Biology subjects

Rashid, M.

Publications and source records attributed to Rashid, M..

At least 19 recordsLinked to original sources

bioRxiv2025

CD33-CD45 Interaction Reveals a Mechanistic Link to Alzheimer's Disease Susceptibility

The innate immune gene CD33, encoding a myeloid inhibitory sialic acid-binding receptor, is associated with Alzheimers disease (AD) susceptibility. The AD-associated rs3865444CC risk variant reduces splicing of the sialic acid-binding domain and increases expression of the full-length (sialic acid-binding) CD33 isoform seven-fold compared to the rs3865444AA protective genotype. Here we identify CD45 as an immune cell-specific sialic acid-dependent cis CD33 binding partner, whose phosphatase activity is inhibited by CD33. Overexpression of CD33 or loss of CD45 contributes to impaired microglial clearance of amyloid beta and amyloid beta-induced loss of dendritic spines in microglial-neuronal co-cultures, aligning with a detrimental effect of CD33-mediated inhibition of CD45. CD33-CD45 interaction frequency was increased in monocytes from individuals with the rs3865444CC risk variant compared to rs3865444AA, as well as in AD compared to controls, independent of genotype. Furthermore, an interaction between CD33 and PTPRC (encoding CD45) gene expression in human brain tissue was associated with a pathological diagnosis of AD and global burden of AD pathology. Our findings thus establish a functional interaction between CD33 and CD45 relevant to AD susceptibility and systemic myeloid dysfunction in this disease.

neuroscience↗

bioRxiv2025

CD33 and Clusterin Interact Biophysically and Genetically to Modulate Alzheimer Risk

We report the results of structural, functional and genetic studies on the CD33 sialic acid- binding receptor that reveal how non-coding variants in CD33 alter risk for Alzheimers disease (AD). The full-length CD33M isoform, whose expression is upregulated by non-coding AD-risk alleles, preferentially forms dimers at the cell surface, where they interact with AD-related proteins (clusterin and A{beta}). This interaction induces CD33M inhibitory signalling and downregulates protective microglial functions including phagocytic removal of amyloid plaques. Human brain expression quantitative trait loci (eQTL) and causal mediation analyses confirm that quantitative interactions between CLU and CD33 genotypes modulate AD phenotypes and suggest that genotypes at these loci might be used to personalise future therapeutic approaches. Our work also highlights several other unexpected aspects of CD33 biology, including a soluble shed extracellular fragment of CD33M and a similar soluble secreted product arising from a truncating mutation in the CD33 extracellular domain (CD33M{Delta}4bp).

neuroscience↗

bioRxiv2025

CloneCoordinate: Open-source software for collaborative DNA construction

Custom DNA constructs have never been more common or important in the life sciences. Many researchers therefore devote substantial time and effort to molecular cloning, aided by abundant computer-aided design tools. However, support for managing and documenting the construction process, and for effectively handling and reducing the frequency of setbacks, is lacking. To address this need, we developed CloneCoordinate, a free, open-source electronic laboratory notebook specifically designed for cloning and fully implemented in Google Sheets. By maintaining a real-time, automatically prioritized task list, a uniform physical sample inventory, and standardized data structures, CloneCoordinate enables productive, collaborative cloning for individuals or teams. We demonstrate how the information captured by CloneCoordinate can be leveraged to troubleshoot assembly problems and provide data-driven insights into cloning efficiency. CloneCoordinate offers a new and uniquely accessible model for how to carry out, and iteratively improve on, real-world DNA assembly.

synthetic biology↗

bioRxiv2025

XVCF: Exquisite Visualization of VCF Data from Genomic Experiments

BackgroundHigh-throughput genomic analyses of germline and cancer genomes facilitate the identification of causal and actionable genetic variants. The recent advances in next-generation sequencing technology generated large-scale genomic and/or multi-omics datasets from different disease types or models. Owing to the huge volume of data coming out of genomic experiments, scientists are facing challenges in handling, manipulating, visualizing, and interpreting the data. Currently, available tools to visualize genetic variants from VCF files are not very user-friendly as most of them require knowledge of command line tools or scripts to install and run those software. Moreover, biologists or clinicians lack this knowledge of computer programming. Therefore, graphical user interface (GUI) based tools or software are needed to effectively summarize and visualize the huge volume of genomic data such as VCF data. MethodsWe have developed a Shiny App, interactive tool using the R programming language that utilizes other R packages like "vcfR" and "maftools" to visualize and generate quality control metrics for genetic data effectively and exquisitely. A key improvement is the addition of a user-friendly interface, providing researchers with an interactive way to explore VCF or Cancer genomics data. Our tool is powered by Shiny, making it even easier for researchers to analyze and visualize genomic variation data using a GUI. Researchers can upload their datasets and customize the analysis parameters to suit their specific research needs. ResultsA user-friendly interactive tool has been developed for the summarization and visualization of data related to genomic variation research. The application features an easy and friendly interface, allowing users to perform various functions such as data loading, summarization, and visualization interactively. XVCF offers an easy-to-use GUI platform to read genetic variant data (annotated or unannotated) and extract useful information such as read depth, mapping quality, genotype, quality control summary, and allele frequency from unannotated data. In the second module of XVCF, the cancer genomic data (annotated, so far supported by ANNOVAR) is analyzed using "maftools" to produce oncoplot, comparison of mutational load across different TCGA datasets, gene summary, etc. XVCF is available for free download from https://github.com/rashidma/XVCF. ConclusionXVCF Shiny web application can serve as a robust visualization and quality control GUI platform for the germline and cancer genomics dataset. We expect this tool will be immensely useful for researchers with less computational or technical knowledge. Being a shiny R package, XVCF can be installed across different operating systems and utilize different computer hardware configurations.

bioinformatics↗

bioRxiv2025

An integrative molecular systems approach unravels mechanisms underlying biphasic nitrate uptake by plant nitrate transporter NRT1.1

Elucidating the mechanisms of transport kinetics in plants is crucial to develop crops that can use nutrients efficiently. The plant nitrate transporter NRT1.1 rapidly switches between high- and low-affinity transport modes to maintain an optimal uptake amidst fluctuations in nitrate levels. This functional switch is regulated by NRT1.1 phosphorylation, but the precise mechanisms remain poorly understood. Here, using an integrated molecular and systems-level modeling, we identify mechanisms underlying biphasic behaviour of NRT1.1. Phosphorylation of NRT1.1 and its binding to nitrate impacts its overall flexibility and synergistically modulates its global conformation, impacting the nitrate transport rate. Integrating these observations with a regulatory network involving kinases CIPK8/CIPK23 and calcium binding proteins CBL1/9, reveals that in high nitrate conditions, CIPK8-mediated sequestration of CBL1 disrupts the CIPK23-CBL complex required for NRT1.1 phosphorylation, switching NRT1.1 to a low-affinity mode. Together, our findings untangle the molecular complexity enabling NRT1.1 phosphorylation switch with broader implications in nitrate sensing and molecular-level adaption to fluctuating external nutrient levels.

systems biology↗

bioRxiv2024

Outer membrane tube formation by Francisella novicida involves extensive envelope modifications and is linked with type VI secretion and alterations to the host phagosomal membrane

Francisella tularensis is a Gram-negative, intracellular pathogen that causes the zoonotic disease tularemia. Due to its ease of dissemination and high lethality, F. tularensis is classified as a Tier 1 Select Agent with potential for misuse as a bioweapon. The mechanisms by which Francisella replicates intracellularly and interacts with the host during infection are not well understood. Francisella produces spherical outer membrane vesicles (OMV) and novel tubular extensions of its cell surface that are also released extracellularly. These OMV and outer membrane tubes (OMT) contain Francisella virulence factors and are produced in response to amino acid starvation and during infection of macrophages. To investigate how the OMT are formed, we used cryogenic electron tomography to examine the model Francisella species, F. novicida, during in vitro culture and within the macrophage phagosome. OMT formation involved progressive alterations of the bacterial envelope, resulting in extensions of both the inner and outer membranes. A dynamic cytoplasmic structure was present at the base of the OMT that extended into the tubes during elongation, together with cytoplasmic material. OMT produced within the macrophage phagosome were associated with changes to the phagosomal membrane, suggesting a role in phagosomal escape. Consistent with this, using confocal microscopy, we observed colocalization of the Francisella type VI secretion system with the OMT, both within bacteria and in released tubular vesicles. These findings reveal the cellular transformations that occur during membrane tubulation by Francisella and provide insights into the function of membrane-derived structures during host-pathogen interactions.

microbiology↗

bioRxiv2024

Decoding the biogenesis of HIV-induced CPSF6 puncta and their fusion with the nuclear speckle

Viruses rely on host cellular machinery for replication. After entering the nucleus, the HIV genome accumulates in nuclear niches where it undergoes reverse transcription and integrates into neighboring chromatin, promoting high transcription rates and new virus progeny. Despite antiretroviral treatment, viral genomes can persist in these nuclear niches and reactivate upon treatment interruption, raising the possibility that they could play a role in the establishment of viral reservoirs. The post-nuclear entry dynamics of HIV remain unclear, and understanding these steps is critical for revealing how viral reservoirs are established. In this study, we elucidate the formation of HIV-induced CPSF6 puncta and the domains of CPSF6 essential for this process. We also explore the roles of nuclear speckle scaffold factors, SON and SRRM2, in the biogenesis of these puncta. Through genetic manipulation and depletion experiments, we demonstrate the key role of the intrinsically disordered region of SRRM2 in enlarging nuclear speckles in the presence of the HIV capsid. We identify the FG domain of CPSF6 as essential for both puncta formation and binding to the viral core, which serves as the scaffold for CPSF6 puncta. While the low-complexity regions (LCRs) modulate CPSF6 binding to the viral capsid, they do not contribute to puncta formation, nor do the disordered mixed charge domains (MCDs) of CPSF6. Interestingly, the FG peptide facilitates viral replication. These results demonstrate how HIV evolved to hijack host nuclear factors, enabling its persistence in the host. Of note, this study provides new insights into the underlying interactions between host factors and viral components, advancing our understanding of HIV nuclear dynamics and offering potential therapeutic targets for preventing viral persistence. HighlightsO_LIThe formation of HIV-induced CPSF6 puncta is critical for restoring HIV-1 nuclear reverse transcription. C_LIO_LICPSF6 protein lacking the FG peptide cannot bind to the viral core, thereby failing to form HIV-induced CPSF6 puncta. C_LIO_LIThe FG peptide, rather than low-complexity regions (LCRs) or the mixed charge domains (MCDs) of the CPSF6 protein, drives the formation of HIV-induced CPSF6 puncta. C_LIO_LIThe presence of the FG peptide plays a pivotal role in facilitating viral replication within macrophage-like cells. C_LIO_LIHIV-induced CPSF6 puncta form individually and later fuse with nuclear speckles (NS) via the intrinsically disordered region (IDR) of SRRM2. C_LI

microbiology↗

bioRxiv2024

Operating principles of interconnected feedback loops underlying cell fate decisions

Interconnected feedback loops are prevalent across biological mechanisms including cell fate transitions enabled by epigenetic mechanisms driving phenotypic plasticity of carcinoma cells. However, the operating principles of these networks remain largely unexplored. Here, we identify numerous coupled feedback loops driving phenotypic transition in cancers and CD4+ T cell lineage decisions. These networks have three generic structures, serial type (ST), hub type (HT), and cyclic, which we discover to be the hallmarks of lower- and higher-order dynamics. While networks having ST or cyclic topology exhibit multiple alternative states, those having HT topology enable at most two states. We also show that topologically distinct networks with equal node or loop count exhibit different steady-state distributions, highlighting the crucial influence of network structure on emergent dynamics. Irrespective of the topology, networks with autoregulated genes exhibit multiple states thereby "liberating" network dynamics from absolute topological control. Finally, we identify precise gene interaction targets to restrict the multistable network dynamics to a unique state. Our results thus reveal design principles of coupled feedback loops in enabling multiple alternative states while also identifying perturbations to restrict it. These findings can serve as crucial inputs to comprehend multi-fate decisions of cells and phenotypic plasticity in carcinomas.

systems biology↗

bioRxiv2024

A ventral hippocampal-lateral septum pathway regulates social novelty preference.

The ability to distinguish strangers from familiar individuals is crucial for the survival of most mammalian species. In humans, an inability to recognize kin and familiar individuals and engage in appropriate behaviors is associated with several types of dementia, including Alzheimers disease. Mice preferentially spend more time investigating a novel individual relative to a familiar individual. Yet, how social novelty related information drives increased investigation of the novel animal remains poorly understood. Recent evidence has implicated the ventral hippocampus (vHPC) as a key node in encoding information about conspecific identity. Of particular interest are vHPC projections to the lateral septum (LS), a region that has been implicated in driving a wide range of motivated social behaviors. In this study using chemogenetics, optogenetics and monosynaptic rabies tracing, we identified a novel vHPC-LS-ventral tegmental area (VTA) pathway that is necessary for mice to preferentially investigate novel conspecifics. Using monosynaptic rabies tracing, we established that LS neurons make direct monosynaptic connections onto dopaminergic neurons in the VTA. Thus, we have identified a potential pathway via which conspecific identity could be transformed to drive motivated social behaviors.

neuroscience↗

bioRxiv2023

Combinatorial cooperativity can facilitate epithelial-mesenchymal transition in a miR200-Zeb feedback network

Carcinoma cells often utilize epithelial-mesenchymal transition (EMT) programs for cancer progression and metastasis. Numerous studies point to the SNAIL-induced miR200/Zeb feedback circuit as crucial in regulating EMT by shifting cancer cells to at least three (epithelial (E), hybrid (h E/M), mesenchymal (M)) phenotypic states arrayed along the epithelial-mesenchymal phenotypic spectrum. However, a coherent molecular-level understanding of how such a tiny circuit controls carcinoma cell entrance into and residence in various states is lacking. Here, we use molecular binding data and mathematical modeling to report that miR200/Zeb circuit can essentially utilize combinatorial cooperativity to control E-M phenotypic plasticity. We identify minimal combinatorial cooperativities that give rise to E, hybrid-E/M, and M phenotypes. We show that disrupting a specific number of miR200 binding sites on Zeb as well as Zeb binding sites on miR200 can have phenotypic consequences - the circuit can dynamically switch between two (E, M) and three (E, hybrid-E/M, M) phenotypes. Further, we report that in both SNAIL-induced and SNAIL knock-out miR200/Zeb circuits, cooperative transcriptional feedback on Zeb as well as Zeb translational inhibition due to miR200 are essential for the occurrence of intermediate hybrid-E/M phenotype. Finally, we demonstrate that SNAIL can be dispensable for EMT, and in the absence of SNAIL, the transcriptional feedback can control cell state transition from E to hybrid-E/M, to M state. Our results thus highlight molecular-level regulation of EMT in miR200/Zeb circuit and we expect these findings to be crucial to future efforts aiming to prevent EMT-facilitated dissemination of carcinomas.

cancer biology↗

bioRxiv2023

RNA- and ATAC-sequencing Reveals a Unique CD83+ Microglial Population Focally Depleted in Parkinson's Disease

All brain areas affected in Parkinsons disease (PD) show an abundance of microglia with an activated morphology together with increased expression of pro-inflammatory cytokines, suggesting that neuroinflammation may contribute to the neurodegenerative process in this common and incurable disorder. We applied a single nucleus RNA- and ATAC-sequencing approach using the 10x Genomics Chromium platform to postmortem PD samples to investigate microglial heterogeneity in PD. We created a multiomic dataset using substantia nigra (SN) tissues from 19 PD donors and 14 non-PD controls (NPCs), as well as three other brain regions from the PD donors which are differentially affected in this disease: the ventral tegmental area (VTA), substantia inominata (SI), and hypothalamus (HypoTs). We identified thirteen microglial subpopulations within these tissues as well as a perivascular macrophage and a monocyte population, of which we characterized the transcriptional and chromatin repertoires. Using this data, we investigated whether these microglial subpopulations have any association with PD and whether they have regional specificity. We uncovered several changes in microglial subpopulations in PD, which appear to parallel the magnitude of neurodegeneration across these four selected brain regions. Specifically, we identified that inflammatory microglia in PD are more prevalent in the SN and differentially express PD-associated markers. Our analysis revealed the depletion of a CD83 and HIF1A-expressing microglial subpopulation, specifically in the SN in PD, that has a unique chromatin signature compared to other microglial subpopulations. Interestingly, this microglial subpopulation has regional specificity to the brainstem in non-disease tissues. Furthermore, it is highly enriched for transcripts of proteins involved in antigen presentation and heat-shock proteins, and its depletion in the PD SN may have implications for neuronal vulnerability in disease.

neuroscience↗

bioRxiv2023

Brain-wide Correspondence Between Neuronal Epigenomics and Long-Distance Projections

Single-cell genetic and epigenetic analyses parse the brains billions of neurons into thousands of "cell-type" clusters, each residing in different brain structures. Many of these cell types mediate their unique functions by virtue of targeted long-distance axonal projections to allow interactions between specific cell types. Here we have used Epi-Retro-Seq to link single cell epigenomes and associated cell types to their long-distance projections for 33,034 neurons dissected from 32 different source regions projecting to 24 different targets (225 source [->]target combinations) across the whole mouse brain. We highlight uses of this large data set for interrogating both overarching principles relating projection cell types to their transcriptomic and epigenomic properties and for addressing and developing specific hypotheses about cell types and connections as they relate to genetics. We provide an overall synthesis of the data set with 926 statistical comparisons of the discriminability of neurons projecting to each target for every dissected source region. We integrate this dataset into the larger, annotated BICCN cell type atlas composed of millions of neurons to link projection cell types to consensus clusters. Integration with spatial transcriptomic data further assigns projection-enriched clusters to much smaller source regions than afforded by the original dissections. We exemplify these capabilities by presenting in-depth analyses of neurons with identified projections from the hypothalamus, thalamus, hindbrain, amygdala, and midbrain to provide new insights into the properties of those cell types, including differentially expressed genes, their associated cis-regulatory elements and transcription factor binding motifs, and neurotransmitter usage.

neuroscience↗

bioRxiv2023

Sex differences in neural representations of social and nonsocial reward in the medial prefrontal cortex.

The reinforcing nature of social interactions is necessary for the maintenance of appropriate social behavior. However, the neural substrates underlying social reward processing and how they might differ based on the sex and internal state of the animal remains unknown. It is also unclear whether these neural substrates are shared with those involved in nonsocial rewarding processing. We developed a fully automated, novel two choice (social-sucrose) operant assay in which mice choose between social and nonsocial rewards to directly compare the reward-related behaviors associated with two competing stimuli. We performed cellular resolution calcium imaging of medial prefrontal cortex (mPFC) neurons in male and female mice across varying states of water restriction and social isolation. We found that mPFC neurons maintain largely non-overlapping, flexible representations of social and nonsocial reward that vary with internal state in a sex-dependent manner. Additionally, optogenetic manipulation of mPFC activity during the reward period of the assay disrupted reward-seeking behavior across male and female mice. Thus, using a novel operant assay, we have identified sex-dependent, non-overlapping neural representations of social and nonsocial reward in the mPFC that vary with internal state and that are essential for appropriate reward-seeking behavior.

neuroscience↗

bioRxiv2022

Network Topology Metrics Explaining Enrichment of Hybrid Epithelial Mesenchymal Phenotypes in Metastasis

Epithelial to mesenchymal transition (EMT) and its reverse mesenchymal to epithelial transition (MET) are hallmarks of metastasis. Cancer cells use this reversible cellular programming to switch among Epithelial (E), Mesenchymal (M), and hybrid Epithelial/Mesenchymal (hybrid E/M) state(s) and seed tumors at distant sites. Hybrid E/M cells are often more aggressive and metastatic than the "pure" E and M cells. Thus, identifying mechanisms to inhibit hybrid E/M cells can be promising in curtailing metastasis. While multiple gene regulatory networks (GRNs) based mathematical models for EMT/MET have been developed recently, identifying topological signatures enriching for hybrid E/M phenotypes remains to be done. Here, we investigate the dynamics of 13 different GRNs and identify which network topologies can enrich for hybrid E/M phenotype(s) across GRNs. We found that increasing negative feedback loops enhances the "hybridness" of these networks, but increasing the number of positive feedback loops or their specific combinations can decrease the frequency of hybrid E/M phenotype. Thus, our analysis identifies network topology based signatures that can give rise to, as well as prevent, the emergence of hybrid E/M phenotype in GRNs underlying EMP. Our results can have implications in terms of targeting specific interactions in GRNs as a potent way to restrict switching to the hybrid E/M phenotype(s) to curtail metastasis.

systems biology↗

bioRxiv2021

Differential expression of H2A isoforms contribute to tissue and lineage specificity with HIST2H2AC as a potential cancer biomarker

Recent advancements in the field of histone biology imply non-redundancy in the function of histone H2A isoforms; however, the expression of H2A isoforms in various normal tissue types, the correlation among organs and tumor/tumor type-specific expression remain poorly investigated. The profiling of sixteen H2A isoforms in eleven different normal human tissue types strongly suggests their tissue-specific or predominant expression. Further, clustering analysis shows a lineage-specific correlation of H2A isoforms. In continuation, the expression analysis in twelve human tumor types shows overexpression of HIST2H2AC. Moreover, overexpression was observed exclusively in tumor samples but not with fetal samples; highlighting the cancer-specific association of HIST2H2AC. Further, in silico analysis of TCGA pan-cancer data also showed tumor-specific over-expression of the HIST2H2AC isoform. Our findings provide insights into tissue-type-specificity of histone H2A isoforms expression patterns and advance our understanding of their importance in lineage specification and cancer.

cancer biology↗

bioRxiv2021

Surveillance of genetic diversity and evolution in locally transmitted SARS-CoV-2 in Pakistan during the first wave of the COVID-19 pandemic

Surveillance of genetic diversity in the SARS-CoV-2 is extremely important to detect the emergence of more infectious and deadly strains of the virus. In this study, we monitored mutational events in the SARS-CoV-2 genome through whole genome sequencing. The samples (n=48) were collected from the hot spot regions of the metropolitan city Karachi, Pakistan during the four months (May 2020 to August 2020) of first wave of the COVID-19 pandemic. The data analysis highlighted 122 mutations, including 120 single nucleotide variations (SNV), and 2 deletions. Among the 122 mutations, there were 71 singletons, and 51 recurrent mutations. A total of 16 mutations, including 5 nonsynonymous mutations, were detected in spike protein. Notably, the spike protein missense mutation D614G was observed in 31 genomes. The phylogenetic analysis revealed majority of the genomes (36) classified as B lineage, where 2 genomes were from B.6 lineage, 5 genomes from B.1 ancestral lineage and remaining from B.1 sub-lineages. It was noteworthy that three clusters of B.1 sub-lineages were observed, including B.1.36 lineage (10 genomes), B.1.160 lineage (11 genomes), and B.1.255 lineage (5 genomes), which represent independent events of SARS-CoV-2 transmission within the city. The sub-lineage B.1.36 had higher representation from the Asian countries and the UK, B.1.160 correspond to the European countries with highest representation from the UK, Denmark, and lesser representation from India, Saudi Arabia, France and Switzerland, and the third sub-lineage (B.1.255) correspond to the USA. Collectively, our study provides meaningful insight into the evolution of SARS-CoV-2 lineages in spatio-temporal local transmission during the first wave of the pandemic.

genomics↗

bioRxiv2021

Unfolding the Apoptotic Mechanism of Antioxidant Enriched-Leaves of Tabebuia pallida in EAC Cells

Targeting apoptosis is a promising approach to inhibit the abnormal cell proliferation of cancer progression. Existing anti-apoptotic drugs, many derived from chemical substances, have often failed to combat cancer development and progression. Therefore, identification of apoptosis-inducing anticancer agents from plant-derived sources has become a key aim in cancer research. The present study was designed to explore the regulation of apoptosis by Tabebuia pallida (T. pallida) using an Ehrlich Ascites Carcinoma (EAC) mouse model and compositional analysis by LC-ESI-MS/MS. Dried and powdered T. pallida leaves (TPL), stem bark (TPSB), root bark (TPRB) and flowers (TPF) were extracted with 80% methanol. Using cultured EAC cells and EAC-bearing mice with and without these extracts, anticancer activities were studied by assessing cytotoxicity and tumor cell growth inhibition, changes in life span of mice, and hematological and biochemical parameters. Apoptosis was analyzed by microscopy and expression of selected apoptosis-related genes (Bcl-2, Bcl-xL, NF{kappa}-B, PARP-1, p53, Bax, caspase-3 and -8) using RT-PCR. LC-ESI-MS analysis was performed to identify the major compounds from the most active extracts. In EAC mice compared with untreated controls, the TPL extract exhibited the highest cytotoxicity with significant tumor cell growth inhibition (p< 0.001), reduced ascites by body weight (p< 0.01), increased the life span (p<0.001), normalized blood parameters (RBC/WBC counts), and increased the levels of superoxide dismutase and catalase. TPL-treated EAC cells showed apoptotic characteristics of membrane blebbing, chromatin condensation and nuclear fragmentation, and caspase-3 activation, compared with untreated EAC cells. Moreover, annexin V-FITC and propidium iodide signals were greatly enhanced in response to TPL treatment, indicating apoptosis induction. Pro- and anti-apoptotic signaling after TPL treatment demonstrated up-regulated p53, Bax and PARP-1, and down-regulated NF{kappa}-B, Bcl-2 and Bcl-xL expression, suggesting that TPL shifts the balance of pro- and anti-apoptotic genes towards cell death. LC-ESI-MS data of TPL showed a mixture of glycosides, lapachol, and quercetin antioxidant and its derivatives that were significantly linked to cancer cell targets. In conclusion, the TPL extract of T. pallida possesses significant anticancer activity. The tumor suppressive mechanism is due to apoptosis induced by activation of antioxidant enzymes and caspases and mediated by a change in the balance of pro- and anti-apoptotic genes that promotes cell death. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/426226v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@1112faaorg.highwire.dtl.DTLVardef@1505d1dorg.highwire.dtl.DTLVardef@1115a2aorg.highwire.dtl.DTLVardef@36a915_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2020

Facial expressions of emotional stress in horses

Horses have the ability to generate a remarkable repertoire of facial expressions, some which have been linked to certain emotional states, for example pain. Studies suggest that facial expressions may be a more honest expression of emotional state in horses than behavioral or physiological parameters. This study sought to describe the facial expressions during stress of healthy horses free of pain, using a standardized method of recording facial expressions in video. Stress was induced in 28 horses by subjecting them to road transport and 10 of these horses were also subjected to social isolation. The horses served as their own control. A body-mounted, remote controlled heart rate monitor provided continuous heart rate measurements during the interventions. The horses facial expressions were video-recorded during the interventions. Frequency and duration of each facial expression were then determined, according to the Equine Facial Action Coding System. Heart rate increased during the stressful interventions (p=0.01), confirming that the interventions were stressful. Using both the human investigation- and the co-occurrence methods, the following facial traits could be observed during stress: eye white increase (p<0.001), nostril dilator (p<0.001), upper eyelid raiser (p<0.001), inner brow raiser (p=0.042), tongue show (p<0.001) along with an increase in ear flicker (p<0.001) and blink frequency (p<0.001). The facial actions were successfully used to train a machine-learning classifier to discriminate between stressed and calm horses, with an accuracy of 74.2 %. Most of the facial features identified correspond well with previous research on the subject, for example flared nostrils, repetitive mouth behaviors, increased eye white, tongue show and ear movements. Some features selected as indicative of emotional pain-free stress are used in face-based pain assessment tools, such as dilated nostrils, eye white increase or inner brow raiser. The relation between facial expressions of stress and pain should therefore further be studied.

animal behavior and cognition↗