Up-regulation of chemokine C-C ligand 2 (CCL2) and C-X-C chemokine 8 (CXCL8) expression by monocytes in chronic idiopathic urticaria.

The disturbed cytokine-chemokine network could play an important role in the onset of diseases with inflammatory processes such as chronic idiopathic urticaria (CIU). Our main objectives were to evaluate the relation between proinflammatory chemokine serum levels from CIU patients and their response to autologous skin test (ASST) and basophil histamine release (BHR). We also aimed to assess the chemokine secretion by peripheral blood mononuclear cells (PBMC) upon polyclonal stimulus and to evaluate chemokine C-C ligand 2/C-X-C chemokine 8 (CCL2/CXCL8) and Toll-like receptor-4 (TLR-4) expression in monocytes. We observed significantly higher serum levels of the CXCL8, CXCL9, CXCL10 and CCL2 in CIU patients compared to the healthy group, regardless of the BHR or ASST response. The basal secretion of CCL2 by PBMC or induced by Staphylococcus aureus enterotoxin A (SEA) was higher in CIU patients than in the control group, as well as for CXCL8 and CCL5 secretions upon phytohaemagglutinin stimulation. Also, up-regulation of CCL2 and CXCL8 mRNA expression was found in monocytes of patients upon SEA stimulation. The findings showed a high responsiveness of monocytes through CCL2/CXCL8 expression, contributing to the creation of a proinflammatory environment in CIU.

Statin effects on regulatory and proinflammatory factors in chronic idiopathic urticaria.

Immunological dysfunction has been described to occur in chronic idiopathic urticaria (CIU), most notably in association with an inflammatory process. Some pharmacological agents as statins--drugs used in hypercholesterolaemia--display a broad effect on the immune response and thus should be tested in vitro in CIU. Our main objectives were to evaluate the effects of statins on the innate and adaptive immune response in CIU. Simvastatin or lovastatin have markedly inhibited the peripheral blood mononuclear cells (PBMC) proliferative response induced by T and B cell mitogens, superantigen or recall antigen. Simvastatin arrested phytohaemaglutinin (PHA)-induced T cells at the G0/G1 phase, inhibiting T helper type 1 (Th1), Th2, interleukin (IL)-10 and IL-17A cytokine secretion in both patients and healthy control groups. Up-regulation of suppressor of cytokine signalling 3 (SOCS3) mRNA expression in PHA-stimulated PBMCs from CIU patients was not modified by simvastatin, in contrast to the enhancing effect in the control group. Statin exhibited a less efficient inhibition effect on cytokine production [IL-6 and macrophage inflammatory protein (MIP)-1α] induced by Toll-like receptor (TLR)-4, to which a statin preincubation step was required. Furthermore, statin did not affect the tumour necrosis factor (TNF)-α secretion by lipopolysaccharide (LPS)-stimulated PBMC or CD14+ cells in CIU patients. In addition, LPS-activated PBMC from CIU patients showed impaired indoleamine 2,3-dioxygenase (IDO) mRNA expression compared to healthy control, which remained at decreased levels with statin treatment. Statins exhibited a marked down-regulatory effect in T cell functions, but were not able to control TLR-4 activation in CIU patients. The unbalanced regulatory SOCS3 and IDO expressions in CIU may contribute to the pathogenesis of the disease.

Impaired IFN-α secretion by plasmacytoid dendritic cells induced by TLR9 activation in chronic idiopathic urticaria.

BACKGROUND: Understanding the early events of the immune response, through the activation of plasmacytoid dendritic cells (pDC) by Toll-like receptor (TLR)9-sensing, could contribute to the evaluation of immune dysregulation in chronic idiopathic urticaria (CIU). OBJECTIVES: We decided to investigate innate immunity in CIU and the mechanisms implicated in the modulation of interferon (IFN)-α production by pDC upon TLR9 activation. METHODS: Patients with CIU (n = 31) and healthy control subjects (HC, n = 36) were enrolled in the study. Leucocytes cultured with the TLR9 ligand, CpG type A, or with inhibitory-oligodeoxynucleotide (ODN) were used to determine IFN-α secretion by enzyme-linked immunosorbent assay. Enumeration of pDC, intracellular IFN-α and signal transducers and activators of transcription protein (STAT) (1 and 4) phosphorylation were assessed by flow cytometry. TLR9 and regulatory factor-7 mRNA transcripts were evaluated by real-time polymerase chain reaction. Evidence of pDC in the skin lesions of patients was analysed with immunohistochemistry staining. RESULTS: The findings show a decreased IFN-α secretion induced by CpG A by leucocytes, due to the diminished IFN-α expression on pDC in CIU. It was mediated by TLR9-activation since inhibitory-ODN further suppressed TLR9-induced IFN-α secretion. A normal pDC percentage and degree of activation by the expression of costimulatory molecules was observed in CIU, with the rare presence of pDC in the skin lesion. In addition, an increased constitutive STAT1 phosphorylation on nonstimulated lymphocytes and a downregulation of TLR9 mRNA transcripts after CpG A activation were verified in patients with CIU. CONCLUSIONS: The findings showed an innate immune response in CIU disturbed by impairment of the pDC response to TLR9 activation.

IgA response in serum and gut secretion in sensitized mice fed with the dust mite Dermatophagoides pteronyssinus extract.

Induced oral tolerance to mucosal-exposed antigens in immunized animals is of particular interest for the development of immunotherapeutic approaches to human allergic diseases. This is a unique feature of mucosal surfaces which represent the main contact interface with the external environment. However, the influence of oral tolerance on specific and natural polyreactive IgA antibodies, the major defense mechanism of the mucosa, is unknown. We have shown that oral administration of an extract of the dust mite Dermatophagoides pteronyssinus (Dp) to primed mice caused down-regulation of IgE responses and an increase in tumor growth factor-beta secretion. In the present study, we observed that primed inbred female A/Sn mice (8 to 10 weeks old) fed by gavage a total weight of 1.0-mg Dp extract on the 6th, 7th and 8th days post-immunization presented normal secretion of IL-4 and IL-10 in gut-associated lymphoid tissue and a decreased production of interferon gamma induced by Dp in the draining lymph nodes (13,340 +/- 3,519 vs 29,280 +/- 2,971 pg/ml). Mice fed the Dp extract also showed higher levels of serum anti-Dp IgA antibodies and an increase of IgA-secreting cells in mesenteric lymph nodes (N = 10), reflecting an increase in total fecal IgA antibodies (N = 10). The levels of secretory anti-Dp IgA antibodies increased after re-immunization regardless of Dp extract feeding. Oral tolerance did not interfere with serum or secretory IgA antibody reactivity related to self and non-self antigens. These results suggest that induction of oral tolerance to a Dp extract in sensitized mice triggered different regulatory mechanisms which inhibited the IgE response and stimulated systemic and secretory IgA responses, preserving the natural polyreactive IgA antibody production.

IgA response in serum and gut secretion in sensitized mice fed with the dust mite Dermatophagoides pteronyssinus extract

Induced oral tolerance to mucosal-exposed antigens in immunized animals is of particular interest for the development of immunotherapeutic approaches to human allergic diseases. This is a unique feature of mucosal surfaces which represent the main contact interface with the external environment. However, the influence of oral tolerance on specific and natural polyreactive IgA antibodies, the major defense mechanism of the mucosa, is unknown. We have shown that oral administration of an extract of the dust mite Dermatophagoides pteronyssinus (Dp) to primed mice caused down-regulation of IgE responses and an increase in tumor growth factor-á secretion. In the present study, we observed that primed inbred female A/Sn mice (8 to 10 weeks old) fed by gavage a total weight of 1.0-mg Dp extract on the 6th, 7th and 8th days post-immunization presented normal secretion of IL-4 and IL-10 in gut-associated lymphoid tissue and a decreased production of interferon gamma induced by Dp in the draining lymph nodes (13,340 ñ 3,519 vs 29,280 ñ 2,971 pg/ml). Mice fed the Dp extract also showed higher levels of serum anti-Dp IgA antibodies and an increase of IgA-secreting cells in mesenteric lymph nodes (N = 10), reflecting an increase in total fecal IgA antibodies (N = 10). The levels of secretory anti-Dp IgA antibodies increased after re-immunization regardless of Dp extract feeding. Oral tolerance did not interfere with serum or secretory IgA antibody reactivity related to self and non-self antigens. These results suggest that induction of oral tolerance to a Dp extract in sensitized mice triggered different regulatory mechanisms which inhibited the IgE response and stimulated systemic and secretory IgA responses, preserving the natural polyreactive IgA antibody production.

Bone involvement in idiopathic hypercalciuria.

BACKGROUND: To evaluate bone involvement in idiopathic hypercalciuria, 40 lithiasic patients and 10 controls were studied. METHODS: According to urinary calcium excretion, patients were first classified as hypercalciuric (Hca, n = 22) and normocalciuric (Nca, n = 18). The Hca patients were then subclassified according to bone densitometry (BMD) as osteopenic (HcaO, n = 10) and non-osteopenic (HCaNO, n = 12). Routine biochemistry, dietary records, bone histomorphometry. and cytokines (IL-1beta, IL-6, and TNF) production by peripheral blood mononuclear cell cultures were studied. RESULTS: There were no differences in routine biochemistry between Hca and Nca groups, except for urinary calcium. Inadequate nutrition was observed in Hca group, showing high protein (80.9% of the patients), carbohydrate (76.2%) and sodium (90%) intake. Calcium intake was low in Hca (57%) and Nca (83%) groups. IL-6 and TNF were not different between the Hca and Nca groups. IL-1beta levels were significantly high in both groups when compared to controls. IL-6 and TNF were higher in HcaO than Nca. BMD in femoral neck in HcaO was lower than in HcaNO and Nca groups. Eroded surface (ES/BS) increased in 91% of the Hca group and 36% had a mineralization defect. In the HcaO group serum PTH correlated negatively with trabecular bone volume (BV/TV) and positively with ES/BS. 1,25(OH),D3 levels correlated positively with osteoblastic surface. Calcium intake correlated positively with BV/TV and inversely with ES/BS. A negative correlation was observed between IL-6 levels and Z score of the femoral neck. CONCLUSION: Bone involvement was detected in a young population with nephrolithiasis demonstrating that a strict follow-up is necessary in order to control hypercalciuria.

Oral tolerance induction to Dermatophagoides pteronyssinus and Blomia tropicalis in sensitized mice: occurrence of natural autoantibodies to immunoglobulin E.

BACKGROUND: The dust mites Dermatophagoides pteronyssinus (Dp) and Blomia tropicalis (Bt) are important sources of indoor allergens in tropical and subtropical countries. Murine models allow the analysis of the immune response and regulation of IgE production to Dp and Bt allergens. Oral tolerance induces unresponsiveness in naive animals, but its application in sensitized animals can provide useful information to improve allergy therapy. OBJECTIVE: To study the profile of IgE and IgG subclasses antibody upon oral administration with Bt and Dp extract in previously sensitized mice. Further, the occurrence of autoantibodies IgG anti-IgE in the immunization and in the oral tolerance was investigated. METHODS: A/Sn mice were immunized with Bt or Dp extract in alum, orally administrated with 0.25 mg of Bt or Dp extract or PBS at the 6th, 7th and 8th days after immunization and boosted twice with their respective allergens. To analyse the mice groups, specific IgE antibodies were measured by passive anaphylaxis reaction and specific IgG subclasses and anti-IgE IgG autoantibody by ELISA assay. RESULTS: IgE levels were markedly increased in Bt-immunized mice compared with Dp-immunized mice. A distinct profile of the specific isotypes was verified in Bt-immunized mice with a preferential production of IgG3 and IgA antibodies, whereas Dp-immunized mice developed high titres of anti-Dp IgG1, IgG2a and IgG2b antibodies. The antigen feeding inhibited IgE response in both fed-mice groups but only Dp-fed mice presented decreased levels of IgG antibodies. Free anti-IgE IgG autoantibodies were detected mainly in the Dp-immunization and they correlated with the antibody isotypes found against the allergen. CONCLUSIONS: This is the first time that the murine-type I hypersensitivity is employed to study Bt-immunization, showing a marked IgE production, associated with IgG response, which is at least in part driven by T-independent antigens. The oral tolerance protocol in previously sensitized animals was able to down-modulate IgE response and points out this route as a strategy for allergy therapy.

Oral tolerance induction in dermatophagoides pteronyssinus-sensitized mice induces inhibition of IgE response and upregulation of TGF-beta secretion.

Oral antigen administration induces peripheral tolerance in naive animals. Studies of oral tolerance induction in sensitized mice have clinical relevance as a strategy to modulate allergy. In this study, the A/Sn mice sensitized with extract of Dermatophagoides pteronyssinus (Dp) and submitted to oral Dp administration showed a marked decrease in IgE anti-Dp antibody production compared with sensitized phosphate-buffered saline (PBS)-fed mice. T cells from Dp-fed mice cocultured with spleen cells from PBS-fed mice were able to inhibit IgE anti-Dp antibody production and did not interfere in IgG1 antibody levels. The analysis of cytokine profile after Dp feeding showed a significant decrease in interleukin-4 (IL-4), IL-5, and IL-13 antigen-induced secretion levels by spleen cells, without shifting to IL-2 and interferon-gamma (IFN-gamma) production. Both transforming growth factor-beta (TGF-beta) baseline and TGF-beta antigen-stimulated levels were increased in Dp-fed mice. The effects of regulatory cytokines on anti-Dp IgE antibody production were investigated in vitro. The addition of recombinant TGF-beta (rTGF-beta) to spleen cell cultures stimulated by Dp inhibited IgE antibody secretion in both mouse groups. Neutralizing antibodies to IL-4, but not anti-TGF-beta, induced a marked inhibition of IgE production. Therefore, a negative modulatory effect on IgE response by inhibition of the axis Th2 was observed in sensitized Dp-fed mice, possibly mediated by induction of regulatory cytokines.

Immune response in hemodialysis patients: is there any difference when low and high iPTH levels are compared?

BACKGROUND: Chronic renal failure is frequently associated with secondary hyperparathyroidism and immunological disorders. Recent studies support the hypothesis that high levels of parathyroid hormone (PTH) may contribute to the impairment of the cellular and humoral immune response by an immunosuppressive effect on T- and B-cell functions. However, many studies indicate that excess PTH exerts a stimulatory effect on T lymphocytes. Since reports about the immunomodulatory effect of PTH are controversial, our aim was to compare the effect of low and high levels of intact PTH (iPTH) in hemodialysis patients. METHODS: The study was performed on 14 hemodialysis patients with high levels of iPTH (GI), 12 patients with low levels of iPTH (GII) and 13 volunteers (GIII), for whom time of dialysis, iPTH, total number of lymphocytes, B, CD4+, CD8+, lymphoproliferative response to phytohemagglutin (PHA), pokeweed mitogen (PWM) and candidin, IgG and IgM production in vitro in response to PWM, and interleukin (IL)-2 and IL-6 production in vitro in response to PHA were determined. RESULTS: Patients with high iPTH levels had significantly higher responses to PHA than patients with low iPTH. Lymphocyte transformation by PWM and candidin antigen was similar in both groups of patients, but significantly decreased when compared to controls. CD4+ cell counts were significantly increased in GI, and there was a positive correlation between the lymphoproliferative response to PHA and iPTH levels and CD4+ number. CONCLUSION: The present study suggests that high levels of iPTH in hemodialysis patients affect T-cell function, increasing the lympho-proliferative response to PHA and the CD4+ number.

Restoration of impaired T-cell proliferation after parathyroidectomy in hemodialysis patients.

BACKGROUND/AIMS: Severe secondary hyperparathyroidism is not infrequent in hemodialysis patients and recent studies suggest that parathyroid hormone (PTH) may play a role in the genesis of cell immunity abnormalities in uremia. The aim of the present study is to describe the effect of parathyroidectomy on T- and B-cell functions in hemodialysis patients. METHODS: The study was performed on 6 patients with severe secondary hyperparathyroidism. iPTH, B, CD4(+), CD8(+), total number of lymphocytes, lymphoproliferative response to PHA, PWM and Candidin, and IgG, IgM, IL-2 production in vitro were determined 1 day before and 4 months after parathyroidectomy. RESULTS: The lymphoproliferative response to PHA increased significantly after parathyroidectomy. We also observed a trend to an increase in production of IgG and IgM after PWM stimulation before therapy. CONCLUSION: The present study suggests that patients with extremely high levels of PTH show a complete restoration of impaired T-cell proliferation after parathyroidectomy.

The role of experimental chronic renal failure and aluminium intoxication in cellular immune response.

BACKGROUND: A positive correlation between successful kidney transplantation, few rejection episodes, greater susceptibility to infection and morbidity in patients with high tissue levels of aluminium (Al) indicate that the metal may play a role in the immune response. The aim of this study was to determine if experimental aluminium intoxication could result in significant changes in lymphocyte activity in uraemic and nonuraemic rats. METHODS: Lewis rats were divided into four groups: normals (N), nephrectomized control (U), and Al-treated (N + Al) and nephrectomized Al-treated (U + Al), which received a cumulative dose of 30 mg Al over a 4-week period. Al quantification, histology, histochemical analysis and immunological assays were performed after Al intoxication. RESULTS: High tissue levels of Al and positive histochemical staining in bones were seen in Al-treated rats. Bone histology revealed osteomalacia in U + Al rats. No statistical differences were observed in mixed lymphocyte cultures from controls and Al-treated rats, whereas U and Al-treated rats showed a decrease in lymphoproliferative response to mitogen and natural killer cell cytotoxic activity. A decreased helper T lymphocyte: cytotoxic T lymphocyte cell ratio and a reduction in interleukin-2 production were observed only in the U + Al group. A reduced number of total T lymphocytes was detected in the spleens of all Al-treated rats. CONCLUSIONS: These findings suggest that aluminium toxicity may contribute to immunological impairment in chronic renal failure.