Clamp loading,unloading and intrinsic stability of the PCNA, β and gp45 sliding clamps of human,E. coli and T4 replicases

Background: The high speed and processivity of replicative DNA polymerases reside in a processivity factor which has been shown to be a ring-shaped protein. This protein (‘sliding clamp’) encircles DNA and tethers the catalytic unit to the template. Although in eukaryotic, prokaryotic and bacteriophage-T4 systems, the processivity factors are ring-shaped, they assume different oligomeric states. The Escherichia coli clamp (the β subunit) is active as a dimer while the eukaryotic and T4 phage clamps (PCNA and gp45, respectively) are active as trimers. The clamp can not assemble itself on DNA. Instead, a protein complex known as a clamp loader utilizes ATP to assemble the ring around the primer-template. This study compares properties of the human PCNA clamp with those of E. coli and T4 phage. Results: The PCNA ring is a stable trimer down to a concentration below 100 nm (K_d ≈ 21 nm ). On DNA, the PCNA clamp slides freely and dissociates from DNA slowly (t_1/2 ≈ 24 min). β is more stable in solution (K_d < 60 pm ) and on DNA (t_1/2 ≈ 1 h) than PCNA which may be explained by its simpler oligomeric state. The T4 gp45 clamp is a much less stable trimer than PCNA (K_d ≈ 250 nm ) and requires association with the polymerase to stabilize it on DNA as observed previously. The consequence of this cooperation between clamp and polymerase is that upon finishing a template and dissociation of the polymerase from DNA, the gp45 clamp spontaneously dissociates from DNA without assistance. However, the greater stability of the PCNA and β clamps on DNA necessitates an active process for their removal. The clamp loaders (RF-C and γ complex) were also capable of unloading their respective clamps from DNA in the presence of ATP. Conclusions: The stability of the different clamps in solution correlates with their stability on DNA. Thus, the low stability of the T4 clamp explains the inability to isolate gp45 on DNA. The stability of the PCNA and β clamps predicts they will require an unloading factor to recycle them on and off DNA during replication. The clamp loaders of PCNA and β double as clamp unloaders presumably for the purpose of clamp recycling.     

Histamine concentrations in nasal secretion and secretory activity in allergic rhinitis

The prerequisites for using the assayed histamine concentration in nasal secretion as an objective measure of disease activity in allergic rhinitis were investigated. It was demonstrated that in histamine determination procedures the presence of quenching substances in the nasal secretion could lead to underestimation of the histamine concentration. This bias was eliminated in a modified spectrofluorometric assay. Only an insignificant fraction of the histamine in samples collected by nasal spray washing was bound to unfiltrable particles or cells. The mean histamine concentration in nasal secretions from 15 healthy subjects was 11.2 micrograms/ml and in a group of nine patients with allergic rhinitis out of season 3.36 micrograms/ml. The histamine concentration in the latter group decreased during the pollen season and after positive allergen challenge. It is suggested that this decrease is caused by the increase in volume of the secretion during the allergic response. The use of lithium as an exogenous marker permitted quantitation of the increase in the relative amount of nasal secretion recovered by washing in the symptomatic subjects.     

The undifferentiated malignant neoplasm. Identification of lymphoma arising in skeletal muscle by immunohistochemical analysis

The undifferentiated malignant neoplasm presents a significant problem in the intelligent selection of therapy. Because of advances in chemotherapy, there are cancers that are effectively palliated, and sometimes cured if appropriately treated. Characterization of tumors by immunohistochemical stains drastically reduces the incidence of "undifferentiated" diagnoses and will optimize patient management, as illustrated by two cases of large-cell lymphoma arising in skeletal muscle.     

SARS-CoV-2-Seroprävalenz bei Kindern und Jugendlichen in Deutschland – ein Überblick

Bundesgesundheitsblatt - Gesundheitsforschung - Gesundheitsschutz - SARS-CoV-2-Antikörperstudien ergänzen und erweitern die Erkenntnisse aus der Meldestatistik laborbestätigter...     

宜昌夷陵地区11656例0~6岁婴幼儿25-羟维生素D水平检测分析

目的为了研究宜昌市夷陵区0~6岁儿童维生素D营养状况及其与年龄、性别、季节之间的关系,以便为夷陵区儿童合理补充维生素D提供科学依据。方法对2017年1月至2018年7月来夷陵区妇幼保健院体检的0~6岁11656例儿童,采用荧光免疫层析方法进行末梢血25-(OH)D 3水平检测。结果该区11656例儿童末梢血25-(OH)D 3水平为(29.35±7.59)ng/mL,其中维生素D缺乏组566例(4.86%),维生素D不足组6579例(56.44%),维生素D充足组4511例(38.70%)。1岁以内婴儿组维生素D水平明显高于幼儿组和学龄前组,3岁以后儿童维生素D水平明显下降,各年龄组间差异有统计学意义(χ2=145.846,P<0.05)。不同季节儿童维生素D水平春季最高,冬季最低(χ2=504.007,P<0.05)。不同性别间儿童维生素D水平差异并无统计学意义(t=0.841,P>0.05)。结论我区0~6岁儿童维生素D水平大部分处于不足的状态,应增加该区儿童维生素D的摄入量,尤其加强学龄前组儿童维生素D的补充及冬季户外活动。     

Abdominal aortic aneurysm: epidemiology,screening and work-up for repair

Abdominal aortic aneurysm (AAA) is a dilatation of the infra-renal abdominal aorta to greater than 3 cm. Population screening is offered to men in the year of their 65th birthday in the UK. Patients with small, asymptomatic AAAs (<5.5 cm) are entered into surveillance programmes and have their cardiovascular risk factors managed aggressively. An AAA ≥5.5 cm diameter, or one which is symptomatic, should be considered for surgical repair to prevent rupture. Aneurysm repair can be undertaken using either an open surgical or endovascular approach; the decision should be tailored to the individual patient and made by the surgeon and patient, with input from a multi-disciplinary team.     

Urinary calprotectin,NGAL, and KIM-1 in the differentiation of primarily inflammatory vs. non-inflammatory stable chronic kidney diseases

Introduction It has been demonstrated that urinary neutrophil gelatinase-associated lipocalin (NGAL) and calprotectin are helpful biomarkers in the differentiation of intrinsic and prerenal acute kidney injury.Objective The present cross-sectional study investigates, whether urinary biomarkers are able to differentiate primarily inflammatory from non-inflammatory entities in chronic kidney disease (CKD).Methods Urinary calprotectin, NGAL, and kidney injury molecule-1 (KIM-1) concentrations were assessed in a study population of 143 patients with stable CKD and 29 healthy controls. Stable renal function was defined as an eGFR fluctuation ≤5 ml/min/1.73 m² in the past 12 months. Pyuria, metastatic carcinoma, and renal transplantation were regarded as exclusion criteria. Diabetic nephropathy, hypertensive nephropathy, and polycystic kidney disease were categorized as ‘primarily non-inflammatory renal diseases’ (NIRD), whereas glomerulonephritis and vasculitis were regarded as ‘primarily inflammatory renal diseases’ (IRD).Results Urinary calprotectin and NGAL concentrations significantly differed between CKD and healthy controls (p < 0.05 each), whereas KIM-1 concentrations did not (p = 0.84). The three biomarkers did neither show significant differences in-between the individual entities, nor the two categories of IRD vs. NIRD (calprotectin 155.7 vs. 96.99 ng/ml; NGAL 14 896 vs. 11 977 pg/ml; KIM-1 1388 vs. 1009 pg/ml; p > 0.05 each). Albumin exceeds the diagnostic power of the investigated biomarkers by far.Conclusions The urinary biomarkers calprotectin, NGAL, and KIM-1 have no diagnostic value in the differentiation of primarily inflammatory vs. non-inflammatory etiologies of CKD.     

Risk factors for Epstein–Barr virus reactivation after renal transplantation: Results of a large,multi-centre study

Epstein–Barr virus (EBV) reactivation is a very common and potentially lethal complication of renal transplantation. However, its risk factors and effects on transplant outcome are not well known. Here, we have analysed a large, multi-centre cohort (N = 512) in which 18.4% of the patients experienced EBV reactivation during the first post-transplant year. The patients were characterized pre-transplant and two weeks post-transplant by a multi-level biomarker panel. EBV reactivation was episodic for most patients, only 12 patients showed prolonged viraemia for over four months. Pre-transplant EBV shedding and male sex were associated with significantly increased incidence of post-transplant EBV reactivation. Importantly, we also identified a significant association of post-transplant EBV with acute rejection and with decreased haemoglobin levels. No further severe complications associated with EBV, either episodic or chronic, could be detected. Our data suggest that despite relatively frequent EBV reactivation, it had no association with serious complications during the first post-transplantation year. EBV shedding prior to transplantation could be employed as biomarkers for personalized immunosuppressive therapy. In summary, our results support the employed immunosuppressive regimes as relatively safe with regard to EBV. However, long-term studies are paramount to support these conclusions.     

Association of Insulin Resistance and Higher Oncotype DX™ Recurrence Score

Annals of Surgical Oncology - Black women with breast cancer have a worse overall survival compared with White women; however, no difference in Oncotype DX? (ODX) recurrence scores has been...     

Comparison of donor scores in bilateral lung transplantation—A large single-center analysis

Objectifying donor lung quality is difficult and currently there is no consensus. Several donor scoring systems have been proposed in recent years. They all lack large-scale external validation and widespread acceptance. A retrospective evaluation of 2201 donor lungs offered to the lung transplant program at the Medical University of Vienna between January 2010 and June 2018 was performed. Five different lung donor scores were calculated for each offer (Oto, ET, MALT, UMN-DLQI, and ODSS). Prediction of organ utilization, 1-year graft survival, and long-term outcome were analyzed for each score. 1049 organs were rejected at the initial offer (group I), 209 lungs declined after procurement (group II), and 841 lungs accepted and transplanted (group III). The Oto score was superior in predicting acceptance of the initial offer (AUC: 0.795; CI: 0.776–0.815) and actual donor utilization (AUC: 0.660; CI: 0.618–0.701). Prediction of 1-year graft survival was best using the MALT score, Oto score, and UMN-DLQI. Stratification of early outcome by MALT was significant for length of mechanical ventilation (LMV), PGD3 rates, ICU stay and hospital stay, and in-hospital-mortality, respectively. To the best of our knowledge, this study is the largest validation analysis comparing currently available donor scores. The Oto score was superior in predicting organ utilization, and MALT score and UMN-DLQI for predicting outcome after lung transplantation.