Find research datasets worth reusing

Reactivating PTEN to impair glioma stem cells by inhibiting cytosolic iron-sulfur assembly pathway

Glioblastoma (GBM), the most lethal primary brain tumor, harbors glioma stem cells (GSCs) that not only initiate and maintain malignant phenotypes but also enhance therapeutic resistance. Although frequently mutated in GBMs, the function and regulation of PTEN in PTEN-intact GSCs are unknown. Here we found that PTEN directly interacts with MMS19 and competitively disrupts MMS19-based cytosolic iron-sulfur (Fe-S) cluster assembly (CIA) machinery in the differentiated glioma cells (DGCs). Interrogation of GSCs, when compared with their matched DGCs, revealed that PTEN is specifically succinated at cysteine (C) 211 in GSCs. Isotope tracing coupled with mass spectrometry analysis confirmed that fumarate, generated by adenylosuccinate lyase (ADSL) in de novo purine synthesis pathway which is highly activated in GSCs, promotes PTEN C211 succination. This modification abrogates the interaction between PTEN and MMS19, thereby reactivating CIA machinery pathway in GSCs. Functionally, inhibiting PTEN C211 succination through re-expressing PTEN C211S mutant, depleting ADSL, or consuming fumarate by N-acetylcysteine (NAC), an FDA-approved prescription drug, impairs GSC maintenance. Importantly, re-expressing PTEN C211S or treating with NAC sensitizes GSC-derived brain tumors to temozolomide and irradiation, the standard-of-care treatments for GBM patients, by retarding CIA machinery-mediated DNA damage repair. These findings reveal an immediately practicable strategy to target GSCs for treating GBMs by combined therapy with repurposing NAC.

Data from: LHFPL5 is a key element to the gating spring of cochlear hair cells: comparison of mechanotransduction channel activation in presence or absence of LHFPL5

During auditory transduction, sound-evoked vibrations of the hair cell stereociliary bundles open mechanotransducer (MET) ion channels via tip links extending from one stereocilium to its neighbor. How tension in the tip link is delivered to the channel is not fully understood. The MET channel comprises a pore-forming subunit, transmembrane channel-like protein (TMC1 or TMC2), aided by several accessory proteins, including LHFPL5 (lipoma HMGIC fusion partner-like 5). We investigated the role of LHFPL5 in transduction by comparing MET channel activation in outer hair cells of Lhfpl5-/- knockout mice with those in Lhfpl5+/- heterozygotes. The 10-90 percent working range of transduction in Tmc1+/+; Lhfpl5+/- was 52 nm, from which the single-channel gating force, Z was evaluated as 0.34 pN. However, in Tmc1+/+; Lhfpl5‑/- mice, the working range increased to 123 nm and Z more than halved to 0.13 pN, indicating reduced sensitivity. Tip link tension is thought to activate the channel via a gating spring, whose stiffness is inferred from the stiffness change on tip link destruction. The gating stiffness was ~40 percent of the total bundle stiffness in wild-type but was virtually abolished in Lhfpl5-/-, implicating LHFPL5 as a principal component of the gating spring. The mutation Tmc1 p.D569N reduced the LHFPL5 immunolabeling in the stereocilia and like Lhfpl5-/- doubled the MET working range but other deafness mutations had no effect on the dynamic range. We conclude that tip-link tension is transmitted to the channel primarily via LHFPL5; residual activation without LHFPL5 may occur by direct interaction between PCDH15 and TMC1.

opencc-by-4.0Dec 2023View details →

Dataset and Code for Manuscript "Cell sorting based on pulse shapes from angle resolved detection of scattered light"

Dataset and code for the cell cycle analysis and cluster selection for sorting: <ul> <li>ReadMe file with explanations of the data set and analysis</li> <li>Python scripts for converting the data and to reproduce the sort cluster selection</li> <li>binary data files containing pulse shapes and wavelet transform coefficients</li> <li>FSC data files containing the respective common flow cytometry parameters</li> <li>text files with event indices that represent the gating</li> </ul>

Data from: anterior pituitary transcriptomics following a high fat diet: impact of oxidative stress on cell metabolism

Anterior pituitary cells are highly active with regards to protein synthesis and secretion, processes which depend heavily on mitochondrial ATP production and functional endoplasmic reticula. It is well known that obesity adds an allostatic overload to tissues, requiring them to adapt to inflammation and oxidative stress. Therefore, we hypothesized that the pituitary is highly vulnerable to the stress of high fat diet-induced weight gain. In this study, we utilized a 10-15 week high fat diet (HFD, 60%) plus a thermoneutral housing paradigm, testing both male and female FVB.129P mice. We quantified serum hormones and cytokines, characterized the metabolic phenotype, and defined changes in the pituitary transcriptome using single-cell RNA-seq. Weight gain was significant by 3 weeks in HFD mice, and by 10 weeks all HFD groups had gained 20 g. HFD females (15 weeks) had increased energy expenditure and decreased activity. All HFD groups showed increases in serum leptin, Il-6, resistin, MCP-1, and TNFα. HFD males had increased insulin; both HFD males and females had increased TSH, and HFD females had decreased serum prolactin and growth hormone (GH) pulse amplitude. Pituitary scRNA-seq revealed modest or no changes in pituitary cell gene expression in the different cell types from HFD males after 10 or 15 weeks or HFD females after 10 weeks. However, females exposed to a HFD for 15 weeks showed significant numbers of differentially expressed genes in lactotropes and stem cells. Pathway analyses identified a reduction in pathways that supported protein translation, ribosome biogenesis, and oxidative phosphorylation, indicating mitochondrial dysfunction. Collectively, these studies reveal that pituitary cells from males are more resilient to the oxidative stress of obesity than females and identify the most vulnerable pituitary cell populations in females.

Data from: Illuminating T cell-dendritic cell interactions in vivo by FlAsHing antigens

Delineating the complex network of interactions between antigen-specific T cells and antigen presenting cells (APCs) is crucial for effective precision therapies against cancer, chronic infections, and autoimmunity. However, the existing arsenal for examining antigen-specific T cell interactions is restricted to a select few antigen-T cell receptor pairs, with limited in situ utility. This lack of versatility is largely due to the disruptive effects of reagents on the immune synapse, which hinder real-time monitoring of antigen-specific interactions. To address this limitation, we have developed a novel and versatile immune monitoring strategy by adding a short cysteine-rich tag to antigenic peptides that emits fluorescence upon binding to thiol-reactive biarsenical hairpin compounds. Our findings demonstrate the specificity and durability of the novel antigen-targeting probes during dynamic immune monitoring in vitro and in vivo. This strategy opens new avenues for biological validation of T-cell receptors with newly identified epitopes by revealing the behavior of previously unrecognized antigen-receptor pairs, expanding our understanding of T cell responses.

Induced pluripotent stem cell-derived cardiomyocyte in vitro models: tissue fabrication protocols, assessment methods, and quantitative maturation metrics for benchmarking progress

The advent of human induced pluripotent stem cells (hiPSCs) and techniques to differentiate cardiomyocytes from them has opened a viable path to creating in vitro models of normal and diseased hearts, accelerating more predictive drug screening and therapeutic strategies for cardiac pathologies. Currently, hiPSC-derived cardiomyocytes (hiPSC-CMs) are more similar to fetal than adult cardiomyocytes, leading many in the field to explore approaches to enhance cell and tissue maturation. There are over 2,000 studies utilizing hiPSC-CMs in models composed of various combinations of cell and extracellular matrix components, using a plethora of differentiation protocols, culture formats, and methods for quantifying cardiomyocyte function. To assess the current state of this rapidly growing area, we systematically analyzed 300 studies using hiPSC-CM models for their selection of hiPSC lines, hiPSC-CM differentiation protocols, types of in vitro models, maturation techniques, and metrics used to assess cardiomyocyte functionality and maturity. Here, we provide the data compiled from our analysis of these papers so others in the field can utilize it to inform their research. Based on this analysis, we highlight the diversity of, and current trends in, in vitro model designs and highlight the most common and promising practices for functional assessments. We further analyzed outputs spanning structural maturity, contractile function, electrophysiology, and gene expression and note field-wide improvements over time. Finally, we observe that a persistent lack of coordination amongst investigators is limiting the field's ability to benchmark and advance hiPSC-CM function against previous studies. We discuss opportunities to collectively pursue the common goal of hiPSC-CM model development, maturation, and assessment that we believe are critical to drive the entire community forward in engineering mature cardiac tissue.

The potassium channel subunit Kv1.8 (Kcna10) is essential for the distinctive outwardly rectifying conductances of type I and II vestibular hair cells

In amniotes, head motions and tilt are detected by two types of vestibular hair cells (HCs) with strikingly different morphology and physiology. Mature type I HCs express a large and very unusual potassium conductance, gK,L, which activates negative to resting potential, confers very negative resting potentials and low input resistances, and enhances an unusual non-quantal transmission from type I cells onto their calyceal afferent terminals. Following clues pointing to KV1.8 (KCNA10) in the Shaker K channel family as a candidate gK,L subunit, we compared whole-cell voltage-dependent currents from utricular hair cells of KV1.8-null mice and littermate controls. We found that KV1.8 is necessary not just for gK,L but also for fast-inactivating and delayed rectifier currents in type II HCs, which activate positive to resting potential. The distinct properties of the three KV1.8-dependent conductances may reflect different mixing with other KV1 subunits, such as KV1.4 (KCNA4). In KV1.8-null HCs of both types, residual outwardly rectifying conductances include KV7 (KCNQ) channels. Current clamp records show that in both HC types, KV1.8-dependent conductances increase the speed and damping of voltage responses. Features that speed up vestibular receptor potentials and non-quantal afferent transmission may have helped stabilize locomotion as tetrapods moved from water to land.

N-cadherin dynamically regulates pediatric glioma cell migration in complex environments

Pediatric high-grade gliomas are highly invasive and essentially incurable. Glioma cells migrate between neurons and glia, along axon tracts, and through extracellular matrix surrounding blood vessels and underlying the pia. Mechanisms that allow adaptation to such complex environments are poorly understood. N-cadherin is highly expressed in pediatric gliomas and is associated with shorter survival. We found that inter-cellular homotypic N-cadherin interactions differentially regulate glioma migration according to the microenvironment, stimulating migration on cultured neurons or astrocytes but inhibiting invasion into reconstituted or astrocyte-deposited extracellular matrix. N-cadherin localizes to filamentous connections between migrating leader cells but to epithelial-like junctions between followers. Leader cells have more surface and recycling N-cadherin, increased YAP1/TAZ signaling, and increased proliferation relative to followers. YAP1/TAZ signaling is dynamically regulated as leaders and followers change position, leading to altered N-cadherin levels and organization. Together, the results suggest that pediatric glioma cells adapt to different microenvironments by regulating N-cadherin dynamics and cell-cell contacts.

Defining the role of the miR-145 – KLF4 – αSMA axis in mitral valvular interstitial cell activation in myxomatous mitral valve prolapse using the canine model

Mitral valve prolapse (MVP) is a common valvular disease, affecting 2-3% of the adult human population and is a degenerative condition. 5-10% of the afflicted will develop severe mitral regurgitation, cardiac dysfunction, congestive heart failure, and sudden cardiac death. Naturally occurring myxomatous MVP in dogs closely resembles MVP in humans structurally, and functional consequences are similar. In both species, valvular interstitial cells (VICs) in affected valves exhibit phenotype consistent with activated myofibroblasts with increased αSMA expression. Using VICs collected from normal and MVP-affected valves of dogs, we analyzed the miRNA expression profile of the cells and their associated small extracellular vesicles (sEV) using RNA sequencing to understand the role of non-coding RNAs and sEV in MVP pathogenesis. miR-145 was shown to be upregulated in both the affected VICs and sEV, and overexpression of miR-145 by mimic transfection in quiescent VIC recapitulates the activated myofibroblastic phenotype. Concurrently, KLF4 expression was suppressed by miR-145, and KLF4 overexpression resulted in a decrease in αSMA expression, and its suppression resulted in an increase in αSMA expression, thus confirming the miR-145 – KLF4 – αSMA axis. Targeting this axis may serve as potential therapy in controlling pathologic abnormalities found in MVP valves.

Generation of KRAS knockout pancreatic cancer cell line PANC1

We used CRISPR to inactivate mutant KRAS and STAT3 in PANC1 (KRASG12D) pancreatic cancer cell line. Gene expression analysis of KRAS intact vs. knockout cells identified sets of genes involved in protein synthesis, cell differentiation, and metabolic processes, while the expression of MAPK/ERK target genes remained unperturbed.

Molecular dynamics dataset for pharmacological repositioning in the treatment of non-small-cell lung cancer

Non-small cell lung cancer (NSCLC) is a type of lung cancer associated with translocation of the EML4 and ALK genes on the short arm of chromosome 2. This leads to the development of an aberrant protein kinase with a deregulated catalytic domain, the cdALK+. Currently, different ALK inhibitors (iALKs) have been proposed to treat ALK+ NSCLC patients. However, the recent resistance to iALKs stimulates the exploration of new iALKs for NSCLC. Here, we describe an in silico approach to finding FDA-approved drugs that can be used by pharmacological repositioning as iALK. We used homology modelling to obtain a structural model of cdALK+ protein and then performed molecular docking and molecular dynamics of the complex cdALK+-iALKs to generate the pharmacophore model. The pharmacophore was used to identify potential iALKs from FDA-approved drugs library by ligand-based virtual screening. Four pharmacophores with different atomistic characteristics were generated, resulting in six drugs that satisfied the proposed atomistic positions and coupled at the ATP-binding site. Mitoxantrone, riboflavin and abacavir exhibit the best interaction energies with 228.29, 165.40 and 133.48 kjoul/mol respectively. In addition, the special literature proposed these drugs for other types of diseases due to pharmacological repositioning. This study proposes FDA-approved drugs with ALK inhibitory characteristics. Moreover, we identified pharmacophores sites that can be tested with other pharmacological libraries.

opencc-by-4.0Aug 2023View details →

Single-cell RNA sequencing reveals dysregulated cellular programs in the inflamed epithelium of Crohn's disease patients.

Crohn’s disease (CD) is a complex inflammatory disorder of incompletely understood molecular aetiology. We generated a large single-cell RNA sequencing dataset from the terminal ileal biopsies of two independent cohorts comprising a total of 50 CD patients and 71 healthy controls. We performed transcriptomic analyses to reveal genes, cell types and mechanisms perturbed in CD, leveraging the power of the two cohorts to confirm our findings and assess replicability. In addition to mapping widespread alterations in cytokine signalling, we provide evidence of pan-epithelial upregulation of MHC class I genes and pathways in CD. Using non-negative matrix factorization we revealed intra- and inter-cellular upregulation of expression programs such as G-protein coupled receptor signalling and interferon signalling, respectively, in CD. We observed an enrichment of CD heritability among marker genes for various activated T cell types and myeloid cells, supporting a causal role for these cell-types in CD aetiology. Comparisons between our discovery and replication cohort revealed significant variation in differential gene-expression replicability across cell types. B, T and myeloid cells showed particularly poor replicability, suggesting caution should be exercised when interpreting unreplicated differential gene-expression result in these cell types. Overall, our results provide a rich resource for identifying cell-type specific biomarkers of Crohn’s disease and identifying genes, cell types and pathways that are causally and replicably associated with disease.

opencc-by-4.0Jan 2024View details →

Spotiphy enables single-cell spatial whole transcriptomics across the entire section

Spatial transcriptomics (ST) has advanced our understanding of tissue regionalization by enabling the visualization of gene expression within whole tissue sections, but the approach remains dogged by the challenge of achieving single-cell resolution without sacrificing whole genome coverage. Here we present Spotiphy (Spot imager with pseudo single-cell resolution histology), a novel computational toolkit that transforms sequencing-based ST data into single-cell-resolved whole-transcriptome images. In evaluations with Alzheimer’s disease (AD) and normal mouse brains, Spotiphy delivers the most precise cellular compositions. For the first time, Spotiphy reveals novel astrocyte regional specification in mouse brains. It distinguishes sub-populations of DAM (Disease-Associated Microglia) located in different AD mouse brain regions. Spotiphy also identifies multiple spatial domains as well as changes in the patterns of tumor-tumor microenvironment interactions using human breast ST data. Spotiphy enables visualization of cell localization and gene expression in tissue sections, offering key insights into the function of complex biological systems.

Proteome of peripheral mononuclear cells (PBMCs) from asymptomatic malaria and uninfected individuals and the ensuing malaria episodes

Cumulative malaria parasite exposure in endemic regions often results in the acquisition of partial immunity and asymptomatic infections. There is limited information on how host-parasite interactions mediate maintenance of chronic symptomless infections that sustain malaria transmission. Here, we have determined the gene expression profiles of the parasite population and the corresponding host peripheral blood mononuclear cells (PBMCs) from 21 children (<15 years). We compared children who were defined as uninfected, asymptomatic and those with febrile malaria. Children with asymptomatic infections had a parasite transcriptional profile characterized by a bias toward trophozoite stage (~12 hours-post invasion) parasites and low parasite levels, while earlier ring stage parasites were characteristic of febrile malaria. The host response of asymptomatic children was characterized by downregulated transcription of genes associated with inflammatory responses, compared to children with febrile malaria. Interestingly, the host responses during febrile infections that followed an asymptomatic infection featured stronger inflammatory responses, whereas the febrile host responses from previously uninfected children featured increased humoral immune responses. The priming effect of prior asymptomatic infection may explain the blunted acquisition of antibody responses seen to malaria antigens following natural exposure or vaccination in malaria endemic areas.

Extraocular muscle stem cells exhibit distinct cellular properties associated with non-muscle molecular signatures

The muscle stem cell (MuSC) population is recognized as functionally heterogeneous. Cranial muscle stem cells, which originate from head mesoderm, can have greater proliferative capacity in culture and higher regenerative potential in transplantation assays when compared to those in the limb. The existence of such functional differences in phenotypic outputs remain unresolved as a comprehensive understanding of the underlying mechanisms is lacking. We addressed this issue using a combination of clonal analysis, live imaging, and scRNA-seq, identifying critical biological features that distinguish extraocular (EOM) and limb (Tibialis anterior, TA) MuSC populations. Time-lapse studies using a MyogenintdTomato reporter showed that the increased proliferation capacity of EOM MuSCs is accompanied by a differentiation delay in vitro. Unexpectedly, in vitro activated EOM MuSCs expressed a large array of distinct extracellular matrix (ECM) components, growth factors, and signaling molecules that are typically associated with mesenchymal non-muscle cells. These unique features are regulated by a specific set of transcription factors that constitute a coregulating module. This transcription factor network, which includes Foxc1 as one of the major players, appears to be hardwired to EOM identity as it is present in quiescent adult MuSCs, in the activated counterparts during growth and retained upon passages in vitro. These findings provide insights into how high-performing MuSCs regulate myogenic commitment by active remodeling of their local environment.

Affected cell types for hundreds of Mendelian diseases revealed by analysis of human and mouse single-cell data

Hereditary diseases manifest clinically in certain tissues, however their affected cell types typically remain elusive. Single-cell expression studies showed that overexpression of disease-associated genes may point to the affected cell types. Here, we developed a method that infers disease-affected cell types from the preferential expression of disease-associated genes in cell types (PrEDiCT). We applied PrEDiCT to single-cell expression data of six human tissues, to infer the cell types affected in 1,459 hereditary diseases. Overall, we identified 114 cell types affected by 1,140 diseases. We corroborated our findings by literature text-mining and recapitulation in mouse corresponding tissues. Based on these findings, we explored features of disease-affected cell types and cell classes, highlighted cell types affected by mitochondrial diseases and heritable cancers, and identified diseases that perturb intercellular communication. This study expands our understanding of disease mechanisms and cellular vulnerability.

Supporting information for: Discrimination ability of central visual field testing using stimulus size I, II, and III and relationship with macular ganglion cell thickness in chiasmal compression

Purpose: To compare the relationship between macular ganglion cell layer (mGCL) thickness and 10-2 visual field (VF) sensitivity using different stimulus sizes in patients with temporal hemianopia from chiasmal compression. Methods: A cross-sectional study was conducted involving 30 eyes from 25 patients with temporal VF loss on 24-2 SITA standard automated perimetry due to previous chiasmal compression and 30 healthy eyes (23 controls). Optical coherence tomography (OCT) of the macular area and 10-2 VF testing using Goldmann stimulus size I (GI), II (GII), and III (GIII) were performed in the Octopus 900 perimeter. For the sake of analysis, mGCL thickness and VF data were segregated into four quadrants (two temporal and two nasal) and two halves (temporal and nasal) centered on the fovea, and the groups were compared using generalized estimated equations. The discrimination ability of GI, GII, and GIII was evaluated, as was the correlation between mGCL and 10-2 VF sensitivity using GI, GII, and GIII. Results: All mGCL parameters were significantly reduced in patients compared to controls. 10-2 VF test sensitivity using GI, GII, and GIII was significantly lower in patients than in controls (p≤0.008) for all parameters, except the three nasal divisions when using GI (p=0.41, 0.07 and 0.18) Significant correlations were found between temporal VF sectors (all stimulus sizes) and the corresponding nasal mGCL measurements, with similar discrimination ability. Significant correlations were also observed between all three nasal VF divisions and the corresponding temporal mGCL thickness when using stimulus sizes I and II, but not stimulus size III. Conclusions: On 10-2 VF testing, GII outperformed GI and GIII with regard to discrimination ability and structure-function correlation with mGCL thickness in chiasmal compression. Our findings suggest that the use of GII can enhance the diagnostic power of 10-2 VF testing, although further studies are necessary to support this conclusion.

opencc-zeroFeb 2024View details →

Data from: Value-related learning in the olfactory bulb occurs through pathway-dependent peri-somatic inhibition of mitral cells

Associating values to environmental cues is a critical aspect of learning from experiences, allowing animals to predict and maximise future rewards. Value-related signals in the brain were once considered a property of higher sensory regions, but their wide distribution across many brain regions is increasingly recognised. Here, we investigate how reward-related signals begin to be incorporated, mechanistically, at the earliest stage of olfactory processing, namely, in the olfactory bulb. In head-fixed mice performing Go/No-Go discrimination of closely related olfactory mixtures, rewarded odours evoke widespread inhibition in one class of output neurons, that is, in mitral cells but not tufted cells. The temporal characteristics of this reward-related inhibition suggest it is odour-driven, but it is also context-dependent since it is absent during pseudo-conditioning and pharmacological silencing of the piriform cortex. Further, the reward-related modulation is present in the somata but not in the apical dendritic tuft of mitral cells, suggesting an involvement of circuit component located deep in the olfactory bulb. Depth-resolved imaging from granule cell dendritic gemmules suggests that granule cells that target mitral cells receive a reward-related extrinsic drive. Thus, our study supports the notion that value-related modulation of olfactory signals is a characteristic of olfactory processing in the primary olfactory area and narrows down the possible underlying mechanisms to deeper circuit components that contact mitral cells peri-somatically.

opencc-zeroFeb 2024View details →

Integrin restriction by miR-34 protects germline progenitors from cell death during aging

During aging, regenerative tissues must dynamically balance the two opposing processes of proliferation and cell death. While many microRNAs are differentially expressed during aging, their roles as dynamic regulators of tissue regeneration have yet to be described. We show that in the highly regenerative Drosophila testis, miR-34 levels are significantly elevated during aging. miR-34 modulates germ cell death and protects the progenitor germ cells from accelerated aging. However, miR-34 is not expressed in the progenitors themselves but rather in neighboring cyst cells that kill the progenitors. Transcriptomics followed by functional analysis revealed that during aging, miR-34 modifies integrin signaling by limiting the levels of the heterodimeric integrin receptor αPS2 and βPS subunits. In addition, we found that in cyst cells, this heterodimer is essential for inducing phagoptosis and degradation of the progenitor germ cells. Together, these data suggest that the miR-34 – integrin signaling axis acts as a sensor of progenitor germ cell death to extend progenitor functionality during aging.