Potassium Chloride Pulse Enhances Mitogen-Activated Protein Kinase Activity in Rat Hippocampal Slices

Abstract: Mitogen-activated protein (MAP) kinases have been implicated in multiple responses to extracellular stimuli. In this study we show that MAP kinase activity is enhanced after a KCI pulse. This activation correlates with an increased tyrosine phosphorylation of a 42-kDa protein as determined by antiphosphotyrosine immunoblot. The same band is found in an anti-MAP kinase immunoblot. Activity is enhanced within 1 min, reaches a maximum at 2 min, and returns to basal level after 10 min. A second peak of activity is observed between 12 and 30 min. The activation is completely blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), showing the involvement of the AMPA type of glutamate receptor. Partial inhibition of MAP kinase activation by 2-amino-5-phosphonovalerate (APV) also shows the involvement of the NMDA receptor. Because the KCI pulse used induces long-term potentiation (LTP) in rat hippocampal slice, we conclude that MAP kinase may be involved in neuronal transduction events leading to LTP.     

Beyond Axillary Lymph Node Metastasis,BMI and Menopausal Status Are Prognostic Determinants for Triple-Negative Breast Cancer Treated by Neoadjuvant Chemotherapy

BackgroundTriple-negative breast cancers (TNBC) are a specific subtype of breast cancers with a particularly poor prognosis. However, it is a very heterogeneous subgroup in terms of clinical behavior and sensitivity to systemic treatments. Thus, the identification of risk factors specifically associated with those tumors still represents a major challenge. A therapeutic strategy increasingly used for TNBC patients is neoadjuvant chemotherapy (NAC). Only a subset of patients achieves a pathologic complete response (pCR) after NAC and have a better outcome than patients with residual disease.PurposeThe aim of this study is to identify clinical factors associated with the metastatic-free survival in TNBC patients who received NAC.MethodsWe analyzed 326 cT1-3N1-3M0 patients with ductal infiltrating TNBC treated by NAC. The survival analysis was performed using a Cox proportional hazard model to determine clinical features associated with prognosis on the whole TNBC dataset. In addition, we built a recursive partitioning tree in order to identify additional clinical features associated with prognosis in specific subgroups of TNBC patients.ResultsWe identified the lymph node involvement after NAC as the only clinical feature significantly associated with a poor prognosis using a Cox multivariate model (HR = 3.89 [2.42–6.25], p<0.0001). Using our recursive partitioning tree, we were able to distinguish 5 subgroups of TNBC patients with different prognosis. For patients without lymph node involvement after NAC, obesity was significantly associated with a poor prognosis (HR = 2.64 [1.28–5.55]). As for patients with lymph node involvement after NAC, the pre-menopausal status in grade III tumors was associated with poor prognosis (HR = 9.68 [5.71–18.31]).ConclusionThis study demonstrates that axillary lymph node status after NAC is the major prognostic factor for triple-negative breast cancers. Moreover, we identified body mass index and menopausal status as two other promising prognostic factors in this breast cancer subgroup. Using these clinical factors, we were able to classify TNBC patients in 5 subgroups, for which pre-menopausal patients with grade III tumors and lymph node involvement after NAC have the worse prognosis.     

Cellular Expression of Cyclooxygenase,Aromatase, Adipokines,Inflammation and Cell Proliferation Markers in Breast Cancer Specimen

Current evidences suggest that expression of Ki67, cyclooxygenase (COX), aromatase, adipokines, prostaglandins, free radicals, β-catenin and α-SMA might be involved in breast cancer pathogenesis. The main objective of this study was to compare expression/localization of these potential compounds in breast cancer tissues with tissues collected adjacent to the tumor using immunohistochemistry and correlated with clinical pathology. The breast cancer specimens were collected from 30 women aged between 49 and 89 years who underwent breast surgery following cancer diagnosis. Expression levels of molecules by different stainings were graded as a score on a scale based upon staining intensity and proportion of positive cells/area or individually. AdipoR1, adiponectin, Ob-R, leptin, COX-1, COX-2, aromatase, PGF_2α, F₂-isoprostanes and α-SMA were localised on higher levels in the breast tissues adjacent to the tumor compared to tumor specimens when considering either score or staining area whereas COX-2 and AdipoR2 were found to be higher considering staining intensity and Ki67 on score level in the tumor tissue. There was no significant difference observed on β-catenin either on score nor on staining area and intensity between tissues adjacent to the tumor and tumor tissues. A positive correlation was found between COX-1 and COX-2 in the tumor tissues. In conclusion, these suggest that Ki67, COXs, aromatase, prostaglandin, free radicals, adipokines, β-catenin and α-SMA are involved in breast cancer. These further focus the need of examination of tissues adjacent to tumor, tumor itself and compare them with normal or benign breast tissues for a better understanding of breast cancer pathology and future evaluation of therapeutic benefit.     

Comparing the feasibility, acceptability, clinical-, and cost-effectiveness of mental health e-screening to paper-based screening on the detection of depression, anxiety, and psychosocial risk in pregnant women: a study protocol of a randomized, parallel-group, superiority trial

Background

Juvenile idiopathic arthritis (JIA) is the most common rheumatic disease in children. Children with JIA are at risk of inflammation of the uvea in the eye (uveitis). Overall, 20% to 25% of paediatric uveitis is associated with JIA. Major risk factors for development of uveitis in JIA are oligoarticular pattern of arthritis, an age at onset of arthritis of less than seven years of age, and antinuclear antibody positivity. In the initial stages of mild to moderate inflammation the uveitis is asymptomatic. This has led to current practice of screening all children with JIA for uveitis. Approximately 12% to 38% of patients with JIA develop uveitis in seven years following onset of arthritis. In 30% to 50% of children with JIA-associated uveitis structural complications are present at diagnosis. Furthermore about 50% to 75% of those with severe uveitis will eventually develop visual impairment secondary to ocular complications such as cataract and glaucoma. Defining the severity of inflammation and structural complications in uveitis patients is now possible following Standardised Uveitis Nomenclature (SUN) guidelines, and modified to incorporate the consensus of end point and outcome criteria into the design of randomised trials. Despite current screening and therapeutic options (pre-biologics) 10% to 15% of children with JIA-associated uveitis may develop bilateral visual impairment and certified legally blind. To date, there remains no controlled trial evidence of benefits of biologic therapy.

Methods/design

This study will randomise 154 patients aged 2 to 18 years with active JIA-associated uveitis (despite methotrexate (MTX) treatment for at least 12 weeks). All participants will be treated for 18 months, with follow up of 3 years from randomisation (continuing on MTX throughout). All participants will receive a stable dose of MTX and in addition either adalimumab (20 mg/0.8 ml for patients <30 kg or 40 mg/0.8 ml for patients weighing 30 kg or more, subcutaneous (s/c) injection every 2 weeks based on body weight), or placebo (0.8 ml as appropriate according to body weight) s/c injection every 2 weeks.

Discussion

This is the first randomised controlled trial that will assess the clinical effectiveness, safety and cost effectiveness of adalimumab in combination with methotrexate for the treatment of juvenile idiopathic arthritis associated uveitis.

Trial registration

ISRCTN10065623     

Antibody informatics for drug discovery

More and more antibody therapeutics are being approved every year, mainly due to their high efficacy and antigen selectivity. However, it is still difficult to identify the antigen, and thereby the function, of an antibody if no other information is available. There are obstacles inherent to the antibody science in every project in antibody drug discovery. Recent experimental technologies allow for the rapid generation of large-scale data on antibody sequences, affinity, potency, structures, and biological functions; this should accelerate drug discovery research. Therefore, a robust bioinformatic infrastructure for these large data sets has become necessary. In this article, we first identify and discuss the typical obstacles faced during the antibody drug discovery process. We then summarize the current status of three sub-fields of antibody informatics as follows: (i) recent progress in technologies for antibody rational design using computational approaches to affinity and stability improvement, as well as ab-initio and homology-based antibody modeling; (ii) resources for antibody sequences, structures, and immune epitopes and open drug discovery resources for development of antibody drugs; and (iii) antibody numbering and IMGT. Here, we review “antibody informatics,” which may integrate the above three fields so that bridging the gaps between industrial needs and academic solutions can be accelerated. This article is part of a Special Issue entitled: Recent advances in molecular engineering of antibody.     

G-quadruplex-induced instability during leading-strand replication

G-quadruplexes are four-stranded nucleic acid structures whose biological functions remain poorly understood. In the yeast S. cerevisiae, we report that G-quadruplexes form and, if not properly processed, pose a specific challenge to replication. We show that the G-quadruplex-prone CEB1 tandem array is tolerated when inserted near ARS305 replication origin in wild-type cells but is very frequently destabilized upon treatment with the potent Phen-DC(3) G-quadruplex ligand, or in the absence of the G-quadruplex-unwinding Pif1 helicase, only when the G-rich strand is the template of leading-strand replication. The orientation-dependent instability is associated with the formation of Rad51-Rad52-dependent X-shaped intermediates during replication detected by two-dimensional (2D) gels, and relies on the presence of intact G-quadruplex motifs in CEB1 and on the activity of ARS305. The asymmetrical behaviour of G-quadruplex prone sequences during replication has implications for their evolutionary dynamics within genomes, including the maintenance of G-rich telomeres.