Biology subjects

Dutt, G.

Publications and source records attributed to Dutt, G..

3 recordsLinked to original sources

bioRxiv2026

ACTIVATED CASO₄-INDUCED VACUOLATION AS A QUANTITATIVE PLATFORM FOR PHAGOCYTOSIS-DRIVEN DRUG SCREENING

Cytoplasmic vacuolization is a fundamental process associated with phagocytosis, lysosomal acidification, and autophagy, yet robust in-vitro models for its quantification and pharmacological screening remainlimitedor insufficiently established. In this study, we demonstrate that thermally activated calcium sulfate (ACS) induces extensive vacuolation across mammalian cell lines including HeLa, RAW 264.7, 3T3-L1, and SH-SY5Y, thereby establishing a versatile platform to study vacuole biogenesis. To ensure reproducibility, particle heterogeneity was addressed using sedimentation-based fractionation, with homogeneous suspensions obtained at the 5th minute producing stable and consistent vacuole formation. Vacuolation was subsequently quantified by Neutral Red (NR) uptake, dose and time dependent response analyses confirmed direct correlation between ACS concentration and vacuole induction. The assay was validated with bafilomycin A1 (BFA1), a selective V-ATPase inhibitor, which served as a positive control and demonstrated concentration and time dependent inhibition of vacuole formation and acidification. Building on this framework, ten commercially available drugs were screened, revealing distinct profiles ranging from early cytotoxicity, strong vacuole inhibition to partial suppression or negligible effects. This dual capacity to discriminate between vacuole inhibition and cytotoxic responses highlights the utility of ACS-induced vacuolization as a sensitive and scalable in vitro platform. Collectively, our findings position this system as a tractable assay for mechanistic studies of vacuole biology and a functional screening tool for identifying modulators of lysosomal and phagocytic pathways relevant to infection, Lysosomal Disorders, and Phagocytotic dysfunction disorders. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC="FIGDIR/small/711143v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@e5b920org.highwire.dtl.DTLVardef@1dd1072org.highwire.dtl.DTLVardef@62ae59org.highwire.dtl.DTLVardef@a468bf_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

Optimization of mRNA Synthesis and its Cell Expression for Vaccine Development

In vitro transcription (IVT) is a versatile procedure that facilitates template-directed synthesis of RNA molecules of any sequence, from short oligonucleotides to those of several kilobases, in quantities ranging from micrograms to milligrams. This technique, involving the engineering of a template with a bacteriophage promoter sequence, enables the synthesis of RNA for use in a variety of applications including structural studies, biochemical assays, and as functional molecules. Messenger RNA (mRNA) therapy has tremendous potential in regenerative medicine, disease treatment, and vaccination. Synthetic mRNA leverages the cells natural translation machinery to produce proteins, with the ability to transfect cells and induce expression of target proteins under physiological conditions until it is eventually degraded. In this study, we explore the impact of pseudouridine ({Psi})-modified mRNA in enhancing RNA stability, translational efficiency, and immune compatibility for therapeutic use. By optimizing IVT conditions and employing cellulose-based purification techniques, we successfully synthesized high-quality, modified mRNA that demonstrated superior functional performance. Luciferase and GFP mRNA, synthesized with pseudouridine-modified rNTPs, exhibited improved stability, reduced immune activation, and enhanced translation efficiency in HEK293 cells. A sevenfold increase in luciferase activity and elevated GFP fluorescence confirmed the higher protein expression capabilities of the modified mRNA. Furthermore, cellulose bead purification effectively separated single-stranded RNA from double-stranded RNA contaminants, ensuring minimal immune response and maximizing transfection efficiency. These findings highlight the potential of pseudouridine-modified mRNA and refined purification methods for advancing mRNA-based therapies, from vaccines to protein-replacement treatments, setting the foundation for scalable, clinical-grade RNA production.

cell biology↗

bioRxiv2025

THE CORRELATION BETWEEN REDOX ACTIVITY AND ANTIMICROBIAL PROPERTIES OF PYOCYANIN FROM Pseudomonas aeruginosa

Pseudomonas aeruginosa, a metabolically versatile gram-negative bacterium, has garnered attention for its ability to thrive in diverse environments and its capacity to produce pyocyanin, a secondary metabolite with multifunctional properties. Pyocyanin, a redox-active phenazine compound, plays a critical role in mediating the ecological competitiveness of P. aeruginosa through its antimicrobial, biofilm-modulating, and reactive oxygen species (ROS)-generating activities. Beyond its contributions to bacterial virulence, pyocyanin demonstrates significant potential in various industrial and biomedical applications due to its redox properties and ability to function under diverse environmental conditions. This study investigates the electrochemical behaviour and pH-dependent antimicrobial activity of pyocyanin to evaluate its applicability in environmental and medical fields. Soil-derived P. aeruginosa isolates were cultured for pyocyanin (PYO) production, and the pigment was characterized using UV-visible spectroscopy to get its spectral integrity. Electrochemical analysis through cyclic voltammetry revealed enhanced redox activity in acidic environments and stable functionality in alkaline conditions. Antimicrobial assays demonstrated that pyocyanin exhibited optimal activity at neutral to slightly alkaline pH, effectively inhibiting bacterial and fungal growth, while extreme acidic conditions reduced its efficacy. The findings highlight pyocyanins versatility as both a redox mediator and an antimicrobial agent. In medical contexts, its pH-sensitive activity aligns well with physiological conditions, offering promise for combating multidrug-resistant pathogens. Future optimization of pyocyanin biosynthesis, particularly through cost-effective and scalable methods, could unlock its full potential in biotechnological and therapeutic innovations.

microbiology↗