Abstract
The role of CD28-mediated signals in T helper cell maturation is not fully understood. We tested the requirement for costimulation through CD28 in several systems of CD4+, T cell differentiation. In vivo priming of mice with genetic disruption of CD28 (CD28-/-) yielded normal levels of antigen-specific interferon gamma production but markedly diminished levels of interleukin 4 (IL-4) after in vitro restimulation. In response to the pathogenic microbe, Leishman a major, C57BL6 CD28-/- mice were fully capable of controlling infection and exhibited a normal T helper 1 response. BALB/c CD28-/- mice unexpectedly exhibited normal susceptibility to L. major. BALB/c CD28-/- mice developed high levels of IL-4 mRNA and protein induction in the draining lymph nodes. In addition, susceptibility of BALB/c CD28-/- mice was reversed by neutralization of IL-4 in vivo. We also activated transgenic CD28-bearing T cells from the BALB and C57BL background in vitro in the presence of CTLA4Ig. BALB cells had greater IL-4 producing capacity than C57BL cells in the absence of costimulation. Diverse factors including costimulatory signals, genetic polymorphism, and the nature of the immunogen all influence T helper phenotype commitment, but these results provide evidence that CD28 is not an absolute requirement for generating either Th1 or Th2 responses.
Highlights
Genotypes were determined by flow cytometry using stains for commercially conjugated fluorescent mAbs against CD28 and CD4 (Caltag Laboratories, South San Francisco, CA) and confirmation by PCR using two oligonucleotide pairs spanning the genomic disruption of CD28 and the neomycin resistance marker
To determine how signals delivered through CD28 affect T helper (Th) cell maturation we examined the ability of CD28 - / - mice to generate IFN-3, and interleukin 4 (IL-4)-producing cells after immunization
CD28-deficient mice bred onto the BALB/c or
Summary
CD28 - / - mice were generated as described [8]. The animals used in these experiments were from the fifth back-cross to C57BL/6 and fourth or fifth back-cross to BALB/c. Heterozygote and wild-type littermates were used as controls for BALB/c experiments. Commercial wild-type mice (Jackson Laboratories, Bar Harbor, ME) were used as controls for C57BL/6 experiments. Mice were housed in a specific pathogenfree environment. Genotypes were determined by flow cytometry using stains for commercially conjugated fluorescent mAbs against CD28 and CD4 (Caltag Laboratories, South San Francisco, CA) and confirmation by PCR using two oligonucleotide pairs spanning the genomic disruption of CD28 and the neomycin resistance marker. All work was perfomaed in accordance with University of Chicago guidelines for animal use and care
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