Search for genetic markers of predisposition to psoriasis and psoriatic arthritis

Psoriasis (PS) and psoriatic arthritis (PsA) are interrelated diseases that occur in approximately 30% of patients and are characterized by the presence of a systemic inflammatory reaction that occurs as a result of a violation of the functional state of the immune system. With the advent of new technologies, several new pro-inflammatory cytokines, such as IL-23, IL-31, and IL-33, which play an important role in the pathogenesis of the psoriatic process, have been discovered and characterized. It was determined that single nucleotide polymorphisms (SNPs) in the promoter regions of the IL23, IL31 and IL33 genes play an important role in controlling the expression of relevant cytokines involved in the immunopathogenesis of psoriatic disease. The purpose of the study: to analyze the distribution of genotypes and allelic variants of polymorphisms of the IL23A (rs2066808), IL23R (rs2201841), IL31 (rs7977932) and IL33 (rs7044343), in order to search for genetic markers of predisposition to psoriasis and psoriatic arthritis. Materials and methods. The genotyping of the patients was conducted: psoriasis (PS, n = 77), median age 31.0 years (27.0-43.0), psoriatic arthritis (PsA, n = 99), median age 49.0 years (39.0-56.0) and practically healthy residents of Krasnoyarsk (n = 103), a median age of 32.0 years (24.0-38.0). DNA was isolated from whole venous blood using a standard sorbent kit. Genotyping of single nucleotide polymorphisms IL23A (rs2066808), IL23R (rs2201841), IL31 (rs7977932), IL33 (rs7044343) was carried out using real-time PCR using specific oligonucleotide primers and fluorescentlylabeled probes. Results and discussion. The frequencies of allelic variants of the studied cytokine genes in the control group obtained during the study correspond to their distribution in Caucasoid populations – the alleles IL23A * T, IL23R * T, IL31 * C, IL33 * C prevail. When comparing the distribution frequency of allelic variants of the IL23A, IL23R, IL31, IL33 genes, we did not obtain statistically significant differences between patients and the control group. Conclusions. Despite the fact that when comparing the distribution frequency of allelic variants of the IL23A, IL23R, IL31, IL33 genes, we did not obtain statistically significant differences between the patients and the control group, there are results worthy of attention. So, in patients with PS, the frequency of the C * IL23A allelic variant (rs2066808) is lower than in the population sample, which may indicate its specific role in relation to the development of the disease. All this dictates the need to continue research with the assessment of other SNPs and increase the sample of patients in search of potential genetic markers of psoriatic disease

Genetic markers of children asthma: predisposition to disease course variants

Asthma is a heterogeneous and often difficult to treat condition that results in a disproportionate cost to healthcare systems. Children with severe asthma are at increased risk for adverse outcomes including medication-related side effects, life-threatening exacerbations, and impaired quality of life. An important therapeutic focus is to achieve disease control, which is supposed to involve a personalized approach to treatment of asthma of any severity. Asthma is a multifactorial disease with a significant genetic determinant, however, the inheritance of asthma has not been fully elucidated. Polymorphic genes of inflammatory mediators, including cytokines, play an important role in developing various disease forms. In the current study, large-scale original data on the prevalence of cytokine gene genotypes (IL2, IL4, IL5, IL6, IL10, IL12, IL13, IL17A, IL31, IL33, IFNG, TNFA) among Russian children with asthma in Krasnoyarsk region have been obtained. Genotyping was carried out using real-time PCR. We identified markers predisposing to the development of different variants of the course of childhood asthma: the CT genotype and T allele of IL4 rs2243250 are associated with asthma (p < 0.05), especially in mild asthma and in controlled asthma. The TT genotype and allele T of IL13 rs1800925 are associated with severe and uncontrolled asthma (p < 0.05). The AA genotype of IL17A rs2275913, the TT genotype of IFNG rs2069705 and allelic A variants of TNFA rs1800629 are associated with mild asthma, and the TT genotype of IFNG rs2069705 is additionally associated with controlled asthma. The results obtained will supplement information on the prevalence of polymorphic variants of the cytokine genes in the Russian population and in asthma patients with different disease courses, which is likely to be used in order to shape a plan for Public Health Authority to prevent the development of severe uncontrolled asthma and to optimize personalized therapy

Unequal allelic expression of wild-type and mutated β-myosin in familial hypertrophic cardiomyopathy

Familial hypertrophic cardiomyopathy (FHC) is an autosomal dominant disease, which in about 30% of the patients is caused by missense mutations in one allele of the β-myosin heavy chain (β-MHC) gene (MYH7). To address potential molecular mechanisms underlying the family-specific prognosis, we determined the relative expression of mutant versus wild-type MYH7-mRNA. We found a hitherto unknown mutation-dependent unequal expression of mutant to wild-type MYH7-mRNA, which is paralleled by similar unequal expression of β-MHC at the protein level. Relative abundance of mutated versus wild-type MYH7-mRNA was determined by a specific restriction digest approach and by real-time PCR (RT-qPCR). Fourteen samples from M. soleus and myocardium of 12 genotyped and clinically well-characterized FHC patients were analyzed. The fraction of mutated MYH7-mRNA in five patients with mutation R723G averaged to 66 and 68% of total MYH7-mRNA in soleus and myocardium, respectively. For mutations I736T, R719W and V606M, fractions of mutated MYH7-mRNA in M. soleus were 39, 57 and 29%, respectively. For all mutations, unequal abundance was similar at the protein level. Importantly, fractions of mutated transcripts were comparable among siblings, in younger relatives and unrelated carriers of the same mutation. Hence, the extent of unequal expression of mutated versus wild-type transcript and protein is characteristic for each mutation, implying cis-acting regulatory mechanisms. Bioinformatics suggest mRNA stability or splicing effectors to be affected by certain mutations. Intriguingly, we observed a correlation between disease expression and fraction of mutated mRNA and protein. This strongly suggests that mutation-specific allelic imbalance represents a new pathogenic factor for FHC

Role of IL13 genetic polymorphism in the development of bronchial asthma in children

Bronchial asthma is a multifactorial disease, with both environmental factors and genetic predisposal affecting its development. A number of gene associations have been obtained between polymorphisms of cytokine genes produced by different types of immune cells and asthma development. Interleukin-13 is involved in allergic inflammation, increased bronchial hypersensitivity, regulation of eosinophil levels and IgE production by B cells, thus making it promising for studying IL13 gene polymorphisms in bronchial asthma coupled to development of the disease. The aim of this study was to investigate possible association between asthma and IL13 rs1800925 polymorphism in the children of Caucasian origin in Eastern Siberia. Four groups of patients with asthma were examined (mean age 12.8±1.2 years): with a controlled (n = 95) and uncontrolled course (n = 107), with severe (n = 71) and moderate severity (n = 131) diseases. The control group consisted of healthy individuals: children (n = 33) and adults (n = 102). DNA was isolated with sorbent method; genotyping was carried out using RT-PCR using specific oligonucleotide primers and fluorescent TaqMan probes. The allele and genotype frequencies were compared by the χ-square test using an online calculator. The odds ratio (OR) with a 95% confidence interval (CI) was performed to link genetic markers with pathological phenotypes. The CT IL13 rs1800925 genotype was shown to be associated with moderate asthma and cases of uncontrollable clinical course, whereas the TT genotype was associated with severe asthma. Thus, rs1800925 polymorphism of IL13 gene (the T* variant is known to be associated with increased IL-13 expression) may be associated with bronchial asthma in children. Our data are consistent with results of other authors. E.g., Liu Z. et al. revealed an association between rs1800925 IL13 and the risk of developing asthma in children, with CT and TT genotypes being more common in the patient group. Radhakrishnan A. et al., was studied rs1800925 IL13 in adult population of Malaysia and found that the T* allele frequency in the group of patients significantly exceeds the frequency of this allele in the control group. Thus, the results of our study showed that IL13 rs1800925 polymorphism is associated with bronchial asthma in children, especially, with level of its control and severity of the disease

Myosin Heads Contribute to the Maintenance of Filament Order in Relaxed Rabbit Muscle

Raising the temperature of rabbit skeletal muscle from ∼0°C to ∼20°C has been shown to enhance the helical organization of the myosin heads and to change the intensities of the 10 and 11 equatorial reflections. We show here by time-resolved x-ray diffraction combined with temperature jump that the movement of the heads to enhance the organized myosin helix occurs at the same fast rate as the change in the intensities of the equatorial reflections. However, model calculations indicate that the change in the equatorials cannot be explained simply in terms of the movement of myosin heads. Analysis of electron micrographs of transverse sections of relaxed muscle fibers cryofixed at ∼5°C and ∼35°C shows that in addition to the reorganization of the heads the thin and thick filaments are less constrained to their positions in the hexagonal filament lattice in the warm muscle than in the cold. Incorporating the changes in filament order in model calculations reconciles these with the observed changes in equatorial reflections. We suggest the thin filaments in the cold muscle are boxed into their positions by the thermal movement of the disordered myosin heads. In the warmer muscle, the packed-down heads leave the thin filaments more room to diffuse laterally

De novo designed peptide-based amyloid fibrils

Identification of therapeutic strategies to prevent or cure diseases associated with amyloid fibril deposition in tissue (Alzheimer's disease, spongiform encephalopathies, etc.) requires a rational understanding of the driving forces involved in the formation of these organized assemblies rich in β-sheet structure. To this end, we used a computer-designed algorithm to search for hexapeptide sequences with a high propensity to form homopolymeric β-sheets. Sequences predicted to be highly favorable on this basis were found experimentally to self-associate efficiently into β-sheets, whereas point mutations predicted to be unfavorable for this structure inhibited polymerization. However, the property to form polymeric β-sheets is not a sufficient requirement for fibril formation because, under the conditions used here, preformed β-sheets from these peptides with charged residues form well defined fibrils only if the total net charge of the molecule is ±1. This finding illustrates the delicate balance of interactions involved in the formation of fibrils relative to more disordered aggregates. The present results, in conjunction with x-ray fiber diffraction, electron microscopy, and Fourier transform infrared measurements, have allowed us to propose a detailed structural model of the fibrils

Self-Assembly and Hydrogelation of an Amyloid Peptide Fragment

The self-assembly of a fragment of the amyloid β peptide that has been shown to be critical in amyloid fibrillization has been studied in aqueous solution. There are conflicting reports in the literature on the fibrillization of Aβ (16–20), i.e., KLVFF, and our results shed light on this. In dilute solution, self-assembly of NH2−KLVFF−COOH is strongly influenced by aromatic interactions between phenylalanine units, as revealed by UV spectroscopy and circular dichroism. Fourier transform infrared (FTIR) spectroscopy reveals β-sheet features in spectra taken for more concentrated solutions and also dried films. X-ray diffraction and cryo-transmission electron microscopy (cryo-TEM) provide further support for β-sheet amyloid fibril formation. A comparison of cryo-TEM images with those from conventional dried and negatively stained TEM specimens highlights the pronounced effects of sample preparation on the morphology. A comparison of FTIR data for samples in solution and dried samples also highlights the strong effect of drying on the self-assembled structure. In more concentrated phosphate-buffered saline (PBS) solution, gelation of NH2−KLVFF−COOH is observed. This is believed to be caused by screening of the electrostatic charge on the peptide, which enables β sheets to aggregate into a fibrillar gel network. The rheology of the hydrogel is probed, and the structure is investigated by light scattering and small-angle X-ray scattering