Biology subjects

Ussar, S.

Publications and source records attributed to Ussar, S..

8 recordsLinked to original sources

bioRxiv2025

DISCO-seq: 3D single-cell transcriptomics of intact biological systems

Single-cell transcriptomics has transformed tissue analysis, yet current methods struggle to integrate whole-tissue 3D architecture. Conventional techniques restrict molecular profiling to pre-selected 2D sections, losing systemic context and introducing anatomical bias by sampling less than 0.001% of a whole organism. To overcome these challenges, we developed DISCO-seq, a tissue-clearing chemistry that enables superior RNA accessibility compared to fresh or fixed tissues. DISCO-seq integrates whole-organ or organism 3D imaging with both untargeted and targeted transcriptomics, yielding high-quality RNA from cleared tissues comparable to standard samples. We demonstrate its versatility by investigating tumor heterogeneity in a syngeneic glioblastoma mouse model, using 3D imaging to identify spatially distinct microenvironments and characterize their unique transcriptomic signatures. Moreover, DISCO-seq enabled unbiased, whole-body mapping of SARS-CoV-2 S1 protein deposition in mice, followed by transcriptomic profiling of spatially defined niches. By bridgingmesoscale 3D imaging with single-cell transcriptomics, DISCO-seq establishes a paradigm for anatomically contextualized, hypothesis-free tissue interrogation. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/693352v1_ufig1.gif" ALT="Figure 1"> View larger version (69K): org.highwire.dtl.DTLVardef@758ee5org.highwire.dtl.DTLVardef@1f862c6org.highwire.dtl.DTLVardef@1cf3b9org.highwire.dtl.DTLVardef@c5082e_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIDISCO-seq integrates RNA-preserving tissue-clearing chemistry with whole-organ or organism 3D imaging, enabling anatomically unbiased single-cell transcriptomics. C_LIO_LIDISCO-seq yields RNA quality and transcriptome profiles equivalent to those obtained from matched fresh or fixed tissues. C_LIO_LIDISCO-seq identifies discrete glioblastoma microenvironments and defines their transcriptomic states within the intact brain. C_LIO_LIDISCO-seq enables whole-body mapping of SARS-CoV-2 S1 and uncovers region-specific immune and metabolic responses across anatomical niches. C_LI Supplementary movies can be seen at: http://discotechnologies.org/DISCO-seq/

systems biology↗

bioRxiv2025

LipiGo: A Versatile DNA-Lipid Nanoparticle Hybrid for Precision Drug Delivery

Lipid nanoparticles (LNPs) are established carriers for nucleic acid delivery, however, achieving efficient delivery to non-hepatic tissues remains a major challenge. Here, we present LipiGo, a nanocarrier platform engineered by integrating short single stranded DNA molecules into the lipid nanoparticle structure. Using whole body tissue clearing, advanced imaging and AI-based analysis, we show that LipiGo redirects functional mRNA delivery to lymphoid organs, particularly the spleen. Immune cell profiling further reveals enhanced uptake within key immune populations, including antigen-presenting cells, compared to standard LNPs. Beyond passive redistribution, LipiGo leverages DNA hybridization to enable modular attachment of targeting ligands for active targeting as demonstrated by cell-specific delivery to white adipocytes. Overall, the dual-purpose design principle of LipiGo demonstrates high modularity and efficiency, enabling tissue and cell specific delivery beyond hepatic applications. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/676334v2_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@9b69eorg.highwire.dtl.DTLVardef@1b45bd0org.highwire.dtl.DTLVardef@1f7b79dorg.highwire.dtl.DTLVardef@37a673_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

Igf2 regulates early postnatal DPP4+ preadipocyte pool expansion

Adipose tissue is rapidly expanding early in life. Identifying the queues facilitating this process will advance our understanding of metabolically healthy obesity. Single-cell RNA sequencing revealed compositional differences between pre-wean and adult murine subcutaneous adipose tissue. We identify a dipeptidyl peptidase-4 (Dpp4) positive precursor population in pre-wean mice residing at the reticular interstitium of subcutaneous adipose tissue expressing insulin growth factor 2 (Igf2). We show that IGF2 drives proliferation rather than differentiation in these cells. Moreover, loss of Igf2 in Dpp4+ progenitor cells promotes adipogenesis. We show that expression of Igf2 at a limited timeframe during development promotes preadipocyte expansion.

developmental biology↗

bioRxiv2022

Hypercaloric diet triggers transient molecular rearrangements of astrocytes selectively in the arcuate nucleus

Hypothalamic astrocytes are particularly affected by energy-dense food consumption. How the anatomical location of these glial cells and their spatial molecular distribution in the arcuate nucleus of the hypothalamus (ARC) determine the cellular response to a high caloric diet remains unclear. In this study, we investigated their distinctive molecular responses following the exposure to a high-fat high-sugar (HFHS) diet, specifically in the ARC. Using RNA sequencing and proteomics, we showed that astrocytes have a distinct transcriptomic and proteomic profile dependent on their anatomical location, with a major proteomic reprogramming in hypothalamic astrocytes. By ARC single-cell sequencing, we observed that a HFHS diet dictates time- and cell-specific transcriptomic responses, revealing that astrocytes have the most distinct regulatory pattern compared to other cell types. Lastly, we topographically and molecularly characterized astrocytes expressing glial fibrillary acidic protein and/or aldehyde dehydrogenase 1 family member L1 in the ARC, of which the abundance was significantly increased, as well as the alteration in their spatial and molecular profiles, with a HFHS diet. Together, our results provide a detailed multi-omics view on the spatial and temporal changes of astrocytes particularly in the ARC during different time points of adaptation to a high caloric diet.

neuroscience↗

bioRxiv2022

Single-cell characterization of neovascularization using hiPSC-derived endothelial cells in a 3D microenvironment

The formation of vascular structures is fundamental for in vitro tissue engineering. Vascularization can enable the nutrient supply within larger structures and increase transplantation efficiency, which are currently limiting factors in organoid research. We differentiated human induced pluripotent stem cells toward endothelial cells in 3D suspension culture. To investigate in vitro neovascularization and various 3D microenvironmental approaches, we designed a comprehensive single-cell transcriptomic study. Time-resolved single-cell transcriptomics of the endothelial and co-evolving mural cells gave insights into cell type development, stability, and plasticity. Transfer to a 3D hydrogel microenvironment induced neovascularization and facilitated tracing of sprouting, coalescing, and tubulogenic endothelial cells states. During maturation, we monitored two pericyte subtypes evolving of mural cells. Profiling cell-cell interactions between pericytes and endothelial cells confirmed in vivo angiogenic signaling and emphasized new cytokine signals during tubulogenesis. Our data, analyses, and results provide an in vitro roadmap to guide vascularization in future tissue engineering.

developmental biology↗

bioRxiv2020

Identification and characterization of distinct murine brown adipocyte lineages

Brown adipose tissue (BAT) plays an important role in the regulation of body weight and glucose homeostasis. While increasing evidence supports white adipose tissue heterogeneity, little is known about heterogeneity within murine BAT. Using single cell RNA sequencing of the stromal vascular fraction of murine BAT and analysis of 67 brown preadipocyte and adipocyte clones we unravel heterogeneity within brown preadipocytes. Statistical analysis of gene expression profiles from these clones identifies markers distinguishing brown adipocyte lineages. We confirm the presence of distinct brown adipocyte populations in vivo using three identified markers; Eif5, Tcf25, and Bin1. Functionally, we demonstrate that loss of Bin1 enhances UCP1 expression and mitochondrial respiration, suggesting that Bin1 marks a dormant brown adipocyte type. The existence of multiple brown adipocyte lineages suggests distinct functional properties of BAT depending on its cellular composition, with potentially distinct function in thermogenesis and the regulation of whole body energy homeostasis.

cell biology↗

bioRxiv2020

Active integrins regulate white adipose tissue insulin sensitivity and brown fat thermogenesis

Reorganization of the extracellular matrix is a prerequisite for healthy adipose tissue expansion, whereas fibrosis is a key feature of adipose dysfunction and inflammation. However, very little is known about the direct effects of impaired cell-matrix interaction in adipocyte function and insulin sensitivity. Using adipose selective deletion of {beta}1 integrin (Itgb1adipo-cre) and Kindlin-2 (Kind2adipo-cre), we demonstrate here that active {beta}1 and {beta}3 integrins directly interact with the insulin receptor to regulate white adipocyte insulin action and systemic metabolism. Consequently, loss of adipose integrin activity, similar to loss of adipose insulin receptors, results in lipodystrophy and systemic insulin resistance. Conversely, we find that brown adipose tissue of Kind2adipo-cre and Itgb1adipo-cre mice is chronically hyperactivated, and has increased substrate delivery, reduced endothelial basement membrane thickness, and increased endothelial vesicular transport. Thus, we establish integrin-extracellular matrix interactions as key regulators of white and brown adipose tissue function and whole body metabolism.

physiology↗

bioRxiv2020

PAT2 regulates autophagy through vATPase assembly and lysosomal acidification in brown adipocytes

Brown adipose tissue (BAT) plays a key role in maintaining body temperature as well as glucose and lipid homeostasis by its ability to dissipate energy through mitochondrial uncoupling. To facilitate these tasks BAT needs to adopt its thermogenic activity and substrate utilization to changes in nutrient availability, regulated by a complex network of neuronal, endocrine and nutritional inputs. Amongst this multitude of factors influencing BAT activity changes in the autophagic response of brown adipocytes are an important regulator of its thermogenic capacity and activity. Increasing evidence supports an important role of amino acid transporters in mTORC1 activation and the regulation of autophagy. However, a specific role of amino acid transporters in BAT regulating its function has not been described. Here we show that the brown adipocyte specific proton coupled amino acid transporter PAT2 rapidly translocates from the plasma membrane to the lysosome in response to amino acid withdrawal, where it facilitates the assembly of the lysosomal vATPase. Loss or overexpression of PAT2 therefore impair lysosomal acidification, autophagolysosome formation and starvation induced mTORC1 activation.

cell biology↗