Characterizing locus specific chromatin structure and dynamics with correlative conventional and super-resolution imaging in living cells.

The dynamic rearrangement of chromatin is critical for gene regulation, but mapping both the spatial organization of chromatin and its dynamics remains a challenge. Many structural conformations are too small to be resolved via conventional fluorescence microscopy and the long acquisition time of super-resolution photoactivated localization microscopy (PALM) precludes the structural characterization of chromatin below the optical diffraction limit in living cells due to chromatin motion. Here we develop a correlative conventional fluorescence and PALM imaging approach to quantitatively map time-averaged chromatin structure and dynamics below the optical diffraction limit in living cells. By assigning localizations to a locus as it moves, we reliably discriminate between bound and unbound dCas9 molecules, whose mobilities overlap. Our approach accounts for changes in DNA mobility and relates local chromatin motion to larger scale domain movement. In our experimental system, we show that compacted telomeres move faster and have a higher density of bound dCas9 molecules, but the relative motion of those molecules is more restricted than in less compacted telomeres. Correlative conventional and PALM imaging therefore improves the ability to analyze the mobility and time-averaged nanoscopic structural features of locus specific chromatin with single molecule sensitivity and yields unprecedented insights across length and time scales.

Structure and Piezoelectricity Due to B Site Cation Variation in ABn+Cln+2 Hybrid Histammonium Chlorometallate Materials.

To provide new insights for understanding the influence of B site cations on the structure in chlorometallate materials of the form ABn+Cln+2, we report novel organic-inorganic hybrid metallates (OIHMs) incorporating histammonium (HistNH3) dications and various transition-metal and main group B site cations. Single crystals of OIHMs with the basic formula (HistNH3Mn+Cln+2, M = Fe, Co, Ni, Cu, Zn, Cd, Hg, Sb, Sn, Pb, Bi) were grown and their structures characterized by single-crystal X-ray crystallography. HistNH3CoCl4, HistNH3ZnCl4, and HistNH3SbCl5 were crystallized in a non-centrosymmetric space group and were subsequently studied with piezoresponse force microscopy (PFM). While bulk measurements of crystals and poly(vinylidene difluoride) (PVDF)/metallate composite films exhibited low bulk response values, the surface-measured local response values using PFM were 5.17 pm/V for HistNH3CoCl4, 22.6 pm/V for HistNH3ZnCl4, and 2.9 pm/V for HistNH3SbCl5 compared with 2.50 pm/V for PVDF reference samples. The magnitudes of the d33 coefficient, net dipole, and cation-Cl bond dipole obtained from the density functional theory calculations confirm the higher response in HistNH3ZnCl4 compared to HistNH3CoCl4. Density of states and crystal orbital Hamilton population analysis indicate that the higher net dipole in HistNH3ZnCl4 compared to HistNH3CoCl4 is due to the lower hybridization of the M-Cl bond.

Describing adsorption of benzene, thiophene, and xenon on coinage metals by using the Zaremba-Kohn theory-based model.

Semilocal (SL) density functional approximations (DFAs) are widely applied but have limitations due to their inability to incorporate long-range van der Waals (vdW) interaction. Non-local functionals (vdW-DF, VV10, and rVV10) or empirical methods (DFT+D, DFT+vdW, and DFT+MBD) are used with SL-DFAs to account for such missing interaction. The physisorption of a molecule on the surface of the coinage metals (Cu, Ag, and Au) is a typical example of systems where vdW interaction is significant. However, it is difficult to find a general method that reasonably describes both adsorption energy and geometry of even the simple prototypes of cyclic and heterocyclic aromatic molecules such as benzene (C6H6) and thiophene (C4H4S), respectively, with reasonable accuracy. In this work, we present an alternative scheme based on Zaremba-Kohn theory, called DFT+vdW-dZK. We show that unlike other popular methods, DFT+vdW-dZK and particularly SCAN+vdW-dZK give an accurate description of the physisorption of a rare-gas atom (xenon) and two small albeit diverse prototype organic molecules on the (111) surfaces of the coinage metals.

Self-interaction-corrected Kohn-Sham effective potentials using the density-consistent effective potential method.

Density functional theory (DFT) and beyond-DFT methods are often used in combination with photoelectron spectroscopy to obtain physical insights into the electronic structure of molecules and solids. The Kohn-Sham eigenvalues are not electron removal energies except for the highest occupied orbital. The eigenvalues of the highest occupied molecular orbitals often underestimate the electron removal or ionization energies due to the self-interaction (SI) errors in approximate density functionals. In this work, we adapt and implement the density-consistent effective potential method of Kohut, Ryabinkin, and Staroverov [J. Chem. Phys. 140, 18A535 (2014)] to obtain SI-corrected local effective potentials from the SI-corrected Fermi-Löwdin orbitals and density in the Fermi-Löwdin orbital self-interaction correction scheme. The implementation is used to obtain the density of states (photoelectron spectra) and HOMO-LUMO gaps for a set of molecules and polyacenes. Good agreement with experimental values is obtained compared to a range of SI uncorrected density functional approximations.

Development and Characterization of Novel Conductive Sensing Fibers for In Vivo Nerve Stimulation.

Advancements in electrode technologies to both stimulate and record the central nervous system's electrical activities are enabling significant improvements in both the understanding and treatment of different neurological diseases. However, the current neural recording and stimulating electrodes are metallic, requiring invasive and damaging methods to interface with neural tissue. These electrodes may also degrade, resulting in additional invasive procedures. Furthermore, metal electrodes may cause nerve damage due to their inherent rigidity. This paper demonstrates that novel electrically conductive organic fibers (ECFs) can be used for direct nerve stimulation. The ECFs were prepared using a standard polyester material as the structural base, with a carbon nanotube ink applied to the surface as the electrical conductor. We report on three experiments: the first one to characterize the conductive properties of the ECFs; the second one to investigate the fiber cytotoxic properties in vitro; and the third one to demonstrate the utility of the ECF for direct nerve stimulation in an in vivo rodent model.

Affinity-Selected Bicyclic Peptide G-Quadruplex Ligands Mimic a Protein-like Binding Mechanism.

The study of G-quadruplexes (G4s) in a cellular context has demonstrated links between these nucleic acid secondary structures, gene expression, and DNA replication. Ligands that bind to the G4 structure therefore present an excellent opportunity for influencing gene expression through the targeting of a nucleic acid structure rather than sequence. Here, we explore cyclic peptides as an alternative class of G4 ligands. Specifically, we describe the development of de novo G4-binding bicyclic peptides selected by phage display. Selected bicyclic peptides display submicromolar affinity to G4 structures and high selectivity over double helix DNA. Molecular simulations of the bicyclic peptide-G4 complexes corroborate the experimental binding strengths and reveal molecular insights into G4 recognition by bicyclic peptides via the precise positioning of amino acid side chains, a binding mechanism reminiscent of endogenous G4-binding proteins. Overall, our results demonstrate that selection of (bi)cyclic peptides unlocks a valuable chemical space for targeting nucleic acid structures.

Molecular Engineering of Hydroxide Conducting Polymers for Anion Exchange Membranes in Electrochemical Energy Conversion Technology.

Anion exchange membranes (AEMs) based on hydroxide-conducting polymers (HCPs) are a key component for anion-based electrochemical energy technology such as fuel cells, electrolyzers, and advanced batteries. Although these alkaline electrochemical applications offer a promising alternative to acidic proton exchange membrane electrochemical devices, access to alkaline-stable and high-performing polymer electrolyte materials has remained elusive until now. Despite vigorous research of AEM polymer design, literature examples of high-performance polymers with good alkaline stability at an elevated temperature are uncommon. Traditional aromatic polymers used in AEM applications contain a heteroatomic backbone linkage, such as an aryl ether bond, which is prone to degradation via nucleophilic attack by hydroxide ion. In this Account, we highlight some of the progress our group has made in the development of advanced HCPs for applications in AEMs and electrode ionomers. We propose that a synthetic polymer design with an all C-C bond backbone and a flexible chain-tethered quaternary ammonium group provides an effective solution to the problem of alkaline stability. Because of the critical demand for such a polymer system, we have established new synthetic strategies for polymer functionalization and polycondensation using an acid catalyst. The first approach is to graft a cationic tethered alkyl group to pre-existing, commercially available styrene-based block copolymers. The second approach is to synthesize high-molecular-weight aromatic backbone polymers using acid-catalyzed polycondensation of arene monomers and a functionalized trifluoromethyl ketone substrate. Both strategies involve a simple two-step reaction process and avoid the use of expensive metal-based catalysts and toxic chemicals, thereby making the synthetic processes easily scalable to large industrial quantities. Both polymer systems were found to have excellent alkaline stability, confirmed by the preservation of ion exchange capacity and ion conductivity of the membrane after an alkaline test under conditions of 1 M NaOH at 80-95 °C. In addition, the advantage of good solvent processability and convenient scalability of the reaction process generates considerable interest in these polymers as commercial standard AEM candidates. AEM fuel cell and electrolyzer tests of some of the developed polymer membranes showed excellent performance, suggesting that this new class of HCPs opens a new avenue to electrochemical devices with real-world applications.

The Fermi-Löwdin self-interaction correction for ionization energies of organic molecules.

(Semi)-local density functional approximations (DFAs) suffer from self-interaction error (SIE). When the first ionization energy (IE) is computed as the negative of the highest-occupied orbital (HO) eigenvalue, DFAs notoriously underestimate them compared to quasi-particle calculations. The inaccuracy for the HO is attributed to SIE inherent in DFAs. We assessed the IE based on Perdew-Zunger self-interaction correction on 14 small to moderate-sized organic molecules relevant in organic electronics and polymer donor materials. Although self-interaction corrected DFAs were found to significantly improve the IE relative to the uncorrected DFAs, they overestimate. However, when the self-interaction correction is interiorly scaled using a function of the iso-orbital indicator zσ, only the regions where SIE is significant get a correction. We discuss these approaches and show how these methods significantly improve the description of the HO eigenvalue for the organic molecules.

Stretched or noded orbital densities and self-interaction correction in density functional theory.

Semilocal approximations to the density functional for the exchange-correlation energy of a many-electron system necessarily fail for lobed one-electron densities, including not only the familiar stretched densities but also the less familiar but closely related noded ones. The Perdew-Zunger (PZ) self-interaction correction (SIC) to a semilocal approximation makes that approximation exact for all one-electron ground- or excited-state densities and accurate for stretched bonds. When the minimization of the PZ total energy is made over real localized orbitals, the orbital densities can be noded, leading to energy errors in many-electron systems. Minimization over complex localized orbitals yields nodeless orbital densities, which reduce but typically do not eliminate the SIC errors of atomization energies. Other errors of PZ SIC remain, attributable to the loss of the exact constraints and appropriate norms that the semilocal approximations satisfy, suggesting the need for a generalized SIC. These conclusions are supported by calculations for one-electron densities and for many-electron molecules. While PZ SIC raises and improves the energy barriers of standard generalized gradient approximations (GGAs) and meta-GGAs, it reduces and often worsens the atomization energies of molecules. Thus, PZ SIC raises the energy more as the nodality of the valence localized orbitals increases from atoms to molecules to transition states. PZ SIC is applied here, in particular, to the strongly constrained and appropriately normed (SCAN) meta-GGA, for which the correlation part is already self-interaction-free. This property makes SCAN a natural first candidate for a generalized SIC.

Identification of a novel ligand for the ATAD2 bromodomain with selectivity over BRD4 through a fragment growing approach.

ATAD2 is an ATPase that is overexpressed in a variety of cancers and associated with a poor patient prognosis. This protein has been suggested to function as a cofactor for a range of transcription factors, including the proto-oncogene MYC and the androgen receptor. ATAD2 comprises an ATPase domain, implicated in chromatin remodelling, and a bromodomain which allows it to interact with acetylated histone tails. Dissection of the functional roles of these two domains would benefit from the availability of selective, cell-permeable pharmacological probes. An in silico evaluation of the 3D structures of various bromodomains suggested that developing small molecule ligands for the bromodomain of ATAD2 is likely to be challenging, although recent reports have shown that ATAD2 bromodomain ligands can be identified. We report a structure-guided fragment-based approach to identify lead compounds for ATAD2 bromodomain inhibitor development. Our findings indicate that the ATAD2 bromodomain can accommodate fragment hits (Mr < 200) that yield productive structure-activity relationships, and structure-guided design enabled the introduction of selectivity over BRD4.

Rapid Postgrafting Reaction to Prepare Quaternized Polycarbazoles.

We report a rapid postgrafting reaction to prepare alkyl ammonium functionalized polycarbazoles from a commercially available monomer. This novel synthetic approach provides benefit to preparing the high molecular weight quaternized polycarbazoles within 1 h of Friedel-Crafts polycondensation, avoiding the synthesis and purification step to prepare a functionalized monomer. The postgrafting reaction produces hexyl alkyl ammonium functionalized polycarbazole with 100% grafting degree. However, the postgrafting reaction produced only 60% grafting with propyl alkyl ammonium due to the competitive elimination reaction because of the higher acidity of ß-hydrogen in the propyl alkyl group resulting from the proximity of the bromide and ammonium groups. The hexyl alkyl ammonium functionalized polycarbazole has a high hydroxide conductivity of 103 mS cm-1 at 80 °C and showed excellent alkaline stability with less than 3% loss of ion group after 1 M NaOH treatment at 80 °C for 500 h. This study highlights that the postgrafting reaction provides a pathway for the scale-up synthesis of quaternized aryl ether-free polyaromatics.

Additively Manufactured Silicone Polymer Composite with High Hydrogen Getter Content and Hydrogen Absorption Capacity.

Hydrogen getters consisting of 1,4-bis[phenylethynyl] benzene (DEB) and a carbon-supported palladium catalyst (Pd/C) have been used to mitigate the accumulation of unwanted hydrogen gas in a sealed system. Here, we report the formulation of a composite resin consisting of silicone polymer plus DEB-Pd/C as an active getter material and the additive manufacturing of silicone getter composites with a high getter content (up to 50 wt %). NMR and DSC studies suggest no reaction between the silicone polymer resin and DEB even at elevated curing temperatures (75 °C). Getter composites with varying amounts of getter and filler were formulated, and their rheological properties were studied. The two composite resins with good printability parameters and different getter contents were chosen to make 3D-printed samples. The hydrogen absorption capacity of these samples was studied at a low hydrogen pressure of 750 mTorr of pure hydrogen. The getter composite with 50 wt% of getter showed normalized DEB conversion of 83%, with the hydrogen adsorption capacity of 100.2 mL of H2 per gram of polymer getter composite.

Adverse Events Following COVISHIELD and VERO CELL Vaccination Campaigns Against COVID-19.

BACKGROUND: Vaccination against COVID-19 for Nepalese was initiated in January 2021 for various age groups. People were anxious about receiving the vaccines and were concerned about the safety profile of the vaccine they received. In this study, we have tried to observe the Adverse Events Following Immunization of two different vaccines namely COVISHIELD (ChAdOx1 nCOV-19) and VERO CELL (CZ02 strain), used in different phases of vaccination by the government of Nepal. METHODS: We conducted a cross-sectional study among people who received COVID-19 vaccines in this study using a self-administered questionnaire.  Data was cleaned and then exported to IBM SPSS v.20 for analysis, Chi-square test was used to see the association between different variables and a p-value<0.05 was considered statistically significant. RESULTS: Out of 303 respondents, all had received the first and 270 participants had received the second dose of the COVID-19 vaccine, among which, 133 (43.89%) reported at least one side effect after the first dose of vaccination while 58 (21.48%) had self-reported side effects after the second dose of vaccination. Seventeen percent of the respondents had COVID-19 infection within the past 3 months before receiving COVID-19 vaccine. Three percent of participants had re-infection with COVID-19 after receiving the first or the second dose of the COVID-19 vaccine. Among participants who experienced adverse events, 42% and 62.1% of participants experienced mild adverse events following the first dose and second dose of the vaccine, respectively.  Conclusions: The adverse events following immunization for both vaccines after both doses of vaccination were quite low, with 43.89% of participants reporting side effects after the first dose and 21.48% of participants reporting side effects after the second dose. Adverse events were most frequently reported within 24 hours of vaccination and were mostly mild. There was no statistical significance of adverse events between both vaccines.

Zosyn-Induced Rapid Thrombocytopenia in a Patient With End-Stage Renal Disease and HIV: A Case Report.

Thrombocytopenia is a rare but potentially serious complication associated with the use of various medications, including antibiotics. Piperacillin-tazobactam (Zosyn), a commonly used broad-spectrum antibiotic, has been reported as an infrequent cause of drug-induced thrombocytopenia. We present a case of a 65-year-old female with end-stage renal disease (ESRD) on hemodialysis, HIV, and multiple comorbidities who developed rapid-onset thrombocytopenia shortly after receiving Zosyn. The patient's platelet count dropped from a baseline of 291,000/µL on admission to a nadir of 8,000/µL within 36 hours of starting Zosyn. The administration of Zosyn was promptly discontinued, and the patient's platelet count gradually increased to 134,000/µL within two days after discontinuation. The patient had no apparent bleeding manifestations during her hospital stay. Further workup for other causes of thrombocytopenia, including heparin-induced thrombocytopenia (HIT), was negative. This case highlights the importance of vigilance for drug-induced thrombocytopenia in patients receiving Zosyn and the need for prompt recognition and management to prevent potential complications.

Poisoning among Children Visiting the Paediatric Emergency Department in a Tertiary Care Centre.

Introduction: Poisoning occurs when substances are ingested, inhaled, or absorbed through skin contact in quantities that are harmful to the body. Knowledge of the pattern and prevalence of paediatric poisoning will help us quantify the burden of poisoning. The aim of the study was to find out the prevalence of poisoning among children visiting the Paediatric Emergency Department in a tertiary care centre. Methods: A descriptive cross-sectional study was conducted among children visiting the Paediatric Emergency Department in a tertiary care centre. Data from 1 January 2020 to 31 December 2021 was collected between 15 April 2022 to 25 April 2022 from medical records. Ethical approval was taken from the Institutional Review Committee. Children aged 0 to 14 years old were included in the study. A convenience sampling method was used. The point estimate was calculated at a 95% Confidence Interval. Results: Among 12,488 children, 162 (1.30%) (1.10-1.50, 95% Confidence Interval) had poisoning. Pesticides and insecticides were the most common agents involved in poisoning 39 (24.07%). Conclusions: The prevalence of poisoning was found to be lower than other studies done in similar settings. Keywords: children; Nepal; poisoning.

Burden of enteric fever and antibiotic sensitivity in Nepalese Children Prior to Typhoid Vaccine in National Immunization Program.

BACKGROUND: Enteric fever is a major public health problem in developing and under developed countries. Case fatality rate without treatment is 10-30% and with appropriate treatment is only 1-4%. Gold standard for diagnosis is isolation of Salmonella enterica from blood or bone marrow. Antibiotics resistance is skyrocketing with emergence of multidrug resistance S. typhi and extensively drug resistant S. typhi. METHODS: The blood culture done in Kanti children hospital in last six years were taken from the data base and the culture positive cases were taken from which the salmonella species positive cases along with the drug sensitivity pattern were used in our study. RESULTS: The culture positivity rate was 2.8% and 7.6% (n=136) among the culture positive cases were Salmonella species. Salmonella typhi (121; 88.9%) was the most frequently isolated species, followed by Salmonella paratyphi A (13; 9.5%) and Salmonella paratyphi B (2;1.4%). Children with age 5-10 years was the most affected age group for infection with Salmonella, 50.0% (n=68). Nalidixic acid is resistant in 89.9% Salmonella typhi; followed by ciprofloxacin (31.8%), ofloxacin (18.2%), ampicillin (9.6%), azithromycin (8.4%), chloramphenicol (8.2%), cotrimoxazole (5.4%), cefixime (4%), ceftriaxone (2.5%) and cefotaxime (0.0%). Cefixime, ceftriaxone, cefotaxime are 100% sensitive to Salmonella paratyphi, followed by cotrimoxazole (92.9%), ofloxacin (81.8%), chloramphenicol (75%), azithromycin (66.7%), ampicillin (60%), ciprofloxacin (50%) and Nalidixic acid (23.1%). CONCLUSIONS: Salmonella species culture isolatation are declining every year. Fluoroquinolones have more resistance than first line drugs of typhoid, azithromycin resistance is rising but 3rd generation cephalosporins are sensitive to Salmonella species.

Interplay between Shelf Life and Printability of Silica-Filled Suspensions.

Although fumed silica/siloxane suspensions are commonly employed in additive manufacturing technology, the interplay between shelf life, storage conditions, and printability has yet to be explored. In this work, direct ink writing (DIW) was used to print unique three-dimensional structures that required suspensions to retain shape and form while being printed onto a substrate. Suspensions containing varying concentrations of hydrophobic and hydrophilic silica were formulated and evaluated over a time span of thirty days. Storage conditions included low (8%) and high (50%) relative humidity and temperatures ranging from 4 °C to 25 °C. The shelf life of the suspensions was examined by comparing the print quality of pristine and aged samples via rheology, optical microscopy, and mechanical testing. Results showed a significant decrease in printability over time for suspensions containing hydrophilic fumed silica, whereas the printability of suspensions containing hydrophobic fumed silica remained largely unchanged after storage. The findings in this work established the following recommendations for extending the shelf life and printability of suspensions commonly used in DIW technology: (1) higher fumed silica concentrations, (2) low humidity and low temperature storage environments, and (3) the use of hydrophobic fumed silica instead of hydrophilic fumed silica.

Organelle interactions compartmentalize hepatic fatty acid trafficking and metabolism.

Organelle interactions play a significant role in compartmentalizing metabolism and signaling. Lipid droplets (LDs) interact with numerous organelles, including mitochondria, which is largely assumed to facilitate lipid transfer and catabolism. However, quantitative proteomics of hepatic peridroplet mitochondria (PDM) and cytosolic mitochondria (CM) reveals that CM are enriched in proteins comprising various oxidative metabolism pathways, whereas PDM are enriched in proteins involved in lipid anabolism. Isotope tracing and super-resolution imaging confirms that fatty acids (FAs) are selectively trafficked to and oxidized in CM during fasting. In contrast, PDM facilitate FA esterification and LD expansion in nutrient-replete medium. Additionally, mitochondrion-associated membranes (MAM) around PDM and CM differ in their proteomes and ability to support distinct lipid metabolic pathways. We conclude that CM and CM-MAM support lipid catabolic pathways, whereas PDM and PDM-MAM allow hepatocytes to efficiently store excess lipids in LDs to prevent lipotoxicity.

Conducting a phase III clinical trial in children during the COVID- 19 pandemic: Experience and lessons learnt from a clinical research facility of Nepal.

Clinical trials in humans are vital to test safety and efficacy of new interventions and are accompanied with the complexity of related regulatory guidelines, stringent time frame and financial burden particularly when participants are children. Conducting clinical trials in low and middle income countries, where 90% of global diseases occur, increases the complexity as resources, infrastructures, and experience related to clinical trials may be limited in some countries. During the COVID-19 pandemic, due to multiple infection control measures such as social distancing, lock-down of the societies, and increased work load of hospital workers, conducting clinical trials seemed very challenging. Related guidelines and recommendations on clinical trials required updates to adapt the situation for ongoing clinical trials to be continued and new clinical trials to be initiated. In this review report, we described the lessons learnt through our experiences, challenges we faced, and the mitigation measures implemented as a response while conducting a phase III clinical trial on a non-COVID-19 vaccine at a government children's hospital during the COVID-19 pandemic. We hope this report will contribute in lowering the obstacles to allow the successful completion of future studies, in countries where people live with the burden of vaccine-preventable diseases.

Eumelanin-Inspired Antimicrobial with Biocidal Activity against Methicillin-Resistant Staphylococcus aureus.

The reliance on antibiotics and antimicrobials to treat bacterial infectious diseases is threatened by the emergence of antibiotic resistance and multi-drug-resistant organisms, thus having the potential to greatly impact human health. Thus, the discovery and development of antimicrobials capable of acting on antibiotic-resistant bacteria is a major area of significance in scientific research. Herein, we present the development of a eumelanin-inspired antimicrobial capable of killing methicillin-resistant Staphylococcus aureus (MRSA). By ligating quaternary ammonium-functionalized "arms" to a eumelanin-inspired indole with intrinsic antimicrobial activity, an antimicrobial agent with enhanced activity was prepared. This resulting antimicrobial, EIPE-1, had a minimum inhibitory concentration of 16 µg/mL (17.1 µM) against a clinical isolate of MRSA obtained from an adult cystic fibrosis patient. The biocidal activity occurred within 30 min of exposure and resulted in changes to the bacterial cell surface as visualized with a scanning electron microscope. Taken together, these studies demonstrate that EIPE-1 is effective at killing MRSA.