with the subcutaneously immunized mice. Immunization with N-PmpC EcN BGs via both immunization routes prompted the establishment of an N-PmpC-specific IFNγ immune response. Immunization via the conjunctiva resulted in a decrease in intensity of the transitional inflammatory reaction in conjunctiva of challenged guinea pigs compared with subcutaneously and non-immunized animals. The delivery of the chlamydial subunit vaccine to the ocular mucosa using a particulate carrier, such as BGs, induced both humoral and cellular immune responses. Further investigations are needed to improve the immunization scheme and dosage.     

Antioxidant Properties of Selected <Emphasis Type="Italic">Boletus</Emphasis> Mushrooms

Considering the growing interest for mushrooms and the demand search of natural antioxidants sources, the aim of this study was to investigate the antioxidant properties of two edible widely used Boletus species, Boletus edulis, and Boletus auranticus, collected from Istra region in Croatia in late summer 2007. To evaluate the antioxidant properties and content of antioxidant compounds, scavenging capacity on DPPH˙, OH˙, and O₂˙⁻ radicals, reducing power and capacity to inhibit lipid peroxidation has been investigated. It is determined that content of total phenols (41.82 ± 0.08 mg gallic acid equivalent per gram of dry extract) was higher for B. edulis. Using high performance liquid chromatography/diode array detector analysis, the main antioxidant compound, variegatic acid, has been detected and quantified. 1,1-Diphenyl-2-picryl-hydrazyl-hydrate assay was used as a preliminary free radical–scavenging evaluation. By this assay, it has been found that B. edulis dry mushroom extract exhibits 50% of inhibition value at the extract concentration of 0.016 ± 0.0003 mg/ml. The extracts were capable of reducing iron(III) and, thus, are capable of donating electrons. Using electron paramagnetic resonance spin-trapping and spin-probing techniques, activity against relevant reactive species, ˙OH and O₂˙⁻ radical, was analyzed for both mushroom extracts. Both investigated extracts are determined as good inhibitors for ˙OH radical reduction, and both exhibited significant capacity for scavenging O₂˙⁻ radical and for that could help to prevent or meliorate oxidative damage. Only B. edulis extract prevents lipid peroxidation. Investigated mushroom extracts could represent easily accessible natural antioxidant resource.     

Modulation of granulocyte functions by peptide YY in the rat: Age-related differences in Y receptors expression and plasma dipeptidyl peptidase 4 activity

It has been acknowledged that aging exerts detrimental effects on cells of the innate immune system and that neuropeptides, including neuropeptide Y (NPY) and NPY-related peptides fine-tune the activity of these cells through a receptor specific mechanism. The present study investigated the age-dependent potential of peptide YY (PYY) to modulate different granulocyte functions. The PYY reduced the carrageenan-elicited granulocyte accumulation into the air-pouch of aged (24 months) rats, and markedly decreased the phagocytosis of zymosan, as well as the H₂O₂ production, when applied in vivo (20 μg/air-pouch). The anti-inflammatory effect of PYY was less prominent in adult (8 months) and young (3 months) rats. However, the proportions of granulocytes expressing Y1, Y2 and Y5 receptor subtypes were significantly lower in both aged and young rats when compared to adult rats. Furthermore, the aging was found to be associated with the diminished dipeptidyl peptidase 4 (DP4, an enzyme converting the NPY and PYY to Y2/Y5 receptor selective agonists) activity in plasma. In conclusion, the diverse age-related anti-inflammatory effect of PYY in rats originates from different expression levels of Y1, Y2, and Y5 receptor subtypes in addition to different plasma DP4 activity.     

Internal Binding of Halogenated Phenols in Dehaloperoxidase-Hemoglobin Inhibits Peroxidase Function

Dehaloperoxidase (DHP) from the annelid Amphitrite ornata is a catalytically active hemoglobin-peroxidase that possesses a unique internal binding cavity in the distal pocket above the heme. The previously published crystal structure of DHP shows 4-iodophenol bound internally. This led to the proposal that the internal binding site is the active site for phenol oxidation. However, the native substrate for DHP is 2,4,6-tribromophenol, and all attempts to bind 2,4,6-tribromophenol in the internal site under physiological conditions have failed. Herein, we show that the binding of 4-halophenols in the internal pocket inhibits enzymatic function. Furthermore, we demonstrate that DHP has a unique two-site competitive binding mechanism in which the internal and external binding sites communicate through two conformations of the distal histidine of the enzyme, resulting in nonclassical competitive inhibition. The same distal histidine conformations involved in DHP function regulate oxygen binding and release during transport and storage by hemoglobins and myoglobins. This work provides further support for the hypothesis that DHP possesses an external binding site for substrate oxidation, as is typical for the peroxidase family of enzymes.     

Antropologic factors in prediction of ovary cancer

The aim of the study was to determine a combination of anthropometric variables that would enable better differentiation between benign and malignant ovarian masses. Prospective study has been performed in a two year period in which 208 women with ovarian lesions were analyzed and correlated with histopathologic surgical findings. We examined the relation between self-reported anthropometric and other variables (height, weight, body mass index--BMI, parity, marital status, education, age, rural versus urban residence, menopausal status) and incidence of ovarian cancer. Age, parity, marital status and menopausal status individually showed statistical significance.     

A single-centre experience with octreotide in the treatment of different hypersecretory syndromes in patients with functional gastroenteropancreatic neuroendocrine tumors

The aim of this research was to assess the clinical and biochemical efficacy of the octreotide in the treatment of patients with various functional gastroenteropancreatic neuroendocrine tumors (GEP-NETs). The study included 14 patients treated with octreotide for 6 months. They were diagnosed with VIPoma, glucagonoma, gastrinoma, medullary thyroid carcinoma (solitary and as a part of MEN-II syndrome), pancreatic carcinoids (solitary and as a part of multiple endocrine neoplasia type-1 syndrome-MEN-1 syndrome) and midgut carcinoids. The patients presented with Verner-Morrison, glucagonoma, Zollinger Ellison and carcinoid syndrome respectively. All had a metastatic disease at the time of diagnosis and a positive octreoscan finding. Initially elevated chromogranin A (CgA) levels were detected in 11 (78.6%) and elevated 5-hydroxyindolacetic acid (5-HIAA) levels in 8 (57.1%) patients. Symptomatic efficacy assessments were made by diarrhea reductions during treatment course, and laboratory efficacy was assessed through changes in 5-HIAA and CgA levels. Assessments were made initially and following 6 months of therapy. Median urinary 5-HIAA and the number of stools decreased significantly (p = 0.016 and p = 0.009 respectively, p < 0.05) while CgA levels had the decreasing tendency but not statistically significant (p = 0.14). There was a positive correlation between the 5-HIAA reduction and the decrease in stool number at baseline and during treatment course (p < 0.05). No correlation was observed between 5-HIAA and CgA levels and also there was no correlation between CgA reduction and symptomatic improvement. The results prove octreotide to be effective in reducing symptoms and biochemical markers associated with hypersecretory syndromes of GEP-NETs.     

Organisation of cervical cytology screening in Croatia: past, present and future

This presentation highlights strengths and weaknesses of cervical cytology screening in Croatia, with particular reference to the opportunistic screening, the use of conventional Papanicolaou (Pap) test and the analysis of some organizational, educational and performance issues that are associated with it. Its aim is to propose measures to improve the efficacy of cervical cytology screening in order to reduce cervical cancer mortality. Currently, in excess of 450,000 Pap tests/ year are examined at 35 laboratories scattered throughout the country. All of these laboratories use standard operating procedures including internal and external quality control. They employ a total of 68 cytologists and 91 cytotechnologists. The sensitivity of cervical screening in Croatia is 90.0%, specificity 98.6%, positive predictive value 92.3%, negative predictive value 98.1% and overall diagnostic accuracy 97.2%. The high diagnostic accuracy of cervical cytology is attributed to the long-standing tradition of education and training of cytologists (postgraduate MSc course since 1967, independent residency since 1974) and cytotechnologists (since 1968). This tradition spanning more than half a century means that today in Croatia there is a developed network of cytology laboratories staffed by highly competent cytologists and trained cytotechnologists. The high accuracy of cancer detection through Pap tests provides strong evidence in support of cervical cytology screening remaining the basic method of prevention for cervical carcinoma. However, some modifications to the current situation are needed. These relate primarily to opportunistic screening. The current screening coverage rate is 68%, although there is capacity, which would allow for all women at risk, i.e. those aged 25-64, to be screened once in three years. The screening coverage relates mainly to those women visiting gynecological out patient clinics for unrelated conditions. A properly organized and controlled national screening programme should replace this. This should be accompanied by the introduction of alternative, highly sensitive methods of sample collection and preparation, such as are available through the introduction of new technologies, e.g. liquid based cytology.     

The growth-hormone inducible transmembrane protein (Ghitm) belongs to the Bax inhibitory protein-like family

The conserved protein domain UPF0005 is a protein family signature distributed among many species including fungi and bacteria. Although of unknown functionality this motif has been found in newly identified antiapoptotic proteins comprising the BI-1 family, namely Bax-inhibitory Protein-1 (BI-1), Lifeguard (LFG), and h-GAAP. In a search for vertebrate proteins presumably belonging to the BI-1 family, we found that Growth-hormone inducible transmembrane protein (Ghitm) is another prospective member of the BI-1 family. Here we characterise Ghitm in a first analysis regarding its phylogeny, expression in cancer cell lines, and proteomical properties.