Biology subjects

Alpern, D.

Publications and source records attributed to Alpern, D..

11 recordsLinked to original sources

bioRxiv2026

Growth factor-free synthetic matrices reveal intrinsic skeletal progenitor competence

Clinical translation of human skeletal progenitor cells has been limited by the inability to reliably predict their regenerative performance, as current characterization methods rely on phenotypic markers or instructive differentiation assays that obscure intrinsic cell function. Here we demonstrate that human skeletal progenitor competence can only be revealed within a precisely defined and growth factor-free synthetic microenvironment. By independently tuning matrix mechanics, degradability, adhesion, and cell density within a fully synthetic poly(ethylene glycol) hydrogel, we create a permissive niche that enables human bone marrow stromal cells (hBMSCs) to undergo spontaneous bone formation in vivo without osteoinductive supplementation. These exogenous growth factor-free conditions expose donor-specific differences in bone forming capacity that are otherwise masked. Strikingly, supplementation with even low concentrations of BMP-2 abolished inter-donor variability, directly demonstrating that osteoinductive factors override intrinsic progenitor competence. Single-cell transcriptomics, proteomic profiling, and longitudinal in vivo analyses converge to show that high performing donors are defined by chondrocyte-primed transcriptional states and coordinated extracellular matrix (ECM) remodeling programs that establish a pro-osteochondral niche and drive bone formation via endochondral ossification. In contrast, low performing donors fail to initiate this cascade despite comparable phenotypic profiles and proliferation rates. Together, these findings implicate matrix context as a critical regulator of stem cell competence. Within this permissive synthetic niche, intrinsic ECM remodeling capacity emerges as a defining feature of donor-specific osteogenic potential, one that remains invisible to conventional assays. These matrices thus provide a functional platform for prospective stratification of skeletal progenitor cells, with direct implications for donor selection in regenerative medicine. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/731042v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@6fa9ceorg.highwire.dtl.DTLVardef@f51dddorg.highwire.dtl.DTLVardef@7bb040org.highwire.dtl.DTLVardef@706a36_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

Interdependent Dynamics of mRNA Expression and HIV-1 Viral Load: Insights from Transcriptomics and Mendelian Randomization

The interaction between HIV-1 and the host immune system plays a crucial role in the natural control and progression of the infection. Previous studies have identified APOBEC3G, Tetherin, SAMHD1, and SERINC5 as HIV-1 host restriction factors, which are counteracted by the viral proteins Vif, Vpu, and Nef, respectively. The blood expression levels of some of these host proteins are correlated with HIV load, suggesting that interindividual differences in the spontaneous control of HIV infection might lead to the identification of novel HIV restriction factors. Our study enrolled 150 participants from the Swiss HIV Cohort Study with human genome-wide genotyping data, pre-antiretroviral treatment peripheral blood mononuclear cells (PBMC) aliquots and HIV load measurements. Using BrB-seq, we quantified mRNA expression of all protein-coding genes and found significant associations between 792 genes and HIV load. Pathway analysis revealed that higher viral load associated with the upregulation of innate immune response, proteasome complex, mitochondrial and cell-cycle related activity, and with the downregulation of ribosomal transcripts and genes involved in cytokine-cytokine receptor interaction, including IL4R, IL7R, and TCF7. Mendelian Randomization confirmed the viral restriction activity of TRABD2A and identified new candidates as potential restriction factors. These findings provide novel insights into the host-virus interplay and suggest additional genes that may contribute to the natural control of HIV-1 infection.

immunology↗

bioRxiv2023

Dissection and reconstruction of the colorectal cancer tumor microenvironment

Patient-derived organoids (PDOs) are the reference in vitro human disease models. However, the utility of colorectal cancer (CRC) PDOs is hindered by the lack of a tumor microenvironment (TME). To address this limitation, we built a living biobank of CRC PDOs with autologous stromal and immune TME. We characterized the original tumors and traditional monocultures using single-cell RNA-seq (scRNA-seq) and whole exome sequencing (WES) to obtain insights into cell type selection and phenotypic drift in culture. Subsequently, we developed culture conditions supporting all cell types to recapitulate the CRC-TME around PDOs. From the transcriptomes of >180k cells obtained from 260 such co-cultures, we illuminated the mutual influence of cells within CRC tumors. Based on original tumor data, atlases of predicted interactions and transcriptional networks elucidated why monocultures were altered and suggested that TME reconstruction more accurately reflected original tumor behavior. We found that inflammatory signals were absent in vitro and recovered upon co-culture with tumor-infiltrating lymphocytes (TILs). We also functionally confirmed that stromal, not cancer cells, mediated immune evasion. Additionally, stroma induced an invasive phenotype in cancer cells. From this deep dive into CRC-TME interactions, we built the human CRC-TME atlas (https://crc-tme.com/), an online portal for interactive exploration of gene expression data, prediction of cell-cell interactions at the pathway and receptor/ligand levels, transcriptional networks, and more. We anticipate PDO cultures with reconstructed TMEs will be valuable for discovery efforts, preclinical studies, and personalized medicine, with the atlas as a framework and inspiration for future CRC-TME studies.

cancer biology↗

bioRxiv2023

Transcriptomic profiling of murine GnRH neurons reveals developmental trajectories linked to human reproduction

Gonadotropin-releasing hormone (GnRH) neurons play a crucial role in human reproduction and are associated with a spectrum of conditions. However, the underlying biological mechanisms remain elusive due to their small number and sparse distribution. We performed transcriptomic profiling of GnRH neurons during mouse embryonic development, revealing their molecular identity and gene expression dynamics. Our findings show that GnRH neurons undergo a profound transcriptional shift as they migrate from the nose to the brain and that distinct expression trajectories are associated with critical biological processes, including cell migration, neuronal projections, and synapse formation. Cell-to-cell communication analysis revealed timely and spatially restricted modulation of signaling pathways involving known molecules, such as Semaphorins and Plexins, and novel candidates, such as Neurexins and Endothelins. Using GWAS genes linked to human reproductive onset, we found a specific association with GnRH neuron trajectories rising in late developmental stages and involved in neuron maturation and connectivity. Finally, analysis of the genetic burden in a large cohort of patients with congenital GnRH deficiency revealed specific GnRH neuron trajectories with a significant mutation load compared to controls. In conclusion, this study revealed the gene expression dynamics underlying GnRH neuron embryonic development and provides novel insights linking GnRH neuron biology to human reproduction.

developmental biology↗

bioRxiv2023

A human omentum-specific mesothelial-like stromal population inhibits adipogenesis through IGFBP2 secretion

Adipose tissue plasticity is orchestrated by molecularly and functionally diverse cells within the stromal vascular fraction (SVF). While several mouse and human adipose SVF cellular subpopulations have now been identified, we still lack an understanding of the cellular and functional variability of adipose stem and progenitor cell (ASPC) populations across human fat depots. To address this, we performed single-cell and bulk RNA-seq analyses of >30 Lin-SVF samples across four human adipose depots, revealing two ubiquitous hASPC subpopulations with distinct proliferative and adipogenic properties but also depot- and BMI-dependent proportions. Furthermore, we identified an omental-specific, high IGFBP2- expressing stromal population that transitions between mesothelial and mesenchymal cell states and inhibits hASPC adipogenesis through IGFBP2 secretion. Our analyses highlight the molecular and cellular uniqueness of different adipose niches while our discovery of an anti-adipogenic IGFBP2+ omental-specific population provides a new rationale for the biomedically relevant, limited adipogenic capacity of omental hASPCs.

cell biology↗

bioRxiv2022

Neuro-toxicogenomic mapping of TMT induced neurotoxicity using human minibrain reveals associated adverse outcome events

Genome-wide transcriptomic interrogation of organoids and 3D tissue models are increasingly used for characterizing drug, toxicity responses and neurodevelopmental disorders. We established here a neuro-toxicogenomic assay by utilizing "minibrain", a human in vitro 3D brain model system and a low-cost, highly multiplexable RNA-seq methodology (BRB-seq) for screening the effect of trimethyltin chloride (TMT) induced neurotoxicity. We demonstrate that transcriptomic profiling is insightful to the cellular composition and regional identity of the minibrain. Further, we characterize the transcriptomic changes associated with the dose-time neurotoxic response of minibrain upon exposure to TMT. The distinct gene expression changes and molecular candidates identified with our pipeline provides insight to map the key events involved in the adverse outcome pathway of TMT associated neurotoxicity. We identify processes such as endoplasmic reticulum stress, dysregulation of synaptic genes and downregulation of neuron-morphology associated genes upon exposure to TMT. In response to TMT, we identify activation of an early response homeostatic mechanism in minibrain and an interplay of STAT pathways correlating with the dose severity. In this study, we present a neuro-toxicogenomic assay that demonstrates the power of a low-cost transcriptomic screening to study chemical induced neurotoxicity.

neuroscience↗

bioRxiv2021

Mammalian adipogenesis regulators (Aregs) exhibit robust non- and anti-adipogenic properties that arise with age and involve retinoic acid signalling

Adipose stem and precursor cells (ASPCs) give rise to adipocytes and determine the composition and plasticity of adipose tissue. Recently, several studies have demonstrated that ASPCs partition into at least three distinct cell subpopulations: Dpp4+ stem-like cells, Aoc3+ pre-adipocyte-like cells, and the enigmatic CD142+ cells. A great challenge now is to functionally characterize these distinct ASPC populations. Here, we focus on CD142+ ASPCs since discrepant properties have been assigned to this subpopulation, from adipogenic to non- and even anti-adipogenic. To address these inconsistencies, we comprehensively characterized mammalian subcutaneous CD142+ ASPCs across various sampling conditions. Our findings demonstrate that CD142+ ASPCs exhibit high molecular and phenotypic robustness, firmly supporting their non- and anti-adipogenic properties. However, these properties emerge in an age-dependent manner, revealing surprising temporal CD142+ ASPC behavioural alterations. Finally, using multi-omic and functional assays, we show that the inhibitory nature of these adipogenesis-regulatory CD142+ ASPCs (Aregs) is driven by specifically expressed secretory factors that cooperate with the retinoic acid signalling pathway to transform the adipogenic state of CD142- ASPCs into a non-adipogenic, Areg-like one.

cell biology↗

bioRxiv2020

The Hippo pathway controls myofibril assembly and muscle fiber growth by regulating sarcomeric gene expression

Skeletal muscles are composed of gigantic cells called muscle fibers, packed with force-producing myofibrils. During development the size of individual muscle fibers must dramatically enlarge to match with skeletal growth. How muscle growth is coordinated with growth of the contractile apparatus is not understood. Here, we use the large Drosophila flight muscles to mechanistically decipher how muscle fiber growth is controlled. We find that regulated activity of core members of the Hippo pathway is required to support flight muscle growth. Interestingly, we identify Dlg5 and Slmap as regulators of the STRIPAK phosphatase, which negatively regulates Hippo to enable post-mitotic muscle growth. Mechanistically, we show that the Hippo pathway controls timing and levels of sarcomeric gene expression during development and thus regulates the key components that physically mediate muscle growth. Since Dlg5, STRIPAK and the Hippo pathway are conserved a similar mechanism may contribute to muscle or cardiomyocyte growth in humans.

developmental biology↗

bioRxiv2020

Disparate temperature-dependent virus - host dynamics for SARS-CoV-2 and SARS-CoV in the human respiratory epithelium

Since its emergence in December 2019, SARS-CoV-2 has spread globally and become a major public health burden. Despite its close phylogenetic relationship to SARS-CoV, SARS-CoV-2 exhibits increased human-to-human transmission dynamics, likely due to efficient early replication in the upper respiratory epithelium of infected individuals. Since different temperatures encountered in the human respiratory tract have been shown to affect the replication kinetics of several viruses, as well as host immune response dynamics, we investigated the impact of temperatures during SARS-CoV-2 and SARS-CoV infection in the human airway epithelial cell culture model. SARS-CoV-2, in contrast to SARS-CoV, replicated more efficiently at temperatures encountered in the upper respiratory tract, and displayed higher sensitivity to type I and type III IFNs. Time-resolved transcriptome analysis highlighted a temperature-dependent and virus-specific induction of the IFN-mediated antiviral response. These data reflect clinical features of SARS-CoV-2 and SARS-CoV, as well as their associated transmission efficiencies, and provide crucial insight on pivotal virus - host interaction dynamics.

microbiology↗

bioRxiv2019

A highly parallel, automated platform enabling individual or sequential ChIP of histone marks and transcription factors

Despite its popularity, chromatin immunoprecipitation followed by sequencing (ChIP-seq) remains a tedious (>2d), manually intensive, low-sensitivity and low-throughput approach. Here, we combine principles of microengineering, surface chemistry and molecular biology to address the major limitations of standard ChIP-seq. The resulting approach, FloChIP, automates and miniaturizes ChIP in a beadless fashion while facilitating the downstream library preparation process through on-chip chromatin tagmentation. FloChIP is fast (<2h), has a wide dynamic range (from 106 to 500 cells), is high-throughput (up to 64 parallel, antibody- or sample-multiplexed experiments) and is compatible with both histone mark and transcription factor ChIP. In addition, FloChIPs interconnected design allows for straightforward chromatin re-immunoprecipitation, thus constituting the first example of a microfluidic sequential ChIP-seq system. Finally, we demonstrate FloChIPs high-throughput capacity by performing ChIP-seq of the transcription factor MEF2A in 32 distinct human lymphoblastoid cell lines, providing novel insights into the main factors driving collaborative DNA binding of MEF2A and into its role in B-cell-specific gene regulation. Together, our results validate FloChIP as a flexible and reproducible automated solution for individual or sequential ChIP-seq.

bioengineering↗

bioRxiv2018

Time- and cost-efficient high-throughput transcriptomics enabled by Bulk RNA Barcoding and sequencing

Genome-wide gene expression analyses by RNA sequencing (RNA-seq) have quickly become a standard in molecular biology because of the widespread availability of high throughput sequencing technologies. While powerful, RNA-seq still has several limitations, including the time and cost of library preparation, which makes it difficult to profile many samples simultaneously. To deal with these constraints, the single-cell transcriptomics field has implemented the early multiplexing principle, making the library preparation of hundreds of samples (cells) markedly more affordable. However, the current standard methods for bulk transcriptomics (such as TruSeq Stranded mRNA) remain expensive, and relatively little effort has been invested to develop cheaper, but equally robust methods. Here, we present a novel approach, Bulk RNA Barcoding and sequencing (BRB-seq), that combines the multiplexing-driven cost-effectiveness of a single-cell RNA-seq workflow with the performance of a bulk RNA-seq procedure. BRB-seq produces 3 enriched cDNA libraries that exhibit similar gene expression quantification to TruSeq and that maintain this quality, also in terms of number of detected differentially expressed genes, even with low quality RNA samples. We show that BRB-seq is about 25 times less expensive than TruSeq, enabling the generation of ready to sequence libraries for up to 192 samples in a day with only 2 hours of hands-on time. We conclude that BRB-seq constitutes a powerful alternative to TruSeq as a standard bulk RNA-seq approach. Moreover, we anticipate that this novel method will eventually replace RT-qPCR-based gene expression screens given its capacity to generate genome-wide transcriptomic data at a cost that is comparable to profiling 4 genes using RT-qPCR.\n\n SoftwareWe developed a suite of open source tools (BRB-seqTools) to aid with processing BRB-seq data and generating count matrices that are used for further analyses. This suite can perform demultiplexing, generate count/UMI matrices and trim BRB-seq constructs and is freely available at http://github.com/DeplanckeLab/BRB-seqTools\n\nHighlightsO_LIRapid (~2h hands on time) and low-cost approach to perform transcriptomics on hundreds of RNA samples\nC_LIO_LIStrand specificity preserved\nC_LIO_LIPerformance: number of detected genes is equal to Illumina TruSeq Stranded mRNA at same sequencing depth\nC_LIO_LIHigh capacity: low cost allows increasing the number of biological replicates\nC_LIO_LIProduces reliable data even with low quality RNA samples (down to RIN value = 2)\nC_LIO_LIComplete user-friendly sequencing data pre-processing and analysis pipeline allowing result acquisition in a day\nC_LI

genomics↗