Biology subjects

Guha, R.

Publications and source records attributed to Guha, R..

11 recordsLinked to original sources

bioRxiv2024

Mitochondrial calcium uniporter regulates human fibroblast-like synoviocytes invasion via altering mitochondrial dynamics and dictates rheumatoid arthritis pathogenesis

Rheumatoid arthritis (RA) is characterized by the aggressive migration and invasion of fibroblast-like synoviocytes (FLS) into cartilage and bone, a process that is significantly influenced by mitochondrial calcium uptake. This study highlights the critical role of mitochondrial calcium uniporter (MCU) in regulating FLS migration and mitochondrial dynamics, and its potential as a therapeutic target in RA. Notably, RA-FLS exhibited increased MCU expression and mitochondrial dysfunction compared to controls. Treatment with Ru360, a potent MCU inhibitor significantly reduced RA-FLS migration, calcium influx, and mitochondrial reactive oxygen species (ROS) levels, while restoring mitochondrial morphology and enhancing ATP production. Further analysis of MCU complex expression revealed elevated levels of MCU and other regulatory subunits (EMRE, MICU1, MICU2) in RA-FLS compared to controls, indicating mitochondrial dysfunction in RA. Mechanistically, MCU inhibition altered gene expression related to cytoskeletal dynamics, focal adhesion, and metabolic pathways. In human RA-FLS, MCU inhibition suppressed migration and invasion both in vitro and in vivo. The interaction between MCU and Miro1, essential for mitochondrial transport, was validated, and its disruption impaired FLS migration. Our findings highlight MCU as a promising therapeutic target to inhibit FLS migration and ameliorate RA progression.

immunology↗

bioRxiv2023

The implicit influence of pitch contours and emotional timbre on P300 components in an own-name oddball paradigm

ObjectivePrevious evidence has suggested that paralinguistic features of speech stimuli may influence the characteristics of P300 components when used in clinical evaluation of consciousness. However, it remains unknown what exact acoustic components of speech are influential in such tasks, and whether they are capable of interacting with attentional deployment (as indexed by P300) in an implicit manner. MethodTo study this question, we adapted here an auditory oddball-paradigm used in clinical practice (the "own-name" paradigm), and tested whether systematic transformations of pitch contours (i.e. rising or falling intonation) and emotional timbre (i.e. smiling or rough voices) on the participants names influenced P300 responses in 24 healthy participants. ResultsP300 responses to rising pitch contours were smaller than to falling pitch contours, possibly reflecting an interference of the rising contours with participant attention in the deviant-counting task. No such difference was observed with emotional timbre variations. Conclusion and SignificanceThese results suggest that the cognitive resources involved in pitch contour processing overlap more strongly than timbre with the resources required to count own-name deviants, and that rising pitch contours should be tested prospectively as a way to increase the saliency of consciousness evaluation in unconscious patients.

neuroscience↗

bioRxiv2023

Cortical responses to looming sources are explained away by the auditory periphery

A wealth of behavioral evidence indicate that sounds with increasing intensity (i.e. appear to be looming towards the listener) are processed with increased attentional and physiological resources compared to receding sounds. However, the neurophysiological mechanism responsible for such cognitive amplification remains elusive. Here, we show that the large differences seen between cortical responses to looming and receding sounds are in fact almost entirely explained away by nonlinear encoding at the level of the auditory periphery. We collected EEG mismatch negativity (MMN) data in response to deviant stimuli with both dynamic (looming and receding) and constant level (flat) differences to the standard in the same participants. We then combined a computational model of the auditory periphery with generative EEG methods (temporal response functions, TRFs) to model the single-participant MMN responses to flat deviants, and used them to predict the effect of the same mechanism on looming and receding stimuli. The flat model explained a remarkable 45% variance of the looming response, and 33% of the receding response. This provide striking evidence that MMN responses to looming and receding sounds result from the same cortical mechanism that generate MMN to constant-level deviants: all such differences are the sole consequence of their particular physical morphology getting amplified and integrated by peripheral auditory mechanisms. Thus, not all effects seen cortically proceed from top-down modulations by high-level decision variables, but can rather be performed early and efficiently by feed-forward peripheral mechanisms that evolved precisely to sparing subsequent networks with the necessity to implement such mechanisms.

neuroscience↗

bioRxiv2023

Oral administration of a specific p300/CBP Lysine acetyltransferase activator induces synaptic plasticity and repairing spinal cord injury

TTK21 is a small molecule activator of p300/CBP acetyltransferase activity, which upon conjugation with glucose-derived carbon nanosphere (CSP) can efficiently cross the blood-brain barrier and activate histone acetylation in the brain. Its role in adult neurogenesis and retention of long-term spatial memory upon intraperitoneal (IP) administration is well established. In this study, we have successfully demonstrated that CSP-TTK21 can be effectively administered via oral gavage. Using a combination of molecular biology, microscopy, and electrophysiological techniques, we systematically investigated the comparative efficacy of oral administration of CSP and CSP-TTK21 in wild-type mice, evaluating their functional effects in comparison to intraperitoneal (IP) administration. Our findings indicate that CSP-TTK21, when administered orally, induces long-term potentiation in the hippocampus without significantly altering basal synaptic transmission, a response comparable to that achieved through IP injection. Remarkably, in a spinal cord injury model, oral administration of CSP-TTK21 exhibits equivalent efficacy to IP administration. Furthermore, our research demonstrates that oral delivery of CSP-TTK21 leads to improvements in motor functions, histone acetylation dynamics, and increased expression of regeneration associated genes (RAGs) in a spinal injury rat model, mirroring the effectiveness of IP administration. Collectively, these results underscore the potential utility of CSP as an oral drug delivery system, particularly for targeting the neural system.

neuroscience↗

bioRxiv2021

EP300 (p300) mediated histone butyrylation is critical for adipogenesis

ObjectiveThe master epigenetic enzyme EP300 (p300) besides having lysine acetyltransferase activity can also catalyse other acylation modifications (propionylation, butyrylation, crotonylation etc.), the physiological implications of which are yet to be established fully. We hypothesized that p300 catalysed histone butyrylation may have a causal relationship with adipogenesis and the consequent obesity. MethodsHistone butyrylation pattern was investigated in 3T3L1 cells upon adipogenesis by immunoblotting and chromatin immunoprecipitation experiments. A small molecule modulator that could specifically inhibit p300 catalysed butyrylation without affecting its canonical acetyltransferase activity was screened from a series of compounds and then administered in differentiating 3T3L1 adipocytes as well as high fat diet-induced and genetically obese mice to validate the importance of butyrylation in adipogenesis. ResultsHistone butyrylation was increased upon adipogenesis both globally and locally in the promoters of pro-adipogenic genes along with an upregulation in the expression of acyl CoA generating enzyme Acss2, knockdown of which led to reduced butyrylation. Treatment of differentiating 3T3L1 cells with the p300 specific butyrylation inhibitor LTK-14A led to abrogation of adipogenesis with reduced expression of pro-adipogenic genes and inhibition of H4K5 butyrylation. LTK-14A administration could also attenuate weight gain in both mice models of obesity by preventing adipocyte hypertrophy via H4K5 butyrylation inhibition. ConclusionOur results indicate that p300 catalysed histone butyrylation may have a causal relationship with the process of adipogenesis. Site specific inhibition of butyrylation could lead to adipogenesis repression and hence this epigenetic modification could be targeted for obesity treatment. HighlightsO_LIHistone butyrylation has been established as a new epigenetic signature in the context of adipogenesis. C_LIO_LITo the best of our knowledge, this is the first report of a selective inhibitor of p300 catalysed histone acylation (butyrylation) without affecting its canonical acetyltransferase activity. C_LIO_LISpecific inhibition of H4K5 butyrylation could be a possible mechanism for inhibiting adipogenesis and hepatic steatosis leading to better control of obesity. C_LIO_LILTK-14A class of molecule could be developed as anti-obesity therapeutics. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=195 HEIGHT=200 SRC="FIGDIR/small/454641v3_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@1f29b35org.highwire.dtl.DTLVardef@abc5fcorg.highwire.dtl.DTLVardef@1534388org.highwire.dtl.DTLVardef@176d706_HPS_FORMAT_FIGEXP M_FIG Proposed model for the role of p300-mediated histone butyrylation in adipogenesis: In pre-adipocytes, there exists a basal level of histone acetylation while butyrylation is present to a much lesser extent owing to low stoichiometric levels of butyryl CoA. Induction of adipogenesis causes a simultaneous upregulation of histone acetylation and butyrylation marks leading to increased rate of adipogenesis and concomitant transcriptional activation of pro-adipogeneic genes. Onset of obesity in mice, either due to excess energy intake through high fat diet consumption or increased de novo synthesis of fatty acids due to leptin receptor gene mutation leading to hyperphagic behavior, is accompanied by adipocyte hyperplasia and hypertrophy. Both the organs of adipose tissue and liver were found to have enhanced levels of H4K5 butyrylation during obesity. LTK-14A, a butyrylation specific inhibitor could efficiently prevent the processes of adipogenesis and adipocyte hypertrophy due to inhibition of H4K5 butyrylation in these organs. Thus the compound could attenuate weight gain by selective inhibition of butyrylation without affecting acetylation, thereby highlighting the importance of histone butyrylation in adipogenesis and obesity. C_FIG

molecular biology↗

bioRxiv2020

Plasmodium falciparum malaria drives epigenetic reprogramming of human monocytes toward a regulatory phenotype

In malaria-naive children and adults, Plasmodium falciparum-infected red blood cells (Pf-iRBCs) trigger fever and other symptoms of systemic inflammation. However, in endemic areas where individuals experience repeated Pf infections over many years, the risk of Pf-iRBC-triggered inflammatory symptoms decreases with cumulative Pf exposure. The molecular mechanisms underlying these clinical observations remain unclear. Age-stratified analyses of monocytes collected from uninfected, asymptomatic Malian individuals before the malaria season revealed an inverse relationship between age and Pf-iRBC-inducible inflammatory cytokine (IL-1{beta}, IL-6 and TNF) production, whereas Malian infants and malaria-naive U.S. adults produced similarly high levels of inflammatory cytokines. Accordingly, monocytes of Malian adults produced more IL-10 and expressed higher levels of the regulatory molecules CD163, CD206, Arginase-1 and TGM2. These observations were recapitulated in an in vitro system of monocyte to macrophage differentiation wherein macrophages re-exposed to Pf-iRBCs exhibited attenuated inflammatory cytokine responses and a corresponding decrease in the epigenetic marker of active gene transcription, H3K4me3, at inflammatory cytokine gene loci. Together these data indicate that Pf induces epigenetic reprogramming of monocytes/macrophages toward a regulatory phenotype that attenuates inflammatory responses during subsequent Pf exposure. These findings also suggest that past malaria exposure could mitigate monocyte-associated immunopathology induced by other pathogens such as SARS-CoV-2. Author SummaryThe malaria parasite is mosquito-transmitted and causes fever and other inflammatory symptoms while circulating in the bloodstream. However, in regions of high malaria transmission the parasite is less likely to cause fever as children age and enter adulthood, even though adults commonly have malaria parasites in their blood. Monocytes are cells of the innate immune system that secrete molecules that cause fever and inflammation when encountering microorganisms like malaria. Although inflammation is critical to initiating normal immune responses, too much inflammation can harm infected individuals. In Mali, we conducted a study of a malaria-exposed population from infants to adults and found that participants monocytes produced less inflammation as age increases, whereas monocytes of Malian infants and U.S. adults, who had never been exposed to malaria, both produced high levels of inflammatory molecules. Accordingly, monocytes exposed to malaria in the laboratory became less inflammatory when re-exposed to malaria again later, and these monocytes turned down their inflammatory genes. This study helps us understand how people become immune to inflammatory symptoms of malaria and may also help explain why people in malaria-endemic areas appear to be less susceptible to the harmful effects of inflammation caused by other pathogens such as SARS-CoV-2.

immunology↗

bioRxiv2019

Moving Targets: Monitoring Target Trends in Drug Discovery by Mapping Targets, GO Terms, and Diseases

Drug Discovery is a lengthy and costly process and has faced a period of declining productivity within the last two decades. As a consequence, integrative data-driven approaches are nowadays on the rise in pharmaceutical research, making use of an inter-connected (network) view on diseases. In addition, evidence-based decisions are alleviated by studying the time evolution of innovation trends in drug discovery.\n\nIn this paper a new approach leveraging data mining and data integration for inspecting target innovation trends protein family-wise is presented. The study highlights protein families which are receiving emerging interest in the drug discovery community (mainly kinases and G protein coupled receptors) and those with areas of interest in target space that have just emerged in the scientific literature (mainly kinases and transporters) highlighting novel opportunities for drug intervention.\n\nIn order to delineate the evolution of target-driven research interest from a biological perspective, trends in biological process annotations from Gene Ontology (GO) and disease annotations from DisGeNet for major target families are captured. The analysis reveals an increasing interest in targets related to immune system processes, and a recurrent trend for targets involved in circulatory system processes. At the level of disease annotations, targets associated to e.g., cancer-related pathologies as well as to intellectual disability and schizophrenia are increasingly investigated nowadays.\n\nCan this knowledge be used to study the \"movement of targets\" in a network view and unravel new links between diseases and biological processes? We tackled this question by creating dynamic network representations considering data from different time periods. The dynamic network for immune system process-associated targets suggest that e.g. breast cancer as well as schizophrenia are linked to the same targets (cannabinoid receptor CB2 and VEGFR2) thus suggesting similar treatment options which could be confirmed by literature search. The methodology has the potential to identify other drug repurposing candidates and enables researchers to capture trends in research attention in target space at an early stage.\n\nThe KNIME workflows and R scripts used in this study are publicly available from https://github.com/BZdrazil/Moving_Targets.\n\nAuthor summaryIn this study we have investigated target innovation in drug discovery over a period of 22 years (1995-2016) by extracting time trends of research interest (as published in the scientific literature and stored in the ChEMBL database) in certain protein classes inspecting different measures (numbers of pharmacological measurements, targets, papers, and drugs). Focusing on the most relevant protein classes in drug discovery (G protein-coupled receptors, kinases, ion channels, nuclear receptors, proteases, and transporters), we further linked single targets to Gene Ontology (GO) biological process annotations and inspected steep increasing or decreasing trends of GO annotations within target families over time. We also tracked trends in disease annotations from DisGeNET by filtering out diseases linked to targets with emerging trends in research interest. Finally, targets, GO terms, and diseases are interconnected in network representations and shifts in research foci are investigated over time. This new methodology which utilizes data mapping and data analysis can be used to explore trends in research attention target family-wise, to uncover previously unknown links between diseases and biological processes and to identify potential candidates for drug repurposing.

bioinformatics↗

bioRxiv2019

Non-uniform Crowding Enhances Transport: Relevance to Biological Environments

The cellular cytoplasm is crowded with macromolecules and other species that occupy up to 40% of the available volume. Previous studies have reported that for high crowder molecule concentrations, colloidal tracer particles have a dampened diffusion due to the higher solution viscosity. However, these studies employed uniform distributions of crowder molecules. We report a scenario, previously unexplored experimentally, of increased tracer transport driven by a non-uniform concentration of crowder macromolecules. In gradients of polymeric crowder, tracer particles undergo transport several times higher than that of their bulk diffusion rate. The direction of the transport is toward regions of lower crowder concentration. Mechanistically, hard-sphere interactions and the resulting volume exclusion between the tracer and crowder increases the effective diffusion by inducing a convective motion of tracers. Strikingly, soft deformable particles show even greater enhancement in transport in crowder gradients compared to similarly sized hard particles. We propose a model that supports the data and quantifies a \"diffusiophoretic buoyancy force\" when a tracer is in a gradient of crowder concentration. Overall, this demonstration of enhanced transport in non-uniform distributions of crowder is anticipated to clarify aspects of multi-component intracellular transport.

biophysics↗

bioRxiv2019

A High-Throughput Screen of a Library of Therapeutics Identifies Substrates of P-glycoprotein

The ATP-binding cassette transporter P-glycoprotein (P-gp) is known to limit brain penetration of many chemotherapy drugs. Although Food and Drug Administration guidelines require that potential interactions of investigational drugs with P-gp be explored, often this information does not enter into the literature. As such, we developed a high-throughput screen (HTS) to identify substrates of P-gp from a series of chemical libraries, testing a total of 10,804 compounds, most of which have known mechanisms of action. We used the CellTiter-Glo viability assay to test library compounds against parental KB-3-1 human cervical adenocarcinoma cells and the colchicine-selected sub-line KB-8-5-11 that over-expresses P-gp. KB-8-5-11 cells were also tested in the presence of a P-gp inhibitor (tariquidar) to assess reversability of transporter-mediated resistance. Of the tested compounds, a total of 90 P-gp substrates were identified including 55 newly identified P-gp substrates. Substrates were confirmed using an orthogonal killing assay against HEK-293 cells transfected with P-gp. We confirmed that AT7159 (cyclin-dependent kinase inhibitor); AT9283, (Janus kinase 2/3 inhibitor); ispinesib (kinesin spindle protein inhibitor); gedatolisib (PKI-587, phosphoinositide 3-kinase/mammalian target of rampamycin inhibitor); GSK-690693 (AKT inhibitor); and KW-2478 (heat shock protein 90 inhibitor) were substrates, and direct ATPase stimulation was assessed. ABCG2 was also found to confer high levels of resistance to AT9283, GSK-690693 and gedatolisib, while ispinesib, AT7519 and KW-2478 were weaker substrates. Combinations of P-gp substrates and inhibitors were assessed to demonstrate on-target synergistic cell killing. This data will be of use in determining understanding how chemotherapeutic agents will cross the blood-brain barrier.

pharmacology and toxicology↗

bioRxiv2018

Cytotoxic Profiling of Annotated and Diverse Chemical Libraries Using Quantitative High-Throughput Screening

Cell-based phenotypic screening is a commonly used approach to discover biological pathways, novel drug targets, chemical probes and high-quality hit-to-lead molecules. Many hits identified from high-throughput screening campaigns are ruled out through a series of follow-up potency, selectivity/specificity, and cytotoxicity assays. Prioritization of molecules with little or no cytotoxicity for downstream evaluation can influence the future direction of projects, so cytotoxicity profiling of screening libraries at an early stage is essential for increasing the likelihood of candidate success. In this study, we assessed cell-based cytotoxicity of nearly 10,000 compounds in NCATS annotated libraries, and over 100,000 compounds in a diversity library, against four normal cell lines (HEK 293, NIH 3T3, CRL-7250 and HaCat) and one cancer cell line (KB 3-1, a HeLa subline). This large-scale library profiling was analyzed for overall screening outcomes, hit rates, pan-activity and selectivity. For the annotated library, we also examined the primary targets and mechanistic pathways regularly associated with cell death. To our knowledge, this is the first study to use high-throughput screening to profile a large screening collection (>100,000 compounds) for cytotoxicity in both normal and cancer cell lines. The results generated here constitutes a valuable resource for the scientific community and provides insight on the extent of cytotoxic compounds in screening libraries, identifying and avoiding compounds with cytotoxicity during high-throughput screening campaigns.

cell biology↗

bioRxiv2017

Drug Target Ontology to Classify and Integrate Drug Discovery Data

BackgroundOne of the most successful approaches to develop new small molecule therapeutics has been to start from a validated druggable protein target. However, only a small subset of potentially druggable targets has attracted significant research and development resources. The Illuminating the Druggable Genome (IDG) project develops resources to catalyze the development of likely targetable, yet currently understudied prospective drug targets. A central component of the IDG program is a comprehensive knowledge resource of the druggable genome.\n\nResultsAs part of that effort, we have been developing a framework to integrate, navigate, and analyze drug discovery data based on formalized and standardized classifications and annotations of druggable protein targets, the Drug Target Ontology (DTO). DTO was constructed by extensive curation and consolidation of various resources. DTO classifies the four major drug target protein families, GPCRs, kinases, ion channels and nuclear receptors, based on phylogenecity, function, target development level, disease association, tissue expression, chemical ligand and substrate characteristics, and target-family specific characteristics. The formal ontology was built using a new software tool to auto-generate most axioms from a database while also supporting manual knowledge acquisition. A modular, hierarchical implementation facilitates development and maintenance and makes use of various external ontologies, thus integrating the DTO into the ecosystem of biomedical ontologies. As a formal OWL-DL ontology, DTO contains asserted and inferred axioms. Modeling data from the Library of Integrated Network-based Cellular Signatures (LINCS) program illustrates the potential of DTO for contextual data integration and nuanced definition of important drug target characteristics. DTO has been implemented in the IDG user interface Portal, Pharos and the TIN-X explorer of protein target disease relationships.\n\nConclusionsDTO was built based on the need for a formal semantic model for druggable targets including various related information such as protein, gene, protein domain, protein structure, binding site, small molecule drug, mechanism of action, protein tissue localization, disease association, and many other types of information. DTO will further facilitate the otherwise challenging integration and formal linking to biological assays, phenotypes, disease models, drug poly-pharmacology, binding kinetics and many other processes, functions and qualities that are at the core of drug discovery. The first version of DTO is publically available via the website http://drugtargetontology.org/, Github (https://github.com/DrugTargetOntology/DTO), and the NCBO Bioportal (https://bioportal.bioontology.org/ontologies/DTO). The long-term goal of DTO is to provide such an integrative framework and to populate the ontology with this information as a community resource.

bioinformatics↗