A Neuroprotective Bovine Colostrum Attenuates Apoptosis in Dexamethasone-Treated MC3T3-E1 Osteoblastic Cells

Glucocorticoid-induced osteoporosis (GIO) is one of the most common secondary forms of osteoporosis. GIO is partially due to the apoptosis of osteoblasts and osteocytes. In addition, high doses of dexamethasone (DEX), a synthetic glucocorticoid receptor agonist, induces neurodegeneration by initiating inflammatory processes leading to neural apoptosis. Here, a neuroprotective bovine colostrum against glucocorticoid-induced neuronal damage was investigated for its anti-apoptotic activity in glucocorticoid-treated MC3T3-E1 osteoblastic cells. A model of apoptotic osteoblastic cells was developed by exposing MC3T3-E1 cells to DEX (0–700 μM). Colostrum co-treated with DEX was executed at 0.1–5.0 mg/mL. Cell viability was measured for all treatment schedules. Caspase-3 activation was assessed to determine both osteoblast apoptosis under DEX exposure and its potential prevention by colostrum co-treatment. Glutathione reduced (GSH) was measured to determine whether DEX-mediated oxidative stress-driven apoptosis is alleviated by colostrum co-treatment. Western blot was performed to determine the levels of p-ERK1/2, Bcl-XL, Bax, and Hsp70 proteins upon DEX or DEX plus colostrum exposure. Colostrum prevented the decrease in cell viability and the increase in caspase-3 activation and oxidative stress caused by DEX exposure. Cells, upon colostrum co-treated with DEX, exhibited higher levels of p-ERK1/2 and lower levels of Bcl-XL, Bax, and Hsp70. Our data support the notion that colostrum may be able to reduce DEX-induced apoptosis possibly via the activation of the ERK pathway and modulation of the Hsp70 system. We provided preliminary evidence on how bovine colostrum, as a complex and multi-component dairy product, in addition to its neuroprotective action, may affect osteoblastic cell survival undergoing apoptosis

The influence of prior VATS experience in the learning curvefor single-port VATS lobectomy: a multicenter comparative study

OBJECTIVES: Competency in VATS lobectomy is estimated to be complete after 50 cases. We aimed to ex¬plore the impact of previous multiport VATS lobectomy competency in completing this work¬load of Single-Port procedures ...postprin

Spectrum and Inoculum Size Effect of a Rapid Antigen Detection Test for Group A Streptococcus in Children with Pharyngitis

BACKGROUND: The stability of the accuracy of a diagnostic test is critical to whether clinicians can rely on its result. We aimed to assess whether the performance of a rapid antigen detection test (RADT) for group A streptococcus (GAS) is affected by the clinical spectrum and/or bacterial inoculum size. METHODS: Throat swabs were collected from 785 children with pharyngitis in an office-based, prospective, multicenter study (2009-2010). We analysed the effect of clinical spectrum (i.e., the McIsaac score and its components) and inoculum size (light or heavy GAS growth) on the accuracy (sensitivity, specificity, likelihood ratios and predictive values) of a RADT, with laboratory throat culture as the reference test. We also evaluated the accuracy of a McIsaac-score-based decision rule. RESULTS: GAS prevalence was 36% (95CI: 33%-40%). The inoculum was heavy for 85% of cases (81%-89%). We found a significant spectrum effect on sensitivity, specificity, likelihood ratios and positive predictive value (p<0.05) but not negative predictive value, which was stable at about 92%. RADT sensitivity was greater for children with heavy than light inoculum (95% vs. 40%, p<0.001). After stratification by inoculum size, the spectrum effect on RADT sensitivity was significant only in patients with light inoculum, on univariate and multivariate analysis. The McIsaac-score-based decision rule had 99% (97%-100%) sensitivity and 52% (48%-57%) specificity. CONCLUSIONS: Variations in RADT sensitivity only occur in patients with light inocula. Because the spectrum effect does not affect the negative predictive value of the test, clinicians who want to rule out GAS can rely on negative RADT results regardless of clinical features if they accept that about 10% of children with negative RADT results will have a positive throat culture. However, such a policy is more acceptable in populations with very low incidence of complications of GAS infection

WW Domains of the Yes-Kinase-Associated-Protein (YAP) Transcriptional Regulator Behave as Independent Units with Different Binding Preferences for PPxY Motif-Containing Ligands

YAP is a WW domain-containing effector of the Hippo tumor suppressor pathway, and the object of heightened interest as a potent oncogene and stemness factor. YAP has two major isoforms that differ in the number of WW domains they harbor. Elucidating the degree of co-operation between these WW domains is important for a full understanding of the molecular function of YAP. We present here a detailed biophysical study of the structural stability and binding properties of the two YAP WW domains aimed at investigating the relationship between both domains in terms of structural stability and partner recognition. We have carried out a calorimetric study of the structural stability of the two YAP WW domains, both isolated and in a tandem configuration, and their interaction with a set of functionally relevant ligands derived from PTCH1 and LATS kinases. We find that the two YAP WW domains behave as independent units with different binding preferences, suggesting that the presence of the second WW domain might contribute to modulate target recognition between the two YAP isoforms. Analysis of structural models and phage-display studies indicate that electrostatic interactions play a critical role in binding specificity. Together, these results are relevant to understand of YAP function and open the door to the design of highly specific ligands of interest to delineate the functional role of each WW domain in YAP signaling.This work was supported by the Spanish Ministry of Education and Science [grant BIO2009-13261-CO2], the Spanish Ministry of Economy and Competitivity [grant BIO2012-39922-CO2] including FEDER (European Funds for Regional Development) funds and the Governement of Andalusia [grant CVI-5915]. Marius Sudol was supported by PA Breast Cancer Coalition Grants (#60707 and #920093) plus the Geisinger Clinic

Metabolomic Profiling of Drug Responses in Acute Myeloid Leukaemia Cell Lines

Combined bezafibrate (BEZ) and medroxyprogesterone acetate (MPA) exert unexpected antileukaemic activities against acute myeloid leukaemia (AML) and these activities are associated with the generation of reactive oxygen species (ROS) within the tumor cells. Although the generation of ROS by these drugs is supported by preceding studies including our own, the interrelationship between the cellular effects of the drugs and ROS generation is not well understood. Here we report the use of NMR metabolomic profiling to further study the effect of BEZ and MPA on three AML cell lines and to shed light on the underlying mechanism of action. For this we focused on drug effects induced during the initial 24 hours of treatment prior to the onset of overt cellular responses and examined these in the context of basal differences in metabolic profiles between the cell lines. Despite their ultimately profound cellular effects, the early changes in metabolic profiles engendered by these drugs were less pronounced than the constitutive metabolic differences between cell types. Nonetheless, drug treatments engendered common metabolic changes, most markedly in the response to the combination of BEZ and MPA. These responses included changes to TCA cycle intermediates consistent with recently identified chemical actions of ROS. Notable amongst these was the conversion of α-ketoglutarate to succinate which was recapitulated by the treatment of cell extracts with exogenous hydrogen peroxide. These findings indicate that the actions of combined BEZ and MPA against AML cells are indeed mediated downstream of the generation of ROS rather than some hitherto unsuspected mechanism. Moreover, our findings demonstrate that metabolite profiles represent highly sensitive markers for genomic differences between cells and their responses to external stimuli. This opens new perspectives to use metabolic profiling as a tool to study the rational redeployment of drugs in new disease settings