Early Ultrasound Surveillance of Newly-Created Hemodialysis Arteriovenous Fistula.

Introduction: We assess if ultrasound surveillance of newly-created arteriovenous fistulas (AVFs) can predict nonmaturation sufficiently reliably to justify randomized controlled trial (RCT) evaluation of ultrasound-directed salvage intervention. Methods: Consenting adults underwent blinded fortnightly ultrasound scanning of their AVF after creation, with scan characteristics that predicted AVF nonmaturation identified by logistic regression modeling. Results: Of 333 AVFs created, 65.8% matured by 10 weeks. Serial scanning revealed that maturation occurred rapidly, whereas consistently lower fistula flow rates and venous diameters were observed in those that did not mature. Wrist and elbow AVF nonmaturation could be optimally modeled from week 4 ultrasound parameters alone, but with only moderate positive predictive values (PPVs) (wrist, 60.6% [95% confidence interval, CI: 43.9-77.3]; elbow, 66.7% [48.9-84.4]). Moreover, 40 (70.2%) of the 57 AVFs that thrombosed by week 10 had already failed by the week 4 scan, thus limiting the potential of salvage procedures initiated by that scan's findings to alter overall maturation rates. Modeling of the early ultrasound characteristics could also predict primary patency failure at 6 months; however, that model performed poorly at predicting assisted primary failure (those AVFs that failed despite a salvage attempt), partly because patency of at-risk AVFs was maintained by successful salvage performed without recourse to the early scan data. Conclusion: Early ultrasound surveillance may predict fistula maturation, but is likely, at best, to result in only very modest improvements in fistula patency. Power calculations suggest that an impractically large number of participants (>1700) would be required for formal RCT evaluation.

Doppler ultrasound surveillance of recently formed haemodialysis arteriovenous fistula: the SONAR observational cohort study.

Background: Arteriovenous fistulas are considered the best option for haemodialysis provision, but as many as 30% fail to mature or suffer early failure. Objective: To assess the feasibility of performing a randomised controlled trial that examines whether, by informing early and effective salvage intervention of fistulas that would otherwise fail, Doppler ultrasound surveillance of developing arteriovenous fistulas improves longer-term arteriovenous fistula patency. Design: A prospective multicentre observational cohort study (the 'SONAR' study). Setting: Seventeen haemodialysis centres in the UK. Participants: Consenting adults with end-stage renal disease who were scheduled to have an arteriovenous fistula created. Intervention: Participants underwent Doppler ultrasound surveillance of their arteriovenous fistulas at 2, 4, 6 and 10 weeks after creation, with clinical teams blinded to the ultrasound surveillance findings. Main outcome measures: Fistula maturation at week 10 defined according to ultrasound surveillance parameters of representative venous diameter and blood flow (wrist arteriovenous fistulas: ≥ 4 mm and > 400 ml/minute; elbow arteriovenous fistulas: ≥ 5 mm and > 500 ml/minute). Mixed multivariable logistic regression modelling of the early ultrasound scan data was used to predict arteriovenous fistula non-maturation by 10 weeks and fistula failure at 6 months. Results: A total of 333 arteriovenous fistulas were created during the study window (47.7% wrist, 52.3% elbow). By 2 weeks, 37 (11.1%) arteriovenous fistulas had failed (thrombosed), but by 10 weeks, 219 of 333 (65.8%) of created arteriovenous fistulas had reached maturity (60.4% wrist, 67.2% elbow). Persistently lower flow rates and venous diameters were observed in those fistulas that did not mature. Models for arteriovenous fistulas' non-maturation could be optimally constructed using the week 4 scan data, with fistula venous diameter and flow rate the most significant variables in explaining wrist fistula maturity failure (positive predictive value 60.6%, 95% confidence interval 43.9% to 77.3%), whereas resistance index and flow rate were most significant for elbow arteriovenous fistulas (positive predictive value 66.7%, 95% confidence interval 48.9% to 84.4%). In contrast to non-maturation, both models predicted fistula maturation much more reliably [negative predictive values of 95.4% (95% confidence interval 91.0% to 99.8%) and 95.6% (95% confidence interval 91.8% to 99.4%) for wrist and elbow, respectively]. Additional follow-up and modelling on a subset (n = 192) of the original SONAR cohort (the SONAR-12M study) revealed the rates of primary, assisted primary and secondary patency arteriovenous fistulas at 6 months were 76.5, 80.7 and 83.3, respectively. Fistula vein size, flow rate and resistance index could identify primary patency failure at 6 months, with similar predictive power as for 10-week arteriovenous fistula maturity failure, but with wide confidence intervals for wrist (positive predictive value 72.7%, 95% confidence interval 46.4% to 99.0%) and elbow (positive predictive value 57.1%, 95% confidence interval 20.5% to 93.8%). These models, moreover, performed poorly at identifying assisted primary and secondary patency failure, likely because a subset of those arteriovenous fistulas identified on ultrasound surveillance as at risk underwent subsequent successful salvage intervention without recourse to early ultrasound data. Conclusions: Although early ultrasound can predict fistula maturation and longer-term patency very effectively, it was only moderately good at identifying those fistulas likely to remain immature or to fail within 6 months. Allied to the better- than-expected fistula patency rates achieved (that are further improved by successful salvage), we estimate that a randomised controlled trial comparing early ultrasound-guided intervention against standard care would require at least 1300 fistulas and would achieve only minimal patient benefit. Trial Registration: This trial is registered as ISRCTN36033877 and ISRCTN17399438. Funding: This award was funded by the National Institute for Health and Care Research (NIHR) Health Technology Assessment programme (NIHR award ref: NIHR135572) and is published in full in Health Technology Assessment; Vol. 28, No. 24. See the NIHR Funding and Awards website for further award information.

For people with advanced kidney disease, haemodialysis is best provided by an 'arteriovenous fistula', which is created surgically by joining a vein onto an artery at the wrist or elbow. However, these take about 2 months to develop fully ('mature'), and as many as 3 out of 10 fail to do so. We asked whether we could use early ultrasound scanning of the fistula to identify those that are unlikely to mature. This would allow us to decide whether it would be practical to run a large, randomised trial to find out if using early ultrasound allows us to 'rescue' fistulas that would otherwise fail. We invited adults to undergo serial ultrasound scanning of their fistula in the first few weeks after it was created. We then analysed whether we could use the data from the early scans to identify those fistulas that were not going to mature by week 10. Of the 333 fistulas that were created, about two-thirds reached maturity by week 10. We found that an ultrasound scan 4 weeks after fistula creation could reliably identify those fistulas that were going to mature. However, of those fistulas predicted to fail, about one-third did eventually mature without further intervention, and even without knowing what the early scans showed, another third were successfully rescued by surgery or X-ray-guided treatment at a later stage. Performing an early ultrasound scan on a fistula can provide reassurance that it will mature and deliver trouble-free dialysis. However, because scans are poor at identifying fistulas that are unlikely to mature, we would not recommend their use to justify early surgery or X-ray-guided treatment in the expectation that this will improve outcomes.

The Interplay between Heat Shock Proteins and Cancer Pathogenesis: A Novel Strategy for Cancer Therapeutics.

Heat shock proteins (HSPs) are developmentally conserved families of protein found in both prokaryotic and eukaryotic organisms. HSPs are engaged in a diverse range of physiological processes, including molecular chaperone activity to assist the initial protein folding or promote the unfolding and refolding of misfolded intermediates to acquire the normal or native conformation and its translocation and prevent protein aggregation as well as in immunity, apoptosis, and autophagy. These molecular chaperonins are classified into various families according to their molecular size or weight, encompassing small HSPs (e.g., HSP10 and HSP27), HSP40, HSP60, HSP70, HSP90, and the category of large HSPs that include HSP100 and ClpB proteins. The overexpression of HSPs is induced to counteract cell stress at elevated levels in a variety of solid tumors, including anticancer chemotherapy, and is closely related to a worse prognosis and therapeutic resistance to cancer cells. HSPs are also involved in anti-apoptotic properties and are associated with processes of cancer progression and development, such as metastasis, invasion, and cell proliferation. This review outlines the previously mentioned HSPs and their significant involvement in diverse mechanisms of tumor advancement and metastasis, as well as their contribution to identifying potential targets for therapeutic interventions.

Crossroad between the Heat Shock Protein and Inflammation Pathway in Acquiring Drug Resistance: A Possible Target for Future Cancer Therapeutics.

The development of multidrug resistance (MDR) against chemotherapeutic agents has become a major impediment in cancer therapy. Understanding the underlying mechanism behind MDR can guide future treatment for cancer with better therapeutic outcomes. Recent studies evidenced that crossroads interaction between the heat shock proteins (HSP) and inflammatory responses under the tumor microenvironment plays a pivotal role in modulating drug responsiveness and drug resistance through a complex cytological process. This review aims to investigate the interrelationship between inflammation and HSP in acquiring multiple drug resistance and investigate strategies to overcome the drug resistance to improve the efficacy of cancer treatment. HSP plays a dual regulatory effect as an immunosuppressive and immunostimulatory agent, involving the simultaneous blockade of multiple signaling pathways in acquiring MDR. For example, HSP27 shows biological effects on monocytes by causing IL10 and TNFα secretion and blocking monocyte differentiation to normal dendritic cells and tumor-associated macrophages to promote cancer progression and chemoresistance. Thus, the HSP function and immune-checkpoint release modalities provide a therapeutic target for a therapeutically beneficial approach for enhancing anti-tumor immune responses. The interconnection between inflammation and HSP, along with the tumor microenvironment in acquiring drug resistance, has become crucial for rationalizing the effect of HSP immunomodulatory activity with immune checkpoint blockade. This relationship can overcome drug resistance and assist in the development of novel combinatorial cancer immunotherapy in fighting cancer with decreasing mortality rates.

ß-Cyclodextrin: a green supramolecular catalyst assisted eco-friendly one-pot three-component synthesis of biologically active substituted pyrrolidine-2-one.

A green, novel and eco-efficient synthetic route towards the synthesis of highly substituted bio-active pyrrolidine-2-one derivatives was demonstrated using ß-cyclodextrin, a water-soluble supramolecular solid as a green and eco-benign catalyst at room temperature under water-ethanol solvent medium. The exploration of the green catalyst ß-cyclodextrin for the metal-free one-pot three-component synthesis of a wide range of highly functionalized bio-active heterocyclic pyrrolidine-2-one moieties from easily available aldehydes and amines explains the superiority and uniqueness of this protocol.

Proteins and their functionalization for finding therapeutic avenues in cancer: Current status and future prospective.

Despite the remarkable advancement in the health care sector, cancer remains the second most fatal disease globally. The existing conventional cancer treatments primarily include chemotherapy, which has been associated with little to severe side effects, and radiotherapy, which is usually expensive. To overcome these problems, target-specific nanocarriers have been explored for delivering chemo drugs. However, recent reports on using a few proteins having anticancer activity and further use of them as drug carriers have generated tremendous attention for furthering the research towards cancer therapy. Biomolecules, especially proteins, have emerged as suitable alternatives in cancer treatment due to multiple favourable properties including biocompatibility, biodegradability, and structural flexibility for easy surface functionalization. Several in vitro and in vivo studies have reported that various proteins derived from animal, plant, and bacterial species, demonstrated strong cytotoxic and antiproliferative properties against malignant cells in native and their different structural conformations. Moreover, surface tunable properties of these proteins help to bind a range of anticancer drugs and target ligands, thus making them efficient delivery agents in cancer therapy. Here, we discuss various proteins obtained from common exogenous sources and how they transform into effective anticancer agents. We also comprehensively discuss the tumor-killing mechanisms of different dietary proteins such as bovine α-lactalbumin, hen egg-white lysozyme, and their conjugates. We also articulate how protein nanostructures can be used as carriers for delivering cancer drugs and theranostics, and strategies to be adopted for improving their in vivo delivery and targeting. We further discuss the FDA-approved protein-based anticancer formulations along with those in different phases of clinical trials.

Silica sand-supported nano zinc oxide-graphene oxide composite induced rapid photocatalytic decolorization of azo dyes under sunlight and improved antimicrobial activity.

Here, silica sand-supported heterojunction composite of nano zinc oxide (nZnO) and graphene oxide nanosheet (nZnO-GO@SS) was prepared, and its potential as an efficient photocatalyst for the degradation of methylene blue (MB) and Rhodamine-B (Rh-B) under sunlight was demonstrated. Transmission electron microscopy confirmed the uniform distribution of spherically shaped nZnO of average size of approximately 8 nm over graphene oxide nanosheet (GO) in the composites. Photodegradation yields of 95.3% and 97.5% for 100 ppm of MB and Rh-B dye within 150 and 220 min, respectively, were achieved under sunlight by the prepared nanocatalyst (nZnO-GO), while sand microparticle-supported nanocatalyst (nZnO-GO@SS) demonstrated faster degradation of MB and Rh-B, i.e., within 120 and 160 min, respectively. Furthermore, when the recyclability of the photocatalyst was studied, the nZnO-GO exhibited more than 80% degradation efficiency after five cycles for both the dyes and nZnO-GO@SS demonstrated 10% higher (~90%) removal capability after five cycles of reuse. Furthermore, the antibacterial assay showed complete inactivation of Escherichia coli and Staphylococcus aureus bacterial strain by nZnO-GO@SS. Hence, our proposed strategy for the removal of toxic dyes from the aquatic environment under sunlight proved that sand microparticle-supported nanocatalyst (nZnO-GO@SS) might be a superior, cost-effective, and suitable photocatalytic system for industrial applications toward toxic dye removal and decontamination from industrial wastewater.

Surface conjugation of titanium dioxide nanoparticles on nano-graphene oxide enhances photocatalytic degradation of azo dyes under sunlight.

Here, conjugate of nano-graphene oxide (GO) and titanium dioxide nanoparticle (nTiO2) was proposed for the photocatalytic degradation of two toxic azo dyes, Congo red (CR) and Methylene blue (MB), under sunlight irradiation. Furthermore, the optimized weight ratio between GO and nTiO2 of 1:5 demonstrated the highest degradation efficiency. The nanoconjugate induced 85% degradation of 40 ppm of CR in 60 min and a complete degradation within 70 min, while it degraded more than 90% of 20 ppm of MB in 80 min. The photocatalyst can be reused for five cycles of photocatalysis. Thus, the photocatalytic potential of GO/nTiO2 under visible light may be used as a very suitable and cost-effective photocatalyst industrially for the removal of toxic dyes from water.

Post-extubation Dysphagia-Early Detection by Serial Tongue Pressure Measurements: Is it Practical?

How to cite this article: Paul S, Pande RK. Post-extubation Dysphagia-Early Detection by Serial Tongue Pressure Measurements: Is it Practical? Indian J Crit Care Med 2022;26(12):1233-1234.

Application of Nanoscale Materials and Nanotechnology Against Viral Infection: A Special Focus on Coronaviruses.

INTRODUCTION: In recent years, viral infections and associated diseases have become a big challenge for humanity due to high morbidity rates globally. However, timely, accurate, and rapid detection of viral infection may lead to the control of morbidity as well as provide enough time for vaccine preparation and early antiviral therapy. Existing virus detection methods based on immunological and molecular diagnosis found drawbacks, such as its time-consuming and costly one. Recently, the introduction of nanomaterials having multiple unique properties with a series of smart and innovative nano-based technologies have been under investigation for rapid viral detection. This chapter aims to critically review recent literature to illustrate the encompassing applications of nano-engineered materials and further highlighting the role of their active surface in improving the virus detection with high sensitivity and detection range, and in a short time. METHODS: The authors review the research findings related to emerging nanotechnology-based virus detection systems and their applicability for diagnostics of infectious viruses. RESULTS: Recent advances in nanotechnology allow for the development of robust, rapid, and sensitive detection of infectious virus to overcome deficiencies of conventional detection technologies. Nanoparticles have several distinctive physical and chemical characteristics such as unique optical, electronic, and magnetic properties compared to their bulk form enabling them the detection of biological agents like viruses. Further, high surface area to volume ratios of nanoparticles also provides a platform for multi-functionalization with various organic or biological ligands for the selective binding and detection of biological targets like viruses. For instance, colloidal gold nanoparticle-based lateral-flow (AuNP-LF) provides rapid diagnosis and on-site diagnosis of SARS-CoV-2 virus via the IgM detection using the indirect immune-chromatography method. CONCLUSION: The distinct properties of nanomaterials such as plasmon resonance absorption, conductivity, redox behavior, etc. along with surface functionalization might be used in the development of the nano-sensing system with high accuracy and rapid detection of infectious viral diagnosis at the point of care application.

Secretory clusterin promotes oral cancer cell survival via inhibiting apoptosis by activation of autophagy in AMPK/mTOR/ULK1 dependent pathway.

AIMS: Secretory clusterin (sCLU) plays an important role in tumor development and cancer progression. However, the molecular mechanisms and physiological functions of sCLU in oral cancer is unclear. We examined the impact of sCLU-mediated autophagy in cell survival and apoptosis inhibition in oral cancer. MAIN METHODS: Immunohistochemical analysis was performed to analyze protein expression in patient samples. Autophagy and mitophagy was studied by immunofluorescence microscopy and Western blot. The gain and loss of function was studied by overexpression of plasmid and siRNA approaches respectively. Cellular protection against nutrient starvation and therapeutic stress by sCLU was studied by cell viability, caspase assay and meta-analysis. KEY FINDINGS: The data from oral cancer patients showed that the expression levels of sCLU, ATG14, ULK1, and PARKIN increased in grade-wise manners. Interestingly, sCLU overexpression promoted autophagy through AMPK/Akt/mTOR signaling pathway leading to cell survival and protection from long exposure serum starvation induced-apoptosis. Additionally, sCLU was demonstrated to interact with ULK1 and inhibition of ULK1 activity by SBI206965 was found to abolish sCLU-induced autophagy indicating critical role of ULK1 in induction of autophagy. Furthermore, sCLU was observed to promote expression of mitophagy-associated proteins in serum starvation conditions to protect cells from nutrient deprivation. The meta-analysis elucidated that high CLU expression is associated with therapy resistance in cancer and we demonstrated that sCLU-mediated mitophagy was revealed to inhibit cell death by cisplatin. SIGNIFICANCE: The present investigation has highlighted the probable implications of the clusterin-induced autophagy in cell survival and inhibition of apoptosis in oral cancer.

Ultrasound-Guided Serratus Anterior Plane Block for Rib Fracture-Associated Pain Management in Emergency Department.

CONTEXT: Traumatic Rib fractures are common and painful conditions to present in the emergency department. Ultrasound-guided serratus anterior plane block (SAPB) is a newer technique which is being used for managing postthoracotomy, thoracoscopic surgery, or post mastectomy pain by the anesthetists. However, we have recently started utilizing this novel technique in our emergency department for rib fracture patients with severe pain. SETTINGS AND METHODS: We present a case series of 10 patients of multiple rib fractures (MFRs) with persistent Defense and Veterans Pain Rating Scale 7 or more even after intravenous analgesics where this block was applied by trained emergency physicians (EP). RESULTS: Following SAPB median (± IQR) pain score reduction was 5 (±4) at 30 min and 7.5 (±2) after 60 min of administering the block. There were no incidences of block failure or block-related complications in our series. CONCLUSIONS: Ultrasound-guided SAPB can be used safely by trained EP in the emergency department to relieve acute severe analgesic-resistant pain in MFR patients.

YouTube as a Source of Medical and Epidemiological Information During COVID-19 Pandemic: A Cross-Sectional Study of Content Across Six Languages Around the Globe.

Introduction The current coronavirus disease 19 (COVID-19) outbreak has been declared to be a pandemic by the World Health Organization (WHO). It is evolving daily and has jeopardized life globally across social and economic fronts. One of the six key strategic objectives identified by the WHO to manage COVID-19 is to communicate critical information to all communities and prevent the spread of misinformation. We analyzed content on YouTube.com, a widely used web-based platform for medical and epidemiological information. Methods YouTube search results using two keywords were analyzed each in six languages - English, Arabic, Bengali, Dutch, Hindi, and Nigerian Pidgin on April 8, 2020. Forty videos in each of the six languages (i.e., a total of 240 videos) were included for analysis in the study. Two reviewers conducted independent analyses for each language. The inter-observer agreement was evaluated with the kappa coefficient (κ). Modified DISCERN index and Medical Information and Content Index (MICI) scores were used for the reliability of content presented in the videos and information quality assessment, respectively. Analysis of variance, Kruskal-Wallis, Mann-Whitney test, and chi-square tests were done appropriately for data analysis. A p-value of less than 0.05 was considered statistically significant. All calculations were performed using SPSS Statistics for Windows, Version 21.0 (IBM Corp, Armonk, NY). Results The videos cumulatively attracted 364,080,193 views. Altogether, 52.5% of videos were Informative, 23.75% were News Updates, and 8.33% were Personal Experiences. Ten percent of videos were found to present medically misleading information. Independent Users contributed 75% of the misleading content. The overall Mean DISCERN score, an index of content reliability, was 2.62/5. The overall Mean MICI Score was 5.68/25. Videos had better scores in the Transmission component of the MICI scale and scored low on the Screening/Testing component. Conclusion The reliability and quality of the content of most videos about COVID-19 and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were found to be unsatisfactory. Videos with misleading content were found across all six languages, and sometimes garnered a higher percentage of views than those from credible sources. The share of videos contributed by Government and Health Agencies was low. Medical institutions and health agencies should produce content on widely used platforms like YouTube for quality medical and epidemiological information dissemination.

Supramolecular nanoassembly of lysozyme and α-lactalbumin (apo α-LA) exhibits selective cytotoxicity and enhanced bioavailability of curcumin to cancer cells.

Hybrid supramolecular spherical nanoassembly of hen egg white lysozyme and bovine apo α lactalbumin (SNLYZ-BLA) was prepared with a mean size of ˜55.2 nm using an optimized desolvation method via chemical crosslinking. The nanoassembly, SNLYZ-BLA demonstrated dose-dependent reactive oxygen species (ROS) mediated cytotoxicity in multiple cancer cells such as MCF-7, MDA-MB231, HeLa and MG 63. It also demonstrated high loading capacity of a phytochemical based anticancer agent, curcumin (248.8 mg/g) and target-based pH-responsive in vitro drug release with around 85.8% curcumin release observed under acidic condition. Moreover, curcumin loaded SNLYZ-BLA (SNLYZ-BLA-CUR) induced cell viability reduction in all cancer cells including mouse melanoma (B16F10) by more than 90% within 24 h. Further, SNLYZ-BLA and SNLYZ-BLA-CUR when conjugated with folic acid enhanced the cytotoxicity via folate receptor-based targeting. Both drug loading and release induced conformational change and folding reconstitution of the protein nano-assembly, respectively, which made the whole system an efficient therapeutic agent that works via a dual mode of action. We demonstrated that SNLYZ-BLA and SNLYZ-BLA-CUR were highly biocompatible in vitro. Therefore, our supramolecular protein nanoassembly loaded with curcumin could emerge as a comprehensive cancer therapeutics that acts via a strategic mode of dual therapeutic mechanisms.

Elucidating the Role of Val-Asn 95 and Arg-Gly 52 Mutations on Structure and Stability of Fibroblast Growth Factor Homologous Factor 2.

BACKGROUND: Fibroblast growth Factor Homologous Factors (FHFs) belong to a subclass of Fibroblast Growth Factor (FGF) family owing to their high sequence and structural similarities with FGFs. However, despite these similarities, there are properties which set them apart from FGFs. FHFs lack the secretion signal sequence unlike other FGF members, except FGF1 and 2. Unlike FGFs, FHFs are not able to bind to FGF Receptors (FGFRs) and instead have been implicated in binding to Voltage-Gated Sodium Channels (VGSCs), neuronal MAP kinase scaffold protein and islet-brain-2 (IB2). The two amino acids Arg-52 and Val95 are conserved in all FHFs and mutation of these residues lead to its inability to bind with VGSC/IB2. However, it is not clear whether the loss of binding is due to destabilization of the protein on mutation or due to involvement of Arg52 and Val95 in conferring functionality to FHFs. OBJECTIVE: In the present study, we have mutated these two conserved residues of FHF2 with its corresponding FGF counterpart amino acids and studied the effects of the mutations on the structure and stability of the protein. METHODS: Several biophysical methods like isothermal equilibrium denaturation study, ANS fluorescence, intrinsic fluorescence, acrylamide quenching, circular dichroism studies as well as using computational approaches were employed. RESULTS: The single mutations were found to affect the overall stability, conformation and functionality of the protein. CONCLUSION: Thus, the studies throw light on the role of specific amino acids in deciding the stability, structure and functionality of proteins and will be useful for development of therapeutically engineered proteins.

Graphene Oxide Quantum Dot Alters Amyloidogenicity of Hen Egg White Lysozyme via Modulation of Protein Surface Character.

A series of neurodegenerative disorders are caused by intracellular or extracellular amyloid deposition, including Alzheimer's disease, Parkinson's disease, Prion disease, and so on. To prevent the progress of such amyloid-mediated disorders, various agents have been tested including nanoparticles. Among different nanomaterials, graphene oxide shows unique electrochemical properties, which have potential applications in various biomedical fields. In our present investigation, we explored the effect of graphene oxide quantum dots (GOQDs) in amyloid ß-fibrillation of hen egg white lysozyme (HEWL) under various conditions. Electron microscopy imaging showed that administration of GOQD inhibited HEWL amyloid ß-fibrillation via producing thin and small fragments of fibrils. ζ-Potential measurement and 8-anilino-1-naphthalenesulfonic fluorescence study of lysozyme amyloid demonstrated a significant drop in surface hydrophobicity and an increase of surface charge of protein molecules. The reduced hydrophobic interaction and enhanced surface charge inhibit the hydrophobic assembly and colloidal stability of the protein. Circular dichroism and thioflavin-T fluorescence demonstrated that GOQD also interfered at the secondary structure level and prevented amyloid ß-sheet formation and assembly of a protein by reducing the amount of amyloid ß-sheet formation. Further, cellular toxicity analysis with HaCaT and 3T3 cells showed reduced toxicity of amyloid samples prepared with GOQD. Therefore, GOQD might be used to be a potential amyloid-preventive agent in various neurodegenerative diseases.

Lapachol inhibits glycolysis in cancer cells by targeting pyruvate kinase M2.

Reliance on aerobic glycolysis is one of the hallmarks of cancer. Although pyruvate kinase M2 (PKM2) is a key mediator of glycolysis in cancer cells, lack of selective agents that target PKM2 remains a challenge in exploiting metabolic pathways for cancer therapy. We report that unlike its structural analog shikonin, a known inhibitor of PKM2, lapachol failed to induce non-apoptotic cell death ferroxitosis in hypoxia. However, melanoma cells treated with lapachol showed a dose-dependent inhibition of glycolysis and a corresponding increase in oxygen consumption. Accordingly, in silico studies revealed a high affinity-binding pocket for lapachol on PKM2 structure. Lapachol inhibited PKM2 activity of purified enzyme as well as in melanoma cell extracts. Blockade of glycolysis by lapachol in melanoma cells led to decreased ATP levels and inhibition of cell proliferation. Furthermore, perturbation of glycolysis in melanoma cells with lapachol sensitized cells to mitochondrial protonophore and promoted apoptosis. These results present lapachol as an inhibitor of PKM2 to interrogate metabolic plasticity in tumor cells.

Formation of the chaperonin complex studied by 2D NMR spectroscopy.

We studied the interaction between GroES and a single-ring mutant (SR1) of GroEL by the NMR titration of 15N-labeled GroES with SR1 at three different temperatures (20, 25 and 30°C) in the presence of 3 mM ADP in 100 mM KCl and 10 mM MgCl2 at pH 7.5. We used SR1 instead of wild-type double-ring GroEL to precisely control the stoichiometry of the GroES binding to be 1:1 ([SR1]:[GroES]). Native heptameric GroES was very flexible, showing well resolved cross peaks of the residues in a mobile loop segment (residue 17-34) and at the top of a roof hairpin (Asn51) in the heteronuclear single quantum coherence spectra. The binding of SR1 to GroES caused the cross peaks to disappear simultaneously, and hence it occurred in a single-step cooperative manner with significant immobilization of the whole GroES structure. The binding was thus entropic with a positive entropy change (219 J/mol/K) and a positive enthalpy change (35 kJ/mol), and the binding constant was estimated at 1.9×105 M-1 at 25°C. The NMR titration in 3 mM ATP also indicated that the binding constant between GroES and SR1 increased more than tenfold as compared with the binding constant in 3 mM ADP. These results will be discussed in relation to the structure and mechanisms of the chaperonin GroEL/GroES complex.

Protein functionalization of ZnO nanostructure exhibits selective and enhanced toxicity to breast cancer cells through oxidative stress-based cell death mechanism.

Zinc oxide nanostructure (ZnONS) was chemically synthesized and functionalized (FZnONSBLA) with a small protein bovine α-lactalbumin (BLA) by chemical cross-linking methods. Both nano-structures were characterized using various techniques such as electron microscopy, dynamic light scattering (DLS), UV-Vis spectroscopy, FT-IR, photo-luminescence and X-ray diffraction. Electron microscopy and DLS analysis revealed their (ZnONS and FZnONSBLA) average size of 200nm and 450nm, respectively. When cytotoxicity of both the nanostructures were assessed in breast cancer cells MCF-7 and MDAMB231 by MTT assay and PI/Annexin V staining (FACS), FZnONSBLA demonstrated higher cell death than ZnONS primarily due to generation of intracellular reactive oxygen species (ROS). Our experimental results also suggested that such enhanced toxicity was due to the lethal structural variant of BLA in FZnONSBLA as well as higher cellular uptake than ZnONS by cancer cells. The death kinetics study with time in cancer cells further proved that FZnONSBLA caused toxicity much faster than ZnONS, thus suggested a strong role of lethal variant of BLA in FZnONSBLA as a cytotoxic agent in cancer cells. Furthermore, FZnONSBLA demonstrated excellent cytocompatibility (normal cells) and hemocompatibility compared to ZnONS. Hence, considering the biodegradable nature of ZnO nonmaterial, our results demonstrated that BLA functionalized ZnONS could be used to develop a suitable therapeutic strategy in cancer.

Abrus Agglutinin, a type II ribosome inactivating protein inhibits Akt/PH domain to induce endoplasmic reticulum stress mediated autophagy-dependent cell death.

Abrus agglutinin (AGG), a type II ribosome-inactivating protein has been found to induce mitochondrial apoptosis. In the present study, we documented that AGG-mediated Akt dephosphorylation led to ER stress resulting the induction of autophagy-dependent cell death through the canonical pathway in cervical cancer cells. Inhibition of autophagic death with 3-methyladenine (3-MA) and siRNA of Beclin-1 and ATG5 increased AGG-induced apoptosis. Further, inhibiting apoptosis by Z-DEVD-FMK and N-acetyl cysteine (NAC) increased autophagic cell death after AGG treatment, suggesting that AGG simultaneously induced autophagic and apoptotic death in HeLa cells. Additionally, it observed that AGG-induced autophagic cell death in Bax knock down (Bax-KD) and 5-FU resistant HeLa cells, confirming as an alternate cell killing pathway to apoptosis. At the molecular level, AGG-induced ER stress in PERK dependent pathway and inhibition of ER stress by salubrinal, eIF2α phosphatase inhibitor as well as siPERK reduced autophagic death in the presence of AGG. Further, our in silico and colocalization study showed that AGG interacted with pleckstrin homology (PH) domain of Akt to suppress its phosphorylation and consequent downstream mTOR dephosphorylation in HeLa cells. We showed that Akt overexpression could not augment GRP78 expression and reduced autophagic cell death by AGG as compared to pcDNA control, indicating Akt modulation was the upstream signal during AGG's ER stress mediated autophagic cell death. In conclusion, we established that AGG stimulated cell death by autophagy might be used as an alternative tumor suppressor mechanism in human cervical cancer. © 2016 Wiley Periodicals, Inc.