Exact Distribution of the Quantal Content in Synaptic Transmission.

During electrochemical signal transmission through synapses, triggered by an action potential (AP), a stochastic number of synaptic vesicles (SVs), called the "quantal content," release neurotransmitters in the synaptic cleft. It is widely accepted that the quantal content probability distribution is a binomial based on the number of ready-release SVs in the presynaptic terminal. But the latter number itself fluctuates due to its stochastic replenishment, hence the actual distribution of quantal content is unknown. We show that exact distribution of quantal content can be derived for general stochastic AP inputs in the steady state. For fixed interval AP train, we prove that the distribution is a binomial, and corroborate our predictions by comparison with electrophysiological recordings from MNTB-LSO synapses of juvenile mice. For a Poisson train, we show that the distribution is nonbinomial. Moreover, we find exact moments of the quantal content in the Poisson and other general cases, which may be used to obtain the model parameters from experiments.

Synthesis of substituted 2H-Chromenes via Pd-catalyzed C-H activation and thermal cyclization.

A proficient approach has been developed for the synthesis of substituted 2H-chromenes from C1-substituted glucal. The key step of our synthetic methodology was C-H activation in propylene carbonate solvent followed by 6π-electrocyclization aromatization in ethylene glycol as greener substitutes to toxic aprotic solvents, to obtain 2H-chromenes in a stepwise manner. The application of the developed methodology was further explored with the synthesis of a small library of substituted 2H-chromenes in good yields.

Orthogonal outlier detection and dimension estimation for improved MDS embedding of biological datasets.

Conventional dimensionality reduction methods like Multidimensional Scaling (MDS) are sensitive to the presence of orthogonal outliers, leading to significant defects in the embedding. We introduce a robust MDS method, called DeCOr-MDS (Detection and Correction of Orthogonal outliers using MDS), based on the geometry and statistics of simplices formed by data points, that allows to detect orthogonal outliers and subsequently reduce dimensionality. We validate our methods using synthetic datasets, and further show how it can be applied to a variety of large real biological datasets, including cancer image cell data, human microbiome project data and single cell RNA sequencing data, to address the task of data cleaning and visualization.

Efficient and stereoselective synthesis of sugar fused pyrano[3,2-c]pyranones as anticancer agents.

A highly stereoselective, efficient and facile route was achieved for the synthesis of novel and biochemically potent sugar fused pyrano[3,2-c]pyranone derivatives starting from inexpensive, naturally occurring d-galactose and d-glucose. First, ß-C-glycopyranosyl aldehydes were synthesized from these d-hexose sugars in six steps, with overall yields 41-55%. Next, two different 1-C-formyl glycals were synthesized from these ß-C-glycopyranosyl aldehydes by treatment in basic conditions. The optimization of reaction conditions was carried out following reactions between 1-C-formyl galactal and 4-hydroxycoumarin. Next, 1-C-formyl galactal and 1-C-formyl glucal were treated with nine substituted 4-hydroxy coumarins at room temperature (25 °C) in ethyl acetate for ∼1-2 h in the presence of l-proline to obtain exclusively single diastereomers of pyrano[3,2-c]pyranone derivatives in excellent yields. Four compounds were found to be active for the MCF-7 cancer cell line. The MTT assay, apoptosis assay and migration analysis showed significant death of the cancer cells induced by the synthesized compounds.

C-Glycopyranosyl aldehydes: emerging chiral synthons in organic synthesis.

Herein, we have summarized the vast array of synthetic processes that have been developed for the synthesis of C-glycopyranosyl aldehydes and diverse C-glycoconjugates derived from them by covering the literature reported from 1979 to 2023. Notwithstanding its challenging chemistry, C-glycosides are considered stable pharmacophores and are used as important bioactive molecules. The discussed synthetic methodologies to access C-glycopyranosyl aldehydes take advantage of seven key intermediates, viz. allene, thiazole, dithiane, cyanide, alkene, and nitromethane. Furthermore, the integration of complex C-glycoconjugates derived from varied C-glycopyranosyl aldehydes involves nucleophilic addition/substitution, reduction, condensation, oxidation, cyclo condensation, coupling, and Wittig reactions. In this review, we have categorized the synthesis of C-glycopyranosyl aldehydes and C-glycoconjugates on the basis of the methodology used for their synthesis and on types of C-glycoconjugates, respectively.

Triazole-linked nucleic acids: Synthesis, therapeutics and synthetic biology applications.

This article covers the triazole-linked nucleic acids where the triazole linkage (TL) replaces the natural phosphate backbone. The replacement is done at either a few selected linkages or all the phosphate linkages. Two triazole linkages, the four-atom TL1 and the six-atom TL2, have been discussed in detail. These triazole-modified oligonucleotides have found a wide range of applications, from therapeutics to synthetic biology. For example, the triazole-linked oligonucleotides have been used in the antisense oligonucleotide (ASO), small interfering RNA (siRNA) and clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 technology as therapeutic agents. Due to the ease of the synthesis and a wide range of biocompatibility, the triazole linkage TL2 has been used to assemble a functional 300-mer DNA from alkyne- and azide-functionalized 100-mer oligonucleotides as well as an epigenetically modified variant of a 335 base-pair gene from ten short oligonucleotides. These outcomes highlight the potential of triazole-linked nucleic acids and open the doors for other TL designs and artificial backbones to fully exploit the vast potential of artificial nucleic acids in therapeutics, synthetic biology and biotechnology.

Chemoenzymatic synthesis of bridged homolyxofuranosyl pyrimidine nucleosides: Bicyclic AZT analogues.

A greener chemo-enzymatic methodology has been developed for the synthesis of conformationally restricted diastereomeric homolyxofuranosyl pyrimidines (AZT analogue), i.e., (5'R)-3'-azido-3'-deoxy-2'-O,5'-C-bridged-ß-d-homolyxofuranosyl-uracil and thymine starting from inexpensive diacetone-d-glucofuranose in 18% and 21% overall yields, respectively. In one of the key steps in multistep synthesis of bicyclic AZT analogues, the primary hydroxyl group of 3'-azido-3'-deoxy-ß-d-glucofuranosyl pyrimidines has been acetylated using Novozyme® 435 in THF in 92% and 97% yields, respectively. The monoacetylated nucleoside was converted to desired bicyclic AZT analogue in two steps in an overall yield of 82% and 83%, respectively.

The role of activated androgen receptor in cofilin phospho-regulation depends on the molecular subtype of TNBC cell line and actin assembly dynamics.

Triple negative breast cancer (TNBC) is highly metastatic and of poor prognosis. Metastasis involves coordinated actin filament dynamics mediated by cofilin and associated proteins. Activated androgen receptor (AR) is believed to contribute to TNBC tumorigenesis. Our current work studied roles of activated AR and cofilin phospho-regulation during migration of three AR+ TNBC cell lines to determine if altered cofilin regulation can explain their migratory differences. Untreated or AR agonist-treated BT549, MDA-MB-453, and SUM159PT cells were compared to cells silenced for cofilin (KD) or AR expression/function (bicalutamide). Cofilin-1 was found to be the only ADF/cofilin isoform expressed in each TNBC line. Despite a significant increase in cofilin kinase caused by androgens, the ratio of cofilin:p-cofilin (1:1) did not change in SUM159PT cells. BT549 and MDA-MB-453 cells contain high p-cofilin levels which underwent androgen-induced dephosphorylation through increased cofilin phosphatase expression, but surprisingly maintain a leading-edge with high p-cofilin/total cofilin not found in SUM159PT cells. Androgens enhanced cell polarization in all lines, stimulated wound healing and transwell migration rates and increased N/E-cadherin mRNA ratios while reducing cell adhesion in BT549 and MDA-MB-453 cells. Cofilin KD negated androgen effects in MDA-MB-453 except for cell adhesion, while in BT549 cells it abrogated androgen-reduced cell adhesion. In SUM159PT cells, cofilin KD with and without androgens had similar effects in almost all processes studied. AR dependency of the processes were confirmed. In conclusion, cofilin regulation downstream of active AR is dependent on which actin-mediated process is being examined in addition to being cell line-specific. Although MDA-MB-453 cells demonstrated some control of cofilin through an AR-dependent mechanism, other AR-dependent pathways need to be further studied. Non-cofilin-dependent mechanisms that modulate migration of SUM159PT cells need to be investigated. Categorizing TNBC behavior as AR responsive and/or cofilin dependent can inform on decisions for therapeutic treatment.

Immune-modulation by 7, 8-diacetoxy-4-methylthiocoumarin in total body-irradiated mice: Implications for the mitigation of radiation-induced hematopoietic injury.

AIMS: Development of novel medical countermeasures (MCMs) against acute radiation syndrome (ARS) and the associated lethality involves protection from and/or mitigation of radiation-induced hematopoietic injury, a critical clinical component of ARS. We earlier identified the molecule 7,8-diacetoxy-4-methylthiocoumarin (DAMTC) as a potent mitigator of hematopoietic injury and mortality in C57BL/6 mice when administered 24 h following total body irradiation (TBI). In the present study, we investigated mechanisms and functional relevance of immune modulation by DAMTC during the mitigation of hematopoietic injury. MAIN METHODS: C57BL/6 mice were subjected to TBI doses of 3 and 7.6Gy; administered DAMTC intra-peritoneally 24 h post TBI. Isolation, characterization, intra-cellular cytokine analysis of myeloid cells from bone marrow and spleen accompanied by flow cytometric determination and characterization of B-lymphocytes, serum isolation from peripheral blood and cytokine analysis. KEY FINDINGS: Results showed that DAMTC induced stimulation of pro-inflammatory myeloid subsets in the bone marrow and spleen of TBI mice. Further, it promoted a favorable transition from Th2 to Th1 immunity, triggered humoral immunity, and activated an intricately balanced inflammatory response that appear to contribute to immune-modulation. SIGNIFICANCE: Thus, the present study shows that immune-modulation maybe one of the contributing factors for the mitigation of hematopoietic injury by DAMTC and underscores its efficacy as a potent mitigator of hematopoietic injury that merits to be developed further as a novel MCM to combat H-ARS.

Transition metal-catalyzed double Cvinyl-H bond activation: synthesis of conjugated dienes.

Conjugated dienes have occupied a pivotal position in the field of synthetic organic chemistry and medicinal chemistry. They act as important synthons for the synthesis of various biologically important molecules and therefore, gain tremendous attention worldwide. A wide range of synthetic routes to access these versatile molecules have been developed in the past decades. Transition metal-catalyzed cross-dehydrogenative coupling (CDC) has emerged as one of the utmost front-line research areas in current synthetic organic chemistry due to its high atom economy, efficiency, and viability. In this review, an up-to-date summary including scope, limitations, mechanistic studies, stereoselectivities, and synthetic applications of transition metal-catalyzed double Cvinyl-H bond activation for the synthesis of conjugated dienes has been reported since 2013. The literature reports mentioned in this review have been classified into three different categories, i.e. (a) Cvinyl-Cvinyl bond formation via oxidative homo-coupling of terminal alkenes; (b) Cvinyl-Cvinyl bond formation via non-directed oxidative cross-coupling of linear/cyclic alkenes and terminal/internal alkenes, and (c) Cvinyl-Cvinyl bond formation via oxidative cross-coupling of directing group bearing alkenes and terminal/internal alkenes. Overall, this review aims to provide a concise overview of the current status of the considerable development in this field and is expected to stimulate further innovation and research in the future.

Morphological signatures of actin organization in single cells accurately classify genetic perturbations using CNNs with transfer learning.

The actin cytoskeleton plays essential roles in countless cell processes, from cell division to migration to signaling. In cancer cells, cytoskeletal dynamics, cytoskeletal filament organization, and overall cell morphology are known to be altered substantially. We hypothesize that actin fiber organization and cell shape may carry specific signatures of genetic or signaling perturbations. We used convolutional neural networks (CNNs) on a small fluorescence microscopy image dataset of retinal pigment epithelial (RPE) cells and triple-negative breast cancer (TNBC) cells for identifying morphological signatures in cancer cells. Using a transfer learning approach, CNNs could be trained to accurately distinguish between normal and oncogenically transformed RPE cells with an accuracy of about 95% or better at the single cell level. Furthermore, CNNs could distinguish transformed cell lines differing by an oncogenic mutation from each other and could also detect knockdown of cofilin in TNBC cells, indicating that each single oncogenic mutation or cytoskeletal perturbation produces a unique signature in actin morphology. Application of the Local Interpretable Model-Agnostic Explanations (LIME) method for visually interpreting the CNN results revealed features of the global actin structure relevant for some cells and classification tasks. Interestingly, many of these features were supported by previous biological observation. Actin fiber organization is thus a sensitive marker for cell identity, and identification of its perturbations could be very useful for assaying cell phenotypes, including disease states.

Correction: Morphological signatures of actin organization in single cells accurately classify genetic perturbations using CNNs with transfer learning.

Correction for 'Morphological signatures of actin organization in single cells accurately classify genetic perturbations using CNNs with transfer learning' by Sydney Alderfer et al., Soft Matter, 2022, https://doi.org/10.1039/d2sm01000c.

Intensity dependence of sub-harmonics in cortical response to photic stimulation.

Objective. Periodic photic stimulation of human volunteers at 10 Hz is known to entrain their electroencephalography (EEG) signals. This entrainment manifests as an increment in power at 10, 20, 30 Hz. We observed that this entrainment is accompanied by the emergence of sub-harmonics, but only at specific frequencies and higher intensities of the stimulating signal. Thereafter, we describe our results and explain them using the physiologically inspired Jansen and Rit neural mass model (NMM).Approach. Four human volunteers were separately exposed to both high and low intensity 10 Hz and 6 Hz stimulation. A total of four experiments per subject were therefore performed. Simulations and bifurcation analysis of the NMM were carried out and compared with the experimental findings.Main results.High intensity 10 Hz stimulation led to an increment in power at 5 Hz across all the four subjects. No increment of power was observed with low intensity stimulation. However, when the same protocol was repeated with a 6 Hz photic stimulation, neither high nor low intensity stimulation were found to cause a discernible change in power at 3 Hz. We found that the NMM was able to recapitulate these results. A further numerical analysis indicated that this arises from the underlying bifurcation structure of the NMM.Significance. The excellent match between theory and experiment suggest that the bifurcation properties of the NMM are mirroring similar features possessed by the actual neural masses producing the EEG dynamics. NMMs could thus be valuable for understanding properties and pathologies of EEG dynamics, and may contribute to the engineering of brain-computer interface technologies.

Synthesis and applications of bicyclic sugar modified locked nucleic acids: A review.

A large number of Locked Nucleic Acids (LNAs) with variety of modifications and restricted conformations have been developed in the last few decades. These modifications have significantly improved the biological properties of oligonucleotides, when LNAs moieties were incorporated into them. Herein, the synthesis and applications of these modified locked nucleic acids as antisense oligonucleotides are discussed.Supplemental data for this article is available online at https://doi.org/10.1080/15257770.2022.2052316 .

The Longevity-Frailty Hypothesis: Evidence from COVID-19 Death Rates in Europe.

By the end of spring (31 May), the COVID-19 death rate was remarkably unevenly distributed across the countries in Europe. While the risk of COVID-19 mortality is known to increase with age, age-specific COVID-19 death rates across Europe were similarly unevenly distributed. To explain these mortality distributions, we present a simple model where more favorable survival environments promote longevity and the accumulation of health frailty among the elderly while less favorable survival environments induce a mortality selection process that results in lower health frailty. Because the age-related conditions of frailty render the elderly less resistant to SARS-CoV-2, pre-existing survival environments may be non-obviously positively related to the COVID-19 death rate. To quantify the survival environment parameter of our model, we leveraged historic cohort- and period-based age-specific probabilities of death and life expectancies at age 65 across Europe. All variables are significantly correlated with indicators of frailty like elderly dependence on others for personal and household care for a subset of European countries. With respect to COVID-19 death rates, we find significant positive relationships between our survival indicators and COVID-19 death rates across Europe, a result that is robust to statistical control for the capacity of a healthcare system to treat and survive infected persons, the timing and stringency of non-pharmaceutical interventions, population density, age structure, case rates and the volume of inbound international travelers, among other factors. To address possible concerns over reporting heterogeneity across countries, we show that results are robust to the substitution of our response variable for a measure of cumulative excess mortality. Also consistent with the intuition of our model, we also show a strong negative association between age-specific COVID-19 death rates and pre-existing all-cause age-specific mortality rates for a subset of European countries. Overall, results support the notion that variation in pre-existing frailty, resulting from heterogeneous survival environments, partially accounts for striking differences in COVID-19 death during the first wave of the pandemic.

Exact Distribution of Threshold Crossing Times for Protein Concentrations: Implication for Biological Timekeeping.

Stochastic protein accumulation up to some concentration threshold sets the timing of many cellular physiological processes. Here we obtain the exact distribution of first threshold crossing times of protein concentration, in either Laplace or time domain, and its associated cumulants: mean, variance, and skewness. The distribution is asymmetric, and its skewness nonmonotonically varies with the threshold. We study lysis times of E. coli cells for holin gene mutants of bacteriophage-λ and find a good match with theory. Mutants requiring higher holin thresholds show more skewed lysis time distributions as predicted. The theory also predicts a linear relationship between infection delay time and host doubling time for lytic viruses, that has recently been experimentally observed.

Chemical and chemoenzymatic routes to bridged homoarabinofuranosylpyrimidines: Bicyclic AZT analogues.

Conformationally restricted diastereomeric homoarabinofuranosylpyrimidines (AZT analogue), i.e., (5'R)-3'-azido-3'-deoxy-2'-O,5'-C-bridged-ß-ᴅ-homoarabinofuranosylthymine and -uracil had been synthesized starting from diacetone ᴅ-glucofuranose following chemoenzymatic and chemical routes in 34-35% and 24-25% overall yields, respectively. The quantitative and diastereoselective acetylation of primary hydroxy over two secondary hydroxy groups present in the key nucleoside precursor was mediated with Lipozyme® TL IM in 2-methyltetrahydrofuran following a chemoenzymatic pathway. Whereas, the protection of the primary hydroxy over the lone secondary hydroxy group in the key azido sugar precursor was achieved using bulky tert-butyldiphenylsilyl chloride (TBDPS-Cl) in pyridine in 92% yield following a chemical synthetic pathway. The chemoenzymatic method was found to be superior over the chemical method in respect of the number of synthetic steps and overall yield of the final product.

Radiosensitization of calreticulin-overexpressing human glioma cell line by the polyphenolic acetate 7, 8-diacetoxy-4-methylcoumarin.

BACKGROUND: Calreticulin (CRT), an endoplasmic reticulum-resident protein generally overexpressed in cancer cells, is associated with radiation resistance. CRT shows higher transacetylase activity, as shown by us earlier, in the presence of the polyphenolic acetates (like 7, 8-diacetoxy-4-methylcoumarin, DAMC) and modifies the activity of a number of proteins, thereby influencing cell signaling. AIM: To investigate the relationship between CRT expression and radiation response in a human glioma cell line and to evaluate the radiomodifying effects of DAMC. METHODS AND RESULTS: Studies were carried out in an established human glioma cell line (BMG-1) and its isogenic clone overexpressing CRT (CROE, CRT-overexpressing cells) by analyzing clonogenic survival, cell proliferation, micronuclei analysis, and protein levels by Western blotting as parameters of responses. CRT overexpression conferred resistance against radiation-induced cell death in CROE cells (D37  = 7.35 Gy, D10  = 12.6 Gy and D0  = 7.25 Gy) as compared to BMG-1 cells (D37  = 5.70 Gy, D10  = 9.2 Gy and D0  = 5.6 Gy). A lower level of radiation-induced micronuclei formation observed in CROE cells suggested that reduced induction and/or enhanced DNA repair partly contributed to the enhanced radioresistance. Consistent with this suggestion, we noted that CRT-mediated radioresistance was coupled with enhanced grp78 level and reduced P53 activation-mediated prodeath signaling, while no changes were noted in acetylation of histone H4. DAMC-enhanced radiation-induced delayed (secondary) apoptosis, which was higher in CROE cells. CONCLUSION: CRT overexpression confers resistance against radiation-induced death of human glioma cells, which can be overcome by the polyphenolic acetate DAMC.

Microwave Assisted Cu-Mediated Trifluoromethylation of Pyrimidine Nucleosides.

Trifluoromethylated nucleosides, such as trifluridine, have widespread applications in pharmaceuticals as anticancer and antiviral agents. However, site-selective addition of a trifluoromethyl group onto a nucleobase typically requires either inconvenient multi-step synthesis or expensive trifluoromethylation reagents, or results in low yield. This article describes a simple, scalable, and high-yielding protocol for late-stage direct trifluoromethylation of pyrimidine nucleosides via a microwave-irradiated pathway. First, 5-iodo pyrimidine nucleosides undergo complete benzoylation to obtain N3 -benzoyl-3',5'-di-O-benzoyl-5-iodo-pyrimidine nucleosides as key precursors. Next, trifluoromethylation is carried out under both conventional and microwave heating using an inexpensive and commercially accessible Chen's reagent, i.e., methyl fluorosulfonyldifluoroacetate, to produce N3 -benzoyl-3',5'-di-Obenzoyl-5-trifluoromethyl-pyrimidine nucleosides. The microwave-assisted transformation accentuates its simplicity, mild reaction conditions, and dominance, providing a facile route to access trifluoromethylation. Finally, the envisioned 5-trifluoromethyl pyrimidine nucleosides are obtained by a routine debenzoylation procedure. This concludes a convenient three-step synthesis to obtain trifluridine and its 2'-modified analogs on a gram scale with consistently high yields, starting from their respective iodo-precursors, and requires only one chromatographic purification at the trifluoromethylation step. Furthermore, this operationally simple protocol can be utilized as a definitive methodology to produce various other trifluoromethylated therapeutics. © 2021 Wiley Periodicals LLC. Basic Protocol: Synthesis of 5-trifluoromethyl pyrimidine nucleosides 4a-c Alternate Protocol: Conventional trifluoromethylation: Synthesis of N3-benzoyl-3',5'-di-O-benzoyl-5-trifluoromethyl pyrimidine nucleosides (3a-c).