Clinical significance of correlation between persistent pulmonary hypertension and 8-hydroxy-2-desoxyguanosine level in premature newborns.

BACKGROUND: Persistent pulmonary hypertension is a problem that leads to high morbidity and mortality in preterm infants. In clinical studies, oxidative stress (OS) contributes to the development of pulmonary hypertension (PH). The most specific biomarker of OS in preterm infants is urinary 8-hydroxy-2-deoxyguanosine (8-OHdG).The aim of the study was to determine the clinical correlation between the value of 8-OHdG and the level of a mean pressure in the pulmonary artery (mPAP) in premature infants with respiratory distress syndrome (RDS) and asphyxia in the early neonatal period. METHODS: Determination of the urinary 8-OHdG value and PH in 96 premature infants born at gestational age of 26-32 weeks on the 1st and the 3rd-5th days of life in two groups: group I -52 children with respiratory distress syndrome; II -44 children with RDS associated with perinatal asphyxia. RESULTS: The 2nd group of children had higher average mPAP level, mmHg, both in the 1st and in the 3rd-5th day of life compared with the 1st group. The value of the urinary 8-OHdG correlated with the manifestation of PH that required prolonged respiratory support in group II. CONCLUSION: Perinatal asphyxia in preterm infants with RDS on the 1st day of life complicates the course of PH, as indicated by a higher level of the urinary 8-OHdG and correlated to mPAP. Gender characteristics of the dynamics of 8-OHdG levels in children with perinatal pathology reveal reduced adaptability and reactivity of boys to OS at birth.

[THE ROLE OF SEXUAL DYMORPHISM AND THE STATE OF THE ARTERIAL DUCT IN THE FORMATION OF OXIDATIVE STRESS DISEASES IN PRETERM INFANTS].

Oxidative stress is a state of imbalance between the production of reactive oxygen species and their inactivation by the body's antioxidant systems. Premature babies are susceptible to oxidative stress due to an imperfect antioxidant system. Oxygen resuscitation, respiratory therapy, surfactant use, and other intensive nursing techniques increase the production of reactive oxygen species and increase oxidative stress. The placenta of male and female fetuses have different expression of proteins and genes, which is especially important when exposed to unfavorable factors, because creates the conditions for a survival advantage for female fetuses. The aim of the study was to study the features of the formation of oxidative stress diseases in preterm infants, depending on gender. The results of the management of 324 preterm infants at 24-36 weeks of gestation were analyzed. At the 1st stage of the study, in order to identify the significance of sexual dimorphism in the formation of oxidative stress diseases, the observed children were divided into 2 groups depending on gender. And at the 2nd stage, in order to identify the patterns of the formation of oxidative stress diseases in preterm infants, depending on the state of the ductus arteriosus and gender, each of the groups was divided into 2 subgroups (A and B) depending on the state of the ductus arteriosus: subgroup A - children with the ductus arteriosus spontaneously closed in the neonatal period, and subgroup B - children in whom the ductus arteriosus remained open throughout the child's stay in the perinatal center, including those with a PDA that required surgical closure. At the 1st stage of the study, both groups were relevant in terms of the frequency and severity of respiratory distress syndrome, the frequency of use of surfactant therapy, the frequency and duration of respiratory oxygen therapy, the frequency and severity of BPD, ROP, the frequency of PVL, long-term persistence of the ductus arteriosus and the formation of hemodynamically significant PDA. Group I in comparison with II had a significantly longer gestation period 30,0±0,2 vs 29,2±0,2 (p=0,009), body weight 1429,2±33,6 vs 1230.7±34,8 (p=0,0001) and a higher Apgar score at the 1st minute of life 4,2±0,1 vs 3,8±0,1 (p=0,016). At the 2nd stage of the study, severe RDS was observed significantly more often in group I-B compared with I-A: 59,8% vs 37,4% (p=0,001). The need for respiratory therapy was observed significantly more often among girls with spontaneously closed PDA: in group II-A - 97,2% compared with I-A - 90,9% (p=0,040) and with II-B - 91,9% (p=0,024). The duration of respiratory support was significantly shorter in the group of boys with spontaneously closed ductus arteriosus: in group I-A - 11,8±1,5 compared with II-A - 18,0±2,3 (p=0,019) and with I-B - 17,9±1,9 (p=0,014). The frequency of diagnosis and a severe BPD were recorded among boys with PDA (in group I-B compared with group I-A): 55,0% vs 34,1% (p=0,002) and 21,8% vs 3,3% (p=0,004), respectively. A significantly higher prevalence of ROP, regardless of gender, was observed in the groups of children with spontaneously closed PDA: among boys 89,8% vs 55,0% (p=0.000), among girls 91,7% vs 61,3% (p=0.000). The highest prevalence of grade 3 or more ROP was observed in children with PDA regardless of gender: among boys 43,6% vs 29,1% (p=0,042), among girls 52,6% vs 34,8 (p=0,038). The same incidence of oxidative stress diseases (BPD, PVL, ROP), despite the fact that the group of preterm girls had a significantly shorter gestational age and body weight at birth, indicates a more stable antioxidant system in female newborns with perinatal pathology. When considering the effect of PDA on the formation of oxidative stress diseases in preterm infants, signs of sexual dimorphism were also noted. In the group of boys with PDA, compared with children with spontaneously closed ductus arteriosus, a higher incidence of severe RDS, a longer duration of oxygen therapy, and a higher incidence of severe BPD were revealed. In the group of girls, no similar significant differences were found. ROP grade 3 or more, which requires surgical methods of treatment, was more often observed in children with PDA, regardless of gender.

[ALGORITHM FOR PREDICTING THE DURATION OF TREATMENT FOR CONGENITAL PNEUMONIA IN PRETERM INFANTS].

Congenital pneumonia is one of the most common diseases of infectious pathology in the fetus and newborns. Early prediction of the duration of the course of the disease can improve the management of preterm infants with congenital pneumonia, prevent the development of complications and optimize their follow-up. The aim of the study was to improve approaches to the management of preterm infants with congenital pneumonia based on the analysis of clinical-anamnestic and paraclinical criteria and the development of algorithms for predicting the duration of treatment of the disease in order to prevent the development of adverse consequences of congenital pneumonia in preterm infants. We analyzed 36 cases of the disease of preterm infants with congenital pneumonia, which were combined into alternative groups according to the principle of the duration of the course of pneumonia: 1st group - 12 preterm infants with the duration of the course of congenital pneumonia less than 15 days; 2nd group - 24 preterm infants with the duration of the course of congenital pneumonia 15 days or more. Using the heterogeneous sequential Wald-Genkin procedure, algorithms were developed for predicting the duration of the course of congenital pneumonia in preterm infants. The influence of clinical-anamnestic and paraclinical factors on the duration of treatment with congenital pneumonia in preterm infants was analyzed. Predictors of treatment duration for preterm infants with congenital pneumonia were determined using the heterogeneous sequential Wald - Genkin procedure. Each of the identified signs is divided into gradations, for which the predictive coefficients and information content were determined. Based on the data obtained, a scale was compiled to predict the duration of the course of congenital pneumonia in preterm infants. Predictors of longer treatment of congenital pneumonia and the likely development of adverse consequences of pneumonia in preterm infants should be considered gestational age ≤34 weeks, weight at birth ≤2000.0 g, Apgar score at 1 minute of life less than 5 points and 5th - less than 7 points, male sex of the child, complicated course of pregnancy (threatened abortion, cervical incompetence, placental dysfunction, respiratory infections), as well as the need for respiratory support with mechanical ventilation and CPAP for more than 5 days in the complex of therapy congenital in preterm infants. Approbation of the scale for predicting the duration of therapy for congenital pneumonia and preventing the development of adverse consequences in preterm infants revealed 88,9% true cases, 9,5% - undefined and 1,6% - erroneous, which indicates a high (≥95%) reliability of the algorithm.

Lamin B1 regulates somatic mutations and progression of B-cell malignancies.

Somatic hypermutation (SHM) is a pivotal process in adaptive immunity that occurs in the germinal centre and allows B cells to change their primary DNA sequence and diversify their antigen receptors. Here, we report that genome binding of Lamin B1, a component of the nuclear envelope involved in epigenetic chromatin regulation, is reduced during B-cell activation and formation of lymphoid germinal centres. Chromatin immunoprecipitation-Seq analysis showed that kappa and heavy variable immunoglobulin domains were released from the Lamin B1 suppressive environment when SHM was induced in B cells. RNA interference-mediated reduction of Lamin B1 resulted in spontaneous SHM as well as kappa-light chain aberrant surface expression. Finally, Lamin B1 expression level correlated with progression-free and overall survival in chronic lymphocytic leukaemia, and was strongly involved in the transformation of follicular lymphoma. In summary, here we report that Lamin B1 is a negative epigenetic regulator of SHM in normal B-cells and a 'mutational gatekeeper', suppressing the aberrant mutations that drive lymphoid malignancy.

RNA-Seq Analysis of Differentiated Keratinocytes Reveals a Massive Response to Late Events during Human Papillomavirus 16 Infection, Including Loss of Epithelial Barrier Function.

The human papillomavirus (HPV) replication cycle is tightly linked to epithelial cell differentiation. To examine HPV-associated changes in the keratinocyte transcriptome, RNAs isolated from undifferentiated and differentiated cell populations of normal, spontaneously immortalized keratinocytes (NIKS) and NIKS stably transfected with HPV16 episomal genomes (NIKS16) were compared using next-generation sequencing (RNA-Seq). HPV16 infection altered expression of 2,862 cellular genes. Next, to elucidate the role of keratinocyte gene expression in late events during the viral life cycle, RNA-Seq was carried out on triplicate differentiated populations of NIKS (uninfected) and NIKS16 (infected). Of the top 966 genes altered (>log2 = 1.8, 3.5-fold change), 670 genes were downregulated and 296 genes were upregulated. HPV downregulated many genes involved in epithelial barrier function, which involves structural resistance to the environment and immunity to infectious agents. For example, HPV infection repressed expression of the differentiated keratinocyte-specific pattern recognition receptor TLR7, the Langerhans cell chemoattractant CCL20, and proinflammatory cytokines interleukin 1α (IL-1α) and IL-1ß. However, the type I interferon regulator IRF1, kappa interferon (IFN-κ), and viral restriction factors (IFIT1, -2, -3, and -5, OASL, CD74, and RTP4) were upregulated. HPV infection abrogated gene expression associated with the physical epithelial barrier, including keratinocyte cytoskeleton, intercellular junctions, and cell adhesion. Quantitative PCR (qRT-PCR) and Western blotting confirmed changes in expression of seven of the most significantly altered mRNAs. Expression of three genes showed statistically significant changes during cervical disease progression in clinical samples. Taken together, the data indicate that HPV infection manipulates the differentiating keratinocyte transcriptome to create an environment conducive to productive viral replication and egress.IMPORTANCE HPV genome amplification and capsid formation take place in differentiated keratinocytes. The viral life cycle is intimately associated with host cell differentiation. Deep sequencing (RNA-Seq) of RNA from undifferentiated and differentiated uninfected and HPV16-positive keratinocytes showed that almost 3,000 genes were differentially expressed in keratinocytes due to HPV16 infection. Strikingly, the epithelial barrier function of differentiated keratinocytes, comprising keratinocyte immune function and cellular structure, was found to be disrupted. These data provide new insights into the virus-host interaction that is crucial for the production of infectious virus and reveal that HPV infection remodels keratinocytes for completion of the virus replication cycle.

Human Papillomavirus E2 Regulates SRSF3 (SRp20) To Promote Capsid Protein Expression in Infected Differentiated Keratinocytes.

UNLABELLED: The human papillomavirus (HPV) life cycle is tightly linked to differentiation of the infected epithelial cell, suggesting a sophisticated interplay between host cell metabolism and virus replication. Previously, we demonstrated in differentiated keratinocytes in vitro and in vivo that HPV type 16 (HPV16) infection caused increased levels of the cellular SR splicing factors (SRSFs) SRSF1 (ASF/SF2), SRSF2 (SC35), and SRSF3 (SRp20). Moreover, the viral E2 transcription and replication factor that is expressed at high levels in differentiating keratinocytes could bind and control activity of the SRSF1 gene promoter. Here, we show that the E2 proteins of HPV16 and HPV31 control the expression of SRSFs 1, 2, and 3 in a differentiation-dependent manner. E2 has the greatest transactivation effect on expression of SRSF3. Small interfering RNA depletion experiments in two different models of the HPV16 life cycle (W12E and NIKS16) and one model of the HPV31 life cycle (CIN612-9E) revealed that only SRSF3 contributed significantly to regulation of late events in the virus life cycle. Increased levels of SRSF3 are required for L1 mRNA and capsid protein expression. Capsid protein expression was regulated specifically by SRSF3 and appeared independent of other SRSFs. Taken together, these data suggest a significant role of the HPV E2 protein in regulating late events in the HPV life cycle through transcriptional regulation of SRSF3 expression. IMPORTANCE: Human papillomavirus replication is accomplished in concert with differentiation of the infected epithelium. Virus capsid protein expression is confined to the upper epithelial layers so as to avoid immune detection. In this study, we demonstrate that the viral E2 transcription factor activates the promoter of the cellular SRSF3 RNA processing factor. SRSF3 is required for expression of the E4(^)L1 mRNA and so controls expression of the HPV L1 capsid protein. Thus, we reveal a new dimension of virus-host interaction crucial for production of infectious virus. SRSF proteins are known drug targets. Therefore, this study provides an excellent basis for developing strategies to regulate capsid protein production in the infected epithelium and the production of new virions.

Preclinical efficacy of the bioreductive alkylating agent RH1 against paediatric tumours.

BACKGROUND: Despite substantial improvements in childhood cancer survival, drug resistance remains problematic for several paediatric tumour types. The urgent need to access novel agents to treat drug-resistant disease should be expedited by pre-clinical evaluation of paediatric tumour models during the early stages of drug development in adult cancer patients. METHODS/RESULTS: The novel cytotoxic RH1 (2,5-diaziridinyl-3-[hydroxymethyl]-6-methyl-1,4-benzoquinone) is activated by the obligate two-electron reductase DT-diaphorase (DTD, widely expressed in adult tumour cells) to a potent DNA interstrand cross-linker. In acute viability assays against neuroblastoma, osteosarcoma, and Ewing's sarcoma cell lines RH1 IC(50) values ranged from 1-200 nM and drug potency correlated both with DTD levels and drug-induced apoptosis. However, synergy between RH1 and cisplatin or doxorubicin was only seen in low DTD expressing cell lines. In clonogenic assays RH1 IC(50) values ranged from 1.5-7.5 nM and drug potency did not correlate with DTD level. In A673 Ewing's sarcoma and 791T osteosarcoma tumour xenografts in mice RH1 induced apoptosis 24 h after a single bolus injection (0.4 mg/kg) and daily dosing for 5 days delayed tumour growth relative to control. CONCLUSION: The demonstration of RH1 efficacy against paediatric tumour cell lines, which was performed concurrently with the adult Phase 1 Trial, suggests that this agent may have clinical usefulness in childhood cancer.

NXF1/p15 heterodimers are essential for mRNA nuclear export in Drosophila.

The conserved family of NXF proteins has been implicated in the export of messenger RNAs from the nucleus. In metazoans, NXFs heterodimerize with p15. The yeast genome encodes a single NXF protein (Mex67p), but there are multiple nxf genes in metazoans. Whether metazoan NXFs are functionally redundant, or their multiplication reflects an adaptation to a greater substrate complexity or to tissue-specific requirements has not been established. The Drosophila genome encodes one p15 homolog and four putative NXF proteins (NXF1 to NXF4). Here we show that depletion of the endogenous pools of NXF1 or p15 from Drosophila cells inhibits growth and results in a rapid and robust accumulation of polyadenylated RNAs within the nucleus. Fluorescence in situ hybridizations show that export of both heat-shock and non-heat-shock mRNAs, as well as intron-containing and intronless mRNAs is inhibited. Depleting endogenous NXF2 or NXF3 has no apparent phenotype. Moreover, NXF4 is not expressed at detectable levels in cultured Drosophila cells. We conclude that Dm NXF1/p15 heterodimers only (but not NXF2-NXF4) mediate the export of the majority of mRNAs in Drosophila cells and that the other members of the NXF family play more specialized or different roles.

[Accounting for sexual dimorphism in neonatology]. / Do pytannia obliku statevoho dymorfizmu v neonatolohiï.

Results are submitted of the study into the role of sex features of reactivity and adaptivity in the organization and compensation of structural and functional changes in the central nervous system in newborn babies having suffered intrauterine hypoxia and born in asphyxia. The identified sex dimorphism of the neuroendocrine system attests to the need for taking account of sex resistance in neonatology and permits the awareness of better adaptation of newborn girls with cerebral disorders of hypoxic genesis. Further study of sex dimorphism will, we believe, help in working out informative-and-quest systems of clinical, instrumental, biochemical, and morphological diagnosis in neonatology.