# A66 EFFECTS OF HYMENOLEPIS DIMINUTA EXTRACELLULAR VESICLES ON MOUSE MACROPHAGES AND THEIR MITOCHONDRIA

## TL;DR

This study explores how extracellular vesicles from the tapeworm Hymenolepis diminutus affect mouse macrophages and their mitochondria.

## Contribution

The paper introduces a novel characterization of Hymenolepis diminuta extracellular vesicles and their effects on macrophage metabolism.

## Key findings

- Hd-EVs were successfully isolated and characterized with sizes ranging from 100-350 nm.
- Hd-EVs increased oxidative phosphorylation in macrophages but did not reduce TNF-α production.
- HES and Hd-EVs both up-regulated macrophage glycolysis.

## Abstract

In recent years, extracellular vesicles (EVs) have emerged as important means of intercellular communication and immune modulation between helminth and host. Earlier work revealed that excretory/secretory products from the rat tapeworm Hymenolepis diminuta (HES) have anti-inflammatory properties when applied to macrophages. In this study, we have investigated the immunomodulatory properties of H. diminuta EVs (HEVs) on macrophages.

1. To isolate and characterize bacterial contaminant-free H. diminuta EVs (Hd-EVs).

2. To determine the effects of Hd-EVs on murine macrophage metabolism and LPS-evoked cytokine production.

Adult H. diminuta were cultured for 6h in serum-free RMPI and Hd-EVs isolated from the conditioned medium using an exoEasy kit. Hd-EV protein content was measured using a micro-BCA kit, and size and quantity were measured using Nanosight tracking analysis. Murine bone marrow-derived macrophages or the human macrophage THP-1 cell line were treated with LPS (10 ng/ml) ± a crude H. diminuta extract (HdE), HES, or Hd-EVs for 24h. TNF-α levels in the culture medium was measured by ELISA. Macrophage oxidative phosphorylation (OX.PHOS) and glycolysis were measured in real-time using an Agilent Seahorse XF Analyzer.

Analysis of 14 Hd-EV samples revealed 4.1 X 109 - 5.48 X 1010 particles/mL, with Hd-EVs predominantly 100-350 nm in diameter and with a protein content of 64.9-181.6 ng/mL. HdE and HES reduced TNF-α production in LPS-activated macrophages by 39-76% (n=11), while Hd-EVs did not lower TNF-α production. Metabolic analysis suggests that HES and Hd-EVs can up-regulate macrophage glycolysis, while only Hd-EVs exposed macrophages showed increased OX.PHOS (n=6).

We have successfully isolated and begun to characterize EVs from adult H. diminuta. Despite the known abilities of HES, we unexpectedly noted that exposure to Hd-EVs did not result in a statistically significant decrease in macrophage TNF-α release. However, the increase in OX.PHOS macrophages treated with Hd-EVs is something to examine further. The next steps are to determine the contents and immunomodulatory activity of the Hd-EVs, thus advancing knowledge of how helminths can manipulate their hosts.

NSERC

Clinical Practice