Expression of vascular endothelial growth factors VEGF- C and D, VEGFR-3, and comparison of lymphatic vessels density labeled with D2-40 antibodies as a prognostic factors in vulvar intraepithelial neoplasia (VIN) and invasive vulvar cancer

OBJECTIVE: The aim of this study was to compare the immunohistochemical expression of vascular endothelial growth factors VEGF-C and D, as well as the expression of VEGFR-3 in VIN and vulvar invasive cancer and to compare the density of lymphatic marker D2-40 antibody in both groups, and to compare them with different clinicopathologic features. Materials & Methods: The study was performed using tissue material and clinical data from 100 women diagnosed with VIN and 100 women diagnosed with invasive vulvar cancer. Results: No significant differences were found in the expression of VEGF-C and -D or VEGFR-3 between those patients with VIN and those with invasive vulvar cancers. Weak expression of VEGF-C was confirmed only in two cases of the analyzed series; in all cases, expression of VEGF-D and VEGFR-3 was observed. The strongest expression of VEGF-D and VEGFR-3 was observed in the group of invasive cancers. The highest density of lymphatic vessels per 2 mm was observed in VIN. In the cancer group, small lymphatic vessels with a narrow oval lumen were observed. Moreover, in two cases of vulvar cancer, the presence of intratumoral lymphatic vessels was observed. Conclusions: These results suggest that lymphangiogenesis begins at the preinvasive stage of vulvar carcinogenesis and suggests the important role of VEGF-C, VEGF-D, VEGFR-3 and LV (D2-40) as prognostic factors in the process of carcinogenesis in the vulvar area.     

Treatment of hypoxia‐dependent cardiovascular diseases by myo‐inositol trispyrophosphate (ITPP)‐enhancement of oxygen delivery by red blood cells

Heart failure is a consequence of progression hypoxia‐dependent tissue damages. Therapeutic approaches to restore and/or protect the healthy cardiac tissue have largely failed and remain a major challenge of regenerative medicine. The myo‐inositol trispyrophosphate (ITPP) is a modifier of haemoglobin which enters the red blood cells and modifies the haemoglobin properties, allowing for easier and better delivery of oxygen by the blood. Here, we show that this treatment approach in an in vivo model of myocardial infarction (MI) results in an efficient protection from heart failure, and we demonstrate the recovery effect on post‐MI left ventricular remodelling in the rat model. Cultured cardiomyocytes used to study the molecular mechanism of action of ITPP in vitro displayed the fast stimulation of HIF‐1 upon hypoxic conditions. HIF‐1 overexpression was prevented by ITPP when incorporated into red blood cells applied in a model of blood‐perfused cardiomyocytes coupling the dynamic shear stress effect to the enhanced O₂ supply by modification of haemoglobin ability to release O₂ in hypoxia. ITPP treatment appears a breakthrough strategy for the efficient and safe treatment of hypoxia‐ or ischaemia‐induced injury of cardiac tissue.     

Colitis Promotes Adaptation of an Intestinal Nematode: A Heligmosomoides Polygyrus Mouse Model System

The precise mechanism of the very effective therapeutic effect of gastrointestinal nematodes on some autoimmune diseases is not clearly understood and is currently being intensively investigated. Treatment with living helminths has been initiated to reverse intestinal immune-mediated diseases in humans. However, little attention has been paid to the phenotype of nematodes in the IBD-affected gut and the consequences of nematode adaptation. In the present study, exposure of Heligmosomoides polygyrus larvae to the changed cytokine milieu of the intestine during colitis reduced inflammation in an experimental model of dextran sulphate sodium (DSS)- induced colitis, but increased nematode establishment in the moderate-responder BALB/c mouse strain. We used mass spectrometry in combination with two-dimensional Western blotting to determine changes in protein expression and changes in nematode antigens recognized by IgG1 in mice with colitis. We show that nematode larvae immunogenicity is changed by colitis as soon as 6 days post-infection; IgG1 did not recognize highly conserved proteins Lev-11 (isoform 1 of tropomyosin α1 chain), actin-4 isoform or FTT-2 isoform a (14-3-3 family) protein. These results indicate that changes in the small intestine provoked by colitis directly influence the nematode proteome. The unrecognized proteins seem to be key antigenic epitopes able to induce protective immune responses. The proteome changes were associated with weak immune recognition and increased larval adaptation and worm growth, altered localization in the intestine and increased survival of males but reduced worm fecundity. In this report, the mechanisms influencing nematode survival and the consequences of changed immunogenicity that reflect the immune response at the site colonized by the parasite in mice with colitis are described. The results are relevant to the use of live parasites to ameliorate IBD.     

Circadian Rhythm of Outside-Nest Activity in Wild (WWCPS), Albino and Pigmented Laboratory Rats

The domestication process of the laboratory rat has been going on for several hundred generations in stable environmental conditions, which may have affected their physiological and behavioural functions, including their circadian system. Rats tested in our ethological experiments were laboratory-bred wild Norway rats (WWCPS), two strains of pigmented laboratory rats (Brown Norway and Long Evans), and two strains of albino rats (Sprague-Dawley and Wistar). Rats were placed in purpose-built enclosures and their cycle of activity (time spent actively outside the nest) has been studied for one week in standard light conditions and for the next one in round-the-clock darkness. The analysis of circadian pattern of outside-nest activity revealed differences between wild, pigmented laboratory, and albino laboratory strains. During daytime, albino rats showed lower activity than pigmented rats, greater decrease in activity when the light was turned on and greater increase in activity when the light was switched off, than pigmented rats. Moreover albino rats presented higher activity during the night than wild rats. The magnitude of the change in activity between daytime and nighttime was also more pronounced in albino rats. Additionaly, they slept outside the nest more often during the night than during the day. These results can be interpreted in accordance with the proposition that intense light is an aversive stimulus for albino rats, due to lack of pigment in their iris and choroid, which reduces their ability to adapt to light. Pigmented laboratory rats were more active during lights on, not only in comparison to the albino, but also to the wild rats. Since the difference seems to be independent of light intensity, it is likely to be a result of the domestication process. Cosinor analysis revealed a high rhythmicity of circadian cycles in all groups.     

The evaluation of the usefulness of pedigree verification-dedicated SNPs for breed assignment in three polish cattle populations

The breed assignment in cattle is one of the issues of molecular genetics which needs further testing and development. Although several statistical approaches have been developed to enable such application, the obtained results strongly depend on specific populations differentiation and power of markers discrimination or their informativeness. Currently, all breeding animals are being tested for parentage with the use of panel of 12 microsatellite markers, which in near future probably will be replaced by about 100 single nucleotide polymorphisms (SNPs). Despite the fact that SNPs are mainly bi-allelic, the multilocus genotypes can reach the level of polymorphism of a panel of microsatellite markers. In this study we attempted to determine the breed of origin of 741 cattle by using 120 SNPs dedicated for parentage testing and included in the BovineSNP50 BeadChip genotyping assay (Illumina). The applied Bayesian and frequency-based methods allowed such differentiation, however, the reliability of the results was not completely satisfying, suggesting that the studied markers are not the best tool for breed assignment.     

A genome-wide detection of selection signatures in conserved and commercial pig breeds maintained in Poland

Background

Identification of selection signatures can provide a direct insight into the mechanism of artificial selection and allow further disclosure of the candidate genes related to the animals’ phenotypic variation. Domestication and subsequent long-time selection have resulted in extensive phenotypic changes in domestic pigs, involving a number of traits, like behavior, body composition, disease resistance, reproduction and coat color. In this study, based on genotypes obtained from PorcineSNP60 Illumina assay we attempt to detect both diversifying and within-breed selection signatures in 530 pigs belonging to four breeds: Polish Landrace, Pu?awska, Z?otnicka White and Z?otnicka Spotted, of which the last three are a subject of conservative breeding and substantially represent the native populations.

Results

A two largely complementary statistical methods were used for signatures detection, including: pairwise F_ST and relative extended haplotype homozygosity (REHH) test. Breed-specific diversifying selection signals included several genes involved in processes connected with fertility, growth and metabolism which are potentially responsible for different phenotypes of the studied breeds. The diversifying selection signals also comprised PPARD gene that was previously found to have a large effect on the shape of the external ear in pigs or two genes encoding neuropeptide Y receptors (Y2 and Y5) involved in fat deposition and stress response which are important features differentiating the studied breeds. REHH statistics allowed detecting several within-breed selection signatures overlapping with genes connected with a range of functions including, among others: metabolic pathways, immune system response or implantation and development of the embryo.

Conclusions

The study provides many potential candidate genes with implication for traits selected in the individual breeds and gives strong basis for further studies aiming at identification of sources of variation among the studied pig breeds.

     

Effects of heme oxygenase-1 on induction and development of chemically induced squamous cell carcinoma in mice

Heme oxygenase-1 (HO-1) is an antioxidative and cytoprotective enzyme, which may protect neoplastic cells against anticancer therapies, thereby promoting the progression of growing tumors. Our aim was to investigate the role of HO-1 in cancer induction. Experiments were performed in HO-1^+/+, HO-1^+/−, and HO-1^−/− mice subjected to chemical induction of squamous cell carcinoma with 7,12-dimethylbenz[a]anthracene and phorbol 12-myristate 13-acetate. Measurements of cytoprotective genes in the livers evidenced systemic oxidative stress in the mice of all the HO-1 genotypes. Carcinogen-induced lesions appeared earlier in HO-1^−/− and HO-1^+/− than in wild-type animals. They also contained much higher concentrations of vascular endothelial growth factor and keratinocyte chemoattractant, but lower levels of tumor necrosis factor-α and interleukin-12. Furthermore, tumors grew much larger in HO-1 knockouts than in the other groups, which was accompanied by an increased rate of animal mortality. However, pathomorphological analysis indicated that HO-1^−/− lesions were mainly large but benign papillomas. In contrast, in mice expressing HO-1, most lesions displayed dysplastic features and developed to invasive carcinoma. Thus, HO-1 may protect healthy tissues against carcinogen-induced injury, but in already growing tumors it seems to favor their progression toward more malignant forms.     

Characterisation and metabolism of astroglia-rich primary cultures from cathepsin K-deficient mice

Abstract Cathepsin K is important for the brain, because its deficiency in mice is associated with a marked decrease in differentiated astrocytes and changes in neuronal patterning in the hippocampus as well as with learning and memory deficits. As cathepsin K activity is most prominent in hippocampal regions of wild type animals, we hypothesised alterations in astrocyte-mediated support of neurons as a potential mechanism underlying the impaired brain functions in cathepsin K-deficient mice. To address this hypothesis, we have generated and characterised astroglia-rich primary cell cultures from cathepsin K-deficient and wild type mice and compared these cultures for possible changes in metabolic support functions and cell composition. Interestingly, cells expressing the oligodendrocytic markers myelin-associated glycoprotein and myelin basic protein were more frequent in astroglia-rich cultures from cathepsin K-deficient mice. However, cell cultures from both genotypes were morphologically comparable and similar with respect to glucose metabolism. In addition, specific glutathione content, glutathione export and γ-glutamyl-transpeptidase activity remained unchanged, whereas the specific activities of glutathione reductase and glutathione-S-transferase were increased by around 50% in cathepsin K-deficient cultures. Thus, lack of cathepsin K in astroglia-rich cultures appears not to affect metabolic supply functions of astrocytes but to facilitate the maturation of oligodendrocytes.