TLR9 -1486T/C and 2848C/T SNPs Are Associated with Human Cytomegalovirus Infection in Infants

Toll-like receptor 9 (TLR9) recognizes non-methylated viral CpG-containing DNA and serves as a pattern recognition receptor that signals the presence of human cytomegalovirus (HCMV). Here, we present the genotype distribution of single-nucleotide polymorphisms (SNPs) of the TLR9 gene in infants and the relationship between TLR9 polymorphisms and HCMV infection. Four polymorphisms (-1237T/C, rs5743836; -1486T/C, rs187084; 1174G/A, rs352139; and 2848C/T, rs352140) in the TLR9 gene were genotyped in 72 infants with symptomatic HCMV infection and 70 healthy individuals. SNP genotyping was performed by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Digested fragments were separated and identified by capillary electrophoresis. The HCMV DNA copy number was measured by a quantitative real-time PCR assay. We found an increased frequency of heterozygous genotypes TLR9 -1486T/C and 2848C/T in infants with HCMV infection compared with uninfected cases. Heterozygous variants of these two SNPs increased the risk of HCMV disease in children (P = 0.044 and P = 0.029, respectively). In infants with a mutation present in at least one allele of -1486T/C and 2848C/T SNPs, a trend towards increased risk of cytomegaly was confirmed after Bonferroni’s correction for multiple testing (Pc = 0.063). The rs352139 GG genotype showed a significantly reduced relative risk for HCMV infection (Pc = 0.006). In contrast, the -1237T/C SNP was not related to viral infection. We found no evidence for linkage disequilibrium with the four examined TLR9 SNPs. The findings suggest that the TLR9 -1486T/C and 2848C/T polymorphisms could be a genetic risk factor for the development of HCMV disease.     

Characterisation of alpha3beta1 and alpha(v)beta3 integrin N-oligosaccharides in metastatic melanoma WM9 and WM239 cell lines

It is well documented that glycan synthesis is altered in some pathological processes, including cancer. The most frequently observed alterations during tumourigenesis are extensive expression of beta1,6-branched complex type N-glycans, the presence of poly-N-acetyllactosamine structures, and high sialylation of cell surface glycoproteins. This study investigated two integrins, alpha3beta1 and alpha(v)beta3, whose expression is closely related to cancer progression. Their oligosaccharide structures in two metastatic melanoma cell lines (WM9, WM239) were analysed with the use of matrix-assisted laser desorption ionisation mass spectrometry. Both examined integrins possessed heavily sialylated and fucosylated glycans, with beta1,6-branches and short polylactosamine chains. In WM9 cells, alpha3beta1 integrin was more variously glycosylated than alpha(v)beta3; in WM239 cells the situation was the reverse. Functional studies (wound healing and ELISA integrin binding assays) revealed that the N-oligosaccharide component of the tested integrins influenced melanoma cell migration on vitronectin and alpha3beta1 integrin binding to laminin-5. Additionally, more variously glycosylated integrins exerted a stronger influence on these parameters. To the best of our knowledge, this is the first report concerning structural characterisation of alpha(v)beta3 integrin glycans in melanoma or in any cancer cells.     

The insulin-degrading enzyme as a link between insulin and neuropeptides metabolism

We have applied a recently developed HPLC-MS enzymatic assay to investigate the cryptic peptides generated by the action of the insulin-degrading enzyme (IDE) on some neuropeptides (NPs) involved in the development of tolerance and dependence to opioids. Particularly, the tested NPs are generated from the NPFF precursor (pro-NPFF (A)): NPFF (FLFQPQRF) and NPAF (AGEGLSSPFWSLAAPQRF). The results show that IDE is able to cleave NPFF and NPAF, generating specific cryptic peptides. As IDE is also responsible for the processing of many other peptides in the brain (amyloid beta protein among the others), we have also performed competitive degradation assays using mixtures of insulin and the above mentioned NPs. Data show that insulin is able to slow down the degradation of both NPs tested, whereas, surprisingly, NPAF is able to accelerate insulin degradation, hinting IDE as the possible link responsible of the mutual influence between insulin and NPs metabolism.     

Pattern of follicular development in high fecundity Olkuska ewes during the estrous cycle

Folliculogenesis was studied daily in the 18 oestrous cycles in six prolific Olkuska ewes from October to December using transrectal ultrasonography to record the number and size of all ovarian follicles > or =2 mm in diameter. Blood samples were taken once a day and were analyzed for concentrations of FSH, LH, estradiol and progesterone. Follicular and hormonal data were analyzed for associations between different stages of development of the follicular waves and concentrations of FSH and estradiol. The first wave during which at least one follicle reached maximum diameter of > or =4 mm after ovulation, was defined as a wave 1, and the following waves were numbered sequentially. Waves 1, 2, 3, 4 and the ovulatory one emerged on days: -2 to 4, 4 to 8, 6 to 11, 10 to 12 and 11 to 15, respectively. The mean number of follicles per wave that reached diameter of > or =4 mm was 4.15 +/- 1.1 and 16.62 +/- 8.6 follicles per estrous cycle of a total 299 follicles were observed. Significantly more follicles (p> or =0.05) emerged on days 2, 8 and 13 than in other days. Serum FSH concentrations fluctuated from 0.11 ngml(-1) on day 2 to preovulatory maximum 1.81 ngml(-1) on day 17 of the estrous cycle. The emergence of follicular waves was associated with elevations of FSH concentrations in blood serum. The mean increase in FSH concentration was followed by the recruitment of follicles of the next wave. The mean daily FSH concentration and the mean number of follicles emerging each day were negatively correlated. The length of the interwave interval (4.4 +/- 1.6 days) did not differ significantly from the interval between pulses of FSH (4.8 +/- 0.3 days). The mean serum estradiol concentrations showed fluctuations until day 14 and then gradually increased from 5.47 +/- 0.3 pgml(-1) to reach a peak 13.14 +/- 0.2 pgml(-1) on the day before ovulation. To summarize, the growth of ovarian follicles during the estrous cycle in high fecundity Olkuska sheep exhibited a distinct wave-like pattern. Ovarian follicles emerged from the pool of 2 mm follicles. The preovulatory follicles originated from the large follicle population were present in the ovary at the time of luteal regression. The initial stages of the growth of the largest follicles appears to be controlled primarily by increases in FSH secretion.     

Genetic distinctness of parthenogenetic forms of European Polydrusus weevils of the subgenus Scythodrusus

Abstract Among eight species of Polydrusus weevils which belong to subgenus Scythodrusus, at least two possess parthenogenetic forms: P. (S.) inustus and P. (S.) pilifer. Both of these species consist of dioecious populations in the Caspian area and of parthenogenetic populations in Eastern Europe (P. (S.) inustus), the Caucasus region (both species) and Middle Asia (P. (S.) pilifer). The origin of parthenogenesis in this subgenus is unresolved; however some data suggest that the parthenogenetic forms are of hybrid ancestry. The genetic distinctness of parthenogenetic Scythodrusus was assessed on the basis of COII, ITS2, EF1‐α and Wolbachiawsp, 16S ribosomal DNA, ftsZ and hcpA sequence comparisons. Both taxa turned out to be monophyletic for all markers, which is an evidence against hybridization of their dioecious ancestors. On the other hand, a high frequency of heterozygous P. (S.) inustus females suggests an origin resulting from hybridization between genetically distinct dioecious representatives of this species. Very similar strains of Wolbachia supergroup A were found in both species, indicating that they have been either inherited from a common ancestor or were transmitted between parthenogenetic Scythodrusus weevils and probably spread randomly across their ranges.     

PCSK9 Plasma Concentrations Are Independent of GFR and Do Not Predict Cardiovascular Events in Patients with Decreased GFR

Background

Impaired renal function causes dyslipidemia that contributes to elevated cardiovascular risk in patients with chronic kidney disease (CKD). The proprotein convertase subtilisin/kexin type 9 (PCSK9) is a regulator of the LDL receptor and plasma cholesterol concentrations. Its relationship to kidney function and cardiovascular events in patients with reduced glomerular filtration rate (GFR) has not been explored.

Methods

Lipid parameters including PCSK9 were measured in two independent cohorts. CARE FOR HOMe (Cardiovascular and Renal Outcome in CKD 2–4 Patients—The Forth Homburg evaluation) enrolled 443 patients with reduced GFR (between 90 and 15 ml/min/1.73 m²) referred for nephrological care that were prospectively followed for the occurrence of a composite cardiovascular endpoint. As a replication cohort, PCSK9 was quantitated in 1450 patients with GFR between 90 and 15 ml/min/1.73 m² enrolled in the Ludwigshafen Risk and Cardiovascular Health Study (LURIC) that were prospectively followed for cardiovascular deaths.

Results

PCSK9 concentrations did not correlate with baseline GFR (CARE FOR HOMe: r = -0.034; p = 0.479; LURIC: r = -0.017; p = 0.512). 91 patients in CARE FOR HOMe and 335 patients in LURIC reached an endpoint during a median follow-up of 3.0 [1.8–4.1] years and 10.0 [7.3–10.6] years, respectively. Kaplan-Meier analyses showed that PCSK9 concentrations did not predict cardiovascular events in either cohort [CARE FOR HOMe (p = 0.622); LURIC (p = 0.729)]. Sensitivity analyses according to statin intake yielded similar results.

Conclusion

In two well characterized independent cohort studies, PCSK9 plasma levels did not correlate with kidney function. Furthermore, PCSK9 plasma concentrations were not associated with cardiovascular events in patients with reduced renal function.     

Angiotensin type 1 receptor antagonist losartan, reduces MPTP-induced degeneration of dopaminergic neurons in substantia nigra

Background

Recent attention has focused on understanding the role of the brain-renin-angiotensin-system (RAS) in stroke and neurodegenerative diseases. Direct evidence of a role for the brain-RAS in Parkinson's disease (PD) comes from studies demonstrating the neuroprotective effect of RAS inhibitors in several neurotoxin based PD models. In this study, we show that an antagonist of the angiotensin II (Ang II) type 1 (AT₁) receptor, losartan, protects dopaminergic (DA) neurons against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity both in primary ventral mesencephalic (VM) cultures as well as in the substantia nigra pars compacta (SNpc) of C57BL/6 mice (Fig. 1).

Results

In the presence of exogenous Ang II, losartan reduced MPP⁺ (5 μM) induced DA neuronal loss by 72% in vitro. Mice challenged with MPTP showed a 62% reduction in the number of DA neurons in the SNpc and a 71% decrease in tyrosine hydroxylase (TH) immunostaining of the striatum, whereas daily treatment with losartan lessened MPTP-induced loss of DA neurons to 25% and reduced the decrease in striatal TH⁺ immunostaining to 34% of control.

Conclusion

Our study demonstrates that the brain-RAS plays an important neuroprotective role in the MPTP model of PD and points to AT₁ receptor as a potential novel target for neuroprotection.     

Assessment of potential application of binary mixtures of 2,4-d with novel aminophosphonates

A series of new aminoalkane- and aminofluorenephosphonates was synthesized for agrochemical application. The particular compounds had different alkyl substituents at the carbon, nitrogen and phosphorus atoms. Their pesticidal activity was checked by applying various experimental methods. These included the measurements of compounds' potency: to inhibit growth of cucumber and germination of white mustard seeds, to influence on the membrane potential of algae and to damage human erythrocyte membranes resulting in hemolysis. All the aminophosphonates were also used in equimolar binary mixtures with the well-known herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), to check, if using such mixtures, the biological efficiencies found for particular compounds could be enhanced due to interactions between aminophosphonates and 2,4-D. The results demonstrated, that depending on the structural features of the compounds, the final effects differed from antagonistic, through additive to the most promising synergistic ones. However, the type of interaction between 2,4-D and the compounds studied found in different experiments was somewhat different. In order to estimate those effects various statistical methods were used (toxic unit method, isobole method).     

Evidence for an increased distribution range of Dermacentor reticulatus in south-west Poland

The expansion of Dermacentor reticulatus to new geographical areas has been observed in several countries in Europe, including Poland and it’s neighbors. In 2011 and 2012, a total of 148 host-seeking D. reticulatus ticks were collected after flagging the vegetation in Lower Silesia, south-western Poland. Tick monitoring was conducted in mixed and deciduous forest, on meadows, in river valleys and ecotones between forested and grassy areas. The ornate dog ticks were found in 10 out of 33 sites located in five districts: Legnica, Wroclaw, ?roda ?l?ska, Lubin and Boleslawiec County. All sites where D. reticulatus ticks were found are located to the south-west of the Odra River. The greatest distance between these disconnected localities was approximately 90 km. It seems that at present the southern boundary of the range is Wroclaw district. This study indicates that D. reticulatus can be ranked as a typical element of the fauna in Lower Silesia in southwestern Poland.