Investigation of Immunoglobulin Heavy Chain Isotypes in an Ancestral Mucosal Immune Model

• Main Author: Du, Christina
• Other Authors: Criscitiello, Michael
• Format: Others
• Language: en_US
• Published: 2012
• Subjects: IgA; IgX; Xenopus; mucosal; immunization
• Online Access: http://hdl.handle.net/1969.1/ETD-TAMU-2011-08-10140

The importance of gut associated lymphoid tissues has been extensively reported in higher vertebrates, but less is known in lower vertebrates. In mammals immunoglobulin (Ig)A is the primary Ig of mucosal immunity. But no IgA has been identified in cold-blooded animals. In higher vertebrates, antigen must stimulate the lymphoid tissues in the intestines to elicit an IgA response, and cytokines from CD4 positive helper T cells are required for B cell switch. It is not known if this is the case in lower vertebrates, or if T cell help evolved before or after class switch recombination between functional antibody isotypes. My study will fill in these gaps in our knowledge by comparing oral antigen inoculation relative to intraperitoneal antigen inoculation in frogs (Xenopus sp.). Oral immunization is a novel approach to eliciting immune responses in Xenopus. I propose that IgX will increase with oral inoculation compared to intraperitoneal injection. This would be the first demonstration of class switch upon oral immunization to a mucosal isotype in the first vertebrates that employs higher vertebrate Ig heavy chain switch mechanism, which would shed light on the most fundamental aspects of our humoral adaptive immune system. Using a total Ig ELISA protocol, measuring total relative levels of IgM, there was no difference between the first three groups of orally immunized frogs compared to intraperitoneally immunized frogs. However, a response to serum IgX was seen in the first group. On the other hand, the refined Ag-specific ELISA protocol did present a significant increase in serum IgM response in frogs immunized systemically over orally immunized animals, but not an overall IgX response. Phylogenetic analysis suggests that, contrary to initial reports, IgA evolved from IgX. With consideration of entire constant region and individual constant domain analyses as well as synteny and function, we suggest new hypotheses of vertebrate antibody evolution to be tested as immunogenetic coverage of more species continues to expand.