Exogenously-administered leptin increases early incisional wound angiogenesis in an experimental animal model

BACKGROUND: Leptin is a potent direct angiogenic factor that stimulates endothelial cell migration and activation in vitro, as well as angiogenesis in vivo. In addition, leptin seems to play an important role in clinical angiogenesis by promoting the development of new blood vessels. OBJECTIVE: To determine the effect of exogenously administered leptin on incisional wound healing in an experimental animal model. MATERIALS AND METHODS: Sixty-three Sprague-Dawley male mice were used for the study. Full thickness incisional wound was considered as the wound model. The mice were divided into seven groups of nine animals each. Surgical wounds were injected with murine recombinant leptin. Three different leptin doses of 100 pg/ml, 200 pg/ml and 500 pg/ml were used in different animal groups (A, B and C). For each of the three leptin doses used, another animal group was evaluated with a combined injection of leptin and antileptin: 100 pg/ml leptin with 50 pg antileptin, 200 pg/ml leptin with 100 pg antileptin, 500 pg/ml leptin with 250 pg antileptin (A1, B1, and C1), in order to study the inhibitory effect on the leptin factor. Nine mice served as controls. These were injected with 0.3 ml water for injection solution. Mice were sacrificed 3, 7 and 9 days postoperatively. After sacrifice of the animals, the skin was grossly assessed for appearance, colour and texture. Full thickness incisional wounds were dissected for histological examination. A qualitative analysis of angiogenesis in the surgical wound was conducted following a standard hematoxylin and eosin stain. The wound tissue samples from each experimental group underwent immunohistochemical evaluation of microvessel density by endothelial cell staining with mouse anti-rat CD34 monoclonal antibody. RESULTS: The most impressive growth of new blood vessels appeared seven and nine days after treatment with the highest leptin doses. There were no significant differences in microvessel density at seven or nine postoperative days among different groups treated with leptin. None of the wounds from the control group, or those from animal groups treated with the combined injection of leptin and antileptin developed any new vessels. CONCLUSION: Exogenous administration of leptin may increase early tissue angiogenesis in the incisional wound of an experimental animal model.     

Detection of precursor Th cells in mesenteric lymph nodes after oral immunization with protein antigen and cholera toxin

We have characterized the earliest antigen-specific Th cells in murine mesenteric lymph nodes (MLN), following oral immunization with the hen egg lysozyme (HEL) as antigen and cholera toxin (CT) as adjuvant. We did this by analyzing in vitro proliferation and cytokine production in response to HEL by the MLN T cells. MLN cells taken 5 days after a single oral immunization with HEL and CT provided the earliest source of proliferating HEL-specific T cells. This proliferation was completely inhibited by anti-IL-2, but not inhibited by anti-IL-4 antibody. IL-2 protein was detected in culture supernatants but not IL- 4 using ELISA or bioassays. IL-4 mRNA was not found in responding cells using RT-PCR. Some of the day 5 MLN cultures produced IFN-gamma in response to HEL, but isolated T cells from the same MLN did not. Exogenous IL-4 alone did not stimulate day 5 MLN T cells, but IL-4 did synergize with HEL to induce a large proliferative response. The data indicate that the HEL-specific CD4 T cell pool in MLN 5 days after oral immunization is composed of undifferentiated precursor Th cells. These cells have the potential for IL-2 production and IL-4R expression upon re-stimulation in vitro.