Multiple epiphyseal dysplasias (MED) are a clinically and genetically heterogeneous group of skeletal dysplasias with a predominant lesion in the epiphyses of tubular bones. Variants in the SLC26A2 gene cause their autosomal recessive form (rMED or MED type 4). The accumulation of data regarding the genotype–phenotype correlation can help in the diagnosis and proper management of these patients. The aim of this study was to survey the clinical and genetic characteristics of 55 patients with MED type 4 caused by variants in the SLC26A2 gene. Diagnosis confirmation was carried out by radiography and custom panel sequencing consisting of 166 genes responsible for the development of hereditary skeletal pathology. This was followed by the validation of the identified variants using automated Sanger sequencing (for six patients) and the direct automatic Sanger sequencing of the coding sequence and the adjacent intron regions of the SLC26A2 gene for 49 patients. Based on the clinical and genetic analysis of our sample of patients, two main MED type 4 phenotypes with early and late clinical manifestations were identified. An early and more severe form of the disease was observed in patients with the c.835C > T variant (p.Arg279Trp), and the late and milder form of the disease was observed in patients with the c.1957T > A variant (p.Cys653Ser) in the homozygous or compound heterozygous state with c.26 + 2T > C. It was also shown that only three pathogenic variants were found in 95.3% of the alleles of Russian patients with MED type 4: c.1957T > A (p.Cys653Ser), c.835C > T (p.Arg279Trp), and c.26 + 2T > C; thus, it can be assumed that the primary analysis of these variants will contribute to the optimal molecular genetic diagnostics of MED type 4.

Pathogenic variants in the RPE65 gene cause the only known form of inherited retinal degenerations (IRDs) that are prone to gene therapy. The current study is aimed at the evaluation of the prevalence of RPE65-associated retinopathy in the Russian Federation, the characterization of known variants in the RPE65 gene, and the establishment of the specificities of the mutation spectrum in Russian patients. Methods: The analysis was carried out on blood samples obtained from 1053 non-related IRDs patients. The analysis, which consisted of 211 genes, was carried out based on the method of massive parallel sequencing (MPS) for all probands. Variant validation, as well as biallelic status verification, were carried out using direct automated Sanger sequencing. The number of copies of RPE65 exons 1–14 was analyzed with quantitative MLPA using an MRC-Holland SALSA MLPA probemix. Results: Out of 1053 non-related patients, a molecular genetic diagnosis of IRDs has been confirmed in 474 cases, including 25 (5.3%) patients with RPE65-associated retinopathy. We detected 26 variants in the RPE65 gene, nine of which have not been previously described in the literature. The most common mutations in the Russian population were c.304G>T/p.(Glu102*), c.370C>T/p.(Arg124*), and c.272G>A/p.(Arg91Gln), which comprised 41.8% of all affected chromosomes. Conclusions: The current study shows that pathogenic variants in the RPE65 gene contribute significantly to the pathogenesis of IRDs and comprise 5.3% of all patients with a confirmed molecular genetic diagnosis. This study allowed for the formation of a cohort for target therapy of the disorder; such therapy has already been carried out for some patients.

Relevance. The oral hygiene and prevention quality depends on toothpaste. The literature analysis shows the underuse of saliva biochemical parameters for assessing oral hygiene product effectiveness.Materials and methods. Thirty-three subjects aged 19 to 22 years participated in the study. The participants used four kinds of toothpaste by the same manufacturer to evaluate the toothpaste’s effectiveness. We conducted clinical examinations and collected saliva samples on the 1st, 14th and 28th days of the study. The saliva was collected on an empty stomach without stimulation in the morning. Then it was centrifuged. The supernatant was biochemically studied.Results. The study established the criteria for the toothpaste’s low effectiveness, which involve a signifcant increase in hygienic indices and total antioxidant activity, associated with a statistically signifcant trend towards an increase in lactate content. The criteria for a toothpaste with medium effectiveness are a signifcant decrease in one of the hygienic indices, associated with an alkaline pH shift, and an increase in the total antioxidant activity, without signifcant changes in the lactate content. The criteria for a highly effective toothpaste are a signifcant decrease in hygienic indices, and an alkaline pH shift, associated with a signifcant lactate decrease. Determination of total calcium and inorganic phosphorus in the oral fluid may be crucial for assessing the toothpaste remineralizing properties. The glucose content in mixed saliva yields little information for the toothpaste’s hygienic effectiveness assessment.Conclusion. We can recommend saliva biochemical parameters for assessing the oral hygiene status and the effectiveness of oral hygiene products during oral hygiene product comprehensive studies.

В статье представлен алгоритм разработки пошаговой методики получения наночастиц серебра размером 30-35 нм с учетом установления на каждом этапе необходимого соотношения концентраций и объемов реагентов. В работе были использованы нитрат серебра и цитрат натрия безводный фирмы «Sigma-Aldrich» (США). Все исследуемые растворы были приготовлены весовым методом. Для взвешивания реагентов использовали электронные аналитические весы Adventurer («OHAUS», США) (с дискретностью 0,0001 г). Оценка размеров и плотности наночастиц серебра проводилась с помощью электронного трансмиссионного микроскопа JEM-1011 («Jeol», Япония). В ходе исследования была установлена прямая зависимость размера наночастиц от концентраций растворов цитрата натрия и нитрата серебра. По результатам первого этапа для отработки синтеза наночастиц коллоидного серебра было выбрано соотношение концентраций AgNO₃ : Na₃C₆H₅O₇ – 1 : 0,75. На втором этапе проводилась оценка влияния соотношений объемов реагентов на кондиционность полученных серий: наилучший результат был получен в серии с соотношением объемов AgNO₃ : Na₃C₆H₅O₇ – 5 : 1, где наблюдались единичные конгломераты, а форма частиц была правильной и округлой. На следующем этапе экспериментальным путем было определено оптимальное время кипячения коллоидного раствора после появления светло-желтой окраски: при увеличении времени кипячения раствора происходит стабилизация частиц, тем самым наблюдается агрегативная устойчивость. Был подобран наиболее соответствующий режим перемешивания реагентов с использованием магнитной мешалки: после появлении светло-желтой окраски изменялось количество оборотов с 375 до 500 об/мин и уменьшался температурный режим с 300 °С до 200 °С.

Pathogenic variants in the USH2A gene are the primary cause of both non-syndromic autosomal recessive inherited retinitis pigmentosa (RP) and the syndromic form, characterized by retinal degeneration and sensorineural hearing loss. This study presents a comparative assessment of the genetic variant spectrum in the USH2A gene among Russian patients in two clinical groups. A retrospective analysis was conducted on massive parallel panel sequencing data from 2415 blood samples of unrelated patients suspected of having hereditary retinal diseases. The copy number of USH2A exons was determined using the quantitative MLPA method with the MRC-Holland SALSA MLPA kit. Biallelic pathogenic and likely pathogenic variants in the USH2A gene were identified in 69 patients (8.7%). In the group of patients with isolated hereditary RP (55 patients), the most frequent pathogenic variants were p.(Glu4445_Ser4449delinsAspLeu) (20.9%), p.(Trp3955*) (15.5%), and p.(Cys934Trp) (5.5%). In patients with the syndromic form (14 patients), the most frequent variants were p.(Trp3955*) (35.7%) and c.8682-9A>G (17.9%). It was found that patients with isolated vision impairment rarely had two “null” variants (17.8%), whereas this was common among patients with both hearing and vision impairment (71.4%) (p ≤ 0.05), explaining the severity of the disease and the earlier onset of clinical symptoms in the syndromic form of RP. Ten previously undescribed loss-of-function variants were identified. The estimated prevalence of USH2A-associated retinal dystrophy in Russia was 1.9 per 100,000 individuals. The obtained data on the differences in the spectra of genetic variants in the USH2A gene in the two studied groups highlight the importance of establishing genotype–phenotype correlations and predicting disease severity, aiming at potential early cochlear implantation and selection of target therapy.

Inherited retinal diseases (IRDs) are a clinically heterogeneous group of retinal pathologies associated with vision loss due to dysfunction or degeneration of photoreceptor and retinal pigment epithelium. Autosomal recessive forms of IRDs account for more than 55% of all diseases in this group on average worldwide. This study presents data on frequent pathogenic and likely pathogenic variants in recessive genes of IRDs obtained from a retrospective analysis of high-throughput sequencing data from a large Russian cohort of patients with suspected inherited nonsyndromic retinal pathology. Data from 1470 unrelated patients were analyzed. Pathogenic and likely pathogenic variants were identified in the zygosity required for the development of the disease in 643 patients (43.74%). It was found that nine genes (ABCA4, CNGB3, USH2A, RPE65, CRB1, CNGA3, CEP290, GUCY2D, PDE6H) account for 73.3% of all molecularly confirmed cases of IRDs in Russian patients. An analysis of the spectrum of nucleotide variants of these genes was carried out, and 17 variants were identified that occur with an allelic frequency of more than 1% for each gene. In light of the data obtained, diagnostic systems based on the multiplex ligation-dependent probe amplification reaction (MLPA) were developed. The informativity of the two systems for diagnosing autosomal recessive nonsyndromic forms of inherited retinal diseases is 16.4%, the informativity for all forms of nonsyndromic retinal diseases exceeds 7%. For a group of patients with achromatopsia, a study using one of the systems will make it possible to establish a diagnosis in 62.5% of cases.