Significant genetic differentiation between Poland and Germany follows present-day political borders, as revealed by Y-chromosome analysis

To test for human population substructure and to investigate human population history we have analysed Y-chromosome diversity using seven microsatellites (Y-STRs) and ten binary markers (Y-SNPs) in samples from eight regionally distributed populations from Poland (n=913) and 11 from Germany (n=1,215). Based on data from both Y-chromosome marker systems, which we found to be highly correlated (r=0.96), and using spatial analysis of the molecular variance (SAMOVA), we revealed statistically significant support for two groups of populations: (1) all Polish populations and (2) all German populations. By means of analysis of the molecular variance (AMOVA) we observed a large and statistically significant proportion of 14% (for Y-SNPs) and 15% (for Y-STRs) of the respective total genetic variation being explained between both countries. The same population differentiation was detected using Monmoniers algorithm, with a resulting genetic border between Poland and Germany that closely resembles the course of the political border between both countries. The observed genetic differentiation was mainly, but not exclusively, due to the frequency distribution of two Y-SNP haplogroups and their associated Y-STR haplotypes: R1a1*, most frequent in Poland, and R1*(xR1a1), most frequent in Germany. We suggest here that the pronounced population differentiation between the two geographically neighbouring countries, Poland and Germany, is the consequence of very recent events in human population history, namely the forced human resettlement of many millions of Germans and Poles during and, especially, shortly after World War II. In addition, our findings have consequences for the forensic application of Y-chromosome markers, strongly supporting the implementation of population substructure into forensic Y chromosome databases, and also for genetic association studies.     

TLR9 2848 GA Heterozygotic Status Possibly Predisposes Fetuses and Newborns to Congenital Infection with Human Cytomegalovirus

Background

Some single nucleotide polymorphisms (SNP), located in Toll-like receptor (TLR) genes, were reported to be associated with human cytomegalovirus (HCMV) infections. The study was aimed to assess the correlation of SNPs at TLR4 and TLR9 genes with the occurrence of congenital cytomegaly, based on available samples.

Methods

Reported case-control study included both HCMV infected and non-infected fetuses and newborns. The specimens were classified to the molecular analyses, based on serological features of the recent infection and HCMV DNAemia in body fluids. TLR SNPs were studied, using multiplex nested PCR-RFLP assay, and determined genotypes were confirmed by sequencing. Hardy-Weinberg equilibrium was assessed for the identified genotypes. The linkage disequilibrium was also estimated for TLR4 SNPs. A relationship between the status of TLR genotypes and congenital cytomegaly development was estimated, using a logistic regression model.

Results

Hardy Weinberg equilibrium was observed for almost all SNPs, both infected and non-infected patients, with exception of TLR4 896 A>G polymorphism in the control group (P≤0.050). TLR4 896 A>G and 1196 C>T SNPs were found in linkage disequilibrium in both study groups (P≤0.050). The CC genotype at TLR4 1196 SNP and the GA variant at TLR9 2848 G>A SNP were significantly associated with HCMV infection (P≤0.050). The risk of congenital cytomegaly was higher in heterozygotes at TLR9 SNP than in the carriers of other genotypic variants at the reported locus (OR 4.81; P≤0.050). The GC haplotype at TLR4 SNPs and GCA variants at TLR4 and TLR9 SNPs were significantly associated with HCMV infection (P≤0.0001). The ACA variants were more frequent among fetuses and neonates with symptomatic, rather than asymptomatic cytomegaly (P≤0.0001).

Conclusions

TLR4 and TLR9 polymorphisms may contribute to the development of congenital infection with HCMV in fetuses and neonates. The TLR9 2848 GA heterozygotic status possibly predisposes to HCMV infection, increasing the risk of congenital cytomegaly development.     

Effects of genetically modified human skin fibroblasts,stably overexpressing hepatocyte growth factor,on hepatic functions of cocultured C3A cells

Diseases leading to terminal hepatic failure are among the most common causes of death worldwide. Transplant of the whole organ is the only effective method to cure liver failure. Unfortunately, this treatment option is not available universally due to the serious shortage of donors. Thus, alternative methods have been developed that are aimed at prolonging the life of patients, including hepatic cells transplantation and bridging therapy based on hybrid bioartificial liver devices. Parenchymal liver cells are highly differentiated and perform many complex functions, such as detoxification and protein synthesis. Unfortunately, isolated hepatocytes display a rapid decline in viability and liver‐specific functions. A number of methods have been developed to maintain hepatocytes in their highly differentiated state in vitro, amongst them the most promising being 3D growth scaffolds and decellularized tissues or coculture with other cell types required for the heterotypic cell‐cell interactions. Here we present a novel approach to the hepatic cells culture based on the feeder layer cells genetically modified using lentiviral vector to stably produce additional amounts of hepatocyte growth factor and show the positive influence of these coculture conditions on the preservation of the hepatic functions of the liver parenchymal cells' model—C3A cells.     

Assessment of the Anticonvulsant Potency of Ursolic Acid in Seizure Threshold Tests in Mice

Ursolic acid (UA) is a plant derived compound which is also a component of the standard human diet. It possesses a wide range of pharmacological properties, i.e., antioxidant, anti-inflammatory, antimicrobial and antitumor, which have been used in folk medicine for centuries. Moreover, influence of UA on central nervous system-related processes, i.e., pain, anxiety and depression, was proved in experimental studies. UA also revealed anticonvulsant properties in animal models of epilepsy and seizures. The aim of the present study was to investigate the influence of UA on seizure thresholds in three acute seizure models in mice, i.e., the 6 Hz-induced psychomotor seizure threshold test, the maximal electroshock threshold (MEST) test and the timed intravenous pentylenetetrazole (iv PTZ) infusion test. We also examined its effect on the muscular strength (assessed in the grip strength test) and motor coordination (estimated in the chimney test) in mice. UA at doses of 50 and 100 mg/kg significantly increased the seizure thresholds in the 6 Hz and MEST tests. The studied compound did not influence the seizure thresholds in the iv PTZ test. Moreover, UA did not affect the motor coordination and muscular strength in mice. UA displays only a weak anticonvulsant potential which is dependent on the used seizure model.     

EPR Oximetry Sensor—Developing a TAM Derivative for In Vivo Studies

Oxygenation is one of the most important physiological parameters of biological systems. Low oxygen concentration (hypoxia) is associated with various pathophysiological processes in different organs. Hypoxia is of special importance in tumor therapy, causing poor response to treatment. Triaryl methyl (TAM) derivative radicals are commonly used in electron paramagnetic resonance (EPR) as sensors for quantitative spatial tissue oxygen mapping. They are also known as magnetic resonance imaging (MRI) contrast agents and fluorescence imaging compounds. We report the properties of the TAM radical tris(2,3,5,6-tetrachloro-4-carboxy-phenyl)methyl, (PTMTC), a potential multimodal (EPR/fluorescence) marker. PTMTC was spectrally analyzed using EPR and characterized by estimation of its sensitivity to the oxygen in liquid environment suitable for intravenous injection (1?mM PBS, pH?=?7.4). Further, fluorescent emission of the radical was measured using the same solvent and its quantum yield was estimated. An in vitro cytotoxicity examination was conducted in two cancer cell lines, HT-29 (colorectal adenocarcinoma) and FaDu (squamous cell carcinoma) and followed by uptake studies. The stability of the radical in different solutions (PBS pH?=?7.4, cell media used for HT-29 and FaDu cells culturing and cytotoxicity procedure, full rat blood and blood plasma) was determined. Finally, a primary toxicity test of PTMTC was carried out in mice. Results of spectral studies confirmed the multimodal properties of PTMTC. PTMTC was demonstrated to be not absorbed by cancer cells and did not interfere with luciferin-luciferase based assays. Also in vitro and in vivo tests showed that it was non-toxic and can be freely administrated till doses of 250?mg/kg BW via both i.v. and i.p. injections. This work illustrated that PTMTC is a perfect candidate for multimodal (EPR/fluorescence) contrast agent in preclinical studies.     

Exploring the cocrystallization potential of urea and benzamide

The cocrystallization landscape of benzamide and urea interacting with aliphatic and aromatic carboxylic acids was studied both experimentally and theoretically. Ten new cocrystals of benzamide were synthesized using an oriented samples approach via a fast dropped evaporation technique. Information about types of known bi-component cocrystals augmented with knowledge of simple binary eutectic mixtures was used for the analysis of virtual screening efficiency among 514 potential pairs involving aromatic carboxylic acids interacting with urea or benzamide. Quantification of intermolecular interaction was achieved by estimating the excess thermodynamic functions of binary liquid mixtures under supercooled conditions within a COSMO-RS framework. The smoothed histograms suggest that slightly more potential pairs of benzamide are characterized in the attractive region compared to urea. Finally, it is emphasized that prediction of cocrystals of urea is fairly direct, while it remains ambiguous for benzamide paired with carboxylic acids. The two known simple eutectics of urea are found within the first two quartiles defined by excess thermodynamic functions, and all known cocrystals are outside of this range belonging to the third or fourth quartile. On the contrary, such a simple separation of positive and negative cases of benzamide miscibility in the solid state is not observed. The difference in properties between urea and benzamide R2,2(8) heterosynthons is also documented by alterations of substituent effects. Intermolecular interactions of urea with para substituted benzoic acid analogues are stronger compared to those of benzamide. Also, the amount of charge transfer from amide to aromatic carboxylic acid and vice versa is more pronounced for urea. However, in both cases, the greater the electron withdrawing character of the substituent, the higher the binding energy, and the stronger the supermolecule polarization via the charge transfer mechanism.     

Stem cells as therapy for cardiac disease - a review

Acute myocardial infarction (AMI) is one of the most significant causes of morbidity and mortality worldwide. Stem cells represent an enormous chance to rebuild damaged heart tissue. Correct definition of the cardiac progenitors is necessary to understand heart development, and would pave the way for the use of cardiac progenitors in the treatment of heart disease. Identifying, purifying and differentiating native cardiac progenitor cells are indispensable if we are to overcome congenital and adult cardiac diseases. To understand their functions, physiology and action, cells are tested in animal models, and then in clinical trials. But because clinical trials yield variable results, questions about proper cardiac stem cells remain unanswered. Transplanted stem cells release soluble factors, acting in a paracrine fashion, which contributes to cardiac regeneration. Cytokines and growth factors have cytoprotective and neovascularizing functions, and may activate resident cardiac stem cells. Understanding all these mechanisms is crucial to overcoming heart diseases.     

A Mutation in the Mitochondrial Fission Gene Dnm1l Leads to Cardiomyopathy

Mutations in a number of genes have been linked to inherited dilated cardiomyopathy (DCM). However, such mutations account for only a small proportion of the clinical cases emphasising the need for alternative discovery approaches to uncovering novel pathogenic mutations in hitherto unidentified pathways. Accordingly, as part of a large-scale N-ethyl-N-nitrosourea mutagenesis screen, we identified a mouse mutant, Python, which develops DCM. We demonstrate that the Python phenotype is attributable to a dominant fully penetrant mutation in the dynamin-1-like (Dnm1l) gene, which has been shown to be critical for mitochondrial fission. The C452F mutation is in a highly conserved region of the M domain of Dnm1l that alters protein interactions in a yeast two-hybrid system, suggesting that the mutation might alter intramolecular interactions within the Dnm1l monomer. Heterozygous Python fibroblasts exhibit abnormal mitochondria and peroxisomes. Homozygosity for the mutation results in the death of embryos midway though gestation. Heterozygous Python hearts show reduced levels of mitochondria enzyme complexes and suffer from cardiac ATP depletion. The resulting energy deficiency may contribute to cardiomyopathy. This is the first demonstration that a defect in a gene involved in mitochondrial remodelling can result in cardiomyopathy, showing that the function of this gene is needed for the maintenance of normal cellular function in a relatively tissue-specific manner. This disease model attests to the importance of mitochondrial remodelling in the heart; similar defects might underlie human heart muscle disease.     

A sensitive and specific HPLC method for the determination of total pentosidine concentration in plasma

Pentosidine is an advanced glycation end-product (AGE) appearing when arginine and lysine residues in proteins are cross-linked with carbonyl derivatives. This paper presents an improved method for the synthesis of pentosidine and reversed-phase chromatography of this substance with fluorometric detection that enables sensitive (0.01 pmol/mg protein) and specific determination of pentosidine in plasma. Separation is done twice on the same C(18) Vydac 218TP54 column, first with trifluoroacetic acid and next with heptafluorobutyric acid as ion pair. The inter-day coefficient of variation is 6.4% at pentosidine concentration in plasma of 25 pmol/mg protein and 8% at 1.7 pmol/mg protein. Spectral properties of pentosidine exploited during identification of the substance with UV absorption and fluorescence detectors are described. Maximum of absorbance was observed at 325 nm, maximum fluorescence at lambda(ex)/lambda(em)=330/373 nm. The method may prove useful for the study of processes associated with generation and accumulation of pentosidine in the body as a marker of AGE production in healthy subjects and patients with chronic renal failure.