Biology subjects

Wilson, K.

Publications and source records attributed to Wilson, K..

At least 37 records · Page 2Linked to original sources

bioRxiv2022

Functional brain region-specific neural spheroids for modeling neurological diseases and therapeutics screening

3D spheroids have emerged as powerful drug discovery tools given their high-throughput screening (HTS) compatibility. Here, we established a method for generating functional neural spheroids with differentiated human induced pluripotent stem cell (hiPSC)-derived neurons and astrocytes at cell type compositions mimicking specific regions of the human brain. Recordings of intracellular calcium oscillations were used as functional assays, and the utility of this spheroids system was shown through disease modelling, drug testing, and formation of assembloids to model neurocircuitry. We developed disease models for Alzheimers and Parkinsons Disease through incorporating genetically engineered cells into spheroids, and Opioid Use Disorder by chronically treating spheroids with DAMGO. We reversed baseline functional deficits in each disease model with clinically approved treatments and showed that assembloid activity can be chemogenetically manipulated. Here, we lay the groundwork for brain region-specific spheroids as a robust functional assay platform for HTS studies.

bioengineering↗

bioRxiv2022

Long-term temporal trends in gastrointestinal parasite infection in wild Soay sheep

Monitoring the prevalence and abundance of parasites over time is important for addressing their potential impact on host life-histories, immunological profiles, and their influence as a selective force. Only long-term ecological studies have the potential to shed light on both the temporal trends in infection prevalence and abundance and the drivers of such trends, because of their ability to dissect drivers that may be confounded over shorter time scales. Despite this, only a relatively small number of such studies exist. Here, we analysed changes in the prevalence and abundance of gastrointestinal parasites in the wild Soay sheep population of St Kilda across 31 years. The host population density has increased across the study, and population density is known to increase parasite transmission, but we found that density and year explained temporal variation in parasite prevalence and abundance independently. Prevalence of both strongyle nematodes and coccidian microparasites increased during the study, and this effect varied between lambs, yearlings and adults. Meanwhile, abundance of strongyles was more strongly linked to host density than to temporal (yearly) dynamics, while abundance of coccidia showed a strong temporal trend without any influence of density. Strikingly, coccidian abundance increased threefold across the course of the study in lambs, while increases in yearlings and adults were negligible. Our decades-long, intensive, individual-based study will enable the role of environmental change and selection pressures in driving these dynamics to be determined, potentially providing unparalleled insight into the drivers of temporal variation in parasite dynamics in the wild. Key findingsO_LIWe studied temporal trends in gastrointestinal parasites of wild sheep over 31 years C_LIO_LIYear and host population density explained temporal variation in parasites independently C_LIO_LIPrevalence of both strongyle nematodes and coccidia varied across the study period C_LIO_LIAbundance of strongyles was more closely linked to host density than year C_LIO_LIAbundance of coccidia increased threefold over time in lambs, but did not vary in adults C_LI

ecology↗

bioRxiv2022

Evaluation of PRKACA as a Therapeutic Target for Fibrolamellar Carcinoma

Background & AimsFibrolamellar carcinoma (FLC) is a rare, difficult-to-treat liver cancer primarily affecting pediatric and adolescent patients, and for which precision medicine approaches have historically not been possible. The DNAJB1-PRKACA gene fusion was identified as a driver of FLC pathogenesis. We aimed to assess whether FLC tumors maintain dependency on this gene fusion and determine if PRKACA is a viable therapeutic target. MethodsFLC patient-derived xenograft (PDX) shRNA cell lines were implanted subcutaneously into female NOD-SCID mice and tumors were allowed to develop prior to randomization to doxycycline (to induce knockdown) or control groups. Tumor development was assessed every 2 days. To assess the effect of treatment with novel selective PRKACA small molecule kinase inhibitors, BLU0588 and BLU2864, FLC PDX tumor cells were implanted subcutaneously into NOD-SCID mice and tumors allowed to develop. Mice were randomized to treatment (BLU0588 and BLU2864, orally, once daily) or control groups and tumor size determined as above. ResultsKnockdown of DNAJB1-PRKACA reversed a FLC-specific gene signature and reduced PDX tumor growth in mice compared to the control group. Furthermore, FLC PDX tumor growth was significantly reduced with BLU0588 and BLU2864 treatment versus control (P = 0.003 and P = 0.0005, respectively). ConclusionsWe demonstrated, using an inducible knockdown and small molecule approaches, that FLC PDX tumors were dependent upon DNAJB1-PRKACA fusion activity. In addition, this study serves as a proof-of-concept that PRKACA is a viable therapeutic target for FLC and warrants further investigation.

cancer biology↗

bioRxiv2021

Oncogenic RAS commandeers amino acid sensing machinery to aberrantly activate mTORC1 in multiple myeloma

Oncogenic mutations within the RAS pathway are common in multiple myeloma (MM), an incurable malignancy of plasma cells. However, the mechanisms of pathogenic RAS signaling in this disease remain enigmatic and difficult to inhibit therapeutically. We employed an unbiased proteogenomic approach to dissect RAS signaling in MM by combining genome-wide CRISPR-Cas9 screening with quantitative mass spectrometry focused on RAS biology. We discovered that mutant isoforms of RAS organized a signaling complex with the amino acid transporter, SLC3A2, and MTOR on endolysosomes, which directly activated mTORC1 by co-opting amino acid sensing pathways. MM tumors with high expression of mTORC1-dependent genes were more aggressive and enriched in RAS mutations, and we detected interactions between RAS and MTOR in MM patient tumors harboring mutant RAS isoforms. Inhibition of RAS-dependent mTORC1 activity synergized with MEK and ERK inhibitors to quench pathogenic RAS signaling in MM cells. This study redefines the RAS pathway in MM and provides a mechanistic and rational basis to target this novel mode of RAS signaling.

cancer biology↗

bioRxiv2021

Rapid spread of a symbiotic virus in a major crop pest following wide-scale adoption of Bt-cotton in China

Bacillus thuringiensis (Bt) crops have been widely planted and the effects of Bt-crops on populations of the target and non-target insect pests were well studied. However, the effects of Bt-crops exposure on microorganisms that interact with crop pests havent previously been quantified. Here, we use laboratory and field data to show that infection of Helicoverpa armigera with a symbiotic densovirus (HaDV2) is associated with its enhanced growth and resistance to Bt-cotton. Moreover, field monitoring showed a much higher incidence of cotton bollworm infection with HaDV2 in regions cultivated with Bt-cotton than in regions without it, with the rate of densovirus infection increasing with increasing use of Bt-cotton. RNA-seq suggested resistance to both baculovirus and Cry1Ac were enhanced via the immune-related pathways. These suggest that the exposure to Bt-crops has selected for beneficial interactions between the target pest and a symbiotic microorganism that enhances its performance on Bt-crops under field conditions.

ecology↗

bioRxiv2020

Resting-state functional connectivity and psychopathology in Klinefelter syndrome (47, XXY)

Klinefelter syndrome (47, XXY; Henceforth: XXY syndrome) is a high impact but poorly understood genetic risk factor for neuropsychiatric impairment. Here, we provide the first neuroimaging study to map resting-state functional connectivity (rsFC) changes in XXY syndrome and ask how these might relate to brain anatomy and psychopathology. We collected resting state functional magnetic resonance imaging data from 75 individuals with XXY and 84 healthy XY males. We implemented a brain-wide screen to identify regions with altered global rsFC in XXY vs. XY males, and then used seed-based analysis to decompose these alterations. We further compared rsFC changes with regional changes in brain volume from voxel-based morphometry and tested for correlations between rsFC and symptom variation within XXY syndrome. We found that XXY syndrome was characterized by increased global rsFC in the left dorsolateral prefrontal cortex (DLPFC), associated with overconnectivity with diverse rsFC networks. Regional rsFC changes were partly coupled to regional volumetric changes in XXY syndrome. Within the precuneus, variation in DLPFC rsFC within XXY syndrome was correlated with the severity of psychopathology in XXY individuals. Our findings provide the first view of altered functional brain connectivity in XXY syndrome and delineate links between these alterations and those relating to both brain anatomy and psychopathology. Taken together, these insights advance biological understanding of XXY syndrome as a disorder in its own right, and as a model of genetic risk for psychopathology more broadly.

neuroscience↗

bioRxiv2020

Functional interpretation of ATAD3A variants in neuro-mitochondrial phenotypes

BackgroundThe ATPase family AAA-domain containing protein 3A (ATAD3A) is a nuclear-encoded mitochondrial membrane anchored protein involved in diverse processes including mitochondrial dynamics, mitochondrial DNA organization, and cholesterol metabolism. Biallelic deletions (null), recessive missense variants (hypomorph), and heterozygous missense variants or duplications (antimorph) in ATAD3A lead to neurological syndromes in humans. ObjectiveTo expand the mutational spectrum of ATAD3A variants and to provide functional interpretation of missense alleles in trans to deletion alleles. MethodsExome sequencing was used to identify single nucleotide variants (SNVs) and copy number variants (CNVs) in ATAD3A in individuals with neurological and mitochondrial phenotypes. A Drosophila Atad3A Gal4 trap null allele was generated using CRISPR-Cas9 genome editing technology to aid interpretation of variants. ResultsWe report 13 individuals from 8 unrelated families with biallelic ATAD3A variants. Four of the identified missense variants, p.(Leu77Val), p.(Phe50Leu), p.(Arg170Trp), p.(Gly236Val), were inherited in trans to loss-of-function alleles. A fifth missense variant, p.(Arg327Pro), was homozygous. Affected individuals exhibited findings previously associated with ATAD3A pathogenic variation, including developmental delay, hypotonia, congenital cataracts, hypertrophic cardiomyopathy, and cerebellar atrophy. Drosophila studies indicated that Phe50Leu, Gly236Val, and Arg327Pro are severe loss-of-function alleles leading to early developmental lethality and neurogenesis defects, whereas Leu77Val and Arg170Trp are partial loss of function alleles that cause progressive locomotion defects. Moreover, Leu77Val and Arg170Trp expression leads to an increase in autophagy and mitophagy in adult muscles. ConclusionOur findings expand the allelic spectrum of ATAD3A variants, and exemplify the use of a functional assay in Drosophila to aid variant interpretation.

genetics↗

bioRxiv2020

Evaluating institutional open access performance: Methodology, challenges and assessment

Open Access to research outputs is becoming rapidly more important to the global research community and society. Changes are driven by funder mandates, institutional policy, grass-roots advocacy and culture change. It has been challenging to provide a robust, transparent and updateable analysis of progress towards open access that can inform these interventions, particularly at the institutional level. Here we propose a minimum reporting standard and present a large-scale analysis of open access progress across 1,207 institutions world-wide that shows substantial progress being made. The analysis detects responses that coincide with policy and funding interventions. Among the striking results are the high performance of Latin American and African universities, particularly for gold open access, whereas overall open access levels in Europe and North America are driven by repository-mediated access. We present a top-100 of global universities with the worlds leading institutions achieving around 80% open access for 2017 publications.

scientific communication and education↗

bioRxiv2020

Evaluating institutional open access performance: Sensitivity analysis

In the article “Evaluating institutional open access performance: Methodology, challenges and assessment” we develop the first comprehensive and reproducible workflow that integrates multiple bibliographic data sources for evaluating institutional open access (OA) performance. The major data sources include Web of Science, Scopus, Microsoft Academic, and Unpaywall. However, each of these databases continues to update, both actively and retrospectively. This implies the results produced by the proposed process are potentially sensitive to both the choice of data source and the versions of them used. In addition, there remain the issue relating to selection bias in sample size and margin of error. The current work shows that the levels of sensitivity relating to the above issues can be significant at the institutional level. Hence, the transparency and clear documentation of the choices made on data sources (and their versions) and cut-off boundaries are vital for reproducibility and verifiability.View Full Text

scientific communication and education↗

bioRxiv2019

High-depth resequencing reveals hybrid population and insecticide resistance characteristics of fall armyworm (Spodoptera frugiperda) invading China

The rapid wide-scale spread of fall armyworm (Spodoptera frugiperda) has caused serious crop losses globally. However, differences in the genetic background of subpopulations and the mechanisms of rapid adaptation behind the invasion are still not well understood. Here we report a 393.25-M chromosome-level genome assembly of fall armyworm with scaffold N50 of 13.3 M consisting of 23281 annotated protein-coding genes. Genome-wide resequencing of 105 samples from 16 provinces in China revealed that the fall armyworm population comprises a complex inter-strain hybrid, mainly with the corn-strain genetic background and less of the rice-strain genetic background, which highlights the inaccuracy of strain identification using mitochondrial or Tpi genes. An analysis of genes related to pesticide- and Bt-resistance showed that the risk of fall armyworm developing resistance to conventional pesticides is very high, while remaining currently susceptible to Bt toxins. Laboratory bioassay results showed that insects invading China carry resistance to organophosphate and pyrethroid pesticides, but are sensitive to genetically modified maize expressing Cry1Ab in field experiments. Additionally, we found that two mitochondrial fragments are inserted into the nuclear genome, and the insertion event occurred after the differentiation of the two strains. This study represents a valuable advancement toward the analysis of genetic differences among subpopulations and improving management strategies for fall armyworm.

genomics↗

bioRxiv2019

Keeping it clean: the cell culture quality control experience at the National Center for Advancing Translational Sciences

Quality control monitoring of cell lines utilized in biomedical research is of critical importance, and is critical for reproducibility of data. Two key pitfalls in tissue culture are 1) cell line authenticity, and 2) mycoplasma contamination. As a collaborative research institute, the National Center for Advancing Translational Sciences receives cell lines from a range of commercial and academic sources, that are adapted for high-throughput screening. Here, we describe the implementation of a routine NCATS-wide mycoplasma testing and short-tandem repeat (STR) testing for cell lines. Initial testing identified a >10% mycoplasma contamination rate, but the implementation of clearly defined protocols that included immediate destruction of contaminated cell lines wherever possible has led to a much-reduced mycoplasma contamination rate, and data for >2,000 cell line samples tested over 3 years, and case studies are provided. STR testing of 170 cell lines with established STR profiles revealed only 5 mis-identified cell lines received from external labs. The data collected over the three years since implementation of this systematic testing demonstrates the importance of continual vigilance for rapid identification of problem cell lines, for ensuring reproducible data in translational science research.

cell biology↗

bioRxiv2019

Comparison of bibliographic data sources: Implications for the robustness of university rankings

Universities are increasingly evaluated, both internally and externally on the basis of their outputs. Often these are converted to simple, and frequently contested, rankings based on quantitative analysis of those outputs. These rankings can have substantial implications for student and staff recruitment, research income and perceived prestige of a university. Both internal and external analyses usually rely on a single data source to define the set of outputs assigned to a specific university. Although some differences between such databases are documented, few studies have explored them at the institutional scale and examined the implications of these differences for the metrics and rankings that are derived from them. We address this gap by performing detailed bibliographic comparisons between three key databases: Web of Science (WoS), Scopus and, the recently relaunched Microsoft Academic (MSA). We analyse the differences between outputs with DOIs identified from each source for a sample of 155 universities and supplement this with a detailed manual analysis of the differences for fifteen universities. We find significant differences between the sources at the university level. Sources differ in the publication year of specific objects, the completeness of metadata, as well as in their coverage of disciplines, outlets, and publication type. We construct two simple rankings based on citation counts and open access status of the outputs for these universities and show dramatic changes in position based on the choice of bibliographic data sources. Those universities that experience the largest changes are frequently those from non-English speaking countries and those that are outside the top positions in international university rankings. Overall MSA has greater coverage than Scopus or WoS, but has less complete affiliation metadata. We suggest that robust evaluation measures need to consider the effect of choice of data sources and recommend an approach where data from multiple sources is integrated to provide a more robust dataset.

scientific communication and education↗

bioRxiv2019

Hematological and Neurological Expressed 1 (HN1): a predictive and pharmacodynamic biomarker of metformin response in endometrial cancers.

Preoperative use of metformin in obese women with endometrioid endometrial cancer (EEC) reduces tumor proliferation and inhibits the mammalian target of rapamycin (mTOR) pathway, though is only effective in select cases. This study sought to identify a predictive and/or pharmacodynamic proteomic signature of metformin response to tailor its pharmacologic use. Matched pre- and post-metformin treated tumor tissues from a recently completed phase 0 window trial of metformin in EEC patients (ClinicalTrials.gov: NCT01911247), were analyzed by mass spectrometry (MS)-based proteomic and immunohistochemical analyses. Hematological and neurological expressed 1 (HN1) was significantly elevated in metformin responders (n=13) versus non-responders (n=7), which was also found to decrease in abundance in metformin responders following treatment; observations that were verified by immunohistochemical staining for HN1. Metformin response and loss of HN1 was assessed in RL95-2 and ACI-181 endometrial cancer cell lines. We further identified that silencing of HN1 abundance does not alter cellular response to metformin or basal cell proliferation, but that HN1 abundance does decrease in response to metformin treatment in RL95-2 and ACI-181 endometrial cancer cell lines. These data suggest that HN1 represents a predictive and pharmacodynamic biomarker of metformin response that, if validated in larger patient populations, may enable pre-operative EEC patient stratification to metformin treatment and the ability to monitor patient response.\n\nNovelty and ImpactHematological and Neurological Expressed 1 (HN1) is elevated in endometrioid endometrial cancer (EEC) patients that respond to metformin vs non-responders and decreases after treatment. HN1 decreases after metformin treatment of EEC cells in vitro, but is not necessary for metformin response or proliferation. We report HN1 as a novel predictive and pharmacodynamic biomarker of metformin response in EEC that may enable pre-operative EEC patient stratification to metformin treatment and the ability to monitor patient response.

cancer biology↗

bioRxiv2018

Macronutrients modulate resistance to infection and immunity in Drosophila

Immunity and nutrition are two essential modulators of individual fitness However, while the implications of immune function and nutrition on an individual s lifespan and reproduction are known, the interplay between feeding behaviour, infection, and immune function, remains poorly understood. In this study, we used the fruit fly, Drosophila melanogaster, to investigate how infection through septic injury modulates nutritional intake, and how macronutrient balance affects survival to infection by the pathogenic Gram-positive bacterium Micrococcus luteus. Our results show that infected flies maintain carbohydrate intake, but reduce protein intake, thereby shifting from a protein-to-carbohydrate (P:C) ratio of ~1:4 to ~1:10 relative to non-infected and sham-infected flies. Strikingly, we found that the proportion of flies dying after M. luteus infection was significantly lower when flies were fed a low-P high-C diet, revealing that flies shift their macronutrient intake as means of nutritional self-medication against bacterial infection. This is likely due to the effects of macronutrient balance on the regulation of the constitutive expression of innate immune genes, as a low-P high-C diet was linked to an up-regulation in the expression of key antimicrobial peptides. Together, our results reveal the intricate relationship between macronutrient intake and resistance to infection, and integrate the molecular cross-talk between metabolic and immune pathways into the framework of nutritional immunology.

physiology↗

bioRxiv2018

Comparison of microbiome and culture techniques for determination of gastrointestinal microbial communities in ceca of chickens

The use of 16S next generation sequencing (NGS) technology to identify the relative abundance of microbial communities have become the standard when studying the intestinal microbiome. The increased use is due to the ability to identify a proportion of bacteria that cannot be observed with culture-based methods. However, culture-based techniques are acceptable to identify key bacterial groups, yet may grossly underestimate the microbial community in question. Since there is limited research comparing NGS results to colony forming units (CFU), the objective of this study was to compare total Enterobacteriaceae and lactic acid bacteria (LAB) recovery with culture techniques (CFU/g ceca) to total number of reads from operational taxonomic units (OTU) categorized as Enterobacteriaceae or LAB from Illumina MiSeq platform from matched chick cecal samples at three and 10 days of age. Both CFU recovery (1.09x109 {+/-} 2.42x108; 1.37x108 {+/-} 5.57x107) and reads (5460 {+/-} 1164; 282 {+/-} 163) belonging to Enterobacteriaceae decreased by 10 days of age (p < 0.001). Similarly, LAB reads decreased over time (21,128 {+/-} 2262; 6220 {+/-} 817, respectively p < 0.0001). However, LAB CFU recovery increased by 10 days (1.18x108 {+/-} 1.91x107; 1.62x109 {+/-} 5.00x108, respectively p < 0.01). At three days the Pearsons correlation was -0.082 between CFU of culturable Enterobacteriaceae to reads and culturable LAB CFU to reads at 0.097, showing no correlation (p = 0.606, 0.551; respectively). By 10 days, no correlation of reads and CFU occurred with Enterobacteriaceae (r=-0.049; p-value = 0.769) while with LAB the correlation was 0.290 (p = 0.066) at 10 days. The CFU may be appropriate to identify a few families that change due to treatment or product. Without identifying viable cells to DNA recovered from NGS, there will always be the question whether the reads within the binned OTU in the intestinal tract is accurate.

microbiology↗

bioRxiv2018

A conserved role of the insulin-like signaling pathway in uric acid pathologies revealed in Drosophila melanogaster

Elevated uric acid (UA) is a key factor for disorders, including gout or kidney stones and result from abrogated expression of Urate Oxidase (Uro) and diet. To understand the genetic pathways influencing UA metabolism we established a Drosophila melanogaster model with elevated UA using Uro knockdown. Reduced Uro expression resulted in the accumulation of UA concretions and diet-dependent shortening of lifespan. Inhibition of insulin-like signaling (ILS) pathway genes reduced UA and concretion load. In humans, SNPs in the ILS genes AKT2 and FOXO3 were associated with UA levels or gout, supporting a conserved role for ILS in modulating UA metabolism. Downstream of the ILS pathway UA pathogenicity was mediated partly by NADPH Oxidase, whose inhibition attenuated the reduced lifespan and concretion accumulation. Thus, genes in the ILS pathway represent potential therapeutic targets for treating UA associated pathologies, including gout and kidney stones.\n\nHighlightsO_LIIn Drosophila high uric acid (UA) levels shorten lifespan and cause UA aggregation\nC_LIO_LIConserved in flies and humans, the ILS pathway associates with UA pathologies\nC_LIO_LIFoxO dampens concretion formation by reducing UA levels and ROS formation\nC_LIO_LIInhibition of NOX alleviates the lifespan attenuation and UA aggregation\nC_LI

pathology↗

bioRxiv2018

Untargeted Mass Spectrometry-Based Metabolomics Tracks Molecular Changes in Raw and Processed Foods and Beverages

A major aspect of our daily lives is the need to acquire, store and prepare our food. Storage and preparation can have drastic effects on the compositional chemistry of our foods, but we have a limited understanding of the temporal nature of processes such as storage, spoilage, fermentation and brewing on the chemistry of the foods we eat. Here, we performed a temporal analysis of the chemical changes in foods during common household preparations using untargeted mass spectrometry and novel data analysis approaches. Common treatments of foods such as home fermentation of yogurt, brewing of tea, spoilage of meats and ripening of tomatoes altered the chemical makeup through time, through both chemical and biological processes. For example, brewing tea altered its composition by increasing the diversity of molecules, but this change was halted after 4 min of brewing. The results indicate that this is largely due to differential extraction of the material from the tea and not modification of the molecules during the brewing process. This is in contrast to the preparation of yogurt from milk, spoilage of meat and the ripening of tomatoes where biological transformations directly altered the foods molecular composition. Comprehensive assessment of chemical changes using multivariate statistics showed the varied impacts of the different food treatments, while analysis of individual chemical changes show specific alterations of chemical families in the different food types. The methods developed here represent novel approaches to studying the changes in food chemistry that can reveal global alterations in chemical profiles and specific transformations at the chemical level.\n\nO_LSTHighlightsC_LSTO_LIWe created a reference data set for tomato, milk to yogurt, tea, coffee, turkey and beef.\nC_LIO_LIWe show that normal preparation and handling affects the molecular make-up.\nC_LIO_LITea preparation is largely driven by differential extraction.\nC_LIO_LIFormation of yogurt involves chemical transformations.\nC_LIO_LIThe majority of meat molecules are not altered in 5 days at room temperature.\nC_LI

biochemistry↗

bioRxiv2017

Detecting Drug-Target Binding In Cells Using Fluorescence Activated Cell Sorting Coupled With Mass Spectrometry Analysis

The assessment of drug-target engagement for determining the efficacy of a compound inside cells remains challenging, particularly for difficult target proteins. Existing techniques are more suited to soluble protein targets. Difficult target proteins include those with challenging in vitro solubility, stability or purification properties that preclude target isolation. Here, we report a novel technique that measures intracellular compound-target complex formation, as well as cellular permeability, specificity and cytotoxicity - the Toxicity-Affinity-Permeability-Selectivity (TAPS) technique. The TAPS assay is exemplified here using human kynurenine 3-monooxygenase (KMO), a challenging intracellular membrane protein target of significant current interest. TAPS confirmed target binding of known KMO inhibitors inside cells. We conclude that the TAPS assay can be used to facilitate intracellular hit validation on most, if not all intracellular drug targets.

pharmacology and toxicology↗