# Vaccination of mice with Trichinella spiralis serine proteinase enhanced gut epithelial barrier and elicited a high protective immunity

## TL;DR

Vaccinating mice with a protein from Trichinella spiralis improved gut health and protected against infection by boosting immune responses and reducing parasite load.

## Contribution

This study demonstrates that rTsSPc vaccination can block parasite invasion and enhance immunity, making it a promising vaccine candidate.

## Key findings

- rTsSPc vaccination reduced gut and muscle parasite burdens by over 60%.
- Immunized mice showed increased IgG and sIgA levels and a Th2-dominant immune response.
- Vaccination improved gut barrier integrity and reduced intestinal inflammation.

## Abstract

A Trichinella spiralis serine proteinase (TsSPc) was identified in the intestinal infective larva (IIL) surface and excretory-secretory (ES) antigens. Our previous study showed that recombinant TsSPc (rTsSPc) disrupted intestinal epithelial integrity and barrier function, and mediated larval invasion of intestinal mucosa. This study aims to investigate the impact of rTsSPc vaccination on intestinal epithelial integrity and its elicited protective immunity in a mouse model.

ELISA results demonstrated that rTsSPc immunization induced a systemic humoral immune response with serum-specific IgG antibody titer reaching 1:10⁵, and elicited a mixed Th1/Th2 immune response dominated by the Th2 type. In the rTsSPc-immunized mice, the TsSPc-specific intestinal sIgA level was also markedly increased (P < 0.0001); The secretion levels of IFN-γ and IL-4 in spleen, mesenteric lymph node (MLN) and Peyer’s patch (PP) cells were significantly increased (P < 0.0001). rTsSPc immunization blocked the binding of parasite-derived TsSPc and gut epithelial RACK 1 receptor, prevented the activation of MAPK/ERK1/2 pathway and enhanced gut epithelial integrity, and impeded the parasite invasion. Vaccination of mice with rTsSPc exhibited a 65.7% reduction of enteral adult burden with a 61.13% decline of female fecundity, and a 58.10% reduction of muscle larval burden. Intestinal inflammation of rTsSPc-immunized mice was also significantly alleviated, as demonstrated that goblet cell numbers were obviously decreased, expression level of mucins (Muc2 and Muc5ac) and pro-inflammatory cytokines (IL-6 and TNF-α) was evidently declined, while expression level of anti-inflammatory cytokines (IL-10 and TGF-β) was distinctly increased after infection. Moreover, peritoneal macrophages of rTsSPc-immunized mice exhibited a mixed M1/M2 polarization, but shifted to a predominant M2 polarization pattern post infection. ADCC assay confirmed that peritoneal macrophage of immunized mice had a stronger anti-rTsSPc antibodies-mediated cytotoxicity killing newborn larvae (P < 0.0001).

rTsSPc vaccination produced a high protective immunity through multiple synergistic mechanisms: eliciting an obvious humoral and cellular immunity, gut local mucosal sIgA responses; blocking the binding of parasite-derived TsSPc to gut RACK1 receptors and the activation of MAPK/ERK1/2 pathway, improved gut epithelial integrity, inhibiting larval invasion, enhancing macrophages’ ability of ADCC killing larvae, and finally reduced parasite burden and alleviated inflammation of intestines and skeletal muscles. TsSPc might be a promising novel candidate target for anti-T. spiralis vaccine.

T. spiralis serine proteinase (TsSPc) plays a crucial role in larval invasion of host intestinal epithelium and disruption of intestinal epithelial integrity. Previous studies demonstrated that rTsSPc specifically bound to gut epithelial RACK 1 receptor and activated MAPK/ERK1/2 pathway, decreased the tight junctions (TJs) expression and damaged gut barrier function, and mediated larval intrusion of gut mucosa. The purpose of this study was to investigate the impact of rTsSPc vaccination on intestinal epithelial integrity and its triggered protective immunity. The results showed that subcutaneous vaccination of mice with rTsSPc triggered a strong systemic humoral immune response (high levels of serum-specific IgG, IgG1/IgG2a, IgA) and gut mucosal sIgA responses. The levels of Th1/Th2 cytokines (IFN-γ/IL-4) secreted by spleen, mesenteric lymph nodes, and Peyer’s patches were significantly increased at 6 weeks post-vaccination. rTsSPc immunization blocked the binding of parasite-derived TsSPc to gut epithelial RACK1 receptor and activation of MAPK/ERK1/2 pathway, enhanced gut epithelial integrity, and impeded the parasite invasion. The rTsSPc-vaccinated mice exhibited a 65.7% and 58.1% reduction of gut adult burden and muscle larval burdens, respectively. Moreover, the numbers of goblet cells and expression levels of Muc2, Muc5ac and pro-inflammatory cytokines (TNF-α and IL-6) in immunized mice were notably decreased after infection, while the expression of anti-inflammatory cytokines (IL-10 and TGF-β) was obviously elevated. Additionally, peritoneal macrophages of rTsSPc-immunized mice exhibited a mixed M1/M2 polarization, but shifted to a predominant M2 polarization pattern post infection. ADCC assay confirmed that peritoneal macrophage of immunized mice had a stronger anti-rTsSPc antibodies-mediated cytotoxicity killing newborn larvae. TsSPc might be a promising novel candidate target for anti-T. spiralis vaccine.