5683099 research outputs found
Sort by
A lyophilized open-source RT-LAMP assay for molecular diagnostics in resource-limited settings.
No full textA critical bottleneck for equitable access to population-scale molecular diagnostics is the limited availability of rapid, inexpensive point-of-care tests, especially in low- and middle-income countries. Here, we developed an open-source reverse transcription loop-mediated isothermal amplification (RT-LAMP) molecular assay for pathogen detection. It is based on nonproprietary enzymes, namely, HIV-1 reverse transcriptase, Bst LF DNA polymerase, and UDG BMTU thermolabile uracil-DNA glycosylase. Formulated as liquid or lyophilized reaction mixtures, these reagents enable sensitive colorimetric detection of respiratory samples without the need for prior nucleic acid isolation. We evaluated our lyophilized RT-LAMP assay on clinical samples with suspected COVID-19 infection, demonstrating high sensitivity and 100% specificity compared with the gold-standard RT-qPCR. Reaction performance was unaffected by prolonged storage of lyophilized reagents at ambient or elevated temperatures. As a proof of concept, we evaluated the robustness and ease of use of lyophilized RT-LAMP reaction mixes through independent laboratory testing of COVID-19 samples in Ghana. Overall, our open-source RT-LAMP assay provides a flexible and scalable point-of-care test that can be adapted for rapid detection of various pathogens in resource-limited settings.</p
EXO1 as a therapeutic target for Fanconi Anaemia, ZRSR2 and BRCA1-A complex deficient cancers.
No full textExonuclease EXO1 performs multiple roles in DNA replication and DNA damage repair (DDR). However, EXO1 loss is well-tolerated, suggesting the existence of compensatory mechanisms that could be exploited in DDR-deficient cancers. Using CRISPR screening, we find EXO1 loss as synthetic lethal with many DDR genes somatically inactivated in cancers, including Fanconi Anaemia (FA) pathway and BRCA1-A complex genes. We also identify the spliceosome factor and tumour suppressor ZRSR2 as synthetic lethal with loss of EXO1 and show that ZRSR2-deficient cells are attenuated for FA pathway activation, exhibiting cisplatin sensitivity and radial chromosome formation. Furthermore, FA or ZRSR2 deficiencies depend on EXO1 nuclease activity and can be potentiated in combination with PARP inhibitors or ionizing radiation. Finally, we uncover dysregulated replication-coupled repair as the driver of synthetic lethality between EXO1 and FA pathway attributable to defective fork reversal, elevated replication fork speeds, post-replicative single stranded DNA exposure and DNA damage. These findings implicate EXO1 as a synthetic lethal vulnerability and promising drug target in a broad spectrum of DDR-deficient cancers unaddressed by current therapies
Gonadotrophs have a dual origin, with most derived from early postnatal pituitary stem cells.
No full textGonadotrophs are the essential pituitary endocrine cells for reproduction. They produce both luteinizing (LH) and follicle-stimulating (FSH) hormones that act on the gonads to promote germ cell maturation and steroidogenesis. Their secretion is controlled by the hypothalamic gonadotrophin-releasing hormone (GnRH), and gonadal steroid feedback. Gonadotrophs first appear in the embryonic pituitary, along with other endocrine cell types, and all expand after birth. While gonadotrophs may display heterogeneity in their response to GnRH, they appear, at least transcriptionally, as a homogenous population. The pituitary also contains a population of stem cells (SCs), whose contribution to postnatal growth is unclear, in part because endocrine cells maintain the ability to proliferate. Here we show an unsuspected dual origin of the murine adult gonadotroph population, with most gonadotrophs originating from postnatal pituitary stem cells starting early postnatally and up to puberty, while embryonic gonadotrophs are maintained. We further demonstrate that postnatal gonadotroph differentiation happens independently of gonadal signals and is not affected by impairment of GnRH signalling. The division of gonadotrophs based on separate origins has implications for our understanding of the establishment and regulation of reproductive functions, both in health and in disease
“It would be better for those of us who have the disease not to be ashamed”: Insights from people living with chronic hepatitis B virus infection and healthcare workers providing HBV care in Kilifi, Kenya
No full textChronic hepatitis B infection (CHB) causes over 1 million deaths annually, with a large burden of morbidity and mortality in the WHO-African Region (WHO-AFRO) where <5% of people are diagnosed and 0.2% are on treatment. Studies have shown that understanding of hepatitis B virus (HBV) here is often poor, and people living with HBV (PLWHB) can experience stigma and discrimination. However there has been little documentation on the impact of an HBV diagnosis on the lives of PLWHB in the WHO-AFRO region or community involvement in improving care provision. We undertook two focus group discussions (FGDs) with PLWHB and two with healthcare workers (HCWs) providing HBV care at Kilifi County Referral Hospital (KCRH), Kenya to explore experiences of living with HBV and barriers to accessing care. FGDs were conducted primarily in Kiswahili, transcribed verbatim and translated into English. The data were analysed thematically using NVivo version 14. PLWHB and HCWs at KCRH had a good understanding of HBV which was likely influenced by a concurrent research study on HBV, however they reported low awareness in the general community, and there is no local name for the infection. Many PLWHB were shocked at their initial diagnosis with mixed reactions from friends and family. Costs of transport and concerns about lost employment were the biggest barriers to care. Many people suggested decentralised clinics would reduce loss to follow up, however others would rather be treated far from home to preserve anonymity. Stigma was highlighted as a major issue, leading to feelings of isolation, rejection and discrimination. Community education, wider testing and advocacy by well-respected community members were mentioned as key methods to reduce HBV transmission. Decentralisation of clinics may improve access to care; however, this needs to be developed in careful consultation with PLWHB to ensure they are acceptable and accessible to all
Multi-omic analysis reveals astrocytic annexin-A2 as critical for network-level circadian timekeeping in the suprachiasmatic nucleus.
No full textThe mammalian suprachiasmatic nucleus (SCN) orchestrates daily (circadian) rhythms of physiology and behavior by broadcasting timing cues generated autonomously by its mutually reinforcing network of ~10,000 neurons and ~3000 astrocytes. Although astrocytic control of extracellular glutamate and GABA has been implicated in driving circadian oscillations in SCN gene expression and neuronal activity, the full scale of the network-level signaling mechanisms is unknown. To understand better how this astrocyte-neuron network operates, we adopted a multi-omics approach, first using SILAC-based mass spectrometry to generate an SCN proteome where ~7% of identified proteins were circadian. This circadian proteome was analyzed bioinformatically alongside existing single-cell RNAseq transcriptomic data to identify the cell-types and processes to which they contribute. This highlighted "S100 protein binding," tracked to astrocytes, and revealed annexin-A2 (Anxa2) as an astrocyte-enriched circadian protein for further investigation. We show that Anxa2 and its partner S100a10 are co-expressed and enriched in SCN astrocytes. We also show that pharmacological disruption of their association acutely and reversibly dysregulated the circadian cycle of astrocytic calcium levels and progressively compromised SCN neuronal oscillations. Anxa2 and S100a10 interaction therefore constitutes an astrocytic cellular signaling axis that regulates circadian neuronal excitability and ultimately SCN network coherence necessary for circadian timekeeping
AllohubPy: Detecting allosteric signals through an information-theoretic approach.
No full textAllosteric regulation is crucial for biological processes like signal transduction, transcriptional regulation, and metabolism, yet the mechanisms and macromolecular properties that govern it are still not well understood. Several methods have been developed over the years to study allosterism through different angles. Among the possible ways to study allosterism, information-theoretic approaches, like AlloHubMat or GSAtools, can be particularly effective due to their use of robust statistics and the possibility to be combined with graph analysis. These methods capture local conformational changes associated with global motions from molecular dynamics simulations through the use of a Structural Alphabet, which simplifies the complexity of the Cartesian space by reducing the dimensionality down to a string of encoded fragments, representing sets of internal coordinates that still capture the overall conformation changes. In this work, we present "AllohubPy," an improved and standardized methodology of AlloHubMat and GSAtools coded in Python. We analyse the performance, limitations and sampling requirements of AllohubPy by using extensive molecular dynamics simulations of model allosteric systems and apply convergence analysis techniques to estimate result reliability. Additionally, we expand the methodology to use different dimensionality reduction Structural Alphabets, such as the 3DI alphabet, and integrate Protein Language Models (PLMs) to refine allosteric hub communication detection by monitoring the detected evolutionary constraints. Overall, AllohubPy expands its preceding methods and simplifies the use and reliability of the method to effectively capture dynamic allosteric motions and residue pathways. AllohubPy is freely available on GitHub (https://github.com/Fraternalilab/AlloHubPy) as a package and as a Jupyter Notebook
Cleared tissue dual-view oblique plane microscopy
No full textWe present a dual-view oblique plane microscope (dOPM) for imaging thick optically cleared tissue samples using a silicone immersion primary objective. The custom-designed remote refocusing relay utilizes stock optics to achieve remote refocusing in refractive index-matched samples. The spatial resolution of the system was characterized using a series of fluorescent bead phantoms with refractive indices ranging from 1.4-1.5, with the point spread function full width at half maximum measuring ∼0.5 µm laterally and ∼1 µm axially for a refractive index-matched bead sample, with minimal degradation over a > 250 µm axial range. We characterize how the remote refocusing performance at sample refractive indices up to n = 1.5 can be partially compensated for using adjustment of the correction collar on the primary objective. We apply the system to imaging a range of biological samples with varied refractive indices. Combined with tiled acquisition, image stitching, and multi-view image fusion, the microscope enables multicolour imaging of mm-wide and ∼250 µm thick optically cleared mouse ovarian cancer, bone marrow, and colon tissue samples, and a whole Drosophila melanogaster fruit fly brain. The system offers a platform for fast and high-resolution, multicolour volumetric imaging across spatial scales, integrated on a commercially available microscope frame
Differentiating clinically important interstitial lung abnormalities in lung cancer screening.
No full textBACKGROUND: Interstitial lung abnormalities (ILAs) are common incidental findings in lung cancer screening (LCS). However, challenges remain in identifying clinically relevant ILAs as highlighted in a joint statement by a European multidisciplinary task force led by the European Respiratory Society (ERS). To address these challenges, we analysed ILAs identified in one of Europe's largest LCS studies. METHODS: Of 11 635 LCS individuals, 417 screen-detected ILAs were evaluated using a new visual classification system focused on traction bronchiolectasis: non-fibrotic ILA (no traction bronchiolectasis), fibrotic ILA (traction bronchiolectasis in ≤2 lobes); undiagnosed interstitial lung disease (traction bronchiolectasis in >2 lobes). Observer agreement was compared with Fleischner Society ILA classification using Cohen's Kappa. An age, sex and smoking history-matched control group allowed the examination of associations between baseline ILA/UILD and comorbidities, forced vital capacity (FVC), hospitalisations (Student's t-tests) and mortality (univariable and multivariable Cox proportional hazards models). FINDINGS: Our visual ILA classification showed superior interobserver agreement (K=0.76) versus the Fleischner ILA classification (K=0.64). ILA/UILD subjects had more prevalent comorbidities, increasing (vs controls) approximately 10 years prior to ILA/UILD diagnosis. Compared with controls, mortality rates were 6-fold higher for UILD participants and 3-fold higher for fibrotic and non-fibrotic ILA subtypes. On multivariable Cox regression analysis, ILA/UILD presence (HR=4.90, 95% CI =2.36 to 10.10, p<0.001) showed stronger independent associations with mortality than baseline FVC (HR=0.98, 95% CI =0.96 to 1.00, p=0.04). CONCLUSION: We demonstrate a new reproducible classification of clinically important ILA/UILDs in LCS populations. We highlight that FVC shows limited associations with mortality in ILA/UILD subjects. Increased multiorgan comorbidity in ILA/UILD subjects highlights a need for comprehensive early multisystem evaluation
Major transitions in sociocultural evolution
No full textRecent years have seen growing interest in applying the Evolutionary Transitions in Individuality (ETI) framework to human sociocultural evolution. Proponents argue that human societies exhibit features—such as multilevel organisation, cooperation, and division of labour—sufficiently analogous to biological ETIs to warrant theoretical extension. This paper critically assesses such claims and argues that they rest on a fundamental misapplication of the ETI framework. Drawing on recent work in cultural evolution, I show that sociocultural systems typically lack the core conditions required for an ETI, including autonomous reproduction at the group level and the operation of natural selection in the reproductive mode. Attempts to relax these criteria risk undermining the coherence of the framework itself. I conclude that while the broader MET framework may still have value for understanding sociocultural change, the specific explanatory structure of ETI theory does not transfer
Boomerang and bones: Refining the chronology of the Early Upper Paleolithic at Obłazowa Cave, Poland.
No full textBeginning with the Early Aurignacian, Homo sapiens demonstrated an enhanced symbolic capacity, expanding artistic expressions from body decoration to portable art and aesthetically refined tools. These artistic endeavors, often intertwined with utilitarian purposes, have sparked debates regarding their symbolic versus functional roles. Among these remarkable artifacts is a complete mammoth tusk boomerang from Layer VIII of Obłazowa Cave, Poland, found in association with a human phalanx. Determining its precise chronology and cultural context is critical for understanding the emergence and variability of symbolic behaviors among early Homo sapiens groups in Europe. This study refines the chronology of the Early Upper Paleolithic occupation of Layer VIII at Obłazowa Cave through radiocarbon dating of several bones and the human fossil found near the ivory boomerang. Bayesian modeling places the site's main occupation phase between 42,810-38,550 cal BP (95,4% probability). The mammoth-ivory boomerang, calibrated to 42,290-39,280 cal BP with a 95.4% probability, emerges as one of Europe's oldest known examples of this complex tool, exemplifying technological and symbolic innovation at Obłazowa Cave. This multi-disciplinary research underscores the importance of integrating advanced methodologies to explore cultural practices during the Upper Paleolithic. The findings not only deepen our understanding of Homo sapiens' adaptive strategies but also highlight the nuanced interplay of technology, symbolism, and environmental interaction during the earliest phases of human dispersals in Central Europe