Divergent Immunomodulating Effects of Probiotics on T Cell Responses to Oral Attenuated Human Rotavirus Vaccine and Virulent Human Rotavirus Infection in a Neonatal Gnotobiotic Piglet Disease Model

Abstract

Rotaviruses (RVs) are a leading cause of childhood diarrhea. Current oral vaccines are not effective in impoverished countries where the vaccine is needed most. Therefore, alternative affordable strategies are urgently needed. Probiotics can alleviate diarrhea in children and enhance specific systemic and mucosal Ab responses, but the T cell responses are undefined. In this study, we elucidated the T cell and cytokine responses to attenuated human RV (AttHRV) and virulent human RV (HRV) in gnotobiotic pigs colonized with probiotics (Lactobacillus rhamnosus strain GG [LGG] and Bifidobacterium lactis Bb12 [Bb12]), mimicking gut commensals in breastfed infants. Neonatal gnotobiotic pigs are the only animal model susceptible to HRV diarrhea. Probiotic colonized and nonvaccinated (Probiotic) pigs had lower diarrhea and reduced virus shedding postchallenge compared with noncolonized and nonvaccinated pigs (Control). Higher protection in the Probiotic group coincided with higher ileal T regulatory cells (Tregs) before and after challenge, and higher serum TGF-β and lower serum and biliary proinflammatory cytokines postchallenge. Probiotic colonization in vaccinated pigs enhanced innate serum IFN-α, splenic and circulatory IFN-γ−producing T cells, and serum Th1 cytokines, but reduced serum Th2 cytokines compared with noncolonized vaccinated pigs (Vac). Thus, LGG+Bb12 induced systemic Th1 immunostimulatory effects on oral AttHRV vaccine that coincided with lower diarrhea severity and reduced virus shedding postchallenge in Vac+Pro compared with Vac pigs. Previously unreported intestinal CD8 Tregs were induced in vaccinated groups postchallenge. Thus, probiotics LGG+Bb12 exert divergent immunomodulating effects, with enhanced Th1 responses to oral AttHRV vaccine, whereas inducing Treg responses to virulent HRV.

Protein Malnutrition Alters Tryptophan and Angiotensin-Converting Enzyme 2 Homeostasis and Adaptive Immune Responses in Human Rotavirus-Infected Gnotobiotic Pigs with Human Infant Fecal Microbiota Transplant

“…Blood, duodenum, ileum, and spleen were collected to isolate mononuclear cells (MNCs) as described previously (17). Result acquisition was done using a MACSQuant Analyzer 10 (Miltenyi Biotec, Bergisch Gladbach, Germany), and analysis was performed using FlowJo software version 10 (FlowJo LLC, Ashland, OR).…”

“…The frequency of T cells was expressed as the percentage among total MNCs (determined based on forward and side scatter characteristics). The frequencies of natural regulatory T cells (Tregs; CD4 ϩ CD25 ϩ Foxp3 ϩ and CD8 ϩ CD25 ϩ Foxp3 ϩ ) were determined as described previously (17).…”

“…Finally, pigs are outbred, eliminating the strain bias common in mouse model research. We have used the GF pig to study controlled exposure to microorganisms and physiologically relevant and economically feasible intervention strategies (15)(16)(17)(18)(19)(20)(21)(22). Recently, investigators have successfully transplanted GF pigs with human gut microbiota (23) and used this model to examine the effects of probiotics on human rotavirus (HRV) pathogenesis and immunity (24)(25)(26).…”

Comparative In Vitro and In Vivo Studies of Porcine Rotavirus G9P[13] and Human Rotavirus Wa G1P[8]

“…The mean number of days to onset of virus shedding, mean duration (days) of virus shedding, average peak titer of virus shedding, and mean duration (days) of diarrhea in postchallenge pigs were analyzed by oneway analysis of variance (ANOVA). Mean cumulative fecal scores in different treatment groups were compared by using the area under the curve, as previously described (49,50). Statistical analyses were performed with GraphPad Prism 6.0c software (GraphPad Software, La Jolla, CA, USA).…”

In vivo gut transcriptome responses toLactobacillus rhamnosusGG andLactobacillus acidophilusin neonatal gnotobiotic piglets

“…Probiotics that facilitate mucosal repair and maintain gut homeostasis are invaluable treatment modalities for neonatal enteric infections including rotavirus diarrhea. 3 Recent research has highlighted the effective role of probiotics in modulation of the gut microbiota, thereby alleviating intestinal disorders like inflammatory bowel disease, travelers' diarrhea, colitis, Crohn disease, and antibiotic associated diarrhea. 4,5 However, pathways and mechanisms by which probiotics mediate their beneficial effects, especially in neonates, are largely undefined.…”