Characterization of hypersensitivity reactions to polysulfone hemodialysis membranes.

BACKGROUND: In recent years, cases have been reported in which unexpected systemic hypersensitivity reactions occurred in patients dialyzed with polysulfone- or polyethersulfone-biocompatible membranes in the absence of other risk factors. The pathomechanisms involved in these reactions are largely unknown. OBJECTIVE: To characterize hypersensitivity reactions to polysulfone hemodialysis using clinical and laboratory data and to identify biomarkers suitable for endotype identification and diagnosis. METHODS: We prospectively collected data from 29 patients with suspected hypersensitivity reactions to polysulfone hemodialysis membranes. Clinical laboratory parameters such as tryptase, blood cell counts, and complement levels were recorded. Acute samples were obtained from 18 cases for the ex vivo assessment of basophil activation by flow cytometry analysis of CD63, CD203, and FcεRI cell membrane expression. Serum cytokines and anaphylatoxin concentrations were evaluated in 16 cases by Luminex and cytometric bead array analysis. RESULTS: Tryptase was elevated during the acute reaction in 4 cases. Evidence of basophil activation was obtained in 10 patients. Complement activation was found in only 2 cases. However, C5a serum levels tended to increase during the acute reaction in those patients with hypoxemia. Significantly higher serum levels of interleukin-6 were observed during the acute reactions to polysulfone hemodialysis (P = .0103). CONCLUSION: Based on biomarker analysis, various endotypes were identified, including type I-like (with the involvement of mast cells or basophils), complement, and cytokine (interleukin-6) release-related reactions, with some patients showing mixed reactions. Further research is needed to unravel the exact mechanisms involved in the activation of these cellular and molecular pathways.

Pre-transplant assessment of pp65-specific CD4 T cell responses identifies CMV-seropositive patients treated with rATG at risk of late onset infection.

Assessment of CMV-specific T cell immunity might be a useful tool in predicting CMV infection after solid organ transplantation. We have investigated CD4 and CD8 T-cell responses to CMV pp65 and IE-1 antigens in a prospective study of 28 CMV-seropositive kidney transplant recipients who were administered lymphocyte-depleting antibodies (Thymoglobulin®) as induction treatment and with universal prophylaxis for CMV infection. The response was analyzed by intracellular flow cytometry analysis of IFN-γ production in pretransplant samples and at 1, 6, 12 and 24 months post-transplant. Overall, only pretransplant CD4 T-cell responses to pp65 were significantly lower (p = .004) in patients with CMV replication post-transplant. ROC curve analysis showed that pre-transplant frequencies of pp65-specific CD4 + T cells below 0.10% could predict CMV infection with 75% sensitivity and 83.33% specificity (AUC: 0.847; 95% CI: 0.693-1.001; p = .0054) and seem to be mandatory for efficient control of CMV viral replication by the host immune system. In conclusion, the functional assessment of CMV-specific CD4 T-cell immunity pretransplant in seropositive patients may allow the identification of Thymoglobulin®-treated kidney transplant recipients at risk of developing CMV infection post-transplantation.

Mechanisms Involved in Hypersensitivity Reactions to Polysulfone Hemodialysis Membranes.

Several cases of patients with anaphylactic or systemic hypersensitivity reactions to polysulfone (PS) hemodialysis (HD) membranes and tolerance to cellulose triacetate (CTA) membranes have recently been reported. To investigate the mechanisms involved in PS hypersensitivity, basophil, T cell, and complement activation were analyzed in acute-phase samples from two patients with systemic reactions to PS-based membranes. Basophil and T cell activation, as well as higher serum tryptase levels were detected in acute-phase samples compared with basal levels. Complement levels (C3 and C4) were decreased in acute-phase samples from PS-allergic patients to a higher extent than in samples from control donors taken at the same time points, indicating complement activation during the acute reactions. An experimental external circuit was established on pediatric membranes after rinsing with low or high priming volumes of saline solution, to analyze basophils, T cells, and complement activation in blood samples from 10 PS-allergic and 8 nonallergic HD patients upon contact with PS-based or CTA membranes. Predialysis and postdialysis samples were collected. Basophils from PS-allergic patients exhibited increased degranulation, and T cells showed significantly increased activation after contact with PS-based membranes primed with low volumes of saline. No activation was detected in leukocytes from nonallergic patients under the same experimental conditions. Membrane priming with high volumes of saline abrogated activation of basophils and T cells. However, basophils from allergic donors showed significantly higher responses to Fcεc stimulation after contact with PS membranes. Basophil degranulation and elevated serum tryptase levels in allergic patients during acute reactions support the systemic activation of mast cells and basophils during hypersensitivity reactions to PS-based membranes. A leachable component of the membranes might be responsible for cell activation in some patients.

Alternative activation of macrophages in human peritoneum: implications for peritoneal fibrosis.

BACKGROUND: Depending on the cytokine microenvironment, macrophages (Mϕ) can adopt a proinflammatory (M1) or a profibrotic (M2) phenotype characterized by the expression of cell surface proteins such as CD206 and CD163 and soluble factors such as CC chemokine ligand 18 (CCL18). A key role for Mϕ in fibrosis has been observed in diverse organ settings. We studied the Mϕ population in a human model of peritoneal dialysis in which continuous stress due to dialysis fluids and recurrent peritonitis represent a risk for peritoneal membrane dysfunction reflected as ultrafiltration failure (UFF) and peritoneal fibrosis. METHODS: We used flow cytometry and quantitative reverse transcription-polymerase chain reaction to analyse the phenotype of peritoneal effluent Mϕ and tested their ability to stimulate the proliferation of human fibroblasts. Mϕ from non-infected patients were compared with those from patients with active peritonitis. Cytokine production was evaluated by enzyme-linked immunosorbent assay (ELISA) in spent dialysates and cell culture supernatants. RESULTS: CD206(+) and CD163(+) M2 were found within peritoneal effluents by flow cytometry analysis, with increased frequencies of CD163(+) cells during peritonitis (P = 0.003). TGFB1, MMP9 and CCL18 messenger RNA (mRNA) levels in peritoneal macrophages (pMϕ) were similar to those found in M2 cells differentiated in vitro. The ability of pMϕ to stimulate fibroblast proliferation correlated with CCL18 mRNA levels (r = 0.924, P = 0.016). CCL18 production by pMϕ was confirmed by immunostaining of cytospin samples and ELISA. Moreover, CCL18 effluent concentrations correlated with decreased peritoneal function, which was evaluated as dialysate to plasma ratio of creatinine (r = 0.724, P < 0.0001), and were significantly higher in patients with UFF (P = 0.0025) and in those who later developed sclerosing peritonitis (P = 0.024). CONCLUSIONS: M2 may participate in human peritoneal fibrosis through the stimulation of fibroblast cell growth and CCL18 production as high concentrations of CCL18 are associated with functional deficiency and fibrosis of the peritoneal membrane.

Sustained low peritoneal effluent CCL18 levels are associated with preservation of peritoneal membrane function in peritoneal dialysis.

Peritoneal membrane failure (PMF) and, ultimately, encapsulating peritoneal sclerosis (EPS) are the most serious peritoneal dialysis (PD) complications. Combining clinical and peritoneal transport data with the measurement of molecular biomarkers, such as the chemokine CCL18, would improve the complex diagnosis and management of PMF. We measured CCL18 levels in 43 patients' effluent and serum at baseline and after 1, 2, and 3 years of PD treatment by retrospective longitudinal study, and evaluated their association with PMF/EPS development and peritoneal risk factors. To confirm the trends observed in the longitudinal study, a cross-sectional study was performed on 61 isolated samples from long-term (more than 3 years) patients treated with PD. We observed that the patients with no membrane dysfunction showed sustained low CCL18 levels in peritoneal effluent over time. An increase in CCL18 levels at any time was predictive of PMF development (final CCL18 increase over baseline, p = .014; and maximum CCL18 increase, p = .039). At year 3 of PD, CCL18 values in effluent under 3.15 ng/ml showed an 89.5% negative predictive value, and higher levels were associated with later PMF (odds ratio 4.3; 95% CI 0.90-20.89; p = .067). Moreover, CCL18 levels in effluent at year 3 of PD were independently associated with a risk of PMF development, adjusted for the classical (water and creatinine) peritoneal transport parameters. These trends were confirmed in a cross-sectional study of 61 long-term patients treated with PD. In conclusion, our study shows the diagnostic capacity of chemokine CCL18 levels in peritoneal effluent to predict PMF and suggests CCL18 as a new marker and mediator of this serious condition as well as a new potential therapeutic target.

Synthesis and biological evaluation of pyridazino[1',6':1,2]pyrido[3,4-b]indolinium and pyridazino[1,6-a]benzimidazolium salts as anti-inflammatory agents.

Condensed polycyclic heteroaromatic cations bearing a bridgehead nitrogen with pyridazino[1',6':1,2]pyrido[3,4-b]indolinium and pyridazino[1,6-a]benzimidazolium structures were assayed as inhibitors of LPS-induced TNF-α production by THP-1 cells. The hit compound 1e, which had the best IC50 value (4.49 µM) and low toxicity, was further assayed on human PMBCs (IC50 3.91 µM) and monocytes (IC50 1.82 µM). This compound also inhibited TNF-α production following poly I:C stimulation of human monocytes and monocyte-derived dendritic cells; in the latter case, inhibition of IL-12 production was also observed. Compound 1e was also able to inhibit TNF-α expression at the transcriptional level and proved to be effective in vivo. Compound 1e is an interesting potential therapeutic agent in IMIDs.