Marianne Bol

Ultrafine but not fine particulate matter causes airway inflammation and allergic airway sensitization to co‐administered antigen in mice

Background Airborne particulate matter (PM) is an important factor associated with the enhanced prevalence of respiratory allergy. The PM adjuvant activity on allergic sensitization is a possible mechanism of action involved, and the induction of airway inflammation is suggested to be of importance in PM‐induced adjuvant activity.

Lung dendritic cells are stimulated by ultrafine particles and play a key role in particle adjuvant activity

BACKGROUND The adjuvant activity of air pollution particles on allergic airway sensitization is well known, but the cellular mechanisms underlying this adjuvant potential are not clear. OBJECTIVE We sough to study the role of dendritic cells and the costimulatory molecules CD80 and CD86 in the adjuvant activity of ultrafine carbon black particles (CBP). METHODS The proliferation of CFSE-labeled DO11.10 CD4 cells was studied after intranasal exposure to particles and ovalbumin (OVA). Next the frequency of myeloid dendritic cells (mDCs) and plasmacytoid dendritic cells and their expression of CD80 and CD86 were studied in the peribronchial lymph nodes (PBLNs). The expression of costimulatory molecules was also studied on bone marrow-derived mDCs after exposure to CBPs in vitro, and the importance of costimulation in CBP adjuvant activity was assessed by using CD80/CD86-deficient mice or cytotoxic T lymphocyte-associated antigen 4 (CTLA4)-Ig in vivo. RESULTS Our data show that CBPs plus OVA caused proliferation of DO11.10 CD4 cells and high levels of cytokine production in the PBLNs. Furthermore, the combined CBP plus OVA exposure increased the number of mDCs and expression of costimulatory molecules in the PBLNs. In addition, CBPs upregulated the expression of CD80/CD86 molecules on dendritic cells in vitro, which are necessary for the particle adjuvant effects in vivo. CONCLUSION Together this study shows the importance of dendritic cells and costimulation in particle adjuvant activity. Furthermore, we show for the first time that CBPs can also directly induce maturation of dendritic cells.

Effects of Supplementation With Vitamins A, C, and E, Selenium, and Zinc on Immune Function in a Murine Sensitization Model

OBJECTIVE We compared the effects of supplementing with vitamins A, C, and E, selenium, and zinc on a range of innate and specific T-helper 1 (Th1) and Th2-driven adaptive immune responses. METHODS BALB/c mice were fed semi-purified AIN93 diets and randomly assigned to receive a diet supplemented with 120 mg/kg of vitamin A, 2500 mg/kg of vitamin C, 1000 mg/kg of vitamin E, 2 mg/kg of selenium, and 500 mg/kg of zinc (n = 15/group). After 4 wk of supplementation, mice were sensitized by topical application of di-nitro-chlorobenzene (DNCB); 2 wk later mice were challenged; and 5 d later they were killed to assess the effect on a range of innate responses (phagocytic activity, oxidative burst and tumor necrosis factor-alpha), adaptive Th1-driven responses (delayed-type hypersensitivity, DNCB-specific immunoglobulin [Ig] G2a and IgG2b, and interferon-gamma [IFN-gamma]), and adaptive Th2-driven responses (DNCB-specific IgE and IgG1 and interleukin-4 [IL-4]). RESULTS Immune function was affected only in the vitamin A group. These mice gained less weight and were less capable of resolving the inflammatory response elicited during sensitization. The oxidative burst of blood cells was increased, but production of IFN-gamma and IL-4 and the ratio of IFN-gamma to IL-4 were markedly depressed. In concordance with the latter result, production of Th1-driven IgG2a antibodies was decreased, whereas Th2-driven isotypes were not affected (IgG1, IgE) and mucosal IgA was increased. CONCLUSIONS These findings confirmed that supplementary amounts of vitamin A above dietary requirements enhance inflammatory responses accompanied by decreased Th1 and increased mucosal responses. However, supplementation of these sufficiently fed, non-stressed, young adult mice with vitamins C and E, selenium, or zinc had no effect on immune function. We speculate that using this model in aged, physiologically, or nutritionally stressed mice may provide outcomes more similar to those in sensitive human populations. If so, this would improve the usefulness of the model to assess, characterize, and rank effects of foods or nutrients on a range of immune functions, including Th1/Th2 polarization.

Selective immunomodulation by the autoimmunity-inducing xenobiotics streptozotocin and HgCl2.

Exposure to certain drugs and environmental chemicals can provoke the onset of autoimmune disease in susceptible individuals by releasing (self) epitopes for which tolerance has not been established, while simultaneously providing the necessary adjuvant activity. The resulting response type is influenced by the genotype of exposed individuals and relates to susceptibility to the adverse immune effects of the chemicals. Here, we assessed the modulatory role of the chemical compounds themselves. A single injection of streptozotocin (STZ) increased the number of CD8+ cells, macrophages, apoptotic cells, and IFN‐γ‐producing T  helper and T cytotoxic cells, whereas the number of CD4+ cells and B cells was reduced in the draining lymphnode. Coinjection with the reporter antigen TNP‐OVA resulted in primary and secondary production of TNP‐specific antibodies that were predominantly of IgG2a and IgG2b isotype, whereas STZ did not enhance priming for delayed‐type hypersensitivity (DTH) responses to TNP‐OVA. Injection of HgCl2 on the other hand, reduced the number of IFN‐γ‐producing cells, induced accumulation of B cells and CD4+ and CD8+ T cells, enhanced IgG1 and IgE production to TNP‐OVA, and primed for secondary IgG1 and IgE production as well as for DTH reactions. Together these results indicate that a single injection of STZ stimulates type‐1 responses, whereas HgCl2 enhanced mixed type‐1 and −2 responses in BALB/c mice. These response types match the (auto)immune effects elicited to unknown (auto)antigens following multiple injections of these chemicals.

Selective Requirement for CD40-CD154 in Drug-Induced Type 1 Versus Type 2 Responses to Trinitrophenyl-Ovalbumin

CD154 is transiently expressed by activated T cells and interacts with CD40 on B cells, dendritic cells, macrophages, and monocytes. This costimulatory receptor-ligand couple seems decisive in Ag-driven immune responses but may be differentially involved in type 1 vs type 2 responses. We studied the importance of CD40-CD154 in both responses using the reporter Ag popliteal lymph node assay in which selectively acting drugs generate clearly polarized type 1 (streptozotocin) or type 2 (D-penicillamine, diphenylhydantoin) responses to a constant coinjected Ag in the same mouse strain. Treatment of mice with anti-CD154 reduced characteristic immunological parameters in type 2 responses (B and CD4+ T cell proliferation, IgG1 and IgE Abs, and IL-4 secretion) and only slightly affected the type 1 response (small decrease in IFN-γ production, influx of CD11c+ and F4/80+ cells, and prevention of architectural disruption of the lymph node, but no effect on IgG2a Ab and TNF-α secretion or B and CD4+ T cell proliferation). The findings indicate that the CD40-CD154 costimulatory interaction is a prerequisite in drug-induced type 2 responses and is only marginally involved in type 1 responses. The observed expression patterns of CD80 and CD86 on different APC (B cells in type 2 and dendritic cells in type 1) may be responsible for this discrepancy.

Immunotoxic organotins as possible model compounds in studying apoptosis and thymocyte differentiation

In the mid-seventies it appeared that some organotin compounds selectively caused thymus atrophy. From that time onward efforts were made to reveal molecular and cellular mechanisms involved. In this review recent studies into organotin-sensitive stages and processes of thymocyte maturation are discussed. Together these studies resulted in the recognition of organotin compounds as possible model compounds in studying immature thymocyte differentiation and protein synthesis-independent apoptotic cell death of thymocytes.

Cellular and molecular aspects of organotin-induced thymus atrophy

1 Organotin compounds, di-n-butyltin dichloride (DBTC) in particular, have been shown to cause Thymus atrophy in the rat. 2 DBTC-induced thymus atrophy results from a depletion of small CD4+CD8+ thymocytes which is caused by a diminished production of immature CD4-CD8+ and CD4+CD8+ thymoblasts. 3 DBTC inhibits the activation, but not the differentiation of immature CD4-CD8+ thymocytes in vitro and in vivo suggesting a selective anti-proliferative activity of DBTC. 4 DBTC inhibits the adhesion molecule-mediated binding of thymocytes to thymic epithelial cells. 5 DBTC enhances the Ca2+ release elicited by cross-linking of the T cell receptor complex (TcRαβ-CD3) on thymocytes and moreover delays cap formation of the TcRαβ-CD3 receptor. 6 It is concluded that DBTC possibly interferes with the functioning of the cytoskeleton. The relation of the in vitro findings to the inhibition of immature CD4-CD8+ thymocyte activation and the induction of thymus atrophy is unknown as yet.

Organotin-induced apoptosis occurs in small CD4+CD8+ thymocytes and is accompanied by an increase in RNA synthesis

The organotin compounds di-n-butyltin dichloride (DBTC) and tri-n-butyltin chloride (TBTC) induce thymus atrophy in rats. At low doses they inhibit immature thymocyte proliferation, whereas at higher doses in particular TBTC induces apoptotic cell death. In vitro, a similar concentration-effect relationship was observed, i.e. low concentrations inhibit DNA and protein synthesis and higher concentrations induce apoptosis. The mechanism of apoptosis by organotins has been partly investigated, but their capacity to inhibit protein synthesis seems to contradict with the idea that macromolecular synthesis is required for organotin-induced apoptosis. Therefore, we aimed to evaluate the relation between apoptosis and the synthesis of RNA and proteins, with a focus on the apoptosis-sensitive thymocyte subset. Results showed that DBTC increases RNA synthesis in particular in the subset of small CD4(+)CD8(+) thymocytes, which normally shows a high incidence of DNA fragmentation. Moreover, the RNA synthesis inhibitor actinomycin D or the protein synthesis inhibitor cycloheximide protected cells from apoptosis by DBTC or TBTC. Although organotin compounds increase synthesis of the heat shock protein HSC73/HSP72, heat shock treatment did not initiate apoptosis in thymocytes, neither antagonized organotin-induced apoptosis. This indicates that synthesis of heat shock proteins is not related to organotin-induced increase of RNA synthesis, and that other RNA-molecules are probably involved.

Effects of dietary lipids on immune function in a murine sensitisation model

We have tested the effect of dietary fatty acids on aspects of innate and specific adaptive T helper (Th) 1- and Th2-driven immune responses in a murine sensitisation model using dinitrochlorobenzene as sensitiser. Six groups of fifteen BALB/c mice were fed diets containing 30 % fat (by energy) for 8 weeks. Diets were rich in saturated fatty acids, n-6 polyunsaturated fatty acid (PUFA), or n-3 PUFA, each at a sufficient (11, 35 and 68 mg/kg) and a supplemented vitamin E level (1028, 1031 and 1030 mg/kg respectively). Feeding n-6 PUFA marginally decreased % phagocytosing cells at the low vitamin E level, but had no other effects on immune function. The n-3 PUFA diets decreased production of prostaglandin E2 while increasing oxidative burst and tumour necrosis factor alpha production. In addition adaptive Th1-driven responses (immunoglobulin, Ig)G2a, IgG2b, interferon-gamma:interleukin 4) were decreased, whereas Th2-driven and mucosal immune responses were increased (IgE) or unaffected (IgG1, IgA). Combination with high levels of alpha-tocopherol did not affect the reduced prostaglandin E2 production, augmented the increase of tumour necrosis factor alpha production and tended to ameliorate the selective suppressive effects of n-3 PUFA on certain Th1-driven effects (interferon-gamma:interleukin 4 ratio and IgG2a levels). We conclude that the sensitisation model appears useful for application in nutrition research. It allows a broad assessment of the effects of dietary intervention on various aspects of immune responsiveness, and as such provides a valuable model to assess, characterise and rank effects of foods and/or nutrients on a range of immune functions, including Th1-Th2 polarisation.

Differential Requirement for CD28/CTLA-4-CD80/CD86 Interactions in Drug-Induced Type 1 and Type 2 Immune Responses to Trinitrophenyl-Ovalbumin

The use of mAbs to abrogate costimulatory interactions has attracted much attention with regard to prevention and modulation of adverse (auto)immune-like reactions. However, the role of costimulatory molecules and possible therapeutic use of Ab-treatment in drug-induced immunostimulation is poorly elucidated. In the present studies, we show that CD28/CTLA-4-CD80/CD86 costimulatory interactions differently regulate drug-induced type 1 and type 2 responses to an identical bystander Ag, TNP-OVA, in BALB/c mice using the reporter Ag popliteal lymph node assay. The antirheumatic drug D-Penicillamine, which may induce lupus-like side-effects, stimulated type 2 responses against TNP-OVA, characterized by the production of IL-4 and TNP-specific IgG1 and IgE. These responses were abrogated in CD80/CD86-deficient mice and in wild-type mice that were treated with anti-CD80 and anti-CD86, or CTLA-4-Ig. Anti-CTLA-4 intensively enhanced the D-Penicillamine-induced effects. In contrast, the type 1 response (IFN-γ, TNF-α, IgG2a) to TNP-OVA induced by the diabetogen streptozotocin still developed in the absence of CD80/CD86 costimulatory signaling. In addition, it was demonstrated that coadministration of anti-CD80 and anti-CD86 mAbs slightly enhanced streptozotocin-induced type 1 responses, whereas the CTLA-4-Ig fusion protein completely abrogated this response. In conclusion, different drugs may stimulate distinct types of immune responses against an identical bystander Ag, which are completely dependent on (type 2) or independent of (type 1) the CD28/CTLA-4-CD80/CD86 pathway. Importantly, the effects of treatment with anti-CD80/CD86 mAbs and CTLA-4-Ig may be considerably different in responses induced by distinct drugs.