Biology subjects

Shah, S.

Publications and source records attributed to Shah, S..

At least 19 recordsLinked to original sources

bioRxiv2026

How to stand out to a mate - Flash colour of Photinus fireflies maximizes signal contrast against ambient light

During twilight, flying male Photinus fireflies flash in search for conspecific females, which sit on the vegetation below and flash back in response. However, before darkness sets in, the green vegetation in the habitat makes the detection of flashes more difficult. Twilight active fireflies tend to emit more yellow flashes than nocturnal fireflies and it has been proposed that firefly flash colour evolved to enhance the contrast of firefly flashes against their signalling background. There is indirect evidence for this contrast hypothesis, but it remains to be rigorously tested. This is the first study to directly test the contrast hypothesis by recording ambient light spectra in three different habitats throughout the daily flash activity of Photinus pyralis and P. australis and comparing these ambient light spectra to the male flash spectra of the two species. The contrast hypothesis predicts that during peak activity, firefly light colour will maximally contrast with the signalling background. Our data show that male flash spectra fit into a gap in the ambient light spectrum that opens during peak flashing activity of each species, effectively enhancing the visual contrast between male flashes and their background and making males more visible to their potential mates.

evolutionary biology↗

bioRxiv2026

Junctional Nanodomains Organised by Extended Synaptotagmin-1 Underlie Inflammatory Signalling in Sensory Neurons

Endoplasmic reticulum-plasma membrane (ER-PM) junctions coordinate Ca2+ signalling and nociceptor excitability during inflammation, but the molecular scaffold governing assembly of signalling complexes remains unclear. Here, we identify Extended Synaptotagmin-1 (Esyt1) as a key, dynamic regulator of inflammatory pain signalling. In nociceptive DRG neurons, Esyt1 is co-expressed with ANO1 and IP3R1. Time-resolved proximity ligation assays and super-resolution microscopy showed that bradykinin rapidly increased ANO1-Esyt1 nanoscale proximity, while IP3R1-Esyt1 interactions were dynamically reorganised during ANO1-IP3R1 complex formation. Esyt1 knockdown reduced bradykinin-evoked ANO1 activity, impaired store-operated Ca2+ entry (SOCE), decreased STIM1-Orai1 complex formation, and disrupted ER Ca2+ store refilling. Simultaneous imaging of Ca2+ dynamics and ANO1 activity demonstrated functional coupling between SOCE and ANO1 that required Esyt1. Furthermore, in-vivo Esyt1-knockdown significantly reduced bradykinin-induced nocifensive behaviour. Together, these findings identify Esyt1 as a Ca2+-responsive scaffold that organises ER-PM signalling nanodomains and promotes inflammatory pain signalling.

neuroscience↗

bioRxiv2026

Genetic control and tissue specificity of transcriptomic rhythmicity

Circadian rhythms influence molecular traits in most tissues, yet genetic studies mostly treat molecular measurements as static. In particular, genetic effects on gene expression are typically investigated through expression quantitative trait locus (eQTL) studies, which ignore sampling time and therefore miss genetic effects on rhythm parameters such as amplitude and acrophase. In this study, we used a cosinor-based joint genotype-by-rhythm interaction framework applied to GTEx version 10 gene expression data, with donor inferred phase as a proxy for circadian time. We first identified rhythmically expressed genes in 49 of the 50 tissues profiled in GTEx. We then tested common variants within 50 kb of each rhythmic gene's transcription start site for genetic regulation of transcriptomic rhythmicity. We identified 167 circadian expression QTLs (cd-eQTLs), comprising 54 lead signals for 51 genes across 28 tissues. Most cd-eQTLs altered oscillation amplitude, while others affected acrophase or both. We then investigated the tissue specificity of cd-eQTLs and found that those acting on core circadian genes had consistent effects across tissues, whereas those acting on clock-regulated genes were largely tissue-specific. A phenome-wide association study in the Pan-UK Biobank linked 34 cd-eQTLs to 100 phenotypes, including self-reported chronotype, adiposity, blood cell counts and blood biochemistry. These results show that genetic variation shapes circadian gene expression in a tissue-dependent manner, and that accounting for time of day reveals regulatory effects that static eQTL analyses cannot detect.

genomics↗

bioRxiv2026

Breast Cancer-derived PDGFB Remodels the CNS Vascular Niche to Promote Brain, Spinal and Leptomeningeal Metastasis

Background: Central nervous system (CNS) metastases are a major driver of breast cancer (BC) mortality. Recently, our group demonstrated that platelet-derived growth factor-B (PDGFB) and its receptor, platelet-derived growth factor receptor-beta (PDGFR{beta}), promote BC metastasis to the brain; however, the cellular/molecular mechanisms driving this process remain unclear. Here, we investigated tumor-derived PDGFB effects on the CNS vascular niche in regulating metastatic spread. Methods: Triple-negative BC cells were engineered to overexpress PDGFB and used to evaluate endothelial sprouting, blood-brain barrier (BBB) function, vascular permeability and experimental CNS metastasis. A novel retro-orbital (RO) injection model was used to study leptomeningeal/spinal experimental metastasis. Results: PDGFB enhanced human brain microvascular endothelial cell (HBMEC) sprouting, induced cell migration in BBB spheroids, increased permeability in 3D models, and disrupted tight junction proteins in HBMECs. Notably, alterations in vascular permeability were observed in both PDGFR{beta}-positive and PDGFR{beta}-negative endothelial systems, suggesting PDGFB can remodel the CNS vasculature through multiple mechanisms. In vivo, PDGFB increased CNS metastatic burden and altered tumor vasculature in xenograft and allograft models of BC. Furthermore, PDGFB promoted robust colonization of both spinal and leptomeningeal compartments, identifying a previously underappreciated role for this signaling pathway in a clinically devastating form of CNS metastasis. Analysis of patient transcriptomic datasets revealed enrichment of angiogenesis-related pathways in PDGFB-high tumors from patients who developed CNS metastases. Conclusions: Collectively, these findings identify tumor-derived PDGFB as a key regulator of CNS vascular remodeling and metastatic tropism, while introducing a new experimental framework to study spinal metastases and leptomeningeal disease in BC.

cancer biology↗

bioRxiv2026

Sugarcane bZIP16 regulates internode elongation by targeting gibberellic acid biosynthesis pathway

Modern sugarcane cultivars are vegetatively propagated interspecific hybrids developed by extensive breeding and selection method to enhance sugar content, culm dimensions, desired plant architecture and disease resistance. Internode length greatly varies among existing sugarcane cultivars which determines culm length, biomass and sugar storage. Molecular factors responsible for this trait remains largely unknown in sugarcane. In this study, we characterized short and long internode sugarcane varieties and elucidated gene regulatory module interplaying with hormonal-network underneath the internodal growth. The genome size, pigment content and juice characteristics of short and long internode varieties differed with each other and correlated with gene expressions pattern. We established tissue and spatial transcriptome dataset, and comparative analyses demonstrated number of genes involved in cell cycle, cell expansion, transcription factors and hormone pathway including bZIP16 are differentially expressed between short and long internode varieties. Biochemical, DNA-protein interactions and molecular evidences revealed that ScbZIP16 inversely related to GA content by repressing transcripts of ent-isokaurene C2/C3-hydroxylase and GA20 Oxidase in sugarcane. The further genetic evidences confirmed role of ScbZIP16 in reducing GA biosynthesis pathway, flowering time and growth in Arabidopsis. This report of ScbZIP16 regulatory module offers a potential candidate for sugarcane improvement programs.

plant biology↗

bioRxiv2026

Single-cell DNA cytometry with magnetic- and fluorescence-activated bead sorting

Many clinically important cell populations are defined by intracellular DNA or RNA signatures, but isolating and profiling these cells remains difficult. Probe-based in situ hybridization strategies that are compatible with cytometry have difficulty resolving low-abundance transcripts, integrated provirus, or single-copy genomic mutations. In contrast, droplet digital PCR achieves single-molecule sensitivity; however, subsequent cell isolation requires specialized microfluidic sorters that are slow and not widely available. Here, we introduce magniFIND-seq, a nucleic acid cytometry platform that combines the sensitivity of digital PCR with commercial magnetic-activated (MACS) and fluorescence-activated (FACS) sorting instruments. Single-cell genomes and transcriptomes are captured in agarose beads, target sequences are detected by digital PCR, and bead-bound amplicons are labeled with magnetic or fluorescent probes. Controlled evaporation shrinks beads from 55 to 20 m, enabling scalable magnetic separation, fast single-bead FACS sorting, and their combination for high-purity recovery of rare populations. Using magniFIND-seq, we demonstrate multiplexed single-copy detection and FACS-based isolation of simian immunodeficiency virus proviral pol and env targets. Separately, using BCR::ABL1 as a disease-defining target, we enrich chronic myeloid leukemia cells and recover single-cell transcriptomes that resolve tyrosine kinase inhibitor-resistance programs. magniFIND-seq extends the throughput and accessibility of nucleic acid cytometry by engineering compatibility with commercial instruments.

bioengineering↗

bioRxiv2026

Spatial second-order features predict glioma malignant transformation

Isocitrate dehydrogenase (IDH) mutant gliomas often transform from low to high grade aggressive tumors. The genetic and molecular drivers of this Malignant Transformation (MT) are poorly understood, and predicting whether a patient will undergo MT is an unmet challenge of high clinical relevance. To stratify patients by MT risk, we applied integrated spatial DNA and RNA profiling to biopsies from 18 retrospective glioma patients which will either remain stable, undergo MT or have already transformed. The resulting dataset consisted of >600,000 single cells, measuring 962 DNA loci, 1150 RNAs, and their spatial locations. Using this dataset, we found that genetic copy number alterations (CNAs), cell types compositions, and cellular neighborhoods did not predict MT. Instead, second order effects i.e. pairwise interactions, are highly predictive of future transformation. First, we identified abnormal chromosomal contact patterns that clearly separate future stable versus future MT samples. Second, we identified 24 ligand-receptors (LR) pairs cross-expressed in neighboring cells as the main molecular factors predictive of transformation and recurrences. We then validated a cross-expressing pair of ENPP2-LPAR1 interactions with a separate cohort of patient samples. In addition, using the LR+ cell pairs as an anchor, we identified cell signaling-specific gene expression programs that can predict from bulk or single cell RNAseq data the time to recurrence. We used a cell-interaction-based foundation model (CIFM) optimized on the spatial RNA data in forward simulations and identified potential myeloid signaling factors involved in MT. Lastly, we analyzed the effect of detection sensitivity on the ability to capture pertinent LR+ neighboring cells by down-sampling transcript and showed that the ability to detect cross-expressing signaling LR transcripts (typically <10 copies per cell) decays rapidly with lower sensitivity, but is more robust to down-sampling of the areas of the tissue imaged. The importance of second-order features suggests that increased depth and dimensionality of data on a smaller quantity of samples can provide valuable insight, and that high-sensitivity and multiple-modalities spatial approaches can help identify markers to risk-stratify patients, aid in therapeutic decision making, and uncover potential therapeutic targets.

cancer biology↗

bioRxiv2026

Oral administration of dibenzoylmethane (DBM) prevents cognitive decline in a C9ORF72-mediated FTD mouse model

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are two related neurodegenerative disorders that display overlapping features. The hexanucleotide repeat expansion GGGGCC (G4C2) in the C9ORF72 gene is the most common cause of ALS and FTD, which results in the accumulation of dipeptide-repeat protein aggregates. Regulation of protein synthesis at the level of the initiation factor eIF2 has been suggested as a transversal event contributing to neurodegeneration in ALS and FTD. eIF2 phosphorylation blocks protein synthesis to alleviate protein misfolding overload, but conversely it can reduce the expression of synaptic proteins resulting in neuronal dysfunction. Dibenzoylmethane (DBM) is a small molecule that reverses the translational attenuation mediated by eIF2 phosphorylation which has been shown to alleviate neurodegeneration in prion-infected mice and Tau transgenic animals. Here we investigated the efficacy of the oral administration of DBM in protecting a mouse model of C9ORF72 pathogenesis. Treatment of mice with 0.5% of DBM mixture in powdered food ad libitum was sufficient to prevent cognitive impairment in C9ORF72 mice. Unexpectedly, DBM treatment did not modify the content of poly(GA) and poly(GR) protein inclusion in the hippocampus and brain cortex. Proteomic profiling of brain tissue indicated that DBM administration corrected nearly 70% of the changes in gene expression triggered by expanded G4C2, where the main pathways modified by DBM were related to cytoskeleton organization, ALS, and metabolic processes. Most proteins corrected by DBM in our C9ORF72 model were also altered in the brain of human FTD/ALS patients. Overall, our results reinforce the idea that targeting protein synthesis with small molecules in patients carrying C9ORF72 mutations may result in improved cognitive capacity.

molecular biology↗

bioRxiv2026

Multilayered specificity of transcription factor binding at cytokine promoters

Transcription factors (TFs) regulate gene expression through sequence-specific DNA binding, and their genomic occupancy is further influenced by TF expression, activation state, and protein-protein interactions. How these mechanisms determine context-specific gene regulation remains incompletely understood, particularly for tightly controlled immune genes such as cytokines. Here, we use paired yeast one-hybrid (pY1H) assays to systematically examine DNA binding of 236 TFs and 392 TF-pairs across 106 cytokine gene promoters. Of the 1,619 TF-promoter interactions identified, 555 required TF cooperativity and 410 were antagonized by at least one TF partner, suggesting that TF-DNA binding is highly dependent on TF partners. Usage of different partners can drastically alter a TFs target repertoire and may result in the recruitment of different transcriptional cofactors. Integration with existing data on TF expression and activation further showed that cooperativity and antagonism provide additional, underappreciated layers of DNA-binding specificity.

systems biology↗

bioRxiv2026

CD8 T cells with classical and NK-like cytotoxic gene expression programs mediate control of HBV replication and functional cure

Chronic hepatitis B is characterized by a decades-long evolving engagement between host immunity and the hepatitis B virus (HBV). Understanding the molecular characteristics of HBV-specific CD8 T cells linked to control of viral replication and antigenemia is essential to design effective immunotherapeutic modalities. Here we show that HBV-specific CD8 T cells, even during infection stages with extremely high viral loads, lack the features of terminally exhausted CD8 T cells observed in chronic HCV and HIV infection or cancer. Instead, we observe emerging gene expression programs over disease stages that correlate with increasing HBV control, which include a bona fide cytotoxic and T-cell localization program associated with low levels of viral replication, and a second NK-like T-cell program that combines expression of classical NK markers (KIRs, KLRs, FCGR3A, TYROBP, IKZF2) with cytotoxic genes (GZMB, GNLY, PRF1), which emerges with complete control of HBV viremia and antigenemia. We also found enrichment of both CD8 T-cell programs in HIV-specific CD8 T cells from HIV elite controllers, supporting a conserved role in controlling persistent viral infections with viral reservoirs.

immunology↗

bioRxiv2026

Immune Aging is an Independent Risk Factor for Cardiovascular Disease

Cardiovascular disease remains the leading cause of mortality, yet current clinical predictors miss substantial disease-risk. While the immune system contributes to this residual risk, its complexity has hindered broadly applicable, clinically scalable metrics of immune-state. Here, we establish the prognostic relevance of IMM-AGE, a system-level metric of immune-aging, to cardiovascular disease. We learn reference-free, low-dimensional representations of IMM-AGE across cell, protein, and mRNA measurements, enabling high-fidelity quantification across modalities, blood fractions, and platforms, including standard hospital flow cytometers. Among UK-Biobank participants, 56.9% of IMM-AGE variation remained unexplained by routine clinical measures, and across diverse cohorts totaling ~48,000 individuals, elevated IMM-AGE was independently associated with future cardiovascular risk, intervention outcomes, and mortality. Moreover, incorporation of IMM-AGE into the PREVENT 10-year risk equation significantly improved risk stratification. These findings establish immune-aging as an independent biological dimension of cardiovascular disease-risk and support IMM-AGE as a practical tool for precision risk assessment.

immunology↗

bioRxiv2026

Comprehensive Analysis Reveals Adaptive DNA Repair and Replication Stress Networks in Genomically Unstable Breast Cancer

Genomic instability is a defining hallmark of breast cancer, yet the mechanisms by which tumors tolerate persistent DNA damage remain poorly understood. We performed a comprehensive, multi-cohort analysis of breast cancer datasets to define how DNA damage response (DDR) and replication stress tolerance (RST) networks are rewired in genomically unstable tumors. Using fraction of genome altered (FGA) as a chromosomal instability metric, we show that BRCA-mutant tumors exhibit elevated genomic instability coupled with increased expression of homologous recombination, Fanconi anemia, mismatch repair, base excision repair, and alternative end-joining pathways. Strikingly, heightened pathway activity correlates with increased genome alteration, supporting a model of damage tolerance rather than repair restoration. RST programs, including fork remodeling, protection, and single strand DNA gap suppression, further contribute to tumor fitness under replication stress. These adaptive states are enriched in aggressive subtypes, intensified with progression, and associate with pathway-specific mutational burden. Co-occurrence and mutual exclusivity mapping uncovered non-random subtype-relevant genetic interactions states among major drivers and DDR genes, nominating context-specific synthetic lethal opportunities. Our findings identify compensatory genome-maintenance programs as central drivers of tumor resilience and highlight pathway-specific vulnerabilities for targeted therapeutic intervention.

cancer biology↗

bioRxiv2026

scPlOver: inferring DNA content from amplification-free single-cell WGS using fragment overlaps

Correctly inferring copy-number aberrations from single-cell DNA sequencing data requires estimating cellular DNA content, which is unidentifiable from read counts alone. In tagmentation-based sequencing, each fragment represents a distinct DNA molecule, thus fragment overlaps provide an orthogonal signal for copy number. We present a theoretical model of fragment overlaps as a function of copy number and coverage and introduce scPlOver, a method that uses this model to infer DNA content. scPlOver outperforms existing approaches on simulated and experimental datasets and identifies thousands of ovarian cancer cells with higher DNA content than previously estimated across a cohort of 41 patients.

genomics↗

bioRxiv2026

Spatially Distinct Macrophage Subsets Drive Myofibroblast Heterogeneity and Maladaptive Fibrosis in Lupus Nephritis

ObjectivesLupus nephritis (LN) is a severe complication of systemic lupus erythematosus (SLE), leading to progressive renal fibrosis and functional decline. Understanding the interplay between immune cells and stromal cells is needed to develop effective therapeutic strategies. Here, we investigated the landscape of macrophage-fibroblast interactions in human LN and validated these findings in mouse models. MethodsWe characterized distinct fibroblast subsets and their interactions with renal macrophages using single-cell RNA sequencing (scRNAseq) of 156 human LN biopsies and 30 healthy controls from the AMP-SLE cohort, and spatial transcriptomics of biopsies from 6 LN patients. In vitro co-culture studies using mouse models were performed to further define functional consequences of these interactions. ResultsWe identified two myofibroblast subsets: a pro-inflammatory subset (Myofib1) enriched in the tubulointerstitium, and a fibrotic/remodeling subset (Myofib2) in glomeruli, both correlating with the histologic chronicity index. Spatial transcriptomics revealed different colocalization patterns, with Myofib1 interacting with activated resident macrophage (RM) subsets and Myofib2 with glomerular infiltrating disease-associated macrophages. In vitro co-culture studies demonstrated that nephritic RMs promote a pro-inflammatory, remodeling fibroblast phenotype that impairs wound healing and drives a Myofib1-like gene program, whereas disease-associated macrophages generated profibrotic fibroblasts with dysregulated reparative capacity. Cell-cell communication analyses identified key ligand-receptor interactions mediating this crosstalk, including Spp1/integrins, Sema4/PlexinB, and NAMPT/INSR. ConclusionsOur data reveal a spatially and functionally heterogeneous landscape of macrophage-fibroblast crosstalk in LN. These findings advance our understanding of renal fibrogenesis in LN, highlighting specific fibro-inflammatory circuits that may represent therapeutic targets to prevent chronic renal damage.

immunology↗

bioRxiv2026

High-pH NMR to Identify Macromolecular Hydrogen-Bonds and Foldons

Hydrogen bond (H-bond) restraints are critical for NMR structure determination, yet their experimental identification can be challenging for marginally stable structures that afford insufficient protection from (H/D) exchange in D2O. As an alternative, we explored the use of NMR between pH 10 and 11 conditions that promote rapid exchange, for identifying backbone amide protons involved in H-bonds. We analyzed [~]750 amide sites distributed across ten proteins with known structures. Persistence of amide protons at high pH in standard 2D 1H-15N HSQC spectra for 15N-labeled proteins in H2O, or TOCSY for unlabeled proteins, identifies H-bonds with [~]91% accuracy that exceeds the [~]80% accuracy of traditional H/D exchange experiments in D2O. For two -helical coiled coils and three globular proteins, we performed alkaline unfolding experiments taking advantage of amide NMR signal attenuation from unstructured polypeptides. Increasing the sample pH led to a progressive loss of native amide proton NMR signals, revealing an unfolding hierarchy where "foldons" remaining at the highest pH values had the most persistent H-bonds under EX1 exchange conditions. The foldons observed at high pH are consistent with partially folded structures previously characterized near neutral pH by native state hydrogen exchange, equilibrium unfolding, and protein fragment studies. For {beta}-sheet proteins, foldons correspond to regions with high inter-residue contact density, whereas in coiled coils they demarcate regions with high -helical propensity. High-pH NMR experiments provide a sensitive, fast, inexpensive, and broadly applicable approach to map H-bonding in marginally stable or partially folded proteins. Additionally, they offer the opportunity to explore uncharted protein dynamics and unfolding pathways under basic pH conditions.

biophysics↗

bioRxiv2026

AI/ML-Assisted Computational Design and Immunoinformatics Evaluation of a Multi-Epitope Vaccine Targeting Podoplanin in Glioblastoma Multiforme

Glioblastoma Multiforme (GBM) is one of the most malignant forms of brain tumor in humans, with limited treatment options and poor overall survival rates. In the present study, we employed an in-silico workflow that integrated immunoinformatics and 3D structural modelling tools to design a multi-epitope vaccine against Podoplanin (PDPN), a transmembrane glycoprotein primarily involved in tumor invasion and metastasis. The differential expression of PDPN in tumor versus normal cells was investigated using transcriptomics datasets. Once the overexpression was confirmed, it was designated as a Tumor-Associated Antigen (TAA). B-cell, CTL, and HTL epitopes were predicted and screened for antigenicity, non-allergenicity, and non-toxicity. Selected epitopes were linked with appropriate adjuvant and linker sequences to construct a vaccine candidate. Codon optimization and in silico cloning was conducted to evaluate the constructs expression in a mammalian expression vector. The 3D structure of the vaccine candidate was modelled, refined, and validated before molecular docking with immune receptors and immune simulation studies. The results indicated that proposed polypeptide, RasIC-01v, could be a potential vaccine candidate for highly vigorous and dangerous cancer like GBM. Further experimental and immunological validations would be required to validate the commercial feasibility and development of RasIC-01v. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=116 SRC="FIGDIR/small/706629v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@7485b1org.highwire.dtl.DTLVardef@1f551c1org.highwire.dtl.DTLVardef@ca871eorg.highwire.dtl.DTLVardef@6cf53d_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2026

Combination Drug Therapy Reduces Iron Accumulation and Microglia-Mediated Pathologies in Neonatal Intraventricular Hemorrhage: A Biochemical and Transcriptomic Analysis

This study describes the distribution of non-reactive brain-resident microglia densely populated along the borders of the lateral ventricles and choroid plexus in premature rabbit pups during early forebrain development. Following intraventricular hemorrhage (IVH), microglia become activated, proliferate, and migrate deeper into parenchymal regions. During this process, activated microglia exhibit a global expansion with a disproportionally elevated proinflammatory M1 nomenclature phenotype from 25% to 50% of the total; that shift was reduced by sulforaphane (SFN; Nrf2-antioxidant response element [ARE] activator of anti-inflammatory pathways) plus deferoxamine (DFN; iron chelator) treatment. Transcriptome analysis identified over expression of pro-inflammatory calcium-binding proteins S100A8 and S100A12 (intracellular damage signals), as well as chemokines CXCL8 and CXCL10 by neurons and microglia. The combination treatment of SFN-DFN mitigated M1 infiltration, suppressed the magnitude of inflammation and reduced ferroptosis after IVH in the developing postnatal brain. Moreover, SFN-DFN treatment reversed most dysregulated genes in inflammation and iron homeostasis networks, revealing potential molecular targets for additional pharmacologic interventions after IVH. We propose that reducing the toxic microcellular environment will attenuate both the injurious inflammatory responses and improve recovery of the trajectory toward normal brain development. Additionally, suppression of proinflammatory molecules and iron toxicity should promote better survival as well as salutary effects of "living stem cell therapy" as we have previously shown.

neuroscience↗

bioRxiv2026

Machine Learning Identifies Distinct Treg-Mediated Remodeling in HFpEF Hearts Treated with Neonatal Mesenchymal Stem Cells and Their Secretome

BackgroundHeart failure with preserved ejection fraction (HFpEF) remains a major therapeutic challenge due to its complex pathophysiology and pronounced heterogeneity. Regenerative approaches using neonatal mesenchymal stromal cells (nMSCs) and their secretome (SEC) have shown promise in other heart failure contexts. ObjectivesHowever, the effect of these therapies in HFpEF, and the underlying molecular mechanisms and causal pathways remain poorly understood. MethodsHFpEF was established in two distinct murine models, followed by treatment with either nMSCs or SEC. Functional and histological endpoints were assessed. We developed a novel machine learning framework, VIPcell, which integrates data augmentation, Partial Least Squares (PLS) regression, and causal structure inference to identify genes causally linked to cardiac function using single-nucleus RNA sequencing (snRNA-seq) data. VIPcell was applied to heart tissues from treated HFpEF animals to uncover key regulators of cardiac remodeling. ResultsBoth nMSC and SEC therapies significantly improved diastolic function in two independent rodent HFpEF models. These improvements were associated with reduced inflammation, attenuated myocardial fibrosis, and improved exercise capacity. Intercellular communication analysis revealed widespread, system-level signaling in nMSC-treated hearts, compared to more localized endothelial-cardiomyocyte crosstalk in SEC-treated hearts. Causal inference via VIPcell suggested overlapping upstream regulators in both treatment groups, particularly genes involved in regulatory T cell (Treg) biology and immunomodulatory signaling pathways, including FOXO signaling, NLRP3 inflammasome inhibition, and Tie2 activation. In vivo validation confirmed selective expansion of Tregs following nMSC and SEC therapy. In vitro, nMSCs induced significantly greater Treg expansion compared to multiple adult stem cell types. Critically, chemical depletion of Tregs abrogated the therapeutic effects of both treatments, establishing Tregs as central mediators of diastolic function recovery in the HFpEF preclinical model. ConclusionsnMSC and SEC therapies improve diastolic function in HFpEF through distinct remodeling mechanisms converging on Treg-mediated immune modulation. VIPcell supported identification of causal regulators, highlighting Treg-related signaling as a key driver of myocardial recovery in HFpEF. These findings offer mechanistic insight into cellular therapies for HFpEF and support the development of targeted, Treg-focused interventions.

physiology↗