Masks of Albinism: Clinical Spectrum of Hermansky-Pudlak Syndrome.

Hermansky-Pudlak syndrome (HPS) is a rare disease inherited in the autosomal recessive mode, including 11 clinical genetic subtypes. They are associated with impaired function of the BLOC protein complex (Biogenesis of Lysosome-related Organelles Complexes), and the subunits of the AP-3 complex (adaptor protein complex). Each has its own clinical features, but they are all characterized by albinism, bleeding disorder, and visual abnormalities. Eleven patients from eight unrelated families with an incoming diagnosis of albinism were examined and novel and previously described genetic variants in HPS1, HPS6, and BLOC1S6 genes (types HPS1, HPS6, and HPS9) were found. To determine the optimal therapy and recommendations for further follow up, it is necessary to consider the entire clinical spectrum and genetic polymorphism of the disease. An interdisciplinary approach, combined with the use of non-routine diagnostic techniques such as RNA analysis, is essential for achieving accurate diagnoses in certain complex cases.

Dose-Response Effect of Various Concentrations of Cl-Containing Water-Soluble Derivatives of C60 Fullerenes on a Selective Regulation of Gene Expression in Human Embryonic Lung Fibroblasts (HELF).

BACKGROUND: The new synthesized water-soluble derivatives of C60 fullerenes are of a great interest to researchers since they can potentially be promising materials for drug delivery, bioimaging, biosonding, and tissue engineering. Surface functionalization of fullerene derivatives changes their chemical and physical characteristics, increasing their solubility and suitability for different biological systems applications, however, any changes in functionalized fullerenes can modulate their cytotoxicity and antioxidant properties. The toxic or protective effect of fullerene derivatives on cells is realized through the activation or inhibition of genes and proteins of key signaling pathways in cells responsible for regulation of cellular reactive oxygen species (ROS) level, proliferation, and apoptosis. METHODS: The 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay was used to assess cells viability. Flow cytometry analyses was applied to measure proteins levels in human embryonic lung fibroblasts (HELF) cells. HELF is a standard, stable and well described human cell line that can be passaged many times. Quantitation of ROS was assessed using H2DCFH-DA. Fluorescence images were obtained using microscopy. Expression of BCL2, CCND1, CDKN2A, BRCA1, BAX, NFKB1, NOX4, NRF2, TBP (reference gene) was analyzed using real-time Polymerase chain reaction (PCR). RESULTS: We found that high and low concentrations of fullerene C60 derivatives with the five residues of potassium salt of 6-(3-phenylpropanamido)hexanoic (F1) or 6-(2-(thiophen-2-yl)acetamido)hexanoic (F2) acid and a chlorine atom attached directly to the cage cause diametrically opposite activation of genes and proteins of key signaling pathways regulating the level of oxidative stress and apoptosis in HELF. High concentrations of F1 and F2 have a genotoxic effect, causing NADPH oxidase 4 (NOX4) expression activation in 24-72 hours (2-4 fold increase), ROS synthesis induction (increase by 30-40%), DNA damage and breaks (2-2.5 fold 8-oxodG level increases), and activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) (by 40-80%) against the background of reduced NF-E2-related factor 2 (NRF2) expression (by 20-45%). Low concentrations of F1 and F2 produced a cytoprotective effect: in 24-72 hours they reduce the oxidative DNA damage (by 20-40%), decrease the number of double-strand DNA breaks (by 20-30%), increase the level of anti-apoptotic proteins and enhance the antioxidant response activating the NRF2 expression (NRF2 gene expression increases 1.5-2.3 fold, phosphorylated form of the NRF2 protein increases 2-3 fold). CONCLUSIONS: Obtained results show that in low doses studied fullrens may serve as perspective DNA protectors against the damaging genotoxic factors.

The Study of the Inheritance Mechanisms of Myotonic Dystrophy Type 1 (DM1) in Families from the Republic of North Ossetia-Alania.

Myotonic dystrophy type 1 (DM1) is a multisystem disorder with progressive myopathy and myotonia. The clinical study was conducted in the Republic of North Ossetia-Alania (RNOA), and in it 39 individuals from 17 unrelated families were identified with DM1. Clinical presentations varied, including muscle weakness, fatigue, intellectual disability, hypersomnia, ophthalmological abnormalities, and alopecia. Using clinical and genotyping data, we confirmed the diagnosis and enabled the study of CTG-repeat anticipation and DM1 prevalence in the Ossetian and Ingush populations. CTG expansion correlated with age of onset, with clinical severity, and with offspring showing more severe symptoms than parents. In many families, the youngest child had a more severe DM1 phenotype than older siblings. The prevalence was 14.17 per 100,000 in Ossetians and 18.74 per 100,000 in Ingush people, aligning with global data. Segregation analysis showed a higher frequency of maternal transmission. The study highlights the clinical and genetic heterogeneity of DM1 and its dependence on repeat expansion and paternal and maternal age.

Presentation of Rare Phenotypes Associated with the FKBP10 Gene.

Pathogenic variants in the FKBP10 gene lead to a spectrum of rare autosomal recessive phenotypes, including osteogenesis imperfecta (OI) Type XI, Bruck syndrome Type I (BS I), and the congenital arthrogryposis-like phenotype (AG), each with variable clinical manifestations that are crucial for diagnosis. This study analyzed the clinical-genetic characteristics of patients with these conditions, focusing on both known and newly identified FKBP10 variants. We examined data from 15 patients, presenting symptoms of OI and joint contractures. Diagnostic methods included genealogical analysis, clinical assessments, radiography, whole exome sequencing, and direct automated Sanger sequencing. We diagnosed 15 patients with phenotypes due to biallelic FKBP10 variants-4 with OI Type XI, 10 with BS I, and 1 with the AG-like phenotype-demonstrating polymorphism in disease severity. Ten pathogenic FKBP10 variants were identified, including three novel ones, c.1373C>T (p.Pro458Leu), c.21del (p.Pro7fs), and c.831_832insCG (p.Gly278Argfs), and a recurrent variant, c.831dup (p.Gly278Argfs). Variant c.1490G>A (p.Trp497Ter) was found in two unrelated patients, causing OI XI in one and BS I in the other. Additionally, two unrelated patients with BS I and epidermolysis bullosa shared identical homozygous FKBP10 and KRT14 variants. This observation illustrates the diversity of FKBP10-related pathology and the importance of considering the full spectrum of phenotypes in clinical diagnostics.

Genetic Landscape and Clinical Features of Hyperphenylalaninemia in North Ossetia-Alania: High Frequency of P281L and P211T Genetic Variants in the PAH Gene.

This study, conducted in the Republic of North Ossetia-Alania (RNOA), aimed to explore the genetic landscape of hyperphenylalaninemia (HPA) and phenylketonuria (PKU) in the Ossetian population using data from newborn screening (NBS). Through comprehensive molecular genetic analysis of 29 patients with HPA from diverse ethnic backgrounds, two major genetic variants in the PAH gene, P281L and P211T, were identified, constituting 50% of all detected pathogenic alleles in Ossetian patients. Remarkably, these variants exhibited an exceptionally high frequency in the Ossetian population, surpassing global prevalence rates. This study unveiled a notable prevalence of mild forms of HPA (78%), underscoring the importance of genetic counseling for carriers of pathogenic variants in the PAH gene. Moreover, the findings emphasized the necessity for ongoing monitoring of patients with mild forms, as they may lack significant symptoms for diagnosis, potentially impacting offspring. Overall, this research offers valuable insights into the genetic landscape of HPA and PKU in the Ossetian population.

Epidemiology of Spinal Muscular Atrophy Based on the Results of a Large-Scale Pilot Project on 202,908 Newborns.

BACKGROUND: This study presents the findings of a newborn screening (NBS) pilot project for 5q-spinal muscular atrophy (5q-SMA) in multiple regions across Russia for during the year 2022. The aim was to assess the feasibility and reproducibility of NBS for SMA5q in diverse populations and estimate the real prevalence of 5q-SMA in Russia as well as the distribution of patients with different number of SMN2 copies. METHODS: The pilot project of NBS here was based on data, involving the analysis of 202,908 newborns. SMA screening assay was performed using a commercially available real-time polymerase chain reaction kit, the Eonis SCID-SMA. RESULTS: In one year, 202,908 newborns were screened, identifying 26 infants with homozygous deletion of SMN1 exon 7, yielding an estimated 5q-SMA incidence of 1:7804 newborns. It was found that 38.46% had two SMN2 copies, 42.31% had three copies, 15.38% had four copies, and 3.85% had five copies of SMN2. Immediate treatment was proposed for patients with two or three SMN2 copies. Infants with four or more SMN2 copies warranted further investigation on management and treatment. Short-term monitoring after gene therapy showed motor function improvements. Delays in treatment initiation were observed, including the testing for adeno-associated virus 9 antibodies and nonmedical factors. CONCLUSIONS: The study emphasizes the need for a standardized algorithm for early diagnosis and management through NBS to benefit affected families. Overall, the NBS program for 5q-SMA in Russia demonstrated the potential to improve outcomes and transform SMA from a devastating disease to a chronic condition with evolving medical requirements.

Newborn Screening for Severe T and B Cell Lymphopenia Using TREC/KREC Detection: A Large-Scale Pilot Study of 202,908 Newborns.

Newborn screening (NBS) for severe inborn errors of immunity (IEI), affecting T lymphocytes, and implementing measurements of T cell receptor excision circles (TREC) has been shown to be effective in early diagnosis and improved prognosis of patients with these genetic disorders. Few studies conducted on smaller groups of newborns report results of NBS that also include measurement of kappa-deleting recombination excision circles (KREC) for IEI affecting B lymphocytes. A pilot NBS study utilizing TREC/KREC detection was conducted on 202,908 infants born in 8 regions of Russia over a 14-month period. One hundred thirty-four newborns (0.66‰) were NBS positive after the first test and subsequent retest, 41% of whom were born preterm. After lymphocyte subsets were assessed via flow cytometry, samples of 18 infants (0.09‰) were sent for whole exome sequencing. Confirmed genetic defects were consistent with autosomal recessive agammaglobulinemia in 1/18, severe combined immunodeficiency - in 7/18, 22q11.2DS syndrome - in 4/18, combined immunodeficiency - in 1/18 and trisomy 21 syndrome - in 1/18. Two patients in whom no genetic defect was found met criteria of (severe) combined immunodeficiency with syndromic features. Three patients appeared to have transient lymphopenia. Our findings demonstrate the value of implementing combined TREC/KREC NBS screening and inform the development of policies and guidelines for its integration into routine newborn screening programs.

Complex Chromosomal Rearrangement Involving Chromosomes 10 and 11, Accompanied by Two Adjacent 11p14.1p13 and 11p13p12 Deletions, Identified in a Patient with WAGR Syndrome.

Three years ago, our patient, at that time a 16-month-old boy, was discovered to have bilateral kidney lesions with a giant tumor in the right kidney. Chemotherapy and bilateral nephron-sparing surgery (NSS) for Wilms tumor with nephroblastomatosis was carried out. The patient also had eye affection, including glaucoma, eye enlargement, megalocornea, severe corneal swelling and opacity, complete aniridia, and nystagmus. The diagnosis of WAGR syndrome was suspected. De novo complex chromosomal rearrangement with balanced translocation t(10,11)(p15;p13) and a pericentric inversion inv(11)(p13q12), accompanied by two adjacent 11p14.1p13 and 11p13p12 deletions, were identified. Deletions are raised through the complex molecular mechanism of two subsequent rearrangements affecting chromosomes 11 and 10. WAGR syndrome diagnosis was clinically and molecularly confirmed, highlighting the necessity of comprehensive genetic testing in patients with congenital aniridia and/or WAGR syndrome.

Epigenomic Profiling Advises Therapeutic Potential of Leukotriene Receptor Inhibitors for a Subset of Triple-Negative Breast Tumors.

Triple-negative breast cancer (TNBC) is the most aggressive molecular subtype, with a poor survival rate compared to others subtypes. For a long time, chemotherapy was the only systemic treatment for TNBC, and the identification of actionable molecular targets might ultimately improve the prognosis for TNBC patients. We performed a genome-wide analysis of DNA methylation at CpG islands on a collection of one hundred ten breast carcinoma samples and six normal breast tissue samples using reduced representation bisulfite sequencing with the XmaI restriction enzyme (XmaI-RRBS) and identified a subset of TNBC samples with significant hypomethylation at the LTB4R/LTB4R2 genes' CpG islands, including CpG dinucleotides covered with cg12853742 and cg21886367 HumanMethylation 450K microarray probes. Abnormal DNA hypomethylation of this region in TNBC compared to normal samples was confirmed by bisulfite Sanger sequencing. Gene expression generally anticorrelates with promoter methylation, and thus, the promoter hypomethylation detected and confirmed in our study might be revealed as an indirect marker of high LTB4R/LTB4R2 expression using a simple methylation-sensitive PCR test. Analysis of RNA-seq expression and DNA methylation data from the TCGA dataset demonstrates that the expression of the LTB4R and LTB4R2 genes significantly negatively correlates with DNA methylation at both CpG sites cg12853742 (R = -0.4, p = 2.6 × 10-6; R = -0.21, p = 0.015) and cg21886367 (R = -0.45, p = 7.3 × 10-8; R = -0.24, p = 0.005), suggesting the upregulation of these genes in tumors with abnormal hypomethylation of their CpG island. Kaplan-Meier analysis using the TCGA-BRCA gene expression and clinical data revealed poorer overall survival for TNBC patients with an upregulated LTB4R. To this day, only the leukotriene inhibitor LY255283 has been tested on an MCF-7/DOX cell line, which is a luminal A breast cancer molecular subtype. Other studies compare the effects of Montelukast and Zafirlukast (inhibitors of the cysteinyl leukotriene receptor, which is different from LTB4R/LTB4R2) on the MDA-MB-231 (TNBC) cell line, with high methylation and low expression levels of LTB4R. In our study, we assess the therapeutic effects of various drugs (including leukotriene receptor inhibitors) with the DepMap gene effect and drug sensitivity data for TNBC cell lines with hypomethylated and upregulated LTB4R/LTB4R2 genes. LY255283, Minocycline, Silibinin, Piceatannol, Mitiglinide, 1-Azakenpaullone, Carbetocin, and Pim-1-inhibitor-2 can be considered as candidates for the additional treatment of TNBC patients with tumors demonstrating LTB4R/LTB4R2 hypomethylation/upregulation. Finally, our results suggest that the epigenetic status of leukotriene B4 receptors is a novel, potential, predictive, and prognostic biomarker for TNBC. These findings might improve individualized therapy for TNBC patients by introducing new therapeutic adjuncts as anticancer agents.

Epidemiology of PAX6 Gene Pathogenic Variants and Expected Prevalence of PAX6-Associated Congenital Aniridia across the Russian Federation: A Nationwide Study.

This study investigates the distribution of PAX6-associated congenital aniridia (AN) and WAGR syndrome across Russian Federation (RF) districts while characterizing PAX6 gene variants. We contribute novel PAX6 pathogenic variants and 11p13 chromosome region rearrangements to international databases based on a cohort of 379 AN patients (295 families, 295 probands) in Russia. We detail 100 newly characterized families (129 patients) recruited from clinical practice and specialized screening studies. Our methodology involves multiplex ligase-dependent probe amplification (MLPA) analysis of the 11p13 chromosome, PAX6 gene Sanger sequencing, and karyotype analysis. We report novel findings on PAX6 gene variations, including 67 intragenic PAX6 variants and 33 chromosome deletions in the 100 newly characterized families. Our expanded sample of 295 AN families with 379 patients reveals a consistent global PAX6 variant spectrum, including CNVs (copy number variants) of the 11p13 chromosome (31%), complex rearrangements (1.4%), nonsense (25%), frameshift (18%), and splicing variants (15%). No genetic cause of AN is defined in 10 patients. The distribution of patients across the Russian Federation varies, likely due to sample completeness. This study offers the first AN epidemiological data for the RF, providing a comprehensive PAX6 variants spectrum. Based on earlier assessment of AN prevalence in the RF (1:98,943) we have revealed unexamined patients ranging from 55% to 87%, that emphases the need for increased awareness and comprehensive diagnostics in AN patient care in Russia.

Major Contribution of c.[1622T>C;3113C>T] Complex Allele and c.5882G>A Variant in ABCA4-Related Retinal Dystrophy in an Eastern European Population.

Inherited retinal diseases (IRDs) constitute a prevalent group of inherited ocular disorders characterized by marked genetic diversity alongside moderate clinical variability. Among these, ABCA4-related eye pathology stands as a prominent form affecting the retina. In this study, we conducted an in-depth analysis of 96 patients harboring ABCA4 variants in the European part of Russia. Notably, the complex allele c.[1622T>C;3113C>T] (p.Leu541Pro;Ala1038Val, or L541P;A1038V) and the variant c.5882G>A (p.Gly1961Glu or G1961E) emerged as primary contributors to this ocular pathology within this population. Additionally, we elucidated distinct disease progression characteristics associated with the G1961E variant. Furthermore, our investigation revealed that patients with loss-of-function variants in ABCA4 were more inclined to develop phenotypes distinct from Stargardt disease. These findings provide crucial insights into the genetic and clinical landscape of ABCA4-related retinal dystrophies in this specific population.

Co-Occurrence of Congenital Aniridia Due to Nonsense PAX6 Variant p.(Cys94*) and Chromosome 21 Trisomy in the Same Patient.

This study aims to present a clinical case involving the unique co-occurrence of congenital aniridia and Down syndrome in a young girl and to analyze the combined impact of these conditions on the patient's phenotype. The investigation involved comprehensive pediatric and ophthalmological examinations alongside karyotyping and Sanger sequencing of the PAX6 gene. The patient exhibited distinctive features associated with both congenital aniridia and Down syndrome, suggesting a potential exacerbation of their effects. Cytogenetic and molecular genetic analysis revealed the presence of trisomy 21 and a known pathogenic nonsense variant in exon 6 of the PAX6 gene (c.282C>A, p.(Cys94*)) corresponding to the paired domain of the protein. The observation of these two hereditary anomalies offers valuable insights into the molecular pathogenetic mechanisms underlying each condition. Additionally, it provides a basis for a more nuanced prognosis of the complex disease course in this patient. This case underscores the importance of considering interactions between different genetic disorders in clinical assessments and treatment planning.

Extracellular vesicles of human glial cells exert neuroprotective effects via brain miRNA modulation in a rat model of traumatic brain injury.

Stem cell-based therapeutic approaches for neurological disorders are widely studied. Paracrine factors secreted by stem cells in vitro and delivered intranasally might allow bypassing the disadvantages associated with a surgical cell delivery procedure with likely immune rejection of a transplant. In this study, we investigated the therapeutic effect of the extracellular vesicles secreted by glial progenitor cells (GPC-EV) derived from human induced pluripotent stem cell in a traumatic brain injury model. Intranasal administration of GPC-EV to Wistar rats for 6 days improved sensorimotor functions assessed over a 14-day observation period. Beside, deep sequencing of microRNA transcriptome of GPC-EV was estimate, and was revealed 203 microRNA species that might be implicated in prevention of various brain pathologies. Modulation of microRNA pools might contribute to the observed decrease in the number of astrocytes that inhibit neurorecovery processes while enhancing neuroplasticity by decreasing phosphorylated Tau forms, preventing inflammation and apoptosis associated with secondary damage to brain tissue. The course of GPC-EV administration was promoted the increasing protein levels of NF-κB in studied areas of the rat brain, indicating NF-κB dependent mechanisms as a plausible route of neuroprotection within the damaged area. This investigation showed that GPC-EV may be representing a therapeutic approach in traumatic brain injury, though its translation into the clinic would require an additional research and development.

Therapeutic Efficiency of Proteins Secreted by Glial Progenitor Cells in a Rat Model of Traumatic Brain Injury.

Traumatic brain injuries account for 30-50% of all physical traumas and are the most common pathological diseases of the brain. Mechanical damage of brain tissue leads to the disruption of the blood-brain barrier and the massive death of neuronal, glial, and endothelial cells. These events trigger a neuroinflammatory response and neurodegenerative processes locally and in distant parts of the brain and promote cognitive impairment. Effective instruments to restore neural tissue in traumatic brain injury are lacking. Glial cells are the main auxiliary cells of the nervous system, supporting homeostasis and ensuring the protection of neurons through contact and paracrine mechanisms. The glial cells' secretome may be considered as a means to support the regeneration of nervous tissue. Consequently, this study focused on the therapeutic efficiency of composite proteins with a molecular weight of 5-100 kDa secreted by glial progenitor cells in a rat model of traumatic brain injury. The characterization of proteins below 100 kDa secreted by glial progenitor cells was evaluated by proteomic analysis. Therapeutic effects were assessed by neurological outcomes, measurement of the damage volume by MRI, and an evaluation of the neurodegenerative, apoptotic, and inflammation markers in different areas of the brain. Intranasal infusions of the composite protein product facilitated the functional recovery of the experimental animals by decreasing the inflammation and apoptotic processes, preventing neurodegenerative processes by reducing the amounts of phosphorylated Tau isoforms Ser396 and Thr205. Consistently, our findings support the further consideration of glial secretomes for clinical use in TBI, notably in such aspects as dose-dependent effects and standardization.

Antipsychotics Affect Satellite III (1q12) Copy Number Variations in the Cultured Human Skin Fibroblasts.

The fragment of satellite III (f-SatIII) is located in pericentromeric heterochromatin of chromosome 1. Cell with an enlarged f-SatIII block does not respond to various stimuli and are highly stress-susceptible. The fraction of f-SatIII in the cells of schizophrenia patients changed during antipsychotic therapy. Therefore, antipsychotics might reduce the f-SatIII content in the cells. We studied the action of haloperidol, risperidone and olanzapine (3 h, 24 h, 96 h) on human skin fibroblast lines (n = 10). The f-SatIII contents in DNA were measured using nonradioactive quantitative hybridization. RNASATIII were quantified using RT-qPCR. The levels of DNA damage markers (8-oxodG, γ-H2AX) and proteins that regulate apoptosis and autophagy were determined by flow cytometry. The antipsychotics reduced the f-SatIII content in DNA and RNASATIII content in RNA from HSFs. After an exposure to the antipsychotics, the autophagy marker LC3 significantly increased, while the apoptosis markers decreased. The f-SatIII content in DNA positively correlated with RNASATIII content in RNA and with DNA oxidation marker 8-oxodG, while negatively correlated with LC3 content. The antipsychotics arrest the process of f-SatIII repeat augmentation in cultured skin fibroblasts via the transcription suppression and/or through upregulated elimination of cells with enlarged f-SatIII blocks with the help of autophagy.

DNA Methylation and Prospects for Predicting the Therapeutic Effect of Neoadjuvant Chemotherapy for Triple-Negative and Luminal B Breast Cancer.

Despite advances in the diagnosis and treatment of breast cancer (BC), the main cause of deaths is resistance to existing therapies. An approach to improve the effectiveness of therapy in patients with aggressive BC subtypes is neoadjuvant chemotherapy (NACT). Yet, the response to NACT for aggressive subtypes is less than 65% according to large clinical trials. An obvious fact is the lack of biomarkers predicting the therapeutic effect of NACT. In a search for epigenetic markers, we performed genome-wide differential methylation screening by XmaI-RRBS in cohorts of NACT responders and nonresponders, for triple-negative (TN) and luminal B tumors. The predictive potential of the most discriminative loci was further assessed in independent cohorts by methylation-sensitive restriction enzyme quantitative PCR (MSRE-qPCR), a promising method for the implementation of DNA methylation markers in diagnostic laboratories. The selected most informative individual markers were combined into panels demonstrating cvAUC = 0.83 (TMEM132D and MYO15B markers panel) for TN tumors and cvAUC = 0.76 (TTC34, LTBR and CLEC14A) for luminal B tumors. The combination of methylation markers with clinical features that correlate with NACT effect (clinical stage for TN and lymph node status for luminal B tumors) produces better classifiers, with cvAUC = 0.87 for TN tumors and cvAUC = 0.83 for luminal B tumors. Thus, clinical characteristics predictive of NACT response are independently additive to the epigenetic classifier and in combination improve prediction.

Genetic Heterogeneity of X-Linked Ichthyosis in the Republic of North Ossetia-Alania, Case Series Report.

North Caucasus has always been a residence of a lot of different authentic ethnic groups speaking different languages and still living their traditional lifestyle. The diversity appeared to be reflected in the accumulation of different mutations causing common inherited disorders. X-linked ichthyosis represents the second most common form of genodermatoses after ichthyosis vulgaris. Eight patients from three unrelated families of different ethnic origin, Kumyk, Turkish Meskhetians, and Ossetian, with X-linked ichthyosis from the North Caucasian Republic of North Ossetia-Alania were examined. NGS technology was implied for searching for disease-causing variants in one of the index patients. Known pathogenic hemizygous deletion in the short arm of chromosome X encompassing the STS gene was defined in the Kumyk family. A further analysis allowed us to establish that likely the same deletion was a cause of ichthyosis in a family belonging to the Turkish Meskhetians ethnic group. In the Ossetian family, a likely pathogenic nucleotide substitution in the STS gene was defined; it segregated with the disease in the family. We molecularly confirmed XLI in eight patients from three examined families. Though in two families, Kumyk and Turkish Meskhetian, we revealed similar hemizygous deletions in the short arm of chromosome X, but their common origin was not likely. Forensic STR markers of the alleles carrying the deletion were defined to be different. However, here, common alleles haplotype is hard to track for a high local recombination rate. We supposed the deletion could arise as a de novo event in a recombination hot spot in the described and in other populations with a recurrent character. Defined here are the different molecular genetic causes of X-linked ichthyosis in families of different ethnic origins sharing the same residence place in the Republic of North Ossetia-Alania which could point to the existing reproductive barriers even inside close neighborhoods.

An Unusual Presentation of Novel Missense Variant in PAX6 Gene: NM_000280.4:c.341A>G, p.(Asn114Ser).

This study investigates a unique and complex eye phenotype characterized by minimal iris defects, foveal hypoplasia, optic nerve coloboma, and severe posterior segment damage. Through genetic analysis and bioinformatic tools, a specific nonsynonymous substitution, p.(Asn114Ser), within the PAX6 gene's paired domain is identified. Although this substitution is not in direct contact with DNA, its predicted stabilizing effect on the protein structure challenges the traditional understanding of PAX6 mutations, suggesting a gain-of-function mechanism. Contrary to classical loss-of-function effects, this gain-of-function hypothesis aligns with research demonstrating PAX6's dosage sensitivity. Gain-of-function mutations, though less common, can lead to diverse phenotypes distinct from aniridia. Our findings emphasize PAX6's multifaceted influence on ocular phenotypes and the importance of genetic variations. We contribute a new perspective on PAX6 mutations by suggesting a potential gain-of-function mechanism and showcasing the complexities of ocular development. This study sheds light on the intricate interplay of the genetic alterations and regulatory mechanisms underlying complex eye phenotypes. Further research, validation, and collaboration are crucial to unravel the nuanced interactions shaping ocular health and development.

In Vitro Analysis of Biological Activity of Circulating Cell-Free DNA Isolated from Blood Plasma of Schizophrenic Patients and Healthy Controls-Part 2: Adaptive Response.

Oxidized in vitro genomic DNA (gDNA) is known to launch an adaptive response in human cell cultures. The cfDNA extracted from the plasma of schizophrenic patients (sz-cfDNA) and healthy controls (hc-cfDNA) contains increased amounts of 8-oxodG, a DNA-oxidation marker. The aim of the research was answering a question: can the human cfDNA isolated from blood plasma stimulate the adaptive response in human cells? In vitro responses of ten human skin fibroblasts (HSFs) and four peripheral blood mononuclear cell (PBMC) lines after 1-24 h of incubation with sz-cfDNA, gDNA and hc-cfDNA containing different amounts of 8-oxodG were examined. Expressions of RNA of eight genes (NOX4, NFE2L2, SOD1, HIF1A, BRCA1, BRCA2, BAX and BCL2), six proteins (NOX4, NRF2, SOD1, HIF1A, γH2AX and BRCA1) and DNA-oxidation marker 8-oxodG were analyzed by RT-qPCR and flow cytometry (when analyzing the data, a subpopulation of lymphocytes (PBL) was identified). Adding hc-cfDNA or sz-cfDNA to HSFs or PBMC media in equal amounts (50 ng/mL, 1-3 h) stimulated transient synthesis of free radicals (ROS), which correlated with an increase in the expressions of NOX4 and SOD1 genes and with an increase in the levels of the markers of DNA damage γH2AX and 8-oxodG. ROS and DNA damage induced an antioxidant response (expression of NFE2L2 and HIF1A), DNA damage response (BRCA1 and BRCA2 gene expression) and anti-apoptotic response (changes in BAX and BCL2 genes expression). Heterogeneity of cells of the same HSFs or PBL population was found with respect to the type of response to (sz,hc)-cfDNA. Most cells responded to oxidative stress with an increase in the amount of NRF2 and BRCA1 proteins along with a moderate increase in the amount of NOX4 protein and a low amount of 8-oxodG oxidation marker. However, upon the exposure to (sz,hc)-cfDNA, the size of the subpopulation with apoptosis signs (high DNA damage degree, high NOX4 and low NRF2 and BRCA1 levels) also increased. No significant difference between the responses to sz-cfDNA and hc-cfDNA was observed. Sz-cfDNA and hc-cfDNA showed similarly high bioactivity towards fibroblasts and lymphocytes. Conclusion: In cultured human cells, hc-cfDNA and sz-cfDNA equally stimulated an adaptive response aimed at launching the antioxidant, repair, and anti-apoptotic processes. The mediator of the development of the adaptive response are ROS produced by, among others, NOX4 and SOD1 enzymes.

Ribosomal DNA Abundance in the Patient's Genome as a Feasible Marker in Differential Diagnostics of Autism and Childhood-Onset Schizophrenia.

Introduction: Differential diagnostics of early-onset schizophrenia and autism spectrum disorders (ASD) are a problem of child psychiatry. The prognosis and relevant treatment are to a large degree determined by the correctness of diagnosis. We found earlier that leucocyte DNA of adult schizophrenia patients contained significantly larger copy numbers of ribosomal repeats (rDNA) coding for rRNA, than DNA of mentally healthy controls. Aim: To compare the contents of ribosomal repeats in the leucocyte DNA of children with schizophrenia, children with ASD, and healthy age-matched controls to estimate the possibility of using this genetic trait in the differential diagnostics of the two types of disorders. Patients and methods: Blood samples of patients with infantile autism (A­F84.0 according to ICD-10, N = 75) and with childhood-onset schizophrenia (SZ­F20.8 according to ICD-10, N = 43) were obtained from the Child Psychiatry Department of the Mental Health Research Center. The healthy control blood samples (HC, N = 86) were taken from the Research Centre for Medical Genetics collection. The recruitment of cases was based on the clinical psychopathologic approach. DNA was extracted from blood leukocytes with organic solvents. Nonradioactive quantitative hybridization technique was applied for determining the abundance of ribosomal repeats in the genomes. Statistical processing was performed using StatPlus, Statgraphics and MedCalc. Findings: DNA derived from SZ cases contained 565 ± 163 rDNA copies, which is significantly (p < 10−6) higher than the rDNA content in ASD cases (405 ± 109 copies) and controls (403 ± 86 copies). The HC and A groups did not differ by rDNA copy number (p > 0.4). The genetic trait "rDNA copy number in patient's genome" can potentially be applied as an additional marker in differential diagnostics of childhood-onset schizophrenia and autism spectrum disorders.