Effects of genetic variation in the 20th intron of Sansui duck ATP2A2 gene on eggshell quality

In order to explore the influence of the polymorphism in the 20 intron region of the Sansui duck ATP2A2 gene on the eggshell quality, this study used Primer Premier 5 software to design and synthesize a pair of primers in the 20 intron region, two-way direct sequencing and sequence alignment to mine SNPs Sites, SPSS 18.0 software was used to analyze the relationship between SNP sites and eggshell quality of Sansui duck. Results Three SNP sites were found in the 20 intron region of the ATP2A2 gene: g.40874 T>C, g.40920 G>A and g.40990 T=C, all of which were moderately polymorphic, at the site g.40874 T ＞C and g.40920 G＞A both deviated significantly from Hardy-Weinberg equilibrium (P>0.05), position g.40990 T=C accords with Hardy-Weinberg equilibrium (P<0.05), and position g.40874 T＞C There is a strong linkage disequilibrium between g.40990 T=C; a total of 4 haplotypes and 9 double types were detected at 3 SNP loci; the results of association analysis show that g.40874 T＞C mutation has an effect on eggshell strength The eggshell strength of CC genotype was significantly higher than that of TC and TT genotypes (P<0.05), and the eggshell weight of CC genotype was significantly higher than that of TC genotype (P<0.05), g. The 40990 T=C mutation has a significant effect on the eggshell strength, and the eggshell strength of the TC genotype is significantly higher than that of the TT genotype (P<0.05). In summary, the g.40874 T>C and g.40990 T=C found in the 20th intron region of the Sansui duck ATP2A2 gene may be the marker sites that affect the quality of the eggshell.

The association of the expression of calcium handling proteins and their polymorphic variants of genes with the structural and functional state of the heart of patients with atrial fibrillation

Funding Acknowledgements Type of funding sources: Public Institution(s). Main funding source(s): The Russian Science Foundation, the Cardiology Research Institute, Tomsk National Research Medical Center of the Russian Academy of Sciences. Background. The cellular mechanisms of the genesis of atrial fibrillation are associated with impaired intracellular transport of calcium ions, which is determined by the activity of calcium-transporting proteins of the sarcoplasmic reticulum (SR). The aim of the study to investigate the expression of calcium handling proteins of the sarcoplasmic reticulum and their polymorphic variants of genes with the structural and functional state of the heart of patients with permanent atrial fibrillation. Methods. The study included 45 patients aged 29 to 65 years with atrial fibrillation. A complete echocardiographic study was performed according to standard views (parasternal and apical views) by an En Visor CHD Philips sonograph (Netherlands). All patients underwent catheter ablation, during which a biopsy of myocardial tissue from the top of the left ventricle (1-3 mg) was taken. The protein content of SERCA2a and calsequestrin (CASQ2) was determined by immunoblotting in patient’s myocardium. We identified polymorphic variants rs1860561 of the SERCA2a and rs6684209 and rs7521023 of the CASQ2 gene by real-time polymerase chain reaction. Results. The expression of both SERCA2a and CASQ2 proteins correlated with the size of the left atrium (LA). Thus, a higher level of SERCA2a expression in the myocardium corresponded to a larger LA size in patients. Despite the fact that the LV ejection fraction did not correlate with the expression level of the studied proteins, a direct correlation was found between the SERCA2a level and the values of LV end-diastolic and systolic volumes. In addition, the hemodynamic parameters of the heart, characterizing the diastolic function of the heart of patients, such as the rates of early (peak E) and late diastolic filling (peak A) LV were lower in patients with a higher SERCA2a level, although the ratio of these parameters (peak E/peak A) had no significant differences. In the homozygous GG genotype of the ATP2A2 gene, the expression level of the SERCA2a protein was (p = 0.039) higher than in patients with the heterozygous genotype (GA). Analysis of the parameters of the ECHO-KG study showed that the presence of the A allele of the rs1860561 variant of the ATP2A2 gene is associated with an increase in the LV sphericity index. The expression of the CASQ2 protein in patients with CC genotype of the rs6684209 variant of the CASQ2 gene was 2.5 times higher than in patients with CT genotype. The genotypes of the rs7521023 variant of the CASQ2 gene were not associated with the level of expression of the corresponding protein in the myocardium of the studied patients. Conclusions. The different genotypes of the rs1860561 variant of the ATP2A2 gene and rs6684209 of the CASQ2 gene can modulate the expression level of the SERCA2a protein and the CASQ2 protein. Moreover, the expression of calcium handling proteins to a greater extent affects the functional and structural characteristics of the heart of patients with permanent AF.

Relationship between the expression level of genes of Ca(2+)-transporting proteins of the cardiomyocytes' sarcoplasmic reticulum with the severity of heart failure

Introduction The sarcoplasmic reticulum of cardiomyocytes is the main Ca(2+) ion depot, the main functional proteins of which are Ca(2+)-ATPase SERCA2a, which re-traps ions from myoplasma, calsequestrin CASQ2, which binds most of Ca(2+), and ryanodine receptors RyR2 releasing Ca(2+) from the intracellular depot. A change in their functional activity can determine the progression of contractile myocardial dysfunction. The effectiveness of the Ca(2+)-transporting system and, consequently, the risk of development and progression of heart failure, may depend on the expression of the corresponding genes. Purpose To evaluate the association between the level of relative expression of the ATP2A2, CASQ2, RYR2 genes in the myocardium and the clinical parameters of heart failure in patients with coronary heart disease. Methods The study was carried out on the material of 90 patients with chronic heart failure developed on the background of coronary heart disease. Myocardial samples (a fragment of the right atrium appendage) obtained by connecting the cardiopulmonary bypass during planned coronary bypass surgery. The level of expression of the Ca(2+)-ATPase ATP2A2 gene, the ryanodine receptor RYR2 gene, and the calsequestrin CASQ2 gene was estimated. Samples were homogenized, RNA was isolated and cDNA was synthesized, and real-time PCR was performed. As reference, the glyceraldehyde-3-phosphate dehydrogenase GAPDH gene, the beta-actin ACTB gene, and the 18S gene were used. The level of gene expression was calculated automatically using the software of the thermocycler. For the analysis of quantitative data, the Kruskel-Wallis test or the Mann-Whitney test was used. The analysis of the strength of the linear relationship was carried out using the Spearman rank correlation coefficient. Results ATP2A2 gene expression was reduced in the myocardium of patients with left ventricular (LV) hypertrophy (p=0.027 for ATP2A2/GAPDH, p=0.043 for ATP2A2/ACTB, p=0.039 for ATP2A2/18S). A correlation was observed between an increase in the functional class of heart failure (NYHA) and a decrease in the expression level of the RYR2/GAPDH gene (p=0.023). Among patients with heart failure, there was a weak negative linear relationship between the expression level of CASQ2/18S and the LV ejection fraction (r=−0.288, p=0.047). A significant (p=0.040) increase in CASQ2/ACTB gene expression was also found in patients with diastolic dysfunction compared to individuals without it, as well as a similar tendency for CASQ2/GAPDH (p=0.068) and CASQ2/18S (p=0.090). Conclusion In patients with heart failure due to coronary heart disease with LV hypertrophy, there was a decrease in the expression of the Ca (2+)-ATPase ATP2A2 gene. A decrease in the expression of the ryanodine receptor RYR2 gene was found as the heart failure class worsened. However, CASQ2 expression increased with diastolic dysfunction and a decrease in LV ejection fraction. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Russian Foundation for Basic Research

ATP2A2 SINE Insertion in an Irish Terrier with Darier Disease and Associated Infundibular Cyst Formation

A 4-month-old female Irish Terrier presented with a well demarcated ulcerative and crusting lesion in the right ear canal. Histological analysis revealed epidermal hyperplasia with severe acantholysis affecting all suprabasal layers of the epidermis, which prompted a presumptive diagnosis of canine Darier disease. The lesion was successfully treated by repeated laser ablation of the affected epidermis. Over the course of three years, the dog additionally developed three dermal nodules of up to 4 cm in diameter that were excised and healed without complications. Histology of the excised tissue revealed multiple infundibular cysts extending from the upper dermis to the subcutis. The cysts were lined by squamous epithelium, which presented with abundant acantholysis of suprabasal keratinocytes. Infundibular cysts represent a novel finding not previously reported in Darier patients. Whole genome sequencing of the affected dog was performed, and the functional candidate genes for Darier disease (ATP2A2) and Hailey-Hailey disease (ATP2C1) were investigated. The analysis revealed a heterozygous SINE insertion into the ATP2A2 gene, at the end of intron 14, close to the boundary of exon 15. Analysis of the ATP2A2 mRNA from skin of the affected dog demonstrated a splicing defect and marked allelic imbalance, suggesting nonsense-mediated decay of the resulting aberrant transcripts. As Darier disease in humans is caused by haploinsufficiency of ATP2A2, our genetic findings are in agreement with the clinical and histopathological data and support the diagnosis of canine Darier disease.

Darier’s Disease: Report of a Case with Facial Involvement

Darier’s disease is a relatively rare autosomal dominant genodermatosis with a defect in the desmosomal attachment due to a mutation in the ATP2A2 gene. The condition is characterized by multiple hyperkeratotic papules predominantly in seborrheic areas on the head, neck, and trunk, with less frequent involvement of the oral mucosa. Histopathologically, the lesions reveal suprabasal clefts in the epithelium with acantholytic and dyskeratotic cells. Facial involvement in Darier’s disease is one of the common presenting features. However, it has been once reported in a severe, chronic form as leonine facies in a long-standing case. To raise awareness of facial involvement in Darier’s disease, we herein report a 65-year-old female patient with prominent facial lesions.