Temporal analysis of murine lymphocyte subpopulations by monoclonal antibodies and dual-color flow cytometry after burn and nonburn injury

The immune suppression that frequently accompanies severe injury undoubtedly contributes to subsequent infectious complications. Various lymphocyte subpopulations may be identified by surface antigen expression, and alterations in antigen expression by lymphocytes may reflect host immune competence. Using monoclonal antibodies (Moabs) and dual-color flow cytometry, we studied lymphocyte phenotypic expression in mice after either controlled burn injury or hind-limb amputation, with use of peripheral blood, lymph node, and spleen for cell preparation. Moabs were utilized specific for T cells (Lyt-1), helper/inducer cells (L3T4), suppressor/cytotoxic cells (Lyt-2), B cells (IgG), and activated T cells (Ia or IL-2 receptor). The assay techniques called for small amounts of tissue and avoided gradient procedures that might result in selective loss of some lymphocyte populations. The most consistent changes observed were depressions in percentages of L3T4+ and Lyt-2+ cells in spleens of burned mice, accompanied by depression in Ia+ (possibly activated or proliferating) subsets of L3T4+ and Lyt-2+ cells, and the appearance of increased percentages of non-B, non-T lymphocytes. Changes in lymph node cells were minimal. The major alteration seen in peripheral blood was substantial depression of Ia+ subsets, although burned mice had increased circulating Lyt-2+ cells on several late postburn days. Burned mice, unlike limb-trauma mice, had marked splenic hypertrophy with more than a 300% increase in spleen weight after the 30-day postburn period. Eschar excision/implantation experiments indicated that splenic hypertrophy and splenocyte phenotypic changes are related to the presence of burned tissue, which suggests that burned tissue may partially mediate immune changes that accompany severe burn injury.