Biology subjects

Jung, C.

Publications and source records attributed to Jung, C..

At least 19 recordsLinked to original sources

bioRxiv2026

Human iPSCs and iPSC-Derived Endothelial Cells from PAD Patients and Healthy Donors Exhibit Comparable Characteristics and Potency: Implications for Autologous Cell Therapy in Peripheral Artery Disease

BACKGROUND: Peripheral artery disease (PAD) can lead to amputation in advanced cases, making cell therapy using human induced pluripotent stem cells (hiPSCs) a promising therapeutic option. hiPSC-derived endothelial cells (hiPSC-ECs) have shown favorable effects in treating experimental ischemic cardiovascular disease. An autologous approach for PAD patients is preferable to avoid immunological reactions. However, it is yet unknown whether hiPSCs and hiPSC-ECs derived from PAD patients have similar characteristics and potency compared to those derived from non-PAD donors. Therefore, we explored whether there are significant differences in the characteristics and potency of hiPSCs and hiPSC-ECs between non-PAD donors and PAD patients. METHODS: We generated hiPSCs from the blood of non-PAD donors and patients with PAD. We determined the pluripotency of hiPSCs using qRT-PCR, flow cytometry, immunostaining, RNA-seq, and teratoma formation assay. The genetic stability of hiPSCs was confirmed by G-banding karyotyping. We then differentiated hiPSCs into endothelial cells (ECs) in a clinically compatible manner. We compared the characteristics and therapeutic potency of hiPSC-ECs derived from non-PAD and PAD groups using qRT-PCR, flow cytometry, immunostaining, and RNA-seq. In vitro endothelial functions were determined by nitric oxide (NO) production and capillary network formation. Therapeutic potency and incorporation of engrafted hiPSC-ECs were evaluated in a hindlimb ischemia model. RESULTS: We successfully generated hiPSCs from the blood of seven non-PAD donors and eight PAD patients. Both non-PAD and PAD-derived hiPSCs exhibited similar expression levels of pluripotency markers. All hiPSCs, regardless of the group, formed teratomas and showed normal karyotypes. RNA-seq analyses revealed similar gene expression profiles between the groups. hiPSC-ECs derived from both non-PAD and PAD donors exhibited similar expression levels of EC markers at both the gene and protein levels. RNA-seq analyses showed no significant overall differences in gene expression profiles between the groups. Functional analyses demonstrated similar endothelial characteristics and function in hiPSC-ECs from both groups. Both groups showed similar perfusion recovery, limb salvage, and vessel-forming capacity. Engrafted hiPSC-ECs from both groups also exhibited similar angiogenic and vessel-forming capabilities. CONCLUSIONS: Our study demonstrated no significant differences in hiPSCs and hiPSC-ECs derived from non-PAD donors and PAD patients in terms of molecular and cell biological characteristics, therapeutic effects, and vessel-forming capability. Our study indicates that hiPSCs and hiPSC-ECs derived from PAD patients can serve as a novel platform for autologous cell therapy.

cell biology↗

bioRxiv2026

First estimates of the population growth rate of the parasitic honey bee mite Tropilaelaps mercedesae in Apis mellifera colonies

A parasitic mite of honey bee brood (Tropilaelaps mercedesae), is spreading through populations of Apis mellifera honey bees in new regions and poses a major threat to honey bee health. Despite its clear threat, the biology of this mite is poorly understood, with gaps on such fundamental issues as how fast its populations can grow. This leaves the beekeeping world underprepared to plan for its arrival and management. In this study, we documented the growth of T. mercedesae populations in untreated A. mellifera colonies in Thailand and South Korea, and did the same for another parasitic mite (Varroa destructor) when possible. We found that the population growth of T. mercedesae was variable but could reach high levels (daily r of 0.010, 0.036, and 0.057), while the population growth of V. destructor (r = 0.021) matched previous estimates. Our results indicate that T. mercedesae populations can grow rapidly but they do not always attain this potential. Based on our results, humidity should be studied as a potential driver of population growth. If future work can reveal key drivers of T. mercedesae population growth, this would help predict infestations and help design management strategies that exploit the pests biological vulnerabilities.

zoology↗

bioRxiv2026

Laudanosine restricts Ebola virus entry by targeting TPC2-dependent endolysosomal trafficking

Late endosome-dependent viruses, including filo- and arenaviruses, rely on host endolysosomal trafficking for productive infection. Here, we used a dual-colour Vesicular stomatitis virus (VSV) based pseudoparticle screen of CytoSorb-derived fractions to identify inhibitors of the Zaire Ebolavirus glycoprotein (GP)-mediated entry. Iterative chromatographic purification and mass spectrometry identified Laudanosine, a degradation product of the clinically used neuromuscular blocker Atracurium, as the antiviral compound. Laudanosine specifically inhibited entry mediated by Ebola, Marburg, Lymphocytic choriomeningitis and Lassa virus glycoproteins without affecting VSV-G-dependent entry. Importantly, Laudanosine inhibited authentic Ebola virus infection without detectable cytotoxicity in cell culture and embryonic zebrafish. Molecular dynamics simulations suggest stable association of Laudanosine with the allosteric inhibitory pocket of the lysosomal two-pore channel (TPC2). Consistently, Laudanosine impairs autophagic flux and disrupts endolysosomal trafficking. Together, our findings identify Laudanosine as a previously unrecognised inhibitor of TPC2-dependent entry of highly lethal viral pathogens.

microbiology↗

bioRxiv2026

Reproducible and shareable bioinformatics pipelines from natural-language prompts

Large language models (LLMs) are increasingly used to generate bioinformatics pipelines and to carry out analyses from natural-language prompts. However, the resulting analyses are often difficult to reproduce across sessions, owing to the non-deterministic nature of LLM-driven conversations and heterogeneity of local execution environments, and cannot run on remote high-performance computing (HPC) servers or be shared and reused. We present Autopipe, a platform that guides any Model Context Protocol (MCP) - compatible LLM to produce, execute, and publish source-preserved, re-executable containerized pipelines. Autopipe enables users to execute bioinformatics pipelines on any on-premises remote servers - supported by comprehensive setup documentation aimed at researchers without prior server-administration experience - and to visualize results through an extensible web-based viewer. The Autopipe platform comprises four components: a desktop application with an embedded MCP server for pipeline management and remote execution, an online registry for pipeline and plugin discovery, a web-based result viewer, and a CLI tool for customizing viewer plugins. Autopipe turns conversational analysis into re-executable and shareable workflows. Autopipe is freely available at https://autopipe.org/.

bioinformatics↗

bioRxiv2026

Efficient base editing and development in human embryos without chromosomal alterations

Cas9-based tools enable the introduction of genetic lesions to investigate DNA repair outcomes and edit the genome at disease-relevant loci. DNA double-strand breaks (DSBs) induced by CRISPR/Cas9 result in frequent aneuploidy and large deletions, revealing a repair deficiency in early human embryos and limiting the clinical application of this technology. Here we evaluated the DNA repair outcomes of DNA nicks and mismatches introduced using base editors in human embryos at two targets, PCSK9 and HBG. Editing was efficient and, unlike Cas9-induced DSBs, did not result in either chromosomal abnormalities or large deletions. Small insertions or deletions after base editing were rare, and off-target activity was dependent on the guide RNA. Delivering the base editor as a protein at fertilization or at the pronuclear stage allowed normal development to the blastocyst stage and the derivation of edited stem cell lines. In stark contrast, introduction of the editor as RNA resulted in early embryo arrest. Our results demonstrated that, unlike DSBs, DNA nicks and mismatches are efficiently repaired in human embryos, allowing specific on-target changes without genotoxic consequences.

genetics↗

bioRxiv2026

Decoding Promoter Activity from DNA Sequence using Pre-trained Language Models

Promoter architecture plays a central role in transcriptional regulation, yet predicting promoter activity directly from DNA sequence remains challenging. Here, we assess whether transformer-based DNA language models can learn and interpret regulatory logic encoded in Drosophila core promoters. We fine-tuned the 117-million-parameter DNABERT-2 model on [~]700 synthetic promoters assayed by [~]2,600 dual-luciferase measurements in Drosophila S2 cells. A sequence-only model achieved high predictive accuracy (R{superscript 2} {approx} 0.91), demonstrating that core promoter sequence alone strongly constrains transcriptional output. Model interpretability using SHapley Additive exPlanations (SHAP) revealed biologically meaningful sequence features corresponding to canonical core promoter elements. Extending the model to incorporate biological context, including Ecdysone (Ecd) hormonal activation and flanking -1/+1 nucleosomal sequences, preserved strong performance while capturing more complex promoter behavior. Gene-wise cross-validation showed robust generalization for most promoters, and application to independent in vivo embryo data demonstrated that the model still generalizes reasonably well, even in complex biological contexts.

bioinformatics↗

bioRxiv2025

Cas9-enriched nanopore sequencing enables comprehensive and multiplexed detection of repeat expansions

Short tandem repeat (STR) expansion is a major genetic mechanism underlying numerous neurogenetic disorders. However, traditional PCR amplification and short-read next-generation sequencing (NGS)-based methods often fail to detect complex and large expansions including methylation information. Here, we modified an amplification-free nanopore Cas9-targeted sequencing (nCATS) platform and developed a dedicated analysis algorithm, STRiker, for the simultaneous assessment of all currently defined STR loci (56 sites) using a single test with genomic DNA from patient-derived blood cells. We ultimately identified pathogenic repeat expansions in 12 of 37 patients (32.4%) with cerebellar ataxia who remained genetically undiagnosed despite extensive prior genetic testing, in FGF14 (n=4), ATXN8OS, NOP56, RFC1 (n=2 each), and PRNP and NOTCH2NLC (n=1 each). Additionally, family-based cascade screening revealed six relatives with repeat expansions in five families. These results demonstrated a broader diversity of pathogenic repeat structures, particularly in FGF14, and illustrated that CpG methylation can mitigate the pathogenic effects of repeat expansions. This approach offers a powerful strategy for improving the diagnosis of STR-related neurogenetic diseases, for cerebellar ataxia and other STR-related diseases in the future. One Sentence SummaryImproved nanopore Cas9-targeted sequencing with STRiker enhances diagnosis of STR- related neurogenetic diseases and offers broader molecular insights.

bioengineering↗

bioRxiv2025

Adeno-Associated Virus-Induced Neurotoxicity is Prevented by CpG Depletion

Adeno-associated viruses (AAVs) are the vector of choice for gene delivery to the nervous system. While AAVs have a strong safety profile, recent studies show that AAV causes dendritic loss and synaptic weakening in mouse somatosensory cortex, impacts that are prevented by systemic administration of blockers of Toll-like receptor 9 (TLR9), an innate immunoreceptor that detects unmethylated cytosine-guanine (CpG) motifs. However, TLR9 blockers are immunosuppressive and costly. To realize AAVs full potential, it is critical to identify strategies to protect neurons without compromising immunity. Here we find that partially depleting CpG motifs from the AAV genome prevents AAV-induced dendritic loss and synaptic weakening. CpG depletion of transgene and intronic regions via codon-optimized synonymous mutations was protective and did not impair transgene expression in vivo. To facilitate the production and use of lower-CpG AAVs, we created a web-facing application, CpG-Assist Tool (CpG-AT), which allows researchers to quantify the CpG content of any sequence, explore the CpG content of commonly used AAV components, and generate CpG-depleted coding and non-coding sequences. This study identifies CpG depletion as a practical strategy to prevent AAV-induced neural circuit disruption, and provides tools to facilitate CpG reduction to enhance the safety and efficacy of AAV-mediated gene delivery.

neuroscience↗

bioRxiv2025

Site-specific replacement of large-scale DNA fragments in human cells

Despite advances in genome editing1-4, precisely replacing large-scale fragments in human cells remains a significant challenge. Here, we present a site-specific gene replacement tool, named Prime Assembly (PA), which adapts prime editors to produce one or two pairs of 3-flaps on both the genome and donor plasmids. These 3-flaps anneal to each other precisely, similar to "Gibson Assembly" in DNA oligonucleotides5, allowing megabase-scale genomic excision and/or kilobase-scale donor insertion at the gene of interest. The PA system achieves an efficiency of up to 57.8% in replacing endogenous sequences with a 2.9 kbp donor DNA fragment in HEK293T cells, and presents an accuracy of more than 99% for integrated PA fragments. Ultimately, the site-specific replacement of large-scale coding sequences (CDSs) in disease-related genes can restore gene function across numerous patients with different mutations, providing a gene implantation technique for genome editing and a universal therapeutic approach.

molecular biology↗

bioRxiv2025

Engineered ADARs enable single-nucleotide resolution DNA A-to-G editing without bystander effects

The adenine base editor (ABE), which enables A*T-to-G*C base conversion, has emerged as a powerful tool for therapeutic applications. However, conventional ABEs suffer from bystander nucleotide conversions, limiting their utility for precise editing. Here, we present a single-nucleotide resolution ABE (snuABE) created by fusing a nickase Cas9, nCas9(H840A), with the deaminase domain of ADAR, which acts on DNA:RNA hybrids, instead of TadA, which acts on single-stranded DNA in conventional ABEs. snuABE requires a specially designed target-adenine guide RNA (tagRNA) that introduces a mismatch at the target adenine, enabling highly specific A-to-G editing by ADAR. Engineering ADAR from Pediculus humanus using the in silico protein evolution algorithm EvolvePro, along with 3-end protection of the tagRNA, further enhances the editing activity of snuABE in human cells. Moreover, snuABE exhibits significantly reduced DNA off-target activity, highlighting its potential as a safer and more precise base editing technology for therapeutic applications.

bioengineering↗

bioRxiv2025

No decoy effect in bees: rewardless flowers do not increase bumblebees' preference for neighbouring flowers

Nectarless flowers are common among flowering plants, which often retain colour-changed, rewardless flowers instead of shedding them. Yet, how these flowers influence pollinators foraging choices within an inflorescence remains unclear. We hypothesised that rewardless flowers in an inflorescence may act as "decoys", causing the rewarding flowers in the inflorescence to be perceived as more valuable by contrast. Using artificial inflorescences, we presented individual bumblebees (Bombus terrestris) with a binary choice between two equally rewarding inflorescences, one of which included additional unrewarded, differently coloured flowers. We found that the presence of rewardless flowers did not increase bees preference for neighbouring flowers, nor did it affect their overall choice between inflorescences. However, bees quickly learned to avoid the unrewarded flowers, drastically reducing visits and probing within a few foraging bouts. We review research on decoy effects in bees, and find very little support for their presence. Our findings contribute to the growing body of evidence that rewardless flowers do not induce decoy effects in bees, and highlight the need for further research into the ecological role of nectarless flowers within floral patches. It may be time to abandon the search for classical decoy effects in pollinators.

animal behavior and cognition↗

bioRxiv2025

Structural basis for DNA double-strand break sensing by human MRE11-RAD50-NBS1 and its TRF2 complex

The MRE11-RAD50-NBS1 (MRN) complex is a central, multifunctional factor in the detection, signaling and nucleolytic processing of DNA double-strand breaks (DSBs). To clarify how human MRN binds generic and telomeric DNA ends and can separate DNA end sensing from nuclease activities, we determined cryo-electron microscopy structures of human MRN bound to DNA and to DNA and the telomere protection factor TRF2. MRN senses DSBs through a tight clamp-like sensing state with closed coiled-coil domains, but auto-inhibited MRE11 nuclease. NBS1 wraps around the MRE11 dimer, with NBS1s ATM recruitment motif sequestered by binding to the regulatory RAD50 S site, necessitating an allosteric switch for ATM activation. At telomeric DNA, TRF2 blocks the second S site via the iDDR motif to prevent nuclease and ATM activation. Our results provide a structural framework for topological DNA sensing and separation of sensing, signaling and processing activities of mammalian MRN. Highlights- Human MRN senses DNA ends with an autoinhibited nuclease - NBS1s C-terminus binds one RAD50 S site in the sensing state - TRF2 binds MRNs second S site at telomeres - RAD50 and ATM compete for the NBS1 C-terminus

biochemistry↗

bioRxiv2025

Large extracellular vesicles derived from red blood cells in coronary artery disease patients with anemia promote endothelial dysfunction

Background and purposeEndothelial dysfunction (ED) is a hallmark of cardiovascular disease (CVD). We recently showed that anemia is associated with worsening of endothelial function after acute myocardial infarction (AMI). Extracellular vesicles (EVs) are efficient communicators between cells and can functionally contribute to different CVD, including, AMI. However, their specific role of EVs in stable coronary artery disease (CAD)-associated with anemia, particularly their contribution to ED, has not yet been investigated systematically. Experimental approachRed blood cell-derived EVs (REVs) and plasma-derived EVs (PLEVs) from all blood cells and endothelium were isolated from patients with stable CAD. The isolated large REVs and PLEVs were characterized using dynamic light scattering (DLS), nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM), and Western blotting. Uptake assays were performed by co-incubating with fluorescently-labeled REVs and PLEVs with human umbilical vein endothelial cells (ECs). Nitric oxide (NO) consumption ability of REVs was analyzed using a chemiluminescence detector (CLD). After co-incubation of aortic rings explanted from wild-type (WT) mice with REVs and PLEVs from anemic and non-anemic CAD patients, endothelial function was assessed using a wire myograph system. To investigate differences in the content of REVs and PLEVs between anemic and non-anemic CAD patients, proteomic analysis was performed. Key resultsDLS analysis showed that both REVs and PLEVs were within the size distribution range of 100-1000 nm. NTA analysis revealed increased release of REVs in anemic patients compared to non-anemic patients. Co-incubation of labeled REVs and PLEVs with ECs demonstrated their uptake by ECs in vitro which was similar between anemic patients compared to non-anemic patients. REVs from anemic patients showed increased NO consumption compared to those from non-anemic patients. Aortic rings co-incubated with REVs from anemic patients showed attenuated endothelial NO-dependent relaxation responses compared to non-anemic patients. Proteomics analysis of REVs from anemic patients revealed numerous differentially expressed proteins, including decreased abundance of antioxidant proteins such as catalase 1 (CAT1), superoxide dismutase 1 (SOD1) and increased oxidative stress-promoting myeloperoxidase (MPO). Co-incubation of ECs with REVs from anemic patients demonstrated increased ROS production. ConclusionAnemia is associated with increased release of REVs and enhanced NO consumption, which promotes ED. This is further exacerbated by an altered redox balance and increased ROS production, implicating therapeutic importance in anemic patients with CAD. Graphical AbstractAnemia is associated with an increased release of RBC-derived large extracellular vesicles (REVs), which are taken up by endothelial cells (ECs). Anemic REVs show enhanced nitric oxide (NO) consumption, contributing to NO dysregulation in ECs. Additionally, REVs carry various redox enzymes, including the oxidative stress-promoting enzyme myeloperoxidase (MPO), as well as antioxidant enzymes such as superoxide dismutase (SOD) and catalase (CAT). An imbalance in these redox enzymes leads to increased oxidative stress and endothelial nitric oxide synthase (eNOS) uncoupling, resulting in impaired NO-mediated relaxation responses and subsequent endothelial dysfunction (ED). O_FIG O_LINKSMALLFIG WIDTH=166 HEIGHT=200 SRC="FIGDIR/small/642191v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@19d4cccorg.highwire.dtl.DTLVardef@1d1b3forg.highwire.dtl.DTLVardef@e7f144org.highwire.dtl.DTLVardef@190b007_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2025

ReaxFF-guided Optimisation of VIRIP-based HIV-1 Entry Inhibitors

Peptides hold great promise for safe and effective treatment of viral infections. However, their use is often constrained by limited efficacy and high production costs, especially for long or complex peptide chains. Here, we used ReaxFF molecular dynamics (MD) simulations to optimize the size and activity of VIRIP (Virus Inhibitory Peptide), a naturally occurring 20-residue fragment of 1-antitrypsin that binds the HIV-1 GP41 fusion peptide (FP), thereby blocking viral fusion and entry into host cells. Specifically, we used the NMR structure of the complex between an optimized VIRIP derivative (VIR-165) and the HIV-1 gp41 FP for ReaxFF-guided in silico analysis, evaluating the contribution of each amino acid in the interaction of the inhibitor with its viral target. This approach allowed us to reduce the size of the HIV-1 FP inhibitor from 20 to 10 amino acids (2.28 to 1.11 kDa). HIV-1 infection assays showed that the size-optimized VIRIP derivative (soVIRIP) retains its broad-spectrum anti-HIV-1 capability and is non-toxic in the vertebrate zebrafish model. Compared to the original VIRIP, soVIRIP displayed more than 100-fold higher antiviral activity (IC50 of [~]120 nM). Thus, it is more potent than a dimeric 20-residue VIRIP derivative (VIR-576) that was proven safe and effective in a phase I/II clinical trial. Our results show that ReaxFF-based MD simulations represent a suitable approach for the optimization of therapeutic peptides.

molecular biology↗

bioRxiv2024

Uncovering Genes Essential in Domestication and Breeding of Sugar Beet

The genus Beta encompasses economically important root crops such as sugar and table beet. A Beta diversity set including the wild relative B. vulgaris ssp. maritima was grown in the field, and a large phenotypic diversity was observed. The genomes of 290 accessions were sequenced, and more than 10 million high-quality SNPs were employed to study genetic diversity. A genome-wide association study was performed, and marker-trait associations were found for nine phenotypic traits. The candidate gene within the M locus controlling monogermity on chromosome 4 was previously unknown. The most significant association for monogermity was identified at the end of chromosome 4. Within this region, a non-synonymous mutation within the Zinc-Finger domain of the WIP2 gene co-segregated with monogermity. This gene plays a regulatory role in AGL8/FUL in Arabidopsis. Intriguingly, commercial hybrids are in a heterozygous state at this position. Thus, the long-sought gene for monogermity was identified in this study. Red and yellow pigmentation due to betalain accumulation in shoots and roots is an important characteristic of table and leaf beets. The strongest associations were found upstream or downstream of two genes encoding Cytochrome P450 and anthocyanin MYB-like transcription factor proteins involved in betalain biosynthesis. Significant associations for Cercospora leaf spot resistance were identified on chromosomes 1, 2, 7, and 9. The associated regions harbor genes encoding proteins with leucine-rich repeats and nucleotide binding sites whose homologs are major constituents of plant-pathogen defense.

genomics↗

bioRxiv2024

C9orf72 expansion creates the unstable folate-sensitive fragile site FRA9A

The hyper-unstable Chr9p21 locus, harbouring the interferon gene cluster, oncogenes and C9orf72, is linked to multiple diseases. C9orf72 (GGGGCC)n expansions (C9orf72Exp) are associated with incompletely penetrant amyotrophic lateral sclerosis, frontotemporal dementia and autoimmune disorders. C9orf72Exp patients display hyperactive cGAS-STING-linked interferon immune and DNA damage responses, but the source of immuno-stimulatory or damaged DNA is unknown. Here, we show C9orf72Exp in pre-symptomatic and ALS-FTD patient cells and brains cause the folate- sensitive chromosomal fragile site, FRA9A. FRA9A centers on >33kb of C9orf72 as highly-compacted chromatin embedded in an 8.2Mb fragility zone spanning 9p21, encompassing 46 genes, making FRA9A one of the largest fragile sites. C9orf72Exp cells show chromosomal instability, heightened global- and Chr9p-enriched sister-chromatid exchanges, truncated-Chr9s, acentric-Chr9s and Chr9-containing micronuclei, providing endogenous sources of damaged and immunostimulatory DNA. Cells from one C9orf72Exp patient contained highly-rearranged FRA9A-expressing Chr9 with Chr9-wide dysregulated gene expression. Somatic C9orf72Exp repeat instability and chromosomal fragility are sensitive to folate-deficiency. Age-dependent repeat instability, chromosomal fragility, and chromosomal instability can be transferred to CNS and peripheral tissues of transgenic C9orf72Exp mice, implicating C9orf72Exp as the source. Our results highlight unappreciated effects of C9orf72 expansions that trigger vitamin-sensitive chromosome fragility, adding structural variations to the disease-enriched 9p21 locus, and likely elsewhere. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=94 SRC="FIGDIR/small/620312v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@154b23borg.highwire.dtl.DTLVardef@1814a6dorg.highwire.dtl.DTLVardef@13d713eorg.highwire.dtl.DTLVardef@d5d994_HPS_FORMAT_FIGEXP M_FIG C_FIG

genetics↗

bioRxiv2024

Hijacking and recording intracellular RNAs in human cells using eukaryotic reprogrammed tracrRNAs

Recording intracellular RNA expression events is critical for understanding cell states and biological processes, but no method without responsive promoters is available in mammalian cells. Here, we determined the sequence formula of eukaryotic reprogrammed tracrRNA, named euRptr, which can hijack intracellular RNAs to make a complex with CRISPR-associated tools. By leveraging the programmability of euRptr, we devised the system "CHEETAH" to directly capture and record intracellular RNAs in human cells. We validated that CHEETAH induces self-editing by hijacking endogenous mRNAs, that CHEETAH selectively records different viral RNA fragments on a DNA reporter using base editors with a shared-euRptr, and that CHEETAH enables multiplexed, time-resolved recording of viral RNAs on a DNA Tape using prime editors, providing a new avenue for investigating intracellular RNA in eukaryotes.

genetics↗

bioRxiv2024

Distinct effects of acute and chronic blood loss anemia on vascular function after acute myocardial infarction

BackgroundAnemia is frequently observed in patients with cardiovascular diseases (CVD). Anemia alone or in combination with other morbid conditions leads to poor prognosis in acute myocardial infarction (AMI). We recently showed that moderate blood loss anemia is associated with red blood cell (RBC) dysfunction and a compensatory increase in flow-mediated dilation (FMD) responses which are compromised in chronic blood loss anemia However, the effects of acute anemia (AA) and chronic anemia (CA) on endothelial function after AMI are unclear. In this study, we evaluated systemic endothelial function following AMI in established murine models of blood loss acute and chronic anemia. We hypothesize that both AA and CA aggravate systemic endothelial dysfunction (ED) after AMI. Methods and resultsAA or CA was induced in male C57BL/6J mice by repeated blood withdrawal for three consecutive days or six weeks, respectively. Separate groups of anemic and non-anemic mice underwent AMI via left anterior descending artery (LAD) ligation (45 min), followed by reperfusion. Endothelial function was assessed using both in vivo and in vitro methods 24 h post-AMI. Impaired flow-mediated dilation (FMD, in vivo) and endothelium-dependent relaxation (EDR) responses were observed in the aorta, femoral, and saphenous arteries of AA mice compared to their respective control groups 24 h post AMI. The aorta and saphenous arteries from CA mice showed significantly reduced vascular smooth muscle (VSM) contractile responses after AMI. Analysis of oxidative products of nitric oxide (NO) in plasma revealed reduced nitrite and nitrate levels in both AA and CA mice compared to controls 24 h post-AMI. Immunohistochemistry of aortic tissues from both anemic groups showed increased reactive oxygen species (ROS) product 4-Hydroxynonenal (4-HNE). Co-incubation of RBCs from anemic mice or anemic ST-elevation myocardial infarction (STEMI) patients with aortic rings from wild type mice demonstrated attenuated VSM contractile and EDR responses. Supplementation with the ROS scavenger N-acetyl cysteine (NAC) for four weeks improved both in vivo and ex vivo EDR in AA and CA mice 24 h post-AMI. ConclusionAfter AMI, both AA and CA are associated with severe ED, while VSM contractile responses specifically reduced in CA mice. These effects are accompanied by increased ROS and partly mediated by RBCs. Antioxidant supplementation with NAC is a potential therapeutic option to reverse the severe vascular dysfunction in anemia following AMI. Graphical AbstractDistinct effects of acute and chronic anemia on vascular function 24 h post-AMI. After acute myocardial infarction, acute and chronic anemia are associated with increased reactive oxygen species (ROS) and inflammation in endothelial cells (EC), leading to the inhibition of endothelial nitric oxide synthase (eNOS) and subsequent endothelial dysfunction by limiting NO bioavailability. Chronic anemia is additionally associated with decreased vascular smooth muscle cell (VSMC) function due to increased oxidative stress, leading to SMC dysfunction. After N-Acetyl-L-Cysteine (NAC) treatment, vascular function is improved in both anemic groups. O_FIG O_LINKSMALLFIG WIDTH=98 HEIGHT=200 SRC="FIGDIR/small/614629v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@136583aorg.highwire.dtl.DTLVardef@da8708org.highwire.dtl.DTLVardef@d73539org.highwire.dtl.DTLVardef@e7516a_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗