Rapidly progressive IgA vasculitis-associated nephritis successfully treated with immunosuppressive therapy in an adolescent with chronic granulomatous disease.

Chronic granulomatous disease (CGD) is a rare immunodeficiency disorder with genetic defects in the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex in phagocytes, leading to recurrent severe infections and granuloma formation. Genitourinary involvement, including obstructive granulomas, infections, nephrotoxicity of anti-infective agents, and amyloidosis, is frequently observed in patients with CGD, whereas the clinical and pathological details of the less commonly reported glomerular disease remain obscure. Here, we report the case of a patient with CGD who developed rapidly progressive IgA vasculitis-associated nephritis (IgAVN) and review the literature on biopsy-proven glomerular diseases in patients with CGD. A 22-year-old male patient with CGD developed rapidly progressive glomerulonephritis (RPGN) following peripheral purpura and was diagnosed with crescentic IgAVN based on the renal biopsy evaluation. There was no evidence of active infections, and he received pulse intravenous methylprednisolone followed by oral prednisolone. His renal function returned to normal within 4 weeks, and his proteinuria and microhematuria finally resolved. The present case and literature review indicate that IgAVN and IgA nephropathy with RPGN are the most common causes of glomerular disease in patients with CGD. Clinicians should be aware of the possibility of these diseases as causes of RPGN in CGD, because delays in diagnosis and appropriate treatment may affect renal outcomes.

Novel use of rituximab for steroid-dependent nephrotic syndrome in children.

BACKGROUND: Though rituximab (RTX) is effective for childhood steroid-dependent nephrotic syndrome (SDNS), an established regimen does not exist. The relapses tend to occur when the peripheral blood B-cell count re-arises at 3 months upon single RTX infusion. This study was conducted to clarify whether the long-term remission of SDNS can be obtained by repeated RTX administrations. METHODS: RTX was administered 4 times at 3-month intervals at 375 mg/m(2)/time to 5 children with SDNS. The changes in the clinical indicators were analyzed. RESULTS: The median (range) observation period was 6.3 (0.9-8.4) years before RTX and 3.2 (1.9-3.8) years following the commencement of RTX. The changes in the clinical indicators were as follows (median and range): (1) annual number of relapses: before administration 1.4 (1.1-3.5) times/year, after administration 0.0 (0.0-0.0) times/year, and (2) median steroid dosage: before administration 0.80 (0.23-0.96) mg/kg/day, after administration 0.00 (0.00-0.00) mg/kg/day. All changes were significant at p < 0.05. Relapse occurred 3 times following the start of RTX (the period to relapse was 2.2, 1.9, and 2.3 years, respectively). No serious side effects were seen. CONCLUSIONS: Repeated RTX against SDNS in children may be a useful therapeutic option.

Different phenotypes of HNF1ß deletion mutants in familial multicystic dysplastic kidneys.

Multicystic dysplastic kidney (MCDK) is one of the most common congenital abnormalities of the kidney and urinary tract (CAKUT), although its pathophysiology remains unknown. Familial occurrence of MCDK suggests that mutations in genes associated with nephrogenesis are involved in the pathogenesis in at least some cases. Hepatocyte nuclear factor 1ß (HNF1ß) is a member of the homeodomain-containing super family of transcription factors, and is known to regulate tissue-specific gene expression in a number of organs including the kidneys, pancreas and liver. It has been recently postulated to be associated with CAKUT, including MCDK. We recently encountered a family with a deletion mutant of HNF1ß, of which the 2nd son, the proband, developed bilateral MCDK resulting in renal loss of function in infancy while the 1st son developed unilateral MCDK. Their father has two normal kidneys. This family confirmed that mutations in the HNF1ß gene are strongly associated with the development of MCDK. Furthermore, the fact that no clear phenotype-genotype correlation exists suggests that gene(s) other than HNF1ß are also involved in nephrogenesis and the development of MCDK.

Increased production of nitric oxide by phagocytic stimulated neutrophils in patients with chronic granulomatous disease.

Chronic granulomatous disease (CGD) is an inherited disorder in which phagocytic leukocytes fail to generate superoxide (O(2)(-)) and antimicrobial oxidants. Patients with CGD develop recurrent and often life-threatening infections due to catalase-positive microorganisms. We attempted to measure the production of nitric oxide and reactive oxygen species in polymorphonuclear neutrophils (PMNs) from patients with CGD using a flow cytometry technique. Venous blood was obtained from 5 male patients with X-linked CGD and from 10 healthy volunteers. The nitric oxide production by PMNs after phagocytosis was measured using diaminofluorescein-2 diacetate, a fluorescent indicator of intracellular nitric oxide production. After erythrocytes were hypotonically lysed, the cell pellet was applied to a cytofluorometer. Although the production of hydrogen peroxide by PMNs from patients with CGD was almost absent, nitric oxide production was detected at nearly half the level as was seen in the PMNs from healthy volunteers (CGD vs. healthy volunteers=140.1±28.6 vs. 256.4±10.3, mean fluorescence intensity; P<0.01). In conclusion, we demonstrated that human PMNs produces nitric oxide after phagocytic stimulation. Also nitric oxide production after phagocytic stimulation by PMNs of patients with CGD is observed although its amount is lower than that observed on healthy control. Despite the fact that CGD patients cannot produce H(2)O(2) which is essential for intracellular bacteriocidal process after phagocytosis, our data suggested that the phylactic effect in PMNs induced by nitric oxide could be at least partially related to the survival of patients with CGD.

Angiotensin type 1a receptor signaling is not necessary for the production of reactive oxygen species in polymorphonuclear leukocytes.

Background. Although angiotensin II (Ang II) has inflammatory effects, little is known about its role in polymorphonuclear leucocytes (PMLs). To elucidate the role of Ang II in PMLs ROS production, we examined hydrogen peroxide (H2O2), one of the ROS, and NO production in AT1a receptor knockout (AT1KO) mice. Methods and Results. PMLs were analyzed from Ang II type 1a receptor knockout mice (AT1KO) and C57BL/6 wild type mice. Using flow cytometry, we studied hydrogen peroxide (H2O2) production from PMLs after Staphylococcus aureus phagocytosis or phorbol myristate acetate (PMA) stimulation. Nitric oxide (NO) production in the AT1KO was low at basal and after phagocytosis. In the AT1KO, basal H2O2 production was low. After PMA or phagocytosis stimulation, however, H2O2 production was comparable to wild type mice. Next we studied the H2O2 production in C57BL/6 mice exposed to Ang II or saline. H2O2 production stimulated by PMA or phagocytosis did not differ between the two groups. Conclusions. AT1a pathway is not necessary for PMLs H2O2 production but for NO production. There was a compensatory pathway for H2O2 production other than the AT1a receptor.

Diagnosis of autoimmune neutropenia by neutrophil-bound IgG and IgM antibodies.

Autoimmune neutropenia (AIN) in infancy is caused by antineutrophil (granulocyte-specific) autoantibodies. These antibodies are rarely found in circulation because their serum levels are extremely low. We hypothesized that a direct granulocyte immunofluorescence test (D-GIFT) that enables us to detect neutrophil-bound autoantibodies consisting of both immunoglobulin (Ig) G and IgM has better diagnostic value than the detection of circulating autoantibodies. Whole blood (100 µL) was obtained from 50 infants with AIN, 12 infants with transient neutropenia, and 37 control infants. D-GIFT was performed using both fluorescein isothiocyanate-conjugated antihuman IgG Fc portion monoclonal antibodies and fluorescein isothiocyanate antihuman IgM monoclonal antibodies. Results were assessed as relative fluorescence intensity (RFI). The RFIs of antineutrophil IgG-bound and antineutrophil IgM-bound cells in patients with AIN were significantly higher than those in patients with transient neutropenia and in controls. Positive results, as assessed by RFI scores of more than 1.81 in either antineutrophil IgG-bound or antineutrophil IgM-bound cells, showed the sensitivity and specificity of D-GIFT, and the areas under the receiver operating characteristic curve (0.98, 0.98, and 0.997, respectively) in the diagnosis of AIN. D-GIFT detecting both neutrophil-bound IgG autoantibodies and IgM autoantibodies has discriminatory power for identifying patients with AIN and, therefore, can be a useful diagnostic test.

A family with X-linked benign familial hematuria.

Gene mutations in COL4A5 located on Xq22 are believed to cause X-linked Alport syndrome, whereas mutations in COL4A3 and COL4A4 located on chromosome 2 are associated with autosomal inherited Alport syndrome or benign familial hematuria. A family with benign familial hematuria caused by COL4A5 mutation, implying X-linked transmission, is reported here for the first time. This result suggests that COL4A5 should be added to the list of causative genes for benign familial hematuria, although the mechanism(s) by which the same mutation leads to the distinct phenotypes, i.e. X-linked Alport syndrome or benign familial hematuria, remains unknown.

Phagocytosis of heat-killed Staphylococcus aureus by eosinophils: comparison with neutrophils.

Eosinophils are characterized by several functional properties, such as chemotaxis, adhesion, superoxide anion production, and degranulation. In this article, we have studied the role of bacterial ingestion by eosinophils in comparison with that by neutrophils. Eosinophils and neutrophils were purified by using the Percoll gradient method followed by selection with CD16-coated immunomagnetic beads and centrifugation through a Ficoll-Hypaque gradient combined with dextran sedimentation, respectively. Both cells were preincubated with anti-FcgammaRIIa mAb (CD32 mAb), anti-FcgammaRIIIb mAb (CD16 mAb), anti-CR3 (CD11b mAb), or anti-CR1 (CD35 mAb) before being examined for phagocytosis of opsonized heat-killed Staphylococcus aureus (S. aureus). Phagocytosis and production of hydrogen peroxide were simultaneously measured by flow cytometry using S. aureus labeled with propidium iodide and stained with 2',7'-dichlorofluorescein diacetate. Eosinophils showed significantly lower activity than neutrophils in both phagocytosis and hydrogen peroxide production. Phagocytosis by both cells was decreased by heat-inactivated serum. Phagocytosis by neutrophils was significantly inhibited by CD16 mAb and CD32 mAb, whereas that by eosinophils was only inhibited by CD35 mAb. Whereas the mechanism of phagocytosis by neutrophils was mediated by CD16 and CD32, that of eosinophils was modulated by complement receptor 1 (CD35).

Increased nitric oxide production by T- and B-cells in idiopathic nephrotic syndrome.

The pathogenesis of idiopathic nephrotic syndrome (INS) remains unclear. To study the role of nitric oxide (NO) in INS, we measured intracellular NO produced by T- and B-cells using a novel fluorescent indicator. Twelve children with INS (mean age 7.3 years; group A-1: in relapse, group A-2: in remission) were enrolled in the study together with 16 children with other renal diseases (9.5 years; group B) and 42 healthy control children (7.7 years; group C). The amount of NO produced by CD3+ cells (T-cells) and CD19+ cells (B-cells) and of plasma NO(x) was measured by flow cytometry and colorimetry, respectively. The average amount of NO produced by CD3+ and CD19+ cells in group A-1 subjects was significantly higher than that produced by these cells in group A-2 and B patients and the healthy controls (group C), respectively (P < 0.01), and it decreased after the patients achieved remission (P < 0.01). Plasma NO(x) levels in group A-1 patients was also highest among the different groups (P < 0.01). There were no significant differences in intracellular NO and plasma NO(x) among group A-2, B, and C subjects (P > 0.05). A significant correlation between plasma NO(x) and urinary protein excretion was found only in group A patients and not in group B patients. We conclude that an aberrant immune system may exist not only in T-cells but also in B-cells, and NO may play some role in INS.

Oxidative imbalance in idiopathic renal hypouricemia.

An important complication of idiopathic renal hypouricemia is exercise-induced acute renal failure (ARF). The most plausible explanation for this complication is that decreased antioxidant potential leads to kidney injury by reactive oxygen species (ROS). We demonstrated this oxidative imbalance by a concomitant assessment of ROS production and antioxidant system capability in a 15- year-old girl with idiopathic renal hypouricemia caused by a mutation in the urate transporter (URAT1) gene. Her serum level of ROS increased with decreasing antioxidant potential capacity soon after the initiation of anaerobic stress due to treadmill exercise. Thereafter, serum levels of ROS and antioxidant potential showed a parallel course, returning to the baseline values at 240 min after exercise. Some patients with idiopathic renal hypouricemia demonstrate oxidative imbalance soon after exercise with a predisposition to exercise-induced acute renal failure. Antioxidant properties may alter this imbalance by augmenting the antioxidant activity.

Increased nitric oxide production by neutrophils from patients with chronic granulomatous disease on trimethoprim-sulfamethoxazole.

Chronic granulomatous disease (CGD) is an inherited disease characterized by severe and recurrent bacterial and fungal infections. Phagocytic cells of CGD patients are unable to produce superoxide anion, and their efficiency in bacterial killing is significantly impaired. In these patients, the prophylactic and therapeutic validity of a long-term use of trimethoprim-sulfamethoxazole (TMP-SMX) has been well established. However a role of nitric oxide (NO) produced by phagocytic cells from CGD patients is unknown, and the mechanism of TMP-SMX in CGD is unclear. We have directly measured NO production in whole human blood by using 4,5-diaminofluorescein as a novel fluorescent indicator for intracellular NO. Intracellular NO production of gated neutrophils increased time dependently when stimulated by lipopolysaccharide (LPS) and calcium ionophore. Although all polymorphonuclear leukocyte (PMN) specimens from patients with CGD failed to generate hydrogen peroxide, NO production by CGD PMNs was significantly increased compared with that of control PMNs (p<0.05). TMP-SMX with LPS significantly increased compared with LPS-stimulated samples at clinical (n=5, p<0.05) and 10-fold clinical concentrations (n=5, p<0.01). TMP-SMX with LPS in CGD PMNs significantly increased the production of NO in comparison with the LPS stimulation at 10-fold clinical concentrations (n=5, p<0.05). In conclusion, our data indicate the possibility that NO production by neutrophils from patients with CGD treated with TMP-SMX has a role of bactericidal activity instead of O(2)(-) in host defense mechanism.