In vivo transfection of cytokine genes into tumor cells using a synthetic vehicle promotes antitumor immune responses in a visceral tumor model.

The tumor microenvironment (TME) strongly affects the clinical outcomes of immunotherapy. This study aimed to activate the antitumor immune response by manipulating the TME by transfecting genes encoding relevant cytokines into tumor cells using a synthetic vehicle, which is designed to target tumor cells and promote the expression of transfected genes. Lung tumors were formed by injecting CT26.WT intravenously into BALB/c mice. Upon intravenous injection of the green fluorescent protein-coding plasmid encapsulated in the vehicle, 14.2% tumor-specific expression was observed. Transfection of the granulocyte-macrophage colony-stimulating factor (GM-CSF) and CD40 ligand (L)-plasmid combination and interferon gamma (IFNγ) and CD40L-plasmid combination showed 45.5% and 54.5% complete remission (CR), respectively, on day 60; alternate treatments with both the plasmid combinations elicited 66.7% CR, while the control animals died within 48 days. Immune status analysis revealed that the density of dendritic cells significantly increased in tumors, particularly after GM-CSF- and CD40L-gene transfection, while that of regulatory T cells significantly decreased. The proportion of activated killer cells and antitumoral macrophages significantly increased, specifically after IFNγ and CD40L transfection. Furthermore, the level of the immune escape molecule programmed death ligand-1 decreased in tumors after transfecting these cytokine genes. As a result, tumor cell-specific transfection of these cytokine genes by the synthetic vehicle significantly promotes antitumor immune responses in the TME, a key aim for visceral tumor therapy.

Effect of tonsillectomy combined with steroid pulse therapy upon IgA nephropathy depending on proteinuria status at diagnosis: a nationwide multicenter cohort study in Japan.

BACKGROUND: The effects of tonsillectomy combined with steroid pulse (TSP) therapy for IgA nephropathy (IgAN) are little known. Therefore, we examined the effects of TSP therapy on the kidney outcomes of IgAN in a large, nationwide cohort study in Japan. METHODS: Between 2002 and 2004, 632 IgAN patients with ≥ 0.5 g/day proteinuria at diagnosis were divided into three groups with mild (0.50-0.99 g/day; n = 264), moderate (1.00-1.99 g/day, n = 216), or severe (≥ 2.00 g/day; n = 153). Decline in kidney function and urinary remission were compared among the three groups after TSP therapy, corticosteroid (ST) therapy, or conservative therapy during a mean follow-up of 6.2 ± 3.3 years. 10.6% and 5.9% of patients in the ST and conservative therapy group underwent tonsillectomy. RESULTS: The rate of urinary remission at the final observation was significantly higher in the TSP therapy group than in the ST or conservative therapy groups (mild proteinuria: 64%, 43%, and 41%; moderate proteinuria: 51%, 45%, and 28%; severe proteinuria: 48%, 30%, and 22%, respectively). In contrast, the rate of a 50% increase in serum creatinine was lower in groups TSP therapy, than ST or conservative therapy (mild proteinuria: 2.1%, 10.1% and 16.7%; moderate proteinuria: 4.8%, 8.8% and 27.7%; severe proteinuria: 12.0%, 28.9% and 43.1%, respectively). In multivariate analysis, TSP therapy significantly prevented a 50% increase in serum creatinine levels compared with conservative therapy in groups with moderate and severe proteinuria (hazard ratio, 0.12 and 0.22, respectively). CONCLUSION: TSP significantly increased the rate of proteinuria disappearance and urinary remission in IgAN patients with mild-to-moderate urinary protein levels. It may also reduce the decline in kidney function in patients with moderate-to-severe urinary protein levels.

Analysis of the impact of obesity on the prognosis of IgA nephropathy according to renal function and sex.

BACKGROUND: Few studies have observed the direct effect of obesity on renal prognoses in immunoglobulin A nephropathy (IgAN) or separately evaluated its effects according to sex. We aimed to evaluate the direct and indirect effects of obesity on the renal outcomes of IgAN and observe these effects separately according to renal function and sex. METHODS: We extracted patients with body mass index (BMI) descriptions from a multicenter retrospective cohort analysis in Japan, and excluded those with < 30 days of follow-up, diabetes mellitus, and steroid treatment. Patients were divided into normal (n = 720; 18.5 ≤ BMI < 25) and obese (n = 212; BMI ≥ 25) groups, which were then compared. The endpoints were a 1.5-fold increase in serum creatinine levels and the initiation of renal replacement therapy. RESULTS: The obese group was older, included more males, and was more likely have hypertension, dyslipidemia, proteinuria, tubular atrophy, and lower renal function than the normal group. Patients with an eGFR < 60 mL/min/1.73 m2 had well-matched characteristics between the groups; however, hypertension, low high-density lipoprotein cholesterol, and hypertriglyceridemia were more common in the obese group. Obesity contributed to tubular atrophy, even when adjusted for renal function. In addition, it contributed to proteinuria only in females. However, obesity itself was not a significant prognostic factor. CONCLUSIONS: Although no independent effect on renal prognosis was observed during the study period, the obese group had more risk factors for IgAN progression and obesity contributed to tubular atrophy and female proteinuria. Our results suggest that separately analyzing the prognostic effect of obesity according to sex is important.

Prognostic factors of IgA nephropathy presenting with mild proteinuria at the time of diagnosis (a multicenter cohort study).

BACKGROUND: Clinical factors affecting renal prognosis in patients with immunoglobulin A nephropathy (IgAN) and low urinary protein excretion (U-Prot) remain unclear. This study evaluated such factors in patients with clinical grade I (CG-I) IgAN with U-Prot < 0.5 g/day. METHODS: This secondary analysis of a previous retrospective study included 394 patients with CG-I IgAN. The primary outcome was the first occurrence of a 1.5-fold increase in serum creatinine levels from baseline. Factors related to renal prognosis were examined using univariate and multivariate Cox regression analyses. CG-I was divided into C-Grade Ia (CG-Ia) (n = 330) with baseline eGFR ≥ 60 ml/min/1.73 m2, and C-Grade Ib (CG-Ib) (n = 64) with baseline eGFR < 60 ml/min/1.73 m2. Outcome incidence was compared between conservative and aggressive therapy (corticosteroids and/or tonsillectomy) groups. RESULTS: Overall outcome incidence was significantly higher in CG-Ib than in CG-Ia; the cumulative incidence was significantly higher in CG-Ib (hazard ratio, 9.67; 95% confidence interval, 2.90-32.23). Older age, higher IgA levels, eGFR < 60 mL/min/1.73 m2, lower eGFR at baseline were independent prognostic factors for CG-I. Older age, lower eGFR, higher IgA levels at baseline, and U-Prot remission at 1-year post-diagnosis were independent prognostic factors for CG-Ib. Aggressive therapy tended to suppress the cumulative outcome incidence compared with conservative therapy in CG-Ib (p = 0.087). CONCLUSION: An eGFR < 60 mL/min/1.73 m2 is a significant predictor of renal prognosis in patients with IgAN and U-Prot < 0.5 g/day.

Granulomatosis with polyangiitis presenting as a solitary renal mass: a case report.

BACKGROUND: Granulomatosis with polyangiitis (GPA) is characterized by necrotizing granulomatous vasculitis involving small-sized vessels in the upper airways, lower airways, and kidneys. Renal pathology is usually characterized by focal segmental necrotizing glomerulonephritis, which often leads to rapidly progressive renal failure. This type of renal involvement is usually unapparent on radiography. The presence of a renal mass in a patient with GPA, although extremely rare, is recognizable. Herein, we report a rare case of GPA presenting as a solitary renal mass and present a review of the literature. CASE PRESENTATION: A 75-year-old woman presented with a solitary right kidney mass measuring 4 × 3.5 cm detected by enhanced computed tomography. There was no history of sinusitis, rhinitis, cough, or pneumonia suggestive of systemic GPA. Nephrectomy was performed based on the suspicion of renal cell carcinoma or malignant lymphoma. Three months later, she was admitted because her serum creatinine levels increased from 54.81 µmol/L to 405.76 µmol/L accompanied by a high C-reactive protein level of 159 mg/L. Anti-neutrophil cytoplasmic antibodies against myeloperoxidase and anti-proteinase 3 were negative. Histological examinations of the solitary renal mass revealed necrotizing granulomatous arteritis in the cortex and medullary vasa recta, surrounded by interstitial fibrosis, and focal segmental necrotizing glomerulonephritis in the localized lesion; however, signs of vasculitis were not observed in areas other than the solitary mass. Therefore, the patient was diagnosed with granulomatosis with polyangiitis (GPA). Despite treatment with prednisolone (30 mg/day), the patient developed oliguria with an elevation of her serum creatinine level to 583.44 µmol/L, which required hemodialysis within one month after the initiation of steroid therapy. The patient could successfully discontinue hemodialysis 21 months later, following a decrease in her serum creatinine level to 251.06 µmol/L. CONCLUSIONS: GPA should be considered as one of the differential diagnoses of a solitary renal mass. Furthermore, patients with solitary renal masses associated with GPA may exhibit a favorable response to steroid or immunosuppressive treatment.

Literature review of allograft adenovirus nephritis and a case presenting as mass lesions in a transplanted kidney without symptoms of urinary tract infection or acute kidney injury.

Adenovirus (AdV) infection is a common complication in bone marrow/hematopoietic stem cell transplant and solid organ transplant recipients. AdV infection usually presents as hemorrhagic cystitis, but sometimes it can progress to acute kidney injury showing AdV nephritis (AdVN). We present the case of a 52-year-old Japanese female who had received a living kidney transplantation (KT) from her husband. At 21 months post-KT, the patient presented with a fever, but no renal dysfunction and no abnormal urine findings. A contrast-enhanced computed tomography (CT) scan revealed a few mass lesions with hypoperfusion in the transplanted kidney. An enhanced CT-guided biopsy targeting one of these lesions revealed a necrotizing tubulointerstitial nephritis suggesting AdVN. The polymerase chain reaction tests for ADV were negative in a urine sample but positive in the sera and the frozen kidney biopsy samples. AdVN can manifest as an unusual pattern of acute lobar nephritis/acute focal bacterial nephritis-like localization without symptoms of acute kidney injury or urinary tract infection. Enhanced CT can provide clues for clinical diagnosis.

Utility of remission criteria for the renal prognosis of IgA nephropathy.

BACKGROUND: Novel criteria for the remission of Immunoglobulin A nephropathy (IgAN) based on an opinion survey of Japanese nephrologists and literature review were proposed in 2013. This single-center, longitudinal retrospective cohort study was conducted to validate this criteria. METHODS: Present study included the IgAN patients diagnosed between 2001 and 2005 in the Juntendo University Hospital. Remission of hematuria was defined as three consecutive dipstick test results of ( -) to ( ±) or a red blood cell count < 5 in urinary sediment per high-power field during at least 6 months. Remission of proteinuria was defined as three consecutive dipstick results of ( -) to ( ±) during at least 6 months. We categorized four groups according to the remission status which was assessed 2 years after the renal biopsy. The primary outcome was a 50% increase in the serum creatinine over the baseline. We evaluated the slope of eGFR decline (mL/min/1.73 m2/year) and a decrease in the eGFR of 30% from baseline eGFR as the secondary outcome, respectively. RESULTS: A total of 74 patients (male: 47.3%, median age: 30 years) were included and were followed for a median of 86.5 months. During the period, forty-one patients achieved neither remission of proteinuria nor hematuria (NR). Twelve patients met the primary study outcome. A survival analysis revealed that the NR had the worst prognosis and the steepest slope of eGFR decline. CONCLUSION: Although further validation in a large cohort is necessary, these novel remission criteria for IgAN patients appear to predict the renal prognosis.

A cross-sectional study in patients with IgA nephropathy of correlations between clinical data and pathological findings at the time of renal biopsy: a Japanese prospective cohort study.

BACKGROUND: The correlations between clinical data and pathological findings at the time of renal biopsy were investigated in IgA nephropathy patients. METHODS: 771 patients diagnosed with IgA nephropathy by renal biopsy were enrolled. The correlations between clinical variables including eGFR, daily proteinuria, mean arterial pressure (MAP), serum uric acid (UA) values, and pathological parameters were examined. These patients were further divided into three groups: children (< 19 years old), young adults (19-60 years), and elderly patients (> 60 years). RESULTS: Daily proteinuria was moderately correlated with all pathological parameters (Rs = 0.23-0.49). The mesangial score, the percentage of glomeruli that contained endocapillary hypercellularity, cellular/fibrocellular crescents or tuft necrosis, and segmental glomerulosclerosis (GS) affected daily proteinuria most on multiple linear regression analysis (MLRA). eGFR, MAP, and serum UA levels were mainly correlated with the degree of GS and interstitial lesions. In children, the degree of cellular/fibrocellular crescents or tuft necrosis was correlated with not only daily proteinuria, but also decreased eGFR (Rs = 0.51, - 0.24). Endocapillary hypercellularity was the only independent variable related to daily proteinuria on MLRA. CONCLUSION: In all age cohorts of IgA nephropathy patients, daily proteinuria was correlated with all histological parameters, including both acute and chronic glomerular lesions, and the mesangial score. Independent variables for daily proteinuria were the meangial score, acute histological lesions, and segmental GS on MLRA, whereas the remaining independent variable in the pediatric group was endocapillary hypercellurality. The clinical pathological correlation at the time of biopsy varied depending on the age group.

Ratio of serum creatinine to cystatin C is related to leg strength in predialysis CKD patients.

BACKGROUND AND OBJECTIVES: Chronic kidney disease (CKD) patients have lower levels of physical function. Especially, leg strength is important for daily living and preventing falls. However, physical function screenings are difficult to perform at clinical sites. To find clinically useful method to evaluate physical function in predialysis CKD patients, we tried to evaluate the relationship between the ratio of serum creatinine to serum cystatin C (Cre/CysC), and knee extensor muscle strength/body weight (KEMS) which reflects their leg strength. DESIGN, SETTING, PARTICIPANTS, AND MEASUREMENTS: We recruited 147 outpatients with CKD (87 men; mean age, 61.6 ± 9.8 years; mean eGFRcreat, 40.7 ± 12.9 mL/min/1.73m2) in this cross-sectional study. KEMS was assessed using a wire strain gauge dynamometer. Skeletal muscle mass and body fat mass were assessed by bioelectrical impedance analysis. RESULTS: The mean value of Cre/CysC was 1.01 ± 0.18. The mean value of KEMS was 1.60 ± 0.47 Nm/kg. In multivariate linear regression analysis, skeletal muscle mass (p < 0.01), body fat mass (p < 0.01), hemoglobin (p = 0.01), and Cre/CysC (p < 0.01) was independently related to KEMS. The correlation between Cre/CysC and KEMS is stronger in high quantile of Cre/CysC. CONCLUSIONS: In predialysis CKD patients, KEMS showed lower as CKD stage advanced. Cre/CysC is significantly related to KEMS independently. Cre/CysC may be an alternative marker for leg strength in CKD patients and even more valuable to utilize in cases with high Cre/CysC.

Impact of the number of steroid pulses in tonsillectomy combined with steroid pulse therapy: a nationwide retrospective study in Japan.

BACKGROUND: Steroid pulse therapy with tonsillectomy is known as a major treatment for IgA nephropathy (IgAN). However, its protocol was different among institutions and the effects of varying the number of steroid pulses remain unclear. METHODS: From a total of 1,174 IgAN patients in a multicenter retrospective cohort analysis in Japan, 195 patients were treated by tonsillectomy combined with corticosteroid. They were divided into four groups based on the number of administered steroid pulses from 0 to three (TSP0-3), and remission of urinary abnormalities and renal survival until 1.5-fold increase in serum creatinine level from baseline were analyzed among the four groups and between TSP1 and TSP3. RESULTS: Among the four groups, renal function was relatively good when the estimated glomerular filtration rate was approximately 80-90 mL/min/1.73m2 and proteinuria was relatively mild (< 1.0 g/gCre). The ratio of patients who developed renal dysfunction was < 5% in all groups, and the cumulative renal survival rate by Kaplan-Meier analysis was similar among groups (log-rank test, p = 0.37), despite varying clinical backgrounds and treatments. After adjustment of the background variables between TSP1 and TSP3, the remission rates of urinary abnormalities were similar and the renal survival rate also remained similar (66.8 vs. 85.4%, p = 0.45). CONCLUSIONS: In patients with mild proteinuria and good renal function, the number of steroid pulses did not affect the renal outcome in steroid pulse therapy with tonsillectomy. The adaptation and protocols, such as the number of steroid pulses, should be determined for each IgAN patient's background.

Remission of proteinuria under therapeutic intervention and the renal outcomes in Japanese patients with lupus nephritis class III and IV.

AbstractsBackground: Recent studies have identified the significance of proteinuria levels after initial induction therapies on the renal outcomes in patients with proliferative lupus nephritis, but the issue has not been evaluated in Japanese patients.Methods: Based on the ISN/RPS classification, only patients diagnosed with lupus nephritis class III or IV were included. The remission of proteinuria 12 months after diagnosis, as well as the clinicopathological features at diagnosis, on renal outcomes was examined retrospectively. Renal progression was defined as a 50% decrease in the estimated glomerular filtration rate or the development of end-stage renal disease.Results: This study included 82 Japanese patients with a median follow-up period of seven years. Although all patients received intensive induction therapy, 15 patients (18%) showed progression. Proteinuric remission 12 months after diagnosis predicted a good renal outcome by multivariate analysis. A receiver-operating characteristic analysis of 38 patients whose quantitative urinary protein excretion levels at 12 months were available for analysis showed that a cut-off value of 0.8 g/day predicted renal progression most effectively. Neither the renal function nor proteinuria level at diagnosis were associated with the renal outcomes.Conclusion: In Japanese patients with lupus nephritis class III or IV, proteinuria levels after 12 months under intensive therapy predicted renal outcomes more accurately than did factors identified at diagnosis.

A grading system that predicts the risk of dialysis induction in IgA nephropathy patients based on the combination of the clinical and histological severity.

Histological classification is essential in the clinical management of immunoglobulin A nephropathy (IgAN). However, there are limitations in predicting the prognosis of IgAN based on histological information alone, which suggests the need for better prognostic models. Therefore, we defined a prognostic model by combining the grade of clinical severity with the histological grading system by the following processes. We included 270 patients and explored the clinical variables associated with progression to end-stage renal disease (ESRD). Then, we created a predictive clinical grading system and defined the risk grades for dialysis induction by a combination of the clinical grade (CG) and the histological grade (HG). A logistic regression analysis revealed that the 24-h urinary protein excretion (UPE) and the estimated glomerular filtration rate (eGFR) were significant independent variables. We selected UPE of 0.5 g/day and eGFR of 60 ml/min/1.73 m2 as the threshold values for the classification of CG. The risk of progression to ESRD of patients with CG II and III was significantly higher than that of patients with CG I. The patients were then re-classified into nine compartments based on the combination of CG and HG. Furthermore, the nine compartments were grouped into four risk groups. The risk of ESRD in the moderate, high, and super-high-risk groups was significantly higher than that in the low-risk group. Herein, we are giving a detailed description of our grading system for IgA nephropathy that predicted the risk of dialysis based on the combination of CG and HG.

Forced expression of vascular endothelial growth factor-A in podocytes decreases mesangial cell numbers and attenuates endothelial cell differentiation in the mouse glomerulus.

BACKGROUND: Glomerular podocyte-derived vascular endothelial growth factor (VEGF) is indispensable for the migration and proliferation of glomerular endothelial cells. In contrast, podocyte-specific Vegf overexpression leads to the collapse of glomerular tufts; however, the mechanisms underlying this outcome have not yet been reported. METHODS: To further clarify the effects of elevated levels of Vegf expression on glomerular cells, we established a dual transgenic mouse line in which Vegf was exclusively and inducibly expressed in podocytes under the control of the "Tet-on system" (Podocin-rtTA/TetO-Vegf164 mice). RESULTS: Macroscopic and microscopic examination of Podocin-rtTA/TetO-Vegf164 animals following Vegf induction identified the presence of prominent red bloody spots. In addition, the endothelial cell number was increased along with enlargement of the subendothelial spaces. We also observed impaired endothelial fenestrations and aberrant plasmalemmal vesicle-associated protein-1 (PV-1) expression. In contrast, the mesangial cell number markedly decreased, resulting in a glomerular tuft intussusceptive splitting defect. Furthermore, whereas platelet-derived growth factor-B (PDGF-B) expression in the glomerular cells of Podocin-rtTA/TetO-Vegf164 mice was not decreased, phospho-PDGF receptor immunoreactivity in the mesangial cells was significantly decreased when compared to wild-type animals. CONCLUSION: Taken together, the results of this study indicated that the upregulation of podocyte VEGF decreased the number of mesangial cells, likely owing to inhibition of PDGF-B-mediated signaling.

Maternally inherited diabetes and deafness complicated by mesangial galactose-deficient IgA1 deposits: a case report.

BACKGROUND: Maternally inherited diabetes and deafness (MIDD), a mitochondrial genetic disorder, typically affects the kidneys and results in end-stage renal disease. Early diagnosis of MIDD is challenging when renal manifestations precede other key clinical features such as diabetes and deafness and/or when the disease is complicated by other renal pathologies. CASE PRESENTATION: Here, we present the case of a 33-year-old Japanese woman who had initially been diagnosed with IgA nephropathy but was found to have MIDD 6 years later. Two renal biopsies were conducted six years apart. While assessment of the first biopsy specimen with the monoclonal antibody (KM55) revealed mesangial IgA deposits (containing the galactose-deficient IgA1 variant [Gd-IgA1]), examination of the second specimen showed no mesangial IgA deposits and newly-developed glomerular global scleroses and tubular damage. Granular swollen epithelial cells (GSECs), characterised by abnormal mitochondria, were observed among the tubules and collecting ducts in both biopsy specimens. Mitochondrial DNA analysis revealed an m.3243A > G mutation. CONCLUSIONS: We rediscovered the usefulness of GSECs as a pathologically distinctive feature of mitochondrial nephropathy and reviewed the literature regarding MIDD complicated by mesangial IgA deposition. Furthermore, we demonstrate that the mesangial IgA deposits in this patient consisted of the galactose-deficient IgA1 variant. The monoclonal antibody (KM55) might be a useful tool to distinguish IgAN from latent IgA deposits.

Reproducibility for pathological prognostic parameters of the Oxford classification of IgA nephropathy: a Japanese cohort study of the Ministry of Health, Labor and Welfare.

BACKGROUND/AIMS: The Oxford classification of IgA nephropathy (IgAN) was proposed by international working group in 2009. Interobserver reproducibility of each pathological definition was already evaluated, but that of four pathological prognostic parameters score has not yet been assessed. We first assess the reproducibility of each pathological definition in Japanese patients. Our study is aimed to assess that of four pathological prognostic parameters score among the five Japanese pathologists. METHODS: The renal specimens from 411 Japanese patients, aged 3-85 years, with biopsied proven primary IgAN were collected from 50 facilities between 2006 and 2012. The reproducibility of pathological definitions was assessed by the intraclass correlation coefficient (ICC) and that of four pathological prognostic parameters score (mesangial hypercellularity (M), endocapillary hypercellularity (E), segmental glomerulosclerosis (S), and tubular atrophy/interstitial fibrosis (T)) was assessed by kappa statistics. RESULTS: The ICC of M, E, S, T, global sclerosis and cellular crescents and/or fibrocellular crescents were good or moderate agreement among the five pathologists and were well agreed with results of the Oxford study. Kappa statistics was moderate agreement for M and T score assessed with the semi-quantitative method by the Oxford group, but that was poor agreement for S and E score based on a simple "present" or "absent" assessment. CONCLUSION: This is the first report to assess the reproducibility of pathological prognostic parameters score in the Oxford classification. Our study supports the utilization of the pathological lesions in routine diagnosis. The methodological assessment of pathological prognostic parameters score should be reconsidered.

Anaemia is an essential complication of ANCA-associated renal vasculitis: a single center cohort study.

BACKGROUND: Anaemia is a common complication of patients with antineutrophil cytoplasmic antibody (ANCA)-associated renal vasculitis. Nevertheless, the cause and degree of such cases of anaemia have not been elucidated in detail. We aimed to investigate the prevalence, cause, pathogenesis of anaemia and the impact of anaemia on prognosis in patients with ANCA-associated renal vasculitis. METHODS: We identified 45 patients with ANCA-associated renal vasculitis that were clinically and/or histologically diagnosed and treated from 2003 to 2014 at University of Tsukuba Hospital. The relationships between anaemia and various clinicopathological findings were evaluated. RESULTS: At the time of diagnosis of ANCA-associated renal vasculitis, all patients showed anaemia, with a mean haemoglobin level of 7.5 ± 1.3 g/dL. Renal anaemia was diagnosed in 92% of patients, anaemia of chronic disease (ACD) in 56%, and anaemia due to hemorrhage in 20%. Next, the patients were divided into two groups according to anaemia severity: minimum haemoglobin (min Hb) < 7.5 (n = 24) and min Hb ≥ 7.5 (n = 21). A comparison of baseline characteristics showed that serum albumin, maximum serum creatinine, minimum estimated glomerular filtration rate (eGFR), serum cystatin C, and the area of tubulointerstitial damage were significantly different between the haemoglobin groups (p < 0.05). No significant intergroup differences were observed in iron-related or inflammation-related data. With regard to the relationship between anaemia severity and prognosis, patients in the min Hb < 7.5 group tended to have a lower eGFR. Anaemia severity was associated with markedly lower survival (Log-rank test, p = 0.03). CONCLUSIONS: In this cohort of patients with ANCA-associated renal vasculitis, all subjects exhibited anaemia. In regard to the cause and pathogenesis, the most prevalent form of anaemia was renal anaemia, not ACD, and a potential reason for the high prevalence of anaemia in our cohort may have been the interaction between renal anaemia and ACD. Moreover, anaemia severity was significantly associated with the degree of renal dysfunction and life prognosis.

Stoichiometry and geometry of the CXC chemokine receptor 4 complex with CXC ligand 12: molecular modeling and experimental validation.

Chemokines and their receptors regulate cell migration during development, immune system function, and in inflammatory diseases, making them important therapeutic targets. Nevertheless, the structural basis of receptor:chemokine interaction is poorly understood. Adding to the complexity of the problem is the persistently dimeric behavior of receptors observed in cell-based studies, which in combination with structural and mutagenesis data, suggest several possibilities for receptor:chemokine complex stoichiometry. In this study, a combination of computational, functional, and biophysical approaches was used to elucidate the stoichiometry and geometry of the interaction between the CXC-type chemokine receptor 4 (CXCR4) and its ligand CXCL12. First, relevance and feasibility of a 2:1 stoichiometry hypothesis was probed using functional complementation experiments with multiple pairs of complementary nonfunctional CXCR4 mutants. Next, the importance of dimers of WT CXCR4 was explored using the strategy of dimer dilution, where WT receptor dimerization is disrupted by increasing expression of nonfunctional CXCR4 mutants. The results of these experiments were supportive of a 1:1 stoichiometry, although the latter could not simultaneously reconcile existing structural and mutagenesis data. To resolve the contradiction, cysteine trapping experiments were used to derive residue proximity constraints that enabled construction of a validated 1:1 receptor:chemokine model, consistent with the paradigmatic two-site hypothesis of receptor activation. The observation of a 1:1 stoichiometry is in line with accumulating evidence supporting monomers as minimal functional units of G protein-coupled receptors, and suggests transmission of conformational changes across the dimer interface as the most probable mechanism of altered signaling by receptor heterodimers.

Circadian blood pressure abnormalities in patients with primary nephrotic syndrome.

BACKGROUND: Only a few studies have evaluated the abnormalities of ambulatory blood pressure (ABP) in patients with nephrotic syndrome (NS). METHODS: The 24-h ABPs were measured in primary NS patients with acute onset of disease and analyzed in relation to the clinical variables. RESULTS: Our subjects comprised 21 patients: 17 with minimal change disease and 4 with focal segmental glomerulosclerosis. Of these patients, 8 (38%) had daytime hypertension, 13 (62%) had nighttime hypertension, and 13 (62%) were non-dippers (nighttime-to-daytime ratio of ABP: NDR > 0.9). The serum sodium level was correlated with the average 24-h ABP and NDR, after adjustment for other clinical variables, such as the increase in body weight, serum albumin level, and urinary protein excretion. The data from repeated ABP measurements, before and after the achievement of remission, showed a marked decrease in the average 24-h ABP after remission. Furthermore, change in the serum sodium level was significantly correlated with the change in NDR. CONCLUSION: These results suggest that alteration in renal handling of sodium and water, which might be reflected in serum sodium level, is involved in the abnormality of circadian blood pressure in primary NS patients.

Screening of Transient Receptor Potential Canonical Channel Activators Identifies Novel Neurotrophic Piperazine Compounds.

Transient receptor potential canonical (TRPC) proteins form Ca(2+)-permeable cation channels activated upon stimulation of metabotropic receptors coupled to phospholipase C. Among the TRPC subfamily, TRPC3 and TRPC6 channels activated directly by diacylglycerol (DAG) play important roles in brain-derived neurotrophic factor (BDNF) signaling, promoting neuronal development and survival. In various disease models, BDNF restores neurologic deficits, but its therapeutic potential is limited by its poor pharmacokinetic profile. Elucidation of a framework for designing small molecules, which elicit BDNF-like activity via TRPC3 and TRPC6, establishes a solid basis to overcome this limitation. We discovered, through library screening, a group of piperazine-derived compounds that activate DAG-activated TRPC3/TRPC6/TRPC7 channels. The compounds [4-(5-chloro-2-methylphenyl)piperazin-1-yl](3-fluorophenyl)methanone (PPZ1) and 2-[4-(2,3-dimethylphenyl)piperazin-1-yl]-N-(2-ethoxyphenyl)acetamide (PPZ2) activated, in a dose-dependent manner, recombinant TRPC3/TRPC6/TRPC7 channels, but not other TRPCs, in human embryonic kidney cells. PPZ2 activated native TRPC6-like channels in smooth muscle cells isolated from rabbit portal vein. Also, PPZ2 evoked cation currents and Ca(2+) influx in rat cultured central neurons. Strikingly, both compounds induced BDNF-like neurite growth and neuroprotection, which were abolished by a knockdown or inhibition of TRPC3/TRPC6/TRPC7 in cultured neurons. Inhibitors of Ca(2+) signaling pathways, except calcineurin, impaired neurite outgrowth promotion induced by PPZ compounds. PPZ2 increased activation of the Ca(2+)-dependent transcription factor, cAMP response element-binding protein. These findings suggest that Ca(2+) signaling mediated by activation of DAG-activated TRPC channels underlies neurotrophic effects of PPZ compounds. Thus, piperazine-derived activators of DAG-activated TRPC channels provide important insights for future development of a new class of synthetic neurotrophic drugs.

The dependence of chemokine-glycosaminoglycan interactions on chemokine oligomerization.

Both chemokine oligomerization and binding to glycosaminoglycans (GAGs) are required for their function in cell recruitment. Interactions with GAGs facilitate the formation of chemokine gradients, which provide directional cues for migrating cells. In contrast, chemokine oligomerization is thought to contribute to the affinity of GAG interactions by providing a more extensive binding surface than single subunits alone. However, the importance of chemokine oligomerization to GAG binding has not been extensively quantified. Additionally, the ability of chemokines to form different oligomers has been suggested to impart specificity to GAG interactions, but most studies have been limited to heparin. In this study, several differentially oligomerizing chemokines (CCL2, CCL3, CCL5, CCL7, CXCL4, CXCL8, CXCL11 and CXCL12) and select oligomerization-deficient mutants were systematically characterized by surface plasmon resonance to determine their relative affinities for heparin, heparan sulfate (HS) and chondroitin sulfate-A (CS-A). Wild-type chemokines demonstrated a hierarchy of binding affinities for heparin and HS that was markedly dependent on oligomerization. These results were corroborated by their relative propensity to accumulate on cells and the critical role of oligomerization in cell presentation. CS-A was found to exhibit greater chemokine selectivity than heparin or HS, as it only bound a subset of chemokines; moreover, binding to CS-A was ablated with oligomerization-deficient mutants. Overall, this study definitively demonstrates the importance of oligomerization for chemokine-GAG interactions, and demonstrates diversity in the affinity and specificity of different chemokines for GAGs. These data support the idea that GAG interactions provide a mechanism for fine-tuning chemokine function.