Biology subjects

Reddy, S. K.

Publications and source records attributed to Reddy, S. K..

4 recordsLinked to original sources

bioRxiv2026

Genomic repeats for single-cell molecular recording

Genomic recording enables transient biological signals to be indelibly captured through DNA alterations, creating a permanent record of cellular history retrievable by sequencing. However, current methods are limited by scarce writing space, typically targeting only one or a few amenable genomic sites and requiring large cell populations for signal reconstruction. Here, we establish Repeats for Genomic Recording (RGRs): sequences with up to 400 copies targetable by a single CRISPR guide RNA, readable with a common primer pair, and predicted to have minimal functional impact. We demonstrate that RGRs enable both signal deconvolution in single cells and high-resolution recording in cell populations. Individual RGR sites exhibit distinct response kinetics; thus, combining them improves recording resolution beyond what redundancy alone provides, analogous to diversity reception in wireless communication. We develop a computational pipeline for systematic RGR identification, revealing 15,000 to 25,000 candidates per species across human, mouse, and zebrafish, thereby markedly expanding recording capacity and enabling cell-type-specific applications. Finally, we validate RGRs in live mice by recording long-term immediate early gene activity across the brain following epilepsy induction. This work establishes genomic repeats as a high-capacity platform for single-cell molecular recording in vivo.

synthetic biology↗

bioRxiv2026

Fibroblast signaling influences macrophage-dependent, biomaterial-induced tissue remodeling

The ability to induce tissue regeneration on demand using biomaterials remains a major goal in biomedical research, yet significant challenges persist. Among the most advanced biomaterial models, the nanofiber-hydrogel composite has demonstrated a striking ability to induce soft adipose tissue remodeling at the injection site without incorporating exogenous biological cues.1,2 However, the underlying mechanisms that drive such a tissue response remain unclear. Here, we show that biomaterial-induced tissue remodeling is driven by sustained and controlled inflammation mediated by macrophages in strong communication with fibroblasts. Notably, both pro-inflammatory and anti-inflammatory signals remained elevated during this process in the long-term, challenging the prevailing notion that inflammation opposes remodeling. Using macrophage depletion in mice, we demonstrate that macrophages are essential for this process. Single-cell RNA sequencing further revealed robust fibroblast-to-macrophage signaling, contrasting with the conventional macrophage-to-fibroblast paradigm, and identified unique Spp1 macrophages and Ctla2a fibroblasts within the remodeling niche. These findings provide a comprehensive view of the immune landscape in biomaterial-induced tissue remodeling, highlighting key cellular interactions, prolonged kinetics, and unexpected signaling pathways. By defining key targets and fundamental principles, this work has broad implications for advancing biomaterial-induced tissue regeneration.

bioengineering↗

bioRxiv2025

WS6 enables scalable ex vivo expansion and gene editing of epithelial basal stem cells

Modeling human epithelial diseases and developing cell-based therapies require robust methods to expand and manipulate epithelial stem and progenitor cells in vitro. Basal stem/progenitor cells from stratified epithelia can be expanded in 3T3-J2 fibroblast feeder cell co-culture systems, and the addition of the ROCK inhibitor Y-27632 enhances proliferation and culture longevity, a phenomenon described as conditional reprogramming. Here, we present a method incorporating the small molecule WS6 to further improve the proliferation and lifespan of cultured epithelial cells from multiple tissues, including airway, skin, and thymus. Cells maintained in this medium ( EpMED; FAD+Y+WS6) retain basal stem/progenitor cell identity and function, including the capacity to differentiate. We demonstrate their capacity to engraft in vivo in a tracheal transplantation model. In a second application, we generate clonal CRISPR-Cas9 genome edited nasal cultures, introducing targeted knockouts of DNAH5 or DNAI2 to create primary ciliary dyskinesia disease models. We anticipate that our method will have broad applications in epithelial cell biology, disease modeling, and regenerative medicine, while reducing reliance on immortalized or cancer cell lines and animal experimentation.

cell biology↗

bioRxiv2025

mRNA lipid nanoparticle-incorporated nanofiber-hydrogel composite generates a local immunostimulatory niche for cancer immunotherapy

Hydrogel materials have emerged as versatile platforms for various biomedical applications. Notably, the engineered nanofiber-hydrogel composite (NHC) has proven effective in mimicking the soft tissue extracellular matrix, facilitating substantial recruitment of host immune cells and the formation of a local immunostimulatory microenvironment. Leveraging this feature, here we report an mRNA lipid nanoparticle (LNP)-incorporated NHC microgel matrix, termed LiNx, by incorporating LNPs loaded with mRNA encoding tumour antigens. Harnessing the potent transfection efficiency of LNPs in antigen-presenting cells (APCs), LiNx demonstrates remarkable immune cell recruitment, antigen expression and presentation, and cellular interaction. These attributes collectively create an immunostimulating milieu and yield a potent immune response achievable with a single dose, comparable to the conventional three-dose LNP immunization regimen. Further investigations reveal that the LiNx not only generates heightened Th1 and Th2 responses but also elicits a distinctive Type 17 T helper cell-mediated response pivotal for bolstering antitumour efficacy. Our findings elucidate the mechanism underlying LiNxs role in potentiating antigen-specific immune responses, presenting a new strategy for cancer immunotherapy.

bioengineering↗