MHC-class-II-deficiency impairs wound healing.

BACKGROUND: MHC-class-II-deficient mice lack T helper cell dependent immune reactions. T cell related immune functions are critical for normal wound healing. We hypothesized that MHC-II-deficiency compromises wound repair by affecting the normal wound immune response. MATERIAL AND METHODS: Groups of 10 male MHC-class II-knockout mice and wild-type controls underwent dorsal skin incision. Polyvinyl alcohol sponges were then inserted subcutaneously. The mice were sacrificed 10 days later to determine wound breaking strength and reparative collagen deposition. Activity of T cells and macrophages isolated from the spleens and from the healing wounds was investigated. Fibroblasts derived from the wounds were tested ex vivo for proliferative activity and collagen synthesis. RESULTS: Wound collagen deposition and wound breaking strength were impaired in MHC-class-II-knockout mice (P < 0.05). Impaired healing was reflected in diminished mitogen-reactivity of splenic T-cells (P < 0.01), and decreased CD4 expression in wounds. In addition, basal and LPS + IFN-gamma-induced synthesis of TNF-alpha and nitric oxide by wound-derived macrophages was impaired. Exvivo, fibroblast proliferation and fibroblast collagen production from MHC-II-deficient mice was decreased. CONCLUSION: MHC-II-deficiency compromises wound healing. This may be a reflection of impaired wound immune cell function and decreased activity of wound fibroblasts.

Regulation of nitric oxide synthesis in wounds by IFN-gamma depends on TNF-alpha.

Macrophage-derived nitric oxide is a critical mediator in wound healing. Its regulation in vivo, however, remains unclear. We hypothesized that interferon (IFN)-gamma plays an important role in the regulation of nitric oxide in wounds. Groups of 12 male IFN-gamma -knockout mice and wild-type controls underwent dorsal skin incision and polyvinyl alcohol sponges were inserted subcutaneously. Mice were sacrificed 10 days later to determine wound breaking strength and reparative collagen deposition. Synthesis of nitric oxide (NO), tumor necrosis factor (TNF)-alpha, and IFN-gamma was measured in the wound. Wound-derived macrophages were tested for NO synthesis in the presence or absence of IFN-gamma, TNF-alpha, and anti-TNF-alpha antibody. In a separate experiment, IFN-gamma -knockout mice and wild-type controls were treated with molsidomine, a nitric oxide donor. It was found that wound collagen deposition and wound breaking strength were impaired in IFN-gamma-knockout mice (p < .05). Impaired healing was reflected in diminished synthesis of TNF-alpha and NO in wounds (p < .05). In vivo treatment with molsidomine reversed impaired healing in IFN-gamma-deficient mice. Ex vivo, addition of IFN-gamma stimulated the synthesis of TNF-alpha and NO in wound-derived macrophages. IFN-gamma -induced NO synthesis by wound-derived macrophages was abolished by anti-TNF-alpha-antibody-treatment, which could be fully reversed by exogenous TNF-alpha. Thus we conclude that IFN-gamma-deficiency impairs wound healing and diminishes NO synthesis in wound-derived macrophages. The stimulatory effect of IFN-gamma on macrophage NO production depends on endogenous TNF-alpha synthesis.

IFN-gamma and IL-10 mediate parasite-specific immune responses of cord blood cells induced by pregnancy-associated Plasmodium falciparum malaria.

Available evidence suggests that immune cells from neonates born to mothers with placental Plasmodium falciparum (Pf) infection are sensitized to parasite Ag in utero but have reduced ability to generate protective Th1 responses. In this study, we detected Pf Ag-specific IFN-gamma(+) T cells in cord blood from human neonates whose mothers had received treatment for malaria or who had active placental Pf infection at delivery, with responses being significantly reduced in the latter group. Active placental malaria at delivery was also associated with reduced expression of monocyte MHC class I and II in vivo and following short term in vitro coculture with Pf Ag compared with levels seen in neonates whose mothers had received treatment during pregnancy. Given that APC activation and Th1 responses are driven in part by IFN-gamma and down-regulated by IL-10, we examined the role of these cytokines in modulating the Pf Ag-specific immune responses in cord blood samples. Exogenous recombinant human IFN-gamma and neutralizing anti-human IL-10 enhanced T cell IFN-gamma production, whereas recombinant human IFN-gamma also restored MHC class I and II expression on monocytes from cord blood mononuclear cells cocultured with Pf Ag. Accordingly, active placental malaria at delivery was associated with increased frequencies of Pf Ag-specific IL-10(+)CD4(+) T cells in cord blood mononuclear cell cultures from these neonates. Generation and maintenance of IL-10(+) T cells in utero may thus contribute to suppression of APC function and Pf Ag-induced Th1 responses in newborns born to mothers with placental malaria at delivery, which may increase susceptibility to infection later in life.

Functional analysis of a promoter variant of the gene encoding the interferon-gamma receptor chain I.

We analyzed a single nucleotide polymorphism (SNP) at position -56 (T-->C) in the promoter region of the gene encoding the human interferon-gamma receptor ligand-binding chain I (IFN-gammaR1). The mutation was present at similar frequencies in Gabonese children with either mild or severe malaria. Functional investigations of the promoter in a transfected human B-cell line showed lower levels of luciferase reporter gene expression in the presence of the mutation, indicating the importance of this position for promoter activity, and suggesting that this SNP might negatively influence the expression level of IFN-gammaR1 at the cell surface. Further examinations of the DNA sequence at this polymorphic site showed a perfectly matched binding site for the transcription factor activator protein 4 (AP-4) on both strands. Binding sites for other important transcription factors involved in gene expression and regulation of the immune response against infections, including Ikaros 2 (Ik-2), nuclear factor kappaB (NFkappaB), and CETS1p54, are also situated in this region.

Age-dependent enhancement of IFN-gamma responses to Plasmodium falciparum liver stage antigen-1 T cell epitopes.

We assessed the cellular immunological responses to two Plasmodium falciparum liver-stage antigen (LSA)-1-derived T-cell epitopes in healthy Gabonese children and adults. The N-terminal peptide, designated T1, induced interferon (IFN)-gamma production in peripheral blood mononuclear cells (PBMC) from a significantly lower proportion of children compared to adults, but both interleukin (IL)-10 and IL-12 were produced by similar proportions of PBMC from the two groups. The LSA-1 junction region peptide (LSA-J) also induced IFN-gamma in a lower, but in this case statistically non-significant, proportion of PBMC from children compared to adults, whilst the proportions producing either IL-10 or IL-12 were again similar. Higher amounts of both IFN-gamma and IL-10 were induced by LSA-J compared to T1. CD8+ T-cells were shown to be primarily responsible for the production of peptide-driven IFN-gamma. The results suggest a significant age-related increase in the proportion of individuals capable of producing IFN-gamma to the N-terminal T1 epitope, with a shift from a predominantly IL-10-led response in children.

Differential expression of inflammatory mediators in radiation-impaired wound healing.

BACKGROUND: Radiation impairs healing, although the underlying mechanisms are not clearly defined. Normal healing requires a fine balance of promoting and inhibiting factors. We hypothesize that there may be a down-regulation of promoting factors (nitric oxide) and, in turn, an up-regulation of healing inhibiting factors (TNF-alpha and IFN-gamma) in the wound after radiation. MATERIAL AND METHODS: Groups of 10 rats were irradiated using single dose 12 or 24 Gy electron radiation at the dorsal skin. Control rats were sham-irradiated. On Day 5 a skin incision in the irradiated area was performed and polyvinyl alcohol sponges were inserted subcutaneously. Rats were sacrificed 10 days later to determine the wound-breaking strength and reparative collagen deposition. Nitrite and nitrate (index of NO synthesis), TNF-alpha, and IFN-gamma were measured within the wound fluid. Expression of the inducible NO-synthase (iNOS) was investigated by immunohistochemistry. Wound-derived fibroblasts were tested in vitro for NO and collagen synthesis. RESULTS: Irradiation significantly reduced wound collagen deposition and wound-breaking strength (P < 0.05). Impaired healing was reflected in diminished wound NO synthesis and iNOS expression (P < 0.01). TNF-alpha and IFN-gamma were increased in irradiated wounds (P < 0.05). Ex vivo, NO synthesis and collagen deposition by fibroblasts from irradiated rats were decreased (P < 0.01). In vitro irradiation of fibroblasts from nonirradiated rats decreased both NO and collagen production (P < 0.01). CONCLUSION: Radiation-impaired healing is reflected in an imbalance of promoting and inhibiting factors, leading to increased levels of TNF-alpha and IFN-gamma and decreased NO expression in the wound.