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Given its central role in life, DNA is remarkably easy to damage. Double strand breaks (DSBs) are the most toxic form of DNA damage, and DSBs pose the greatest danger to genomic integrity. In higher vertebrates, the non-homologous end joining pathway (NHEJ) is the predominate pathway that repairs DSBs. NHEJ has three steps: 1) DNA end recognition by the DNA dependent protein kinase [DNA-PK], 2) DNA end-processing by numerous NHEJ accessory factors, and 3) DNA end ligation by the DNA ligase IV complex (LX4). Although this would appear to be a relatively simple mechanism, it has become increasingly apparent that it is not. Recently, much insight has been derived regarding the mechanism of non-homologous end joining through a proliferation of cryo-EM studies, structure-function mutational experiments informed by these new structural data, and novel single-molecule imaging approaches. An emerging consensus in the field is that NHEJ progresses from initial DSB end recognition by DNA-PK to synapsis of the two DNA ends in a long-range synaptic complex where ends are held too far apart (115 Å) for ligation, and then progress to a short-range synaptic complex where ends are positioned close enough for ligation. What was surprising from these structural studies was the observation of two distinct types of DNA-PK dimers that represent NHEJ long-range complexes. In this review, we summarize current knowledge about the function of the distinct NHEJ synaptic complexes and align this new information with emerging cellular single-molecule microscopy studies as well as with previous studies of DNA-PK's function in repair., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Non-homologous end joining (NHEJ) is the predominant pathway that repairs DNA double-stranded breaks (DSBs) in vertebrates. However, due to challenges in detecting DSBs in living cells, the repair capacity of the NHEJ pathway is unknown. The DNA termini of many DSBs must be processed to allow ligation while minimizing genetic changes that result from break repair. Emerging models propose that DNA termini are first synapsed ~115Å apart in one of several long-range synaptic complexes before transitioning into a short-range synaptic complex that juxtaposes DNA ends to facilitate ligation. The transition from long-range to short-range synaptic complexes involves both conformational and compositional changes of the NHEJ factors bound to the DNA break. Importantly, it is unclear how NHEJ proceeds in vivo because of the challenges involved in analyzing recruitment of NHEJ factors to DSBs over time in living cells. Here, we develop a new approach to study the temporal and compositional dynamics of NHEJ complexes using live cell single-molecule imaging. Our results provide direct evidence for stepwise maturation of the NHEJ complex, pinpoint key regulatory steps in NHEJ progression, and define the overall repair capacity NHEJ in living cells., Competing Interests: COMPETING INTERESTS None declared.

The genetic material in human cells is continuously exposed to a wide variety of insults that can induce different DNA lesions. To maintain genomic stability and prevent potentially deleterious genetic changes caused by DNA damage, mammalian cells have evolved a number of pathways that repair specific types of DNA damage. These DNA repair pathways vary in their accuracy, some providing high-fidelity repair while others are error-prone and are only activated as a last resort. Adding additional complexity to cellular mechanisms of DNA repair is the DNA damage response which is a sophisticated a signaling network that coordinates repair outcomes, cell-cycle checkpoint activation, and cell fate decisions. As a result of the sheer complexity of the various DNA repair pathways and the DNA damage response there are large gaps in our understanding of the molecular mechanisms underlying DNA damage repair in human cells. A key unaddressed question is how the dynamic recruitment of DNA repair factors contributes to repair kinetics and repair pathway choice in human cells. Methodological advances in live cell single-molecule imaging over the last decade now allow researchers to directly observe and analyze the dynamics of DNA repair proteins in living cells with high spatiotemporal resolution. Live cell single-molecule imaging combined with single-particle tracking can provide direct insight into the biochemical reactions that control DNA repair and has the power to identify previously unobservable processes in living cells. This review summarizes the main considerations for experimental design and execution for live cell single-molecule imaging experiments and describes how they can be used to define the molecular mechanisms of DNA damage repair in mammalian cells., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Repair of DNA double strand breaks (DSBs) is integral to preserving genomic integrity. Therefore, defining the mechanisms underlying DSB repair will enhance our understanding of how defects in these pathways contribute to human disease and could lead to the discovery of new approaches for therapeutic intervention. Here, we established a panel of HaloTagged DNA damage response factors in U2OS cells which enables concentration-dependent protein labeling by fluorescent HaloTag ligands. Genomic insertion of HaloTag at the endogenous loci of these repair factors preserves expression levels and proteins retain proper subcellular localization, foci-forming ability, and functionally support DSB repair. We systematically analyzed total cellular protein abundance, measured recruitment kinetics to laser-induced DNA damage sites, and defined the diffusion dynamics and chromatin binding characteristics by live-cell single-molecule imaging. Our work demonstrates that the Shieldin complex, a critical factor in end-joining, does not exist in a preassembled state and that relative accumulation of these factors at DSBs occurs with different kinetics. Additionally, live-cell single-molecule imaging revealed the constitutive interaction between MDC1 and chromatin mediated by its PST repeat domain. Altogether, our studies demonstrate the utility of single-molecule imaging to provide mechanistic insights into DNA repair, which will serve as a powerful resource for characterizing the biophysical properties of DNA repair factors in living cells., Competing Interests: JH, MM, AB, DB, JS No competing interests declared, (© 2023, Heyza et al.)

MoonProt 3.0 (http://moonlightingproteins.org) is an updated open-access database storing expert-curated annotations for moonlighting proteins. Moonlighting proteins have two or more physiologically relevant distinct biochemical or biophysical functions performed by a single polypeptide chain. Here, we describe an expansion in the database since our previous report in the Database Issue of Nucleic Acids Research in 2018. For this release, the number of proteins annotated has been expanded to over 500 proteins and dozens of protein annotations have been updated with additional information, including more structures in the Protein Data Bank, compared with version 2.0. The new entries include more examples from humans, plants and archaea, more proteins involved in disease and proteins with different combinations of functions. More kinds of information about the proteins and the species in which they have multiple functions has been added, including CATH and SCOP classification of structure, known and predicted disorder, predicted transmembrane helices, type of organism, relationship of the protein to disease, and relationship of organism to cause of disease., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Unlabelled: The reported incidence of the preterm deliveries in Bulgaria is 7-11% and it has increased in the recent years. The neonatal respiratory distress syndrome (NRDS) is the most frequent pulmonary disease caused by the biological immaturity. It affects 40-50% of the neonates born before 32 weeks of gestation and results in high morbidity and mortality. There is plenty of evidence, proving that the antenatal corticosteroid (CS) administration decreases the incidence of NRDS in preterm infants., Objective: To study the influence of the implicated algorithm for antenatal prophylaxis of NRDS on the neonatal morbidity and mortality in premature infants., Materials and Methods: Two groups of preterm neonates were studied. First group included preterm neonates either without or with a single application of CS before delivery. Second group consisted of preterm newborns with completed CS prophylaxis. The applied antenatal corticosteroid treatment included 4 doses of Betamethazon 7 mg i.m. in 12 hour intervals. It was indicated in cases of a preterm labor between 24-32 gestational weeks. The neonatal mortality, the incidence of NRDS and intraventricular hemorrhage (IVH), the neonatal intensive care unit (NICU) stay and treatment were compared between both groups., Results: The incidence of NRDS, the need of exogenous surfactant treatment, and the incidence of severe IVH (grades II-IV) were significantly lower in the group with completed CS prophylaxis compared to the control group (p < 0.05). The neonatal mortality rate and the duration of NICU stay and treatment were also reduced among infants with antenatal CS., Conclusions: The antenatal treatment with CS reduces significantly the risk of development of NRDS and IVH 3d-4th degrees and the neonatal mortality rate of the prematurely born infants. It has also important influence on the duration of NICU stay and treatment and decreases the cost of the neonatal intensive treatment.

Objective: To study the therapeutic application of Gynazol in cases of recurrent vaginal Candida infections during the last trimester of pregnancy., Materials and Methods: 28 pregnant women in third trimester with recurrent vaginal Candida infection were included in the study. All cases were treated with a single vaginal application of Gynazol (butoconazole nitrate 2%). Follow up of reported subjective complaints and control vaginal swab and culture 10-14 days after treatment was performed., Results and Discussion: In 24 (85.72%) cases of recurrent vaginal Candida infection, Candida albicans was isolated. C. tropicalis and C. parapsilosis were found in 3 (10.71%) and 1 (3.57%) cases respectively. Single vaginal application of Gynazol led to alleviation of subjective symptoms 24-48 hours after initiation of treatment. Control vaginal swab and culture were performed in 24 patients. Negative cultures were found in 21 cases (87.5%) and single Candida colonies were isolated in 3 (12.5%) patients. According to our results, application of Gynazol is recommended in cases of recurrent vaginal Candida infection during third trimester of pregnancy.

Unlabelled: The aim of the study is to determine and estimate the absolute values of the distance between the symphysis pubis and the uterine fundus (S-FD) and the rate of uterine involution according to the method of delivery., Material: 150 women included in this prospective study are divided into 3 groups of 50 women with vaginal delivery, caesarean section and re-caesarean section. The absolute values of the distance S-FD and the rate of uterine involution were determined in cm and estimated according to their parity and the gestational age. The measurement of the S-FD was daily performed from the second day till the 5th day for the vaginal deliveries and till the 7th day for the caesarean sections., Results: The uterine involution in primiparous and premature vaginal deliveries starts from lower values of the S-FD than in the multiparous and in cases of term delivery. The rate of uterine involution in primiparous increases gradually in the earliest day after delivery (from 0.95 to 1.6 cm per day), while in multiparous this increasing starts after the 4th day. When caesarean section is performed and in the cases of preterm delivery the rates of uterine involution are delayed and uneven., Conclusions: in cases of vaginal delivery the availability of prematurity and macrosomia of the fetus delays the rates of the uterine involution. The uterine involution after cesarean section is slower and unsteady, which is more conscious in re-sections.

Objective: To establish the incidence of cervicovaginal infections in patients with preterm and term deliveries., Materials and Methods: 88 pregnancies with no previous detectable risk factors have been included in a retrospective study. 48 patients delivered at term and 40 had delivery between 24 and 37 weeks of gestation. A vaginal swab for bacterial infections and a cervical swab for detection of Chlamydia trachomatis infection, using PCR, have been performed in each case. The vaginal ecosystem has been appreciated according Nungent criteria., Results and Conclusions: C. trachomatis has been detected in 5% (2/40) of preterm deliveries and in 2.08% (1/48) of term deliveries. In preterm delivery group bacterial vaginosis has been isolated in 15% (6/40), Enterococcus spp. in12.5% (5/40) and Group B Streptococcus in 12.5% (5/40) of cases. In patients, delivered at term, bacterial vaginosis has been proved in 6.25% (3/48), Enterococcus spp in 8.33% (4/48), Group B Streptococcus in 4.17% (2/48) patients. Significant differences between both groups have been found in GBS and BV (p < 0.05). Although the difference C. trachomatis infection is not significantly higher in preterm delivery group, testing for C. tracomatis, using highly specific methods, along with vaginal swab are recommended as routine tests in pregnancy.

A prospective study, including 60 patients with vaginal discharge has been made at Maternity hospital "Majchin dom"--Sofia for the period November 2006- February 2007. In 32 patients no causative agent has been revealed. They have been counseled to use Yin Care--vaginal lotion for 3 months in prophylactic concentration. 88.4% reported for diminished discharge. No adverse effects have been observed. In 11 patients suffering from bacterial vaginitis, caused by S. Epidermidis, S. Aureus, Enterococcus and E. Coli cure was achieved in 72.7% of cases. 17 women with Candida vaginitis have been included in the study. After standard antifungal treatment, 8 of them continued therapy with Yin Care. Recurrence of disease has been observed in only one case, while in the group treated with antifungal medication only recurrence has been observed in 2 cases.

Unlabelled: Doppler velocimetry is a fast and convenient method for monitoring of fetal well being in high risk pregnancies. Doppler velocimetry is more specific and sensitive method of fetal surveillance than cardiotocography. The study's objective is to prove Doppler's effectiveness in predicting perinatal outcome in preeclamptic pregnancies. A retrospective study is designed, including pregnant women, diagnosed with preeclampsia, admitted to Maternity hospital "Majchin dom" for ten years. Doppler study was made and results were correlated to Apgar score, umbilical artery pH and base excess of the neonate at delivery. A correlation between RI of umbilical artery, uterine arteries and Apgar score at 5th minute was demonstrated. In severe cases correlation between RI of umbilical artery and neonatal pH was found., Conclusion: Doppler velocimetry is an effective method of fetal surveillance in pregnancies, complicated by preeclampsia

No matter the fact that the changes in the blood pressure are quite often the first clinical feature of preeclampsia, is known that they are not early, but are quite late preeclampsia symptom. Close follow-up of the blood pressure trough whole pregnancy and in preeclamptic patients plays an important role in the improvement of the perinatal outcome. Five hundred forty four singleton pregnant women were enrolled in the study. Of them 334 were with mild preeclampsia and 210 were with severe form of the disease. The analysis of correlations shows a correlation between the blood pressure at admititon and before delivery of the women with mild and severe form of preeclampsia and the perinatal fetal outcome. The fifth minute Apgar score is significantly lower in the patients with severe preeclampsia in comparison with women with mild disease. The represented data confirm the hypothesis about the influence of the maternal blood pressure on the perinatal fetal outcome.

Objective: To compare the effectiveness of the different types of cervical cerclage (primary, secondary and tertiary) for prevention of preterm deliveries and second trimester abortions., Materials and Methods: The study is retrospective, including 145 pregnant women with history and/or clinical signs of cervical incompetence. In 86 cases cervical cerclage has been performed while in the remaining 59 cases management has been expectant. Among the patients with cerclage 57 had primary, 25 had secondary and 4--tertiary cervical suture. In 21 cases cervical length was measured by transvaginal ultrasound. In 9 cases cervical length was < 25 mm and in the remaining 12 cases-- > or = 25 mm., Results: Cervical suture placement and expectant management were found to be equally effective in patients with indications for primary cerclage. On the contrary, in patients with indications for secondary cerclage mean gestational age at delivery and the percentage of patients that remained undelivered after 34 w.g. were higher in the subgroup that had cerclage compared to the one that was treated expectantly. In patients with indications for tertiary cerclage pregnancy outcome was similar in the subgroups with and without cerclage and in general was unfavorable. Gestational age at delivery was higher if the cerclage was placed before 18 w.g. and if cervical length was > or = 25 mm.

Cardiotocographic monitoring (NST) is well accepted in obstetrical practice. It provides direct and valuable information about neurological and myocardial function of the fetus. In a retrospective and prospective study, including 544 preeclamptic women for period of 10 years cardiotocographic monitoring of the fetus was applied. At admission in 427 (82.80%) of the cases, CTG readings were found normal. In the rest of 89 (17.20%) preeclamptic patients the NST records were found pathological. A correlation between mean values of resistance indices (RI) of umbilical and uterine arteries (RI = 0.73; RI = 0.75 respectively) and pathological NST was demonstrated. A similar correlation is found for newborn's Apgar score at 5th minute--7.16. Although the difference is found statistically significant, it is not possible to conclude that Doppler velocimetry is more predictive than cardiotocographic records.

The presented study is retrospective and prospective in design. It is based on the medical records of Maternity hospital "Majchin dom" and planned studies during the period 1994-2005. 544 preeclamptic patients with singleton pregnancies were included. 334 (61.3%) of them were diagnosed with mild preeclampsia and 210 (38.7%) had severe preeclampsia. Blood pressure was monitored at least four times daily, depending on the severity of the case with mechanical manometer. In special cases cardiopulsometry was applied. In order to compare the effectiveness of different antihypertensive medications, administered orally, either alone, or in combination (no antihypertensive medication, Methyldopa, Nifedipin, beta-blockers and combinations between them), patients were divided in seven groups. The mean values of systolic and diastolic blood pressure, measured before delivery were compared. No statistically significant difference between the groups was noticed. We were not also able to find statistically significant difference in blood pressure measurements before delivery, between patients with severe preeclampsia, treated with parenteral Nepresol or Chlofazolin.

The represented study is made in order to prove whether the mode of delivery influences the perinatal prognosis of neonates born with preeclamptic mothers, appreciated by pH and the Apgar score at the 5-th minute. There are 544 preeclamptic patients included in the study, admitted to Majchin dom hospital, between 1995 and 2004. The comparison between the mean Apgar and mean pH, demonstrated no difference between neonates delivered through Cesarean Section and neonates delivered through vaginal delivery.

Observations on one of the rare complications in children presenting malignant neoplasms of the abdominal cavity and retroperitoneal space are described. Over a 7-year period, operative treatment is undertaken in 42 children aged 3 to 15 years. In eleven of them (26.42 per cent) it is a matter of mechanical ileus. The type of bowel obstruction in the series of children under study is a follows: obturation-in three and adhesion-in five cases. The obturation involves the large intestine, and is due to pressure of a neoplastic process in advanced stage of development on the colon. Ileus due to adhesions occurs after operative removal of the neoplastic formation. The essential differences in type of intestinal obstruction in children with malignant neoformation in the abdominal region from the one in adult patients justify the report on the observations.

This is a retrospective analysis of 651 children, aged 0 to 14 years, presenting chest and abdominal injuries, treated over a 6-year period in the section of pediatric surgery of the Emergency Medicine Institute "Pirogov". The results of studies on the etiopathogenesis of chest and abdominal trauma, patterns of associated lesions and quality of prehospital aid are discussed. Experience with the application of updated noninvasive and invasive methods of diagnosis, as well as the scope of surgical approach to the variety of lesions by resorting to organ sparing and organ salvaging operative procedures are analyzed. On the basis of observations and experience gained to date, the indications for application of the prospective method of nonoperative treatment of certain organic lesions in the practice of pediatric surgery are broadened. The overall research project provides good reason to work out an algorithm of the therapeutic approach to children presenting abdominal and chest trauma.

Two cases of primary tumors of the stomach in children (7 and 10 years of age) are reported. One of them had non-Hodgkin's lymphoma and the other hamartoma, both radically operated. In the former case subtotal resection of the stomach was made followed by polychemotherapy. For an observation period of one year there was no evidence of recurrence or metastases of the neoplastic process. The incidence, histologic characteristics, clinical course, diagnostic and therapeutic problems in tumors of the stomach in children, with special reference to non-Hodgkin's lymphoma and hamartoma, are discussed.

In a series of 4350 children with abdominal surgical diseases operated at the Department of Pediatric Surgery in Sofia for a period of 3 years (1987-1989) the authors have observed 4 cases of pleuropulmonary complications (0.09 per cent). In two of these children the primary abdominal operation was for liver echinococcus, in 1--rupture of the liver and subphrenic abscess and in 1--perforative appendicitis and purulent peritonitis. Symptoms of respiratory failure developed in addition to the abdominal symptoms. Leading in the complex treatment was sanation of the abdominal cavity from the purulent process and double pleural drainage. The severe pathologic changes in the organs of the two adjacent cavities were responsible for the high case fatality rate (25 per cent).

One of the rare surgical diseases of the organs in the abdominal cavity in children are the splenic cysts. For a period of 20 years (1968-1988) 3 children have been observed and treated at the Clinic of Pediatric Surgery in Sofia. The use of modern diagnostic methods--echography, computer axial tomography, scintigraphy and laparoscopy furnished an opportunity for precise preoperative organ localization of the disease in the patients observed by the authors. Partial resection of the spleen in this pathology with good postoperative results, performed for the first time in Bulgaria, encouraged the authors to recommend organ-preserving operation as the method of choice in nonparasitic cysts of the spleen.

The combination of subphrenic abscess with purulent pericarditis is a rare postoperative complication of perforated appendicitis in children, with severe clinical course, difficult to diagnose and high case fatality rate. A 7-year-old child with this complication, successfully diagnosed and treated is reported. The importance of complex therapy--surgical and intensive and collaboration between pediatric surgeons, anesthesiologists, cardiac surgeons and cardiologists for the favourable outcome is emphasized.

A method of organ-preserving operations in children with injured spleen with the use of glue is described. The method is recommended for a wider use in child surgery.