The combination of the neutrophil-to-lymphocyte and platelet-to-lymphocyte ratios as a novel predictor of intravenous immunoglobulin resistance in patients with Kawasaki disease: a multicenter study.

INTRODUCTION: The neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) have been reported to be a predictor for intravenous immunoglobulin (IVIG) resistance in patients with Kawasaki disease (KD) recently. The objective of the present study was to elucidate the predictive validity of this new marker in a multicenter study. MATERIALS AND METHODS: We retrospectively reviewed the clinical records of 520 consecutive KD patients (development data set) and 332 subsequent patients (validation data set) at 7 hospitals in Japan. RESULTS: Both NLR and PLR were significantly higher in the IVIG-resistant group than in the IVIG-responsive group. When we set the cut-off point as NLR ≥ 4.11 and PLR ≥ 119, multiple logistic regression analyses showed that a high NLR and PLR before initial IVIG were independent predictors of IVIG resistance, and their combination was a stronger predictor than either alone. The sensitivity and specificity of the combination of NLR ≥ 4.11 and PLR ≥ 119 were 0.58 and 0.73 in the development data set. Validated using an independent data set, they were 0.54 and 0.72 in the validation data set. On comparing the AUC of this predictor with those of the Gunma and Kurume scores, the AUC was highest for this predictor, followed by the Gunma score and Kurume score (0.70, 0.68, and 0.64, respectively). DISCUSSION: The predictive validity of the combination of a high NLR and PLR, which is a simple and convenient indicator, was equal to or better than that of the existing scoring systems. The new predictive marker may be a suitable indicator for predicting IVIG resistance in KD patients.

Enhanced formation of neutrophil extracellular traps in Kawasaki disease.

BACKGROUND: Neutrophils contribute to the clearance of pathogens through the formation of neutrophil extracellular traps (NETs) in a process known as NETosis, but the excessive release of NETs has been reported to be involved in the pathogenesis of various diseases, including vasculitis, by inducing tissue injury. The aim of the present study was to investigate whether or not NETosis is enhanced in the acute phase of Kawasaki disease (KD). METHODS: After neutrophils isolated from the peripheral blood of patients with KD and healthy control (HC) were cultured in vitro, the degree of spontaneous NETosis was evaluated by measuring the number of NETs formed and the titers of cell-free DNA (cfDNA) and neutrophil elastase (NE)-DNA complex. RESULTS: Spontaneous NET formation in vitro was observed in neutrophils isolated from KD patients, and the number of NET formations was significantly higher in acute KD than in convalescent KD and HC. The increased levels of cfDNA and NE-DNA complexes in the acute phase of KD tended to decrease in the convalescent phase. CONCLUSIONS: Spontaneous NET formation was enhanced in neutrophils from patients with acute KD, suggesting that circulating neutrophils may be primed to undergo NETosis in KD vasculitis.

A comparison of the predictive validity of the combination of the neutrophil-to-lymphocyte ratio and platelet-to-lymphocyte ratio and other risk scoring systems for intravenous immunoglobulin (ivig)-resistance in Kawasaki disease.

BACKGROUND: We recently reported that the combination of the neutrophil-to-lymphocyte ratio (NLR) and the platelet-to-lymphocyte ratio (PLR) is a novel and useful predictor of intravenous immunoglobulin (IVIG)-resistance in Kawasaki disease (KD). In the present study, to evaluate the effectiveness of the new risk score, we compared its predictive validity to that of previously reported risk scores. MATERIALS AND METHODS: The laboratory records of 437 patients with KD before IVIG therapy were retrospectively analyzed, and the IVIG-responsive (n = 344) and IVIG-resistant (n = 93) patients were compared. The validity of the new score (the combination of NLR≥3.83 and PLR≥150) for predicting IVIG resistance in KD was compared to that of the Kobayashi, Egami and Sano risk scores. RESULTS: The new score and the Kobayashi score displayed high sensitivity (0.72 and 0.70 respectively) and specificity (0.67 and 0.68 respectively), while the Egami and Sano scores showed high specificity (0.71 and 0.81 respectively) but relatively low sensitivity (0.56 and 0.45 respectively). The odds ratios (ORs) for the new score, the Kobayashi score, the Egami score and the Sano score were 5.34 (95% confidence interval [CI] 3.22-8.85), 4.87 (95% CI 2.96-8.01), 3.14 (95% CI 1.96-5.03) and 3.53 (95% CI 2.17-5.77) respectively. CONCLUSIONS: The predictive validity of the combination of NLR≥3.83 and PLR≥150, which is a simple and convenient indicator, was equal to or higher than that of the other risk scores. This suggests that the new score could be a widely available marker for predicting IVIG resistance in KD.

Urinary Lactate Dehydrogenase Activity and Its Isozyme Patterns in Kawasaki Disease.

Abnormal urinary findings, such as sterile pyuria, proteinuria, and microscopic hematuria, are often seen in the acute phase of Kawasaki disease (KD). We investigated the potential significance of urinary lactate dehydrogenase (U-LDH) activity and its isozyme patterns in KD. Total U-LDH activity and its isozymes (U-LDH1-5) levels were compared among 120 patients with KD, 18 patients with viral infection (VI), and 43 patients with upper urinary tract infection (UTI) and additionally compared between intravenous immunoglobulin (IVIG) responders (n = 89) and nonresponders (n = 31) with KD. Total U-LDH activity was higher in KD (35.4 ± 4.8 IU/L, P < 0.05) and UTI patients (66.0 ± 8.0 IU/L, P < 0.01) than in VI patients (17.0 ± 6.2 IU/L). In the isozyme pattern analysis, KD patients had high levels of U-LDH1 and U-LDH2, while UTI patients had high levels of U-LDH3, U-LDH4, and U-LDH5. Furthermore, IVIG nonresponders of KD had significantly higher levels of total U-LDH activity (45.1 ± 4.7 IU/L, P < 0.05), especially U-LDH1 and U-LDH2 (P < 0.05), than IVIG responders (32.0 ± 2.8 IU/L). KD patients have increased levels of total U-LDH activity, especially U-LDH-1 and U-LDH2, indicating a unique pattern of U-LDH isozymes different from that in UTI patients.

The Combined Usefulness of the Neutrophil-to-Lymphocyte and Platelet-to-Lymphocyte Ratios in Predicting Intravenous Immunoglobulin Resistance with Kawasaki Disease.

The laboratory records of 405 patients with Kawasaki disease before and after intravenous immunoglobulin (IVIG) therapy were compared between the IVIG-responsive (n = 320) and IVIG-resistant (n = 85) groups. A high neutrophil-to-lymphocyte ratio and a high platelet-to-lymphocyte ratio before IVIG, especially when combined, were useful predictors for IVIG resistance in Kawasaki disease.

[Use of the urinary trypsin inhibitor ulinastatin for acute Kawasaki disease].

Intravenous immunoglobulin (IVIG) therapy is widely recognized as standard treatment for Kawasaki disease(KD). However, about 20 % of KD patients are resistant to IVIG and are considered to be a high risk group for coronary artery lesions (CAL). Ulinastatin(UTI) is one of the neutrophil elastase inhibitors used for patients with pancreatitis or circulatory shock, and several studies have shown its efficacy for KD. Recently, we demonstrated that initial UTI treatment combined with IVIG decreased the number of patients requiring addi- tional rescue treatment and the occurrence of CAL. In this study, no severe adverse events occurred. Further research and a prospective trial are needed to prove the clinical efficacy and demonstrate the limits of UTI in patients with KD.

Ulinastatin, a urinary trypsin inhibitor, for the initial treatment of patients with Kawasaki disease: a retrospective study.

BACKGROUND: Markedly activated neutrophils or higher plasma levels of neutrophil elastase are involved in the poor response to intravenous immunoglobulin (IVIG) and the formation of coronary artery lesions (CAL) in patients with acute Kawasaki disease. We hypothesized that ulinastatin (UTI), by both direct and indirect suppression of neutrophils, would reduce the occurrence of CAL. METHODS AND RESULTS: We retrospectively analyzed the clinical records of patients with Kawasaki disease between 1998 and 2009. Three hundred sixty-nine patients were treated with a combination of UTI, aspirin, and IVIG as an initial treatment (UTI group), and 1178 were treated with a conventional initial treatment, and IVIG with aspirin (control group). The baseline characteristics did not demonstrate notable differences between the two groups. The occurrence of CAL was significantly lower in the UTI group than in the control group (3% versus 7%; crude odds ratio [OR], 0.46; 95% confidence interval [CI], 0.25-0.86; P=0.01). The OR adjusted for sex, Gunma score (the predictive score for IVIG unresponsiveness), and dosage of initial IVIG (1 or 2 g/kg) was 0.32 (95% CI, 0.17-0.60; P<0.001). In addition, most CAL occurred in patients requiring additional rescue treatment and the proportion of those patients was significantly lower in the UTI group than in the control group (13% versus 22%; crude OR, 0.52; 95% CI, 0.38-0.73; P<0.001). The adjusted OR was 0.30 (95% CI, 0.20-0.44; P<0.001). CONCLUSIONS: UTI was associated with fewer patients requiring additional rescue treatment and reduction of CAL in this retrospective study.

Circulating adiponectin levels in Kawasaki disease.

AIM: To investigate the kinetics of adipocytokine in Kawasaki disease (KD). METHODS: The plasma levels of adiponectin, IL-6 and TNF-alpha were measured in 20 patients with KD, 15 patients with acute febrile diseases (disease control, DC) and 15 healthy children (HC). RESULTS: Plasma adiponectin levels in patients with acute KD were significantly lower (p < 0.01) than those with convalescent KD, DC and HC. The decreased levels of adiponectin negatively correlated with the increased levels of CRP and IL-6 in acute KD. CONCLUSION: Hypoadiponectinaemia was shown during the acute phase of KD vasculitis. The downregulation of adiponectin secreted from adipose tissue may be closely linked to upregulation of systemic pro-inflammatory markers in acute KD.

Standard-dose and short-term corticosteroid therapy in immunoglobulin-resistant Kawasaki disease.

Six patients with Kawasaki disease (KD) were treated with prednisolone (1 to 2 mg/kg/day) for 3 days (from days 10 to 12 after the onset of the illness) after apparently unsuccessful treatment with intravenous immunoglobulin (IVIG, 2 g/kg/dose and additional 1 g/kg/dose). Five patients responded immediately to the first course of prednisolone infusion. One patient failed to respond to the first course of prednisolone therapy, but he did respond to the second 3-day course of therapy. None of the patients demonstrated a further progression of coronary artery dilatation or any adverse effects. Standard-dose and short-term corticosteroid therapy therefore appears to be a safe and effective treatment for patients with IVIG-resistant KD.

Intravenous immunoglobulin preparations promote apoptosis in lipopolysaccharide-stimulated neutrophils via an oxygen-dependent pathway in vitro.

Since prolonged survival of activated neutrophils has an autotoxic potential, neutrophil apoptosis plays an important role in the rapid resolution of inflammation. Intravenous immunoglobulin (IVIG) preparations, which are beneficial therapeutic agents for the treatment of autoimmune diseases and systemic inflammatory diseases, have been reported to induce apoptosis of lymphocytes and endothelial cells in vitro. In the present study, we investigated whether IVIG may induce apoptosis of neutrophils cultured in vitro. After neutrophils prestimulated with or without lipopolysaccharide (LPS) were cultured in the presence or absence of IVIG, the number of apoptotic cells, intracellular H2O2 and GSH were measured by a flow cytometer. IVIG induced apoptosis of LPS-stimulated neutrophils dose dependently, but not in unstimulated neutrophils. Although anti-Fas monoclonal antibodies (mAbs) had no effect on the IVIG-induced apoptosis in the LPS-stimulated neutrophils, anti-Fc gamma receptor (Fc gammaR) II- and III-blocking mAbs significantly inhibited the IVIG-induced apoptosis. IVIG increased the production of intracellular H2O2, while it decreased the production of GSH, in the LPS-stimulated neutrophils. Furthermore, a specific NADPH oxidase inhibitor and anti-oxidants inhibited the IVIG-induced neutrophil apoptosis. Therefore, these findings indicate that IVIG preparations induce apoptosis of LPS-stimulated neutrophils and suggest that the IVIG-induced apoptosis may be mediated by an oxygen-dependent pathway via Fc gammaRII and III.

Intravenous immunoglobulin therapy induces neutrophil apoptosis in Kawasaki disease.

To investigate the effect of high-dose intravenous immunoglobulin (IVIg) on neutrophil apoptosis in Kawasaki disease (KD), we studied the in vitro spontaneous and IVIg-induced apoptosis of neutrophils by analyzing a proportion of annexin V-positive cells and cells with fragmented DNA. The mean number of peripheral neutrophils in the post-IVIg phase decreased significantly (P < 0.01) compared with that in the pre-IVIg phase. The mean proportion of spontaneous apoptotic neutrophils in the post-IVIg phase was significantly higher (P < 0.01) than that in the pre-IVIg phase, and there was a significantly positive correlation (P < 0.01) with the reduction ratio of the circulating neutrophil counts from the pre-IVIg through the post-IVIg phases. IVIg induced a dose-dependent increase in the proportion of apoptotic neutrophils in the pre-IVIg phase. As a result, the present study demonstrated a novel action in which high-dose IVIg therapy decreased the number of circulating neutrophils by accelerating their apoptosis in KD.

Increased production of serum IgA-class antibody to lipid A in Kawasaki disease.

BACKGROUND: The etiology of Kawasaki disease (KD) remains unknown. To investigate whether a conventional bacterial antigen is involved in the pathogenesis of KD, we studied the serum response to lipopolysaccharide (LPS). METHODS: We measured the serum levels of IgG-, IgM- and IgA-class antibodies (Ab) to lipid A, a toxic site of LPS, using enzyme-linked immunosorbent assay in 20 patients with KD, 11 patients with Gram-negative bacterial infection (GNBI), 27 healthy children and 12 healthy adults. RESULTS: The serum levels of anti-lipid A IgG, IgM and IgA tended to increase with advancing age in healthy children older than 6 months of age. The mean level of anti-lipid A IgM in the acute phase of GNBI and the mean levels of anti-lipid A IgM and IgA in the acute phase of KD were found to increase significantly, in comparison to the age-matched controls. Furthermore, the mean level of anti-lipid A IgA also showed a significant increase from the acute to the subacute phases of KD. Regarding the IgA-subclass response, higher titers of anti-lipid A specific Ab were seen in the IgA2 subclass than in the IgA1 subclass. CONCLUSION: These findings indicate that KD patients demonstrate an intense response to lipid A in the IgA, especially IgA2-subclass, thus suggesting that an unusual activation of the mucosal immune response to a ubiquitous antigen derived from Gram-negative bacteria may be involved in the pathogenesis of KD.