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Uptake of Fluorescent Labeled Small Extracellular Vesicles <em>In Vitro</em> and in Spinal Cord

Small extracellular vesicles (sEVs) are 50-150 nm vesicles secreted by all cells and present in bodily fluids. sEVs transfer biomolecules such as RNA, ...

Drexel University College of Medicine

Biological sex influences inflammatory response, as there is a greater incidence of acute inflammation in men and chronic inflammation in women. Here, we report that acute inflammation is attenuated by X-inactive specific transcript (Xist), a female cell-specific nuclear long noncoding RNA crucial for X-chromosome inactivation. Lipopolysaccharide-mediated acute inflammation increased Xist levels in the cytoplasm of female mouse J774A.1 macrophages and human AML193 monocytes. In both cell types, cytoplasmic Xist colocalizes with the p65 subunit of NF-κB. This interaction was associated with reduced NF-κB nuclear migration, suggesting a novel mechanism to suppress acute inflammation. Further supporting this hypothesis, expression of 5' XIST in male cells significantly reduced IL-6 and NF-κB activity. Adoptive transfer of male splenocytes expressing Xist reduced acute paw swelling in male mice indicating that Xist can have a protective anti-inflammatory effect. These findings show that XIST has functions beyond X chromosome inactivation and suggest that XIST can contribute to sex-specific differences underlying inflammatory response by attenuating acute inflammation in women.

Xist attenuates acute inflammatory response by female cells.

Small extracellular vesicles (sEVs) derived from antigen-presenting cells such as macrophages can induce therapeutically relevant immune responses. Anti-inflammatory miRNAs are elevated in sEVs secreted by RAW 264.7 mouse macrophages after lipopolysaccharide (LPS) stimulation. We observed uptake of these sEVs by primary mouse cortical neurons, microglia and astrocytes followed by downregulation of proinflammatory miRNA target genes in recipient cells. Pre-treating primary microglia with these sEVs decreased pro-inflammatory gene expression. A single intrathecal injection of sEVs derived from LPS stimulated RAW 264.7 cells attenuated mechanical hyperalgesia in the complete Freund's adjuvant (CFA) mouse model of inflammatory pain and formalin induced acute pain. Importantly, sEVs did not alter the normal pain threshold in control mice. RNA sequencing of dorsal horn of the spinal cord showed sEVs-induced modulation of immune regulatory pathways. Further, a single prophylactic intrathecal injection of sEVs two weeks prior, attenuated CFA-induced pain hypersensitivity and was ineffective in formalin model. This indicates that prophylactic sEVs administration can be beneficial in attenuating chronic pain without impacting responses to the protective physiological and acute inflammatory pain. Prophylactic administration of sEVs could form the basis for a safe and novel vaccine-like therapy for chronic pain or as an adjuvant, potentially reducing the dose of drugs needed for pain relief.

Richa Gupta

Department of Pharmacology & Physiology

Drexel University College of Medicine

Uptake of Fluorescent Labeled Small Extracellular Vesicles In Vitro and in Spinal Cord

Richa Gupta

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Pharmacology & Physiology

Drexel University College of Medicine

Uptake of Fluorescent Labeled Small Extracellular Vesicles In Vitro and in Spinal Cord

Department of Pharmacology & Physiology