Studies of the light-induced signal transduction in Agrobacterium fabrum

Organisms regardless of their origin or type, monitor their environment and respond differently to any environmental changes that occur. These changes not only can alter biochemical reactions that are taking part in the signal transmission, but also the properties of the proteins involved. Therefore, biochemical, or biophysical analyses are a useful approach to understand bacterial cellular mechanisms such as the ones in the bacterium Agrobacterium fabrum (former Agrobacterium tumefaciens) and its phytochrome system. The biliprotein photoreceptors Agp1 and Agp2, found in A. fabrum, exhibit opposite spectral characteristics regarding light-induced and thermal conversion between the red absorbing Pr form and the far-red absorbing Pfr form and could control various cellular mechanisms. These phytochromes have been shown to be able to act together, regulating various mechanisms or processes, such as bacterial conjugation, plant infection and cell growth, in addition to the phosphorylation, and their photoconversion ability. Understanding a signal transduction pathway requires the understanding of the function of proteins, which in turn requires a detailed knowledge of their structure, dynamics, and the different conformational states they can adopt. Therefore, structures studies by X-ray crystallography of the photosensory core modules of various phytochromes were analysed, revealing insights into their conformational changes upon photoconversion. Various hypothesises were already published, on how the structure of phytochromes varies upon photoconversion and in transducing the light signal. Nonetheless, it is still not clear how this signal transduction pathway occurs and how phytochromes are involved in. For this reason, it is aimed in this research to understand the signal transduction pathways of a bacterial phytochrome. These pathways are protein-driven processes, involving photoisomerization, light induced protein conformational changes and possible protein-protein interactions. Within this work, in vitro studies using Agp1 mutants were performed. These mutants were prepared to introduce a single cysteine residue at defined positions, enabling targeted labelling with a maleimide fluorophore. First, the influence of mutations in Agp1 on the autophosphorylation was examined, to determine that these single mutations did not significantly affect the protein function, serving thus as a control experiment. The autophosphorylation results of Agp1 mutants revealed a similar behaviour as for Agp1 wild type, exhibiting a higher autophosphorylation activity in the Pr form than in the Pfr form. Subsequently, these Agp1 mutants were then investigated by time resolved fluorescence anisotropy, to gain insight into the dynamics of the protein. Results have indeed shown that the dynamics of the phytochrome at the corresponding (sub-) domain, changes between the Pr and Pfr form for both the PCM and HK modules, where mainly the dynamics in the Pfr form is lower than in the Pr form. Thus, emphasizing the conformational changes of the phytochrome upon photoconversion that are involved in the transmission of the light signal. Moreover, interaction studies were also carried out via FRET, where Agp1 and Agp2 were labelled with different fluorophores. Results have indeed shown an interaction between both phytochromes, suggesting that the PAS-GAF bidomain of Agp2 interacts with the histidine kinase module of Agp1 and that the FRET efficiency is light-regulated. A different approach for the study of the signal transduction in A. fabrum was realised through in vivo analysis of the bacterial conjugal transfer, since it was proved to be affected by the red light sensitive phytochromes and by their histidine kinases. The conjugation assay in this work revealed the importance of the TraA gene, which is supposed to be the first protein involved in the conjugation cascade of Agp1. Three TraA homologues are presented in A. fabrum and are encoded by the Ti plasmid (Atu6127), the linear chromosome and the At plasmid. Results showed that the knockout of the TraA encoded by the pTi, inhibited the conjugal transfer in A. fabrum and that phytochromes regulate the expression of the other TraA genes encoded by the linear chromosome and the pAt

In-situ Bioengineering of Arterial Vein Grafts

The autogenous saphenous vein remains the graft of choice for both coronary (500,000 annually in the US) and peripheral (80,000 annually) arterial bypass procedures. Failure of arterial vein grafts (AVGs) remains a major problem, and patients with failed grafts will die or require re-operation. Intimal hyperplasia (IH) accounts for 20% to 40% of all AVG failures. It is believed that this adverse pathological response by AVGs is largely due to their abrupt exposure to the significantly elevated circumferential wall stress (CWS) associated with the arterial system. We believe that if an AVG is given an ample opportunity to adapt and remodel to the stresses of its new environment, cellular injury may be reduced, thus limiting the initiating mechanisms of IH. The goal of this work was to develop a new mechanical conditioning paradigm, in the form of a peri-adventitially placed, biodegradable polymer wrap, to safely and functionally "arterialize" AVGs in situ. The polymer wrap was tuned so that as it degraded over a desired period of time, the mechanical support offered by it was reduced and the vein was exposed to gradually increasing levels of CWS in situ. To investigate the effects of mechanical conditioning on AVGs, we utilized both our well established, validated ex vivo vascular perfusion system (EVPS) as well as an appropriate preclinical animal model. The "engineering" component of this bioengineering study was to enhance our EVPS capabilities. Enhancements were made in the form of rigorous mathematical modeling, via subspace system identification, and automatic feedback control, via proportional integral and derivative control, of the arterial CWS and shear stress waveform generation capabilities of the EVPS. Pairs of freshly harvested porcine internal jugular veins (PIJVs) were perfused ex vivo under several biomechanical conditions. The acute hyperplastic response of PIJVs abruptly exposed to arterial hemodynamic conditions was compared to PIJVs perfused under normal venous conditions. In an attempt to attenuate this acute hyperplastic response, an ex vivo mechanical conditioning paradigm was imposed onto the PIJVs both via manual adjustment of EVPS parameters and via an adventitially placed tuned electrospun biodegradable polymer wrap. Early markers of IH were evaluated post-perfusion, and they included vascular smooth muscle cell apoptosis, proliferation, and phenotypic modulation. Quantification of these markers via immunohistochemical techniques provided the foundation for the final stage of this work. To assess the efficacy of the tuned electrospun biodegradable polymer wrap in attenuating the development of intimal hyperplasia in AVGs, a series of preclinical studies was performed in a pig model.PIJVs abruptly exposed to arterial levels of CWS showed a significant increase in apoptosis and in the number of synthetic smooth muscle cells, as well as a decrease in proliferation. Mechanical conditioning, via both manual adjustment of the EVPS parameters and placement of the biodegradable adventitial wrap, appeared to have beneficial effects on the acute hyperplastic response of PIJVs perfused ex vivo. The beneficial effects of the adventitially placed polymer wrap was also observed in vivo, however the results did not achieve significance over unwrapped controls. Future work should be aimed at enhancing the beneficial effects of the electrospun biodegradable polymer wrap by incorporating the delivery of drugs and/or stem cells in addition to the delivery of structural support to AVGs

Comparing monolithic and fused core HPLC columns for fast chromatographic analysis of fat soluble vitamins

HPLC stationary phases of monolithic and fused core type can be used to achieve fast chromatographic separation as an alternative to UPLC. In this study, monolithic and fused core stationary phases are compared for fast separation of four fat-soluble vitamins. Three new methods on the first and second generation monolithic silica RP-18e columns and a fused core pentafluoro-phenyl propyl column were developed. Application of three fused core columns offered comparable separations of retinyl palmitate, DL-α-tocopheryl acetate, cholecalciferol and menadione in terms of elution speed and separation efficiency. Separation was achieved in approx. 5 min with good resolution (Rs >5) and precision (RSD ≤ 0.6 %). Monolithic columns showed, however, a higher number of theoretical plates, better precision and lower column backpressure than the fused core column. The three developed methods were successfully applied to separate and quantitate fat-soluble vitamins in commercial products

Tracheobronchomalacia: An Unreported Pulmonary Complication of Acute Pancreatitis

Acute Pancreatitis (AP) is a common disease with systemic complications, specifically pulmonary complications that are well-documented [1]. Here we present, to the best of our knowledge, the first reported case of tracheobronchomalacia as a respiratory complication of AP. A 54-year-old white male with multiple chronic comorbidities developed necrotizing acute pancreatitis (NAP) following a surgical procedure. Internal Medicine evaluated and managed his NAP according to protocol. Within one week of NAP onset, the patient developed rapid respiratory distress. Chest radiography and ABGs were unable to diagnose ARDS. A CT scan with IV contrast was completed to investigate a pulmonary embolus and found the tracheal diameter variations during inspiration and expiration of the respiratory cycle consistent with tracheobronchomalacia (TBM). The patient’s respiratory status continued to deteriorate requiring endotracheal intubation and mechanical ventilation with weaning trials proving to be futile. The patient eventually developed fungemia and expired after his family opted for palliative extubation. Airway collapse related to TBM is an under-recognized diagnosis which should be suspected in patients with NAP who develop acute respiratory distress in whom no specific etiology has been determined

Neural Architecture Search for Effective Teacher-Student Knowledge Transfer in Language Models

Large pretrained language models have achieved state-of-the-art results on a variety of downstream tasks. Knowledge Distillation (KD) into a smaller student model addresses their inefficiency, allowing for deployment in resource-constrained environments. However, KD can be ineffective when the student is manually selected from a set of existing options, since it can be a sub-optimal choice within the space of all possible student architectures. We develop multilingual KD-NAS, the use of Neural Architecture Search (NAS) guided by KD to find the optimal student architecture for task agnostic distillation from a multilingual teacher. In each episode of the search process, a NAS controller predicts a reward based on the distillation loss and latency of inference. The top candidate architectures are then distilled from the teacher on a small proxy set. Finally the architecture(s) with the highest reward is selected, and distilled on the full training corpus. KD-NAS can automatically trade off efficiency and effectiveness, and recommends architectures suitable to various latency budgets. Using our multi-layer hidden state distillation process, our KD-NAS student model achieves a 7x speedup on CPU inference (2x on GPU) compared to a XLM-Roberta Base Teacher, while maintaining 90% performance, and has been deployed in 3 software offerings requiring large throughput, low latency and deployment on CPU.Comment: 11 pages, 5 figure

k.p theory beyond standard 8-band theory parametrization strategies and its applicability in electronic and optoelectronic devices design

International audienc

Knowledge Sharing in Higher Education Institutions: A Systematic Review

Purpose – The purpose of this paper is to help in providing a better understanding on knowledge sharing amongst academics in Higher Education Institutions (HEIs). The aim in this study is realized by profiling existing literature to understand the determinants of knowledge sharing, research trends, theories and future research opportunities. Design/methodology/approach – After carefully examining the extant literature, and by utilising relevant academic-based research databases, a total of 73 papers published in peer-reviewed journals over the last decade were reviewed and analysed using well established systematic literature review methodology. Findings – The adopted systematic review revealed that there is limited contributions in understanding knowledge sharing in HEIs when compared with other sectors. The review provides a number of avenues for future research including technological, cultural, organisational and behavioural aspects at different levels. Practical implications – This study helps in offering a focal point to senior management in HEIs for realising the requirements for developing appropriate strategies and programmes to promote knowledge sharing among academics and consequently enhance their institutions’ performance. Originality/value – This study utilised Jesson et al. (2011) in presenting a comprehensive systematic review of knowledge sharing specifically in the context of HEIs. This paper offers some theoretical and practical insights on what contributes towards understating the determinates affecting knowledge sharing practices among academics

FACTORS INFLUENCING POLITICAL PARTICIPATION IN LEBANON: THE MEDIATING ROLE OF PERCEIVED CONGRUENCE

Abstract This study analyzes the factors that influence political participation. These factors include: political information efficacy, political interest, community engagement, political party affiliation and perceived congruence. Moreover, this study addresses the mediating role of perceived congruence on the relationship between political interest, community engagement and political participation. A quantitative survey method was used and structured questionnaire was administered to a convenience sample of 412 respondents. The findings of this study revealed that political interest, community engagement, and political party affiliation have a positive effect on political participation. In addition, the results indicated that perceived congruence has no direct or mediate effect on political participation. The current study enhances marketing literature to understand political behaviors under unusual political situations. In contrast, this research supports both political parties and governments for better understanding the factors that influence political participation which guide them to fulfill their political marketing objectives and gain citizens’ support