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Arya, M.

Publications and source records attributed to Arya, M..

2 recordsLinked to original sources
bioRxiv2026

Corilagin attenuates high glucose-induced neurotoxicity and mitochondrial dysfunction through restoration of the AMPK-SIRT1-PGC1α-TFAM signaling axis

Mitochondrial dysfunction and oxidative stress represent two interconnected, primary causes for Diabetic Neuropathy (DN); however, the majority of currently available anti-diabetic therapies have focused on glucose control as opposed to neurodegenerative downstream effects. Corilagin, is an ellagitannin having high anti-oxidant properties; however, it has not been evaluated against hyperglycemia induced neuronal injury. The present study demonstrates the ability of Corilagin to protect against mitochondrial dysfunction via models of diabetic nephropathy and cerebral ischemia. High glucose (50 mM, 24 hr) was utilized to induce diabetes like conditions in the SH-SY5Y human neuroblastoma Cell Line. High glucose induced significant decreases in cell viability, increases in intracellular and mitochondrial reactive oxygen species, depletion of reduced glutathione reserves, induces apoptosis, and causes mitochondrial depolarization and fragmentation. Corilagin pre-treatment attenuated each of these high-glucose induced effects by protecting against mitochondrial membrane potential loss and maintaining mitochondrial network morphology while reducing apoptotic cell fraction relative to glucose alone. Additionally, these protective effects were accompanied by restoration of AMPK phosphorylation and up-regulation of SIRT1, PGC1 and TFAM, components that are part of the principal signaling pathway that regulates mitochondrial biogenesis; therefore, therefore, this pathway may contribute mechanistically to the cyto-protective effect of Corilagin. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/740444v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@1be4a92org.highwire.dtl.DTLVardef@11d6e9org.highwire.dtl.DTLVardef@1346757org.highwire.dtl.DTLVardef@16c9f1e_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO Proposed mechanism underlying the neuroprotective effects of Corilagin against high glucose-induced mitochondrial dysfunction.High glucose suppresses AMPK phosphorylation, leading to downregulation of the SIRT1-PGC-1-TFAM signaling axis, increased intracellular and mitochondrial reactive oxygen species (ROS), glutathione depletion, apoptosis, mitochondrial depolarization, and mitochondrial fragmentation. Corilagin pretreatment restores AMPK activation and the downstream SIRT1-PGC-1-TFAM pathway, thereby reducing oxidative stress, preserving intracellular glutathione, preventing apoptosis, maintaining mitochondrial membrane potential, and protecting mitochondrial network integrity. C_FIG

cell biology↗
bioRxiv2023

A C-terminal HaloTag Impairs AGO2 Function

A full understanding of RNA silencing requires appropriate molecular biology tools to explore the roles of Argonaute 2 (AGO2) and the RNA-induced silencing complex (RISC). Approaches relying on affinity tagging and antibodies have important limitations that can lead to artificial results. Both the N- and C-terminal domains of AGO2 have been shown to be important for correct activity and yet the consequences of appending tags to either terminus have not been fully investigated. N-terminal tags are frequently used to study AGO2 biology. Recently, an N-terminal HaloTag-Ago2 fusion was reported and examined in mice. While the versatile HaloTag provided new opportunities to study RISC biology, the tagged construct showed certain activity changes compared to unmodified AGO2. CRISPaint, a new CRISPR-Cas9 technique, permits the creation of endogenous C-terminal tag fusions. We used CRISPaint to generate the first reported recombinant AGO2 construct with a C-terminal tag: an endogenous C-terminal HaloTag fusion to AGO2 (AGO2HALO) in human (A549) cells. We found that the AGO2HALO fusion protein has a reduced capacity to interac with the key protein binding partner TNRC6A and that the C-terminal HaloTag does not affect cell viability. However, the AGO2HALO fusion significantly impairs RNA cleavage and RNA silencing activity compared to control cells and reduces nuclear localisation of the fusion protein. Using plasmid constructs and transient transfection, we compared AGO2 tagged with EGFP at the N- or C-terminus in siRNA and miRNA reporter gene assays, and cellular localisation. N-terminally tagged AGO2 functioned and localised similarly to WT untagged AGO2, whereas C-terminally tagged AGO2 was impaired in siRNA and miRNA silencing and exhibited poor nuclear and P-body localisation. We conclude that the fusion of a C-terminal HaloTag to AGO2 is not appropriate for studying AGO2 and RISC. Our results assert the importance of comprehensively validating recombinant tagging strategies to ensure that any experimental results generated do not arise from, or are not obscured by, critical functional defects.

molecular biology↗