Delayed-type hypersensitivity (DTH) reaction to rotavirus EDIM and RRV. Part 2
The rotavirus-specific IgG titers in the pre-serum showed similar results as the rotavirus-specific IgM with titers of about 100 AU in the RRV and RRV+EDIM groups, non-detectable in the EDIM group and low but detectable (20 AU) in the Gastrogard-R® group. Inoculation with EDIM increased the titers of rotavirus-specific IgG in all groups (RRV 350 AU; RRV+EDIM 520 AU; EDIM 50 AU; Gastrogard-R® 125 AU).
Rotavirus-specific serum IgG subclass antibodies
The rotavirus-specific IgG subclasses IgG1, IgG2a, IgG2b and IgG3 were measured in the pool serum, individually collected and then pooled per group, at day 16 (pre-EDIM serum) and in the individual sera of the pups collected at day 28 (post-EDIM serum). Like rotavirus-specific IgG titers, the pre-serum showed detectable levels of all subclasses in the RRV and RRV+EDIM groups, 2 times lower in the Gastrogard-R® group and non-detectable in the EDIM group (data not shown). The rotavirus-specific IgG subclass titers in the post-serum were markedly higher and in Figure 7 the geometric mean titers (GMT) per group are shown. Primary rotavirus inoculation at day 7 of age (RRV; group A) showed the following antibody titers; IgG2a antibodies (GMT 838 AU), IgG3 (GMT 213 AU), IgG2b (GMT 164 AU) and IgG1 (GMT 131 AU). A secondary EDIM inoculation at day 17 of age (RRV+EDIM; group C) showed similar IgG2a (GMT 981 AU), IG2b (GMT 151 AU) and IgG1 (GMT 125 AU) levels, only the amount of IgG3 seemed to be elevated (GMT 555 AU) but this increase was not significant. Primary inoculation with EDIM at day 17 of age (group B) resulted in low antibody levels for all subclasses (range 1-4 AU). Antibody levels in the mice receiving Gastrogard-R® were significantly lower than the RRV+EDIM group (IgG1 p = 0.035, IgG2a p = 0.024, IgG2b p = 0.025, IgG3 p = 0.001). However, this Gastrogard-R® group showed significantly higher levels of antibodies than the EDIM group (IgG1 p = 0.028, IgG2a p = 0.002, IgG2b p = 0.002, IgG3 p = 0.007). These results indicate that although Gastrogard-R® had completely inhibited rotavirus-induced diarrhea during a primary infection, some infection had occurred or stimulation of the immune system because B cells were activated and rotavirus-specific IgG (subclass) antibodies were produced.
Discussion
In this study a neonatal mouse model, originally developed by VanCott et al., was modified to investigate the effect of nutritional intervention (Gastrogard-R®) during a primary (heterologous) rotavirus infection and/or on a secondary (homologous) rotavirus infection. Gastrogard-R® is prepared from the colostrum of hyperimmunised cows and contains high antibody titers against four human rotavirus serotypes. In parallel, the neonatal mouse model could provide better insight into the immunological response to rotavirus since mechanism of rotavirus protection and rotavirus clearance in mice are still not fully understood. The ability of a neonatal mouse or human to generate sufficient immune effectors needed for protection after gastrointestinal virus infection is dependent on its state of immunological maturity. Specific immune cell functions as well as the gastrointestinal tract mature in neonatal mice through the weaning period, while the numbers of immune cells in inductive and effector sites increase gradually. Many studies have been performed to clarify the immune response to rotavirus infection. Clearance of rotavirus can occur T cell independent as well as B cell/antibody independent. As for protection to rotavirus reinfection, B cells are absolutely necessary for long-term protection against rotavirus re-infection. On the other hand, T cells are important for antiviral immunity in mice as well. CD4+ T cells are essential for the development of more than 90% of the rotavirus-specific intestinal IgA and their presence seems to be critical for the establishment of protective long term memory responses. Moreover, murine rotavirus-specific CD8+ T cells can mediate short-term partial protection against reinfection. These data implicates that there is not one specific route that leads to rotavirus clearance and/or protection to rotavirus re-infection, but that both B cell as well as T cell-dependent and independent mechanisms can lead to clearance of infection and long-term maintenance of protection. Within the model as described herein, both aspects of immune responses related towards protection can be studied.