Characterization of the caspase-3 cleavage motif of the Salmonella Typhimurium effector protein SifA and its role in pathogenesis

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a Gram-negative facultative anaerobe that induces severe inflammation resulting in gastroenteritis. In the case of S. Typhimurium infection, induction of an inflammatory response has been linked to its primary virulence mechanism, the type III secretion system (T3SS). The T3SS secretes protein effectors that exploit the host’s cell biology to facilitate bacterial entry and intracellular survival, and to modulate the host immune response. One such effector, SifA, is a bi-functional T3SS effector protein that plays an important role in Salmonella virulence. The N-terminal domain of SifA binds SifA-Kinesin-Interacting-Protein (SKIP), and via an interaction with kinesin, forms tubular membrane extensions called Sif filaments (Sifs) that emanate from the Salmonella Containing Vacuole (SCV). The C-terminal domain of SifA harbors a WxxxE motif that functions to mimic active host cell GTPases. Taken together, SifA functions in inducing endosomal tubulation in order to maintain the integrity of the SCV and promote bacterial dissemination. Since SifA performs multiple, unrelated functions, the objective of this study was to determine how each functional domain of SifA becomes processed. In the present study, we demonstrate that a linker region containing a caspase-3 cleavage motif separates the two functional domains of SifA. To test the hypothesis that processing of SifA by caspase-3 at this particular site is required for function and proper localization of the effector protein domains, we developed two tracking methods to analyze the intracellular localization of SifA. We first adapted a fluorescent tag called phiLOV that allowed for T3SS mediated delivery of SifA and observation of its intracellular colocalization with caspase-3. Additionally, we created a dual-tagging strategy that permitted tracking of each of the SifA functional domains following caspase-3 cleavage to different subcellular locations. The results of this study reveal that caspase-3 cleavage of SifA is required for the proper localization of functional domains and bacterial dissemination. Considering the importance of these events in Salmonella pathogenesis, we conclude that caspase-3 cleavage of effector proteins is a more broadly applicable effector processing mechanism utilized by Salmonella to invade and persist during infection

An Investigation Of Counselor Educators\u27 Attitudes Towards Evidence-based Practices And Perceived Barriers To The Incorporat

The overall purpose of this study was to investigate counselor educators\u27 attitudes towards evidence-based practices (EBPs) and perceived barriers to the inclusion of EBPs in counselor education curricula. Additionally, this study aimed to assess whether counselor educators\u27 level of agreement towards the presence of motivational interviewing (MI) principles in the counseling relationship impacted attitudes towards EBPs. As such, this researcher analyzed four research questions using two instruments and a demographic questionnaire. Two hundred sixty nine counselor educators (39.8% response rate) from the Association of Counselor Education and Supervision responded to an electronic survey, which consisted of the Evidence-Based Practice Attitude Scale (EBPAS; Aarons, 2004), the BARRIERS Scale (Funk, Champagne, Wiese, & Tornquist, 1991), and a demographic questionnaire. Specifically, this study investigated four research questions to determine: (a) the difference in attitude towards adopting EBPs among counselor educators with respect to specific individual factors (i.e. specialized training in evidence-based practices, years of professoriate experience, and primary counselor education focus); (b) the difference in perceived barriers towards adopting EBPs into counselor education curricula among counselor educators with respect to organizational factors (i.e. type of program, status of CACREP accreditation, and faculty position); (c) the influence of EBP attitude on perceived barriers to the inclusion of EBPs in counselor education curricula; and (d) the correlation between counselor educators reported level of agreement towards MI principles\u27 presence in the counseling relationship and their attitude towards EBPs. Multivariate analyses of variance (MANOVA) were computed to analyze the data for the first two research questions, while linear regressions were utilized to compute the data for the last two research questions. In terms of individual factors, study results indicated that neither specialized training in EBPs nor years of professoriate experience resulted in significant differences with regards to attitudes towards EBPs. However, data analysis did reveal a significant difference between counselor educators with a clinical focus and counselor educators with a vocational focus. With regards to organizational factors influence on perceived barriers to the inclusion of EBPs in counselor education curricula, analyses revealed that neither CACREP accreditation nor faculty position resulted in any significant differences. Although, analysis did reveal that counselor educators in masters only programs perceived significantly less barriers to the inclusion of EBPs than did counselor educators in doctorate granting programs. Furthermore, results suggested a negative correlation between attitude towards EBPs and barriers towards the inclusion of EBPs in counselor education curricula, and a positive correlation between counselor educators\u27 agreement towards the inclusion of MI principles in the counseling relationship and their attitudes towards EBPs. Limitations of the study, implications for this study, and recommendations for future research as it relates to EBPs in counselor education and the counseling profession are addressed

‘Islands of understanding’ : Environmental journalism in the South Pacific

"We're expected to know too much. Journalists who specialise in science and the environment have to grasp, communicate and synthesise scientific, political and economic issues. And we have to do it on deadline with accuracy, authority and readability. Sometimes it can be overwhelming...

SYNTHESIS OF FLAVONES FROM 2-HYDROXY ACETOPHENONE AND AROMATIC ALDEHYDE DERIVATIVES BY CONVENTIONAL METHODS AND GREEN CHEMISTRY APPROACH

Objective: Flavones occupy a special place in the realm of natural and synthetic organic chemistry owing to their diversified biological activities. In this study, a series of chalcone derivatives were synthesized and after cyclization of chalcone to synthesized various substituted flavone derivatives (2A-2L). 2 Methods: The reaction of 2-hydroxy acetophenone with substituted aromatic aldehydes produced chalcone by trituration (NaOH) and conventional methods (KOH/EtOH), which upon further cyclization with dimethyl sulfoxide/I  resulted to form flavone derivatives.Results: The purity of compounds was ascertained by melting point and thin-layer chromatography. The synthesized compounds have been characterized by mass, infrared, and1H nuclear magnetic resonance spectral analysis.Conclusion: Based on spectral data, it was proved that all synthesized chalcones and flavones derivatives meet the standard values of various spectral techniques and further it will be evaluated for pharmacological activities.Keywords: Chalcone, Flavone, Trituration, Conventional, Claisen-Schmidt condensation

Identification of chalcone derivatives as putative non-steroidal aromatase inhibitors potentially useful against breast cancer by molecular docking and ADME prediction

Aromatase is an influential target to overcome estrogen receptor positive breast cancer, as the enzyme is responsible for conversion of androstenedione to estrone, a promising drug target for therapeutic management of breast cancer. Chalcones are prominent biosynthetic compounds and parent candidate for the synthesis of heterocycles with diversified biological activities. The prime objective of the present study is to evaluate the binding interaction of 2-hydroxyphenyl- prop-2-en-1-one (1A-1X), 2-hydroxy-4-methoxyphenyl- prop-2-en-1-one (3A-3X), 2,4-dihydroxyphenyl- prop-2-en-1-one (9A-9X) and 1-hydroxynaphthalen-2-yl-prop-2-en-1-one (5A-5X) derivatives with aromatase enzyme by molecular docking study and also check their ADME properties by maestro suit. The designed chalcones derivatives have been docked against our target protein with PDB id 3S7S retrieved from the protein data bank, whereas exemestane has been taken as the positive control. As docking data revealed that docking score of 1K, 1U, 1B 3K 3N, 5K, 5U, 9S, 9K, 9N and 9F compounds found less than exemestane and all of these compounds with appropriate ADME properties have proven their excellent absorption as well as solubility characteristics. The present findings provided valuable information about binding interactions of chalcones derivatives to the active site of aromatase. These compounds may serve as potential lead compound for developing new aromatase inhibitors in breast cancer treatment

Identification of chalcone derivatives as putative non-steroidal aromatase inhibitors potentially useful against breast cancer by molecular docking and ADME prediction

283-293Aromatase is an influential target to overcome estrogen receptor positive breast cancer, as the enzyme is responsible for conversion of androstenedione to estrone, a promising drug target for therapeutic management of breast cancer. Chalcones are prominent biosynthetic compounds and parent candidate for the synthesis of heterocycles with diversified biological activities. The prime objective of the present study is to evaluate the binding interaction of 2-hydroxyphenyl- prop-2-en-1-one (1A-1X), 2-hydroxy-4-methoxyphenyl- prop-2-en-1-one (3A-3X), 2,4-dihydroxyphenyl- prop-2-en-1-one (9A-9X) and 1-hydroxynaphthalen-2-yl-prop-2-en-1-one (5A-5X) derivatives with aromatase enzyme by molecular docking study and also check their ADME properties by maestro suit. The designed chalcones derivatives have been docked against our target protein with PDB id 3S7S retrieved from the protein data bank, whereas exemestane has been taken as the positive control. As docking data revealed that docking score of 1K, 1U, 1B 3K 3N, 5K, 5U, 9S, 9K, 9N and 9F compounds found less than exemestane and all of these compounds with appropriate ADME properties have proven their excellent absorption as well as solubility characteristics. The present findings provided valuable information about binding interactions of chalcones derivatives to the active site of aromatase. These compounds may serve as potential lead compound for developing new aromatase inhibitors in breast cancer treatment

FORMULATION OPTIMIZATION AND EVALUATION OF MOUTH DISSOLVING FILM OF RAMOSETRON HYDROCHLORIDE

Objective: Ramosetron Hydrochloride is found to be more potent and having a longer duration of action with the least side effects, but the major drawback is it undergoes hepatic first-pass metabolism so our aim is to prepare mouth dissolving film (MDF) of Ramosetron hydrochloride for rapid relief in emesis. Methods: The mouth dissolving films of Ramosetron Hydrochloride were prepared by using the solvent casting method. Films were formulated using HPMC E5 (Hydroxy Propyl Methyl Cellulose) as a film-forming agent, PEG400 (Polyethylene glycol) as a plasticizer and Aspartame as the sweetening agent. A 32 full factorial design was applied considering the concentration of HPMC E5 (X1) and concentration of PEG400 (X2) as independent variables and % cumulative drug release (Y1) (CDR), disintegration time (Y2) (DT) and tensile strength (Y3) (TS) as dependent variables. The prepared films were evaluated for thickness, folding endurance, tensile strength, disintegration time, drug content uniformity and taste masking by E-tongue. The results indicated that factors X1 and X2 were found to be having a positive effect on DT and TS and negative effects on CDR. Results: The optimized formulation was found to be the best with 94.00±0.85% in vitro drug release, 33.22±0.75 sec DT and 1.359±0.005 g/mm2 tensile strength. Concentration of aspartame was optimized with E-tongue taking into consideration increased electric potential with decreasing bitterness. Conclusion: Thus, a rapidly dissolving oral film of Ramosetron Hydrochloride with successful taste masking and immediate in vitro drug release was prepared using a solvent casting technique

DEVELOPMENT AND VALIDATION OF A STABILITY INDICATING RP-HPLC METHOD FOR THE DETERMINATION OF POTENTIAL DEGRADATION PRODUCTS OF DIFLUPREDNATE IN OPHTHALMIC EMULSION

Objective: The objective of the current study was to develop and validate a simple, robust, precise and accurate RP-HPLC (reverse phase-high performance liquid chromatography) method for the quantitative determination of potential degradation products of Difluprednate (DIFL) in the ophthalmic emulsion.Methods: Chromatographic separation was achieved on the YMC pack ODS-AQ (150× 4.6) mm, 3μm column with a mobile phase containing a gradient mixture of mobile phase A (0.02M Ammonium formate buffer pH 4.5 adjusted with formic acid) and Acetonitrile as mobile phase B, at flow rate of 1.5 ml/min and with UV detection at 240 nm.Results: The peak retention time of DIFL was found at about 17.2 min, the RRT of degradation product-1 (DP-1), degradation product-2 (DP-2), and degradation product-3 (DP-3), were found to be about 0.49, 0.65 and 0.79 respectively (calculated with respect to Difluprednate). Stress testing was performed in accordance with an ICH (international council for harmonisation) guideline Q1A (R2) [1]. The method was validated as per ICH guideline Q2 (R1)[2]. The calibration curve was found to be linear in the concentration range of 0.1 to 0.75 µg/ml for Difluprednate, DP-1, DP-2 and DP-3. The LOD (Limit of detection) was found to be 0.1µg/ml and LOQ (Limit of quantification) of 0.15µg/ml for Difluprednate, DP-1, DP-2 and DP-3 respectively. The recovery from LOQ to 150% was within 90-110%. The forced degradation data confirms the stability indicating the nature of the method.Conclusion: A simple, robust, precise and accurate RP-HPLC method for the quantitative determination of potential degradation products of Difluprednate in the ophthalmic emulsion was developed and validated.Â

A Software Engineering Team's Perspective on the Switch to Agile Software Development

Healthcare information technology is one of the fastest growing industries in the world. There are millions of errors made in the medical paper chart at hospitals around the world, resulting in mistaken medication and death. Healthcare information technology provides a way to eliminate these errors and to allow patients access to their medical record from anywhere in the world. Corporation X is the foremost leader in the industry and has been providing solutions to hospitals and doctors’ offices for the past 30+ years. Developing software that affects peoples’ lives requires many processes and certifications. As a result, the software development process is instrumental in providing high quality software for Corporation X’s clients. In the past, Corporation X has employed the Waterfall Software Methodology to design and implement their software. This methodology or process has been in use since the beginning of software. There are distinct phases that need to be sequentially followed, requiring much up-front work. There are many advantages and disadvantages to this process. As the healthcare information technology industry grows and new competitors make their way to market, Corporation X has been forced to re-think this process and adapt to the ever-changing external environment. The recent popularity of Agile Software Development has spread across the world. In the past 10 years, Agile has become the most used process for software development. In 2010, Corporation X decided to switch from Waterfall to Agile, to keep up with the industry and improve it’s own processes. The advantages of the switch to Agile have proven to be a great asset to Corporation X’s engineering teams. The new process has increased code quality, increased client satisfaction, and produced more revenue for Corporation X. There were, however, a few disadvantages such as resource pooling and team morale.. A new concept was introduced with Agile, resource pooling. This was Corporation X’s own spin on resourcing, not something that is traditionally a part of Agile practices at most companies. This proved to be a disadvantage because it caused teams to lose that camaraderie and closeness that is typical of a team. Engineers had to start over with people with every project. Corporation X also put a new defect accountability process in place to help drive down the number of defects by introducing the feeling of ownership. And with this new process, engineers became more aware of the number of defects being introduced to clients. All of these factors will be discussed and explained in this study. This study will help dissect the transition from Waterfall to Agile from the software engineers’ viewpoint