Insight into the Interplay of Gd-IgA1, HMGB1, RAGE and PCDH1 in IgA Vasculitis (IgAV).

The pathogenesis of IgAV, the most common systemic vasculitis in childhood, appears to be complex and requires further elucidation. We aimed to investigate the potential role of galactose-deficient immunoglobulin A1 (Gd-IgA1), high-mobility group box 1 (HMGB1), receptor for advanced glycation end products (RAGE) and protocadherin 1 (PCDH1) in the pathogenesis of IgAV. Our prospective study enrolled 86 patients with IgAV and 70 controls. HMGB1, RAGE, Gd-IgA1 and PCDH1 in serum and urine were determined by the enzyme-linked immunosorbent assay (ELISA) method at the onset of the disease and after a six-month interval in patients and once in the control group. Serum concentrations of HMGB1, RAGE and PCDH1 and urinary concentrations of HMGB1, RAGE, Gd-IgA1 and PCDH1 were significantly higher in patients with IgAV than in the control group (p < 0.001). Concentrations of HMGB1 (5573 pg/mL vs. 3477 pg/mL vs. 1088 pg/mL, p < 0.001) and RAGE (309 pg/mL vs. 302.4 pg/mL vs. 201.3 pg/mL, p = 0.012) in the serum of patients remained significantly elevated when the disease onset was compared with the six-month follow-up interval, and thus could be a potential marker of disease activity. Urinary concentration of HMGB1 measured in the follow-up period was higher in patients with nephritis compared to IgAV without nephritis (270.9 (146.7-542.7) ng/mmol vs. 133.2 (85.9-318.6) ng/mmol, p = 0.049) and significantly positively correlated with the urine albumine to creatinine ratio (τ = 0.184, p < 0.05), the number of erythrocytes in urine samples (τ = 0.193, p < 0.05) and with the outcome of nephritis (τ = 0.287, p < 0.05); therefore, HMGB1 could be a potential tool for monitoring patients with IgAV who develop nephritis. Taken together, our results imply a possible interplay of Gd-IgA1, HMGB1, RAGE and PCDH1 in the development of IgAV. The identification of sensitive biomarkers in IgAV may provide disease prevention and future therapeutics.

High mobility group box protein 1 (HMGB1) serum and urinary levels and gene polymorphism in Egyptian patients with systemic lupus erythematosus: A possible relation to lupus nephritis.

OBJECTIVE: The aim of this study was to evaluate the effects of the high mobility group box protein 1 (HMGB1) serum and urinary levels and gene polymorphisms on systemic lupus erythematosus (SLE) development and investigate their link to lupus nephritis (LN). METHODS: We enrolled 120 Egyptian SLE patients and 120 healthy controls. Thorough medical and clinical evaluation were carried out, and SLE disease activity index (SLEDAI) was assessed. Lupus patients were divided into two groups according to the presence of LN. Measurement of HMGB1 serum and urinary levels was done using ELISA and genotyping for HMGB1 (rs1045411) was performed. RESULTS: There were statistically significantly higher HMGB1 serum and urinary levels in SLE patients (p < 0.001). There was a marginally significant association between lupus and alleles (p = 0.059, φ = -0.086). 'C' allele was marginally significant risk allele for SLE. After classifying SLE patients based on the presence or absence of LN, there was no significant difference as regard sex (p = 0.387), age (p = 0.208) and disease duration (p = 0.094).However, there was a significant difference between the 2 groups in regard to the frequency of musculoskeletal manifestations (p = 0.035), SLEDAI score (p < 0.001), both serum (p < 0.001) and urinary HMGB1 levels (p < 0.001) in addition to the frequency of HMGB1 genotypes (p = 0.003). Lupus patients with C/T-T/T HMGB1 genotypes had 3.5-times higher odds to exhibit LN. CONCLUSIONS: Serum and urine HMGB1 measurements are helpful in the diagnosis of SLE and the prediction of LN. There is a link between HMGB1 gene variations and the risk of SLE, with evidence that the C/T-T/T HMGB1 genotype is linked to a significantly greater risk of LN in the Egyptian population.

Cross talk between Hsp72, HMGB1 and RAGE/ERK1/2 signaling in the pathogenesis of bronchial asthma in obese patients.

BACKGROUND: The incidence of obesity-related asthma has shown a remarkable increase. OBJECTIVES: We aimed to explore the role of heat shock protein 72 (Hsp72) and receptor for advanced glycation end products (RAGE) axis with its downstream signaling in the pathogenesis of obesity-related asthma. METHODS: We enrolled a total of 55 subjects and divided them into three groups. Groups I and II included healthy, normal weight (n = 15) and obese (n = 15) subjects, respectively. Twenty-five obese asthmatics (group III) were subdivided into group IIIa (10 patients with mild to moderate asthma) and group IIIb (15 patients with severe asthma). High mobility group box 1 (HMGB1), interleukin 8 (IL-8), monocyte chemoattractant protein 1 (MCP-1), extracellular signal-regulated protein kinases 1 and 2 (ERK1/2), and urinary Hsp72 were immunoassayed. Hydrogen peroxide (H2O2) and free fatty acids (FFAs) levels were photometrically measured. RAGE mRNA expression was relatively quantified by real-time PCR. RESULTS: We found significant elevations of serum HMGB1, IL-8, MCP1, ERK1/2, FFAs, and H2O2 levels as well as urinary Hsp72 levels in obese subjects compared to healthy control. These were more evident in patients with severe asthma (group IIIb). Multivariate regression analysis identified Hsp72 and ERK1/2 as independent predictors of bronchial asthma severity. Receiver operating characteristic (ROC) curve analysis revealed that areas under the curve (AUC) for Hsp72 and ERK1/2 were 0.991 and 0.981, respectively, which denotes a strong predictive value for identifying the severity of bronchial asthma in obese patients. CONCLUSION: The current study highlights the role of Hsp72 and HMGB1/RAGE/ERK1/2 signaling cascade in the pathogenesis of bronchial asthma and its link to obesity, which could be reflected on monitoring, severity grading, and management of this disease.

A novel diagnostic tool for the detection of bladder cancer: Measurement of urinary high mobility group box-1.

OBJECTIVE: We aimed to investigate the diagnostic value of urinary High Mobility Group Box-1 (HMGB1) level as a noninvasive tool that can be potentially used for diagnosis and during follow-up in patients with bladder cancer patients. METHOD: The study was conducted in a total of 121 participants including 61 patients diagnosed with primary bladder cancer, 30 patients with an acute urinary tract infection and 30 healthy controls. Age, gender and urinary HMGB1 levels of the study groups were evaluated. The association of clinical features (tumor diameter, number of foci, pathological grade, muscle invasion) with urinary HMGB1 levels was investigated in patients with bladder cancer. RESULTS: All 3 groups showed a normal age and gender distribution with no significant difference among them (P = 0.775 and P = 0.967, respectively). A significant difference was detected in urinary HMGB1 levels among the 3 groups (P < 0.001). When urinary HMGB1 levels were compared between patients with high grade vs. low grade tumors, the mean HMGB1 level was 44.39 pg/ml (12.1-505.2) in patients with low grade tumors and 280 pg/ml (18.7-2685.3) in patients with high grade tumors (P < 0.001). Patients with a greater number of tumor foci had higher HMGB1 levels in comparison to patients with a single tumor focus (P = 0.008). Urinary HMGB1 levels were higher in patients with a tumor diameter of ≥3 cm than in patients with a tumor diameter less than 3 cm (P = 0.001). Patients with muscle-invasive bladder cancer exhibited higher urinary HMGB1 levels compared to patients with non-muscle-invasive bladder cancer (P = 0.033). The cut-off values derived from the ROC analysis were 63.30 pg/ml for distinguishing bladder cancer from urinary tract infection, 30.94 pg/ml for urinary tract infection versus control group and 38.70 pg/ml for bladder cancer vs. control group, respectively. Sensitivity was 59% and specificity was found 77%. CONCLUSION: In future controlled studies involving larger patient groups, urinary HMGB1 levels can be used for diagnostic and screening purposes in bladder cancer patients.

HMGB1+ microparticles present in urine are hallmarks of nephritis in patients with systemic lupus erythematosus.

Non-classical monocytes infiltrate the kidney parenchyma and participate in tissue damage in patients with lupus nephritis (LN). Circulating microparticles (MPs) seem to play critical roles in the activation of monocytes in systemic lupus erythematosus (SLE) patients. This study aims to characterize the phenotypes of MPs and monocyte subsets in LN patients and to determine their potential to discriminate between SLE patients with and without LN. Blood and urine samples from SLE patients were collected. In monocyte subsets from whole blood samples several phenotypic markers were evaluated. MPs were isolated from platelet-poor plasma and urine by centrifugation. This phenotypic marker characterization was performed using multiparametric flow cytometry. We observed that patients with active LN have lower counts of non-classical monocytes than do those without renal involvement. All monocyte subsets exhibited lower expression of CX3CR1 and ICAM-1 in LN than in patients without LN. High frequencies of MP-HMGB1+ and MP-HLA-DR+ were detected in circulation and urine of LN patients. Although MP-HMGB1+ , MP-HLA-DR+ , and MP-CX3CR1+ from urine were able to discriminate between patients with and without LN, only urinary MP-HMGB1+ were different between patients with active and inactive LN. Therefore, these vesicles may be useful as biomarkers of LN.

Urinary MCP-1、HMGB1 increased in calcium nephrolithiasis patients and the influence of hypercalciuria on the production of the two cytokines.

The study aims to observe the urinary excretion of monocyte chemoattractant-1 (MCP-1) and high-mobility group box 1 (HMGB1) in patients with calcium nephrolithiasis and to determine the influence of hypercalciuria on the production of the two cytokines. 81 cases of patients with calcium nephrolithiasis (group CN) and 30 healthy controls (group C) were involved in this study. To observe the influence of urinary calcium on the excretion of those cytokines, the patients were subdivided according to their 24-h urinary calcium level: ≥4 mg/kg/day (group H) and <4 mg/kg/day (group N). MCP-1 and HMGB1 in urina sanguinis were determined for all subjects. In addition, in vitro study was done to determine the production of the two cytokines and index of apoptosis and oxidative injuries in human kidney epithelial cells (HK-2) exposed to three high levels of calcium. Data showed that both urinary MCP-1 and HMGB1 in group CN were higher than that of group C. When the patients were subdivided, comparisons among the three groups showed that both MCP-1 and HMGB1 in group H and group N were higher than group C, but there was no significant statistical difference between the two stone groups. In vitro study, the apoptosis rate of cells, the lactate dehydrogenase activities, the hydrogen peroxide, and 8-isoprostane concentrations in the medium all increased in accordance with the increased concentration of calcium supplemented. Compared with the control, mRNA expressions of MCP-1 and HMGB1 in cells and the protein concentrations of the two cytokines in the medium of calcium-supplemented groups increased significantly. Results showed that urinary MCP-1 and HMGB1 increased in calcium nephrolithiasis patients and hypercalciuria might affect the identical pathways (through the reactive oxygen species) with other factors in stimulating the production of MCP-1 and HMGB1 in vivo.

Urinary high-mobility group box-1 associates specifically with lupus nephritis class V.

INTRODUCTION: High-mobility group box 1 protein (HMGB-1) has been implicated in the pathogenesis of lupus nephritis (LN). There is increased HMGB-1 expression in the kidneys and increased levels are observed in serum and urine of patients with LN. This study was performed to determine whether the increased urinary HMGB-1 was specific for active lupus or secondary to renal damage. METHODS: Urine from 61 lupus patients (32 had active LN and 29 had systemic lupus erythematosus (SLE) with no evidence of LN) and 14 control proteinuric patients (all with hypertension and eight also with diabetes) were included in this study. HMGB-1 was detected by Western blot. Urine protein was normalized to urine creatinine to account for volume of the specimen. RESULTS: Median normalized urine HMGB-1 levels were significantly elevated in LN patients compared to lupus patients without kidney disease (53.81 vs 9.46, p < 0.001). A difference in median levels was seen between LN classes, with a significant difference between proliferative and membranous disease (33.4 vs 138.8, p = 0.003). Urine protein to urine creatinine ratio (P/C) correlated with urinary HMGB-1 (r = 0.52, p < 0.001), but across the classes this was true only for membranous disease (r = 0.71, p = 0.022, proliferative, p = 0.63; mixed, p = 0.34). CONCLUSIONS: HMGB-1 is elevated in the urine of patients with active LN. Levels are associated with LN class, and higher levels of urinary HMGB-1 are seen in patients with class V when compared to both proliferative and mixed classes. Therefore, urinary HMGB-1 may be suggestive of membranous LN and warrants further evaluation in a large lupus cohort.

Urinary levels of high mobility group box-1 are associated with disease activity in antineutrophil cytoplasmic autoantibody-associated vasculitis.

BACKGROUND: High mobility group box-1 (HMGB1), a kind of pro-inflammatory mediator, is associated with inflammatory conditions and tissue damage. Our previous study demonstrated that the circulating levels of HMGB1 correlated with disease activity of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). In the current study, we aimed to measure urinary levels of HMGB1 in AAV patients, correlated them to clinical activity index and analysed the immunohistochemical HMGB1 staining in kidney specimens. METHODS: 50 patients with AAV in active stage and 56 patients with AAV in remission were recruited. The urinary levels of HMGB1 were determined by enzyme-linked immunosorbent assay. Moreover, renal biopsy specimens from 27 patients with active AAV were randomly collected to evaluate the deposition of HMGB1. RESULTS: Urinary HMGB1 levels in AAV patients in active stage were significantly higher than those in AAV patients in remission and healthy controls (1.46 [0.56-3.43] versus 0.38 [0.10-1.35] mg/µmolCr, P=0.001; 1.46 [0.56-3.43] versus 0.48 [0.40-0.60] mg/µmolCr, P=0.000, respectively). Further analysis found that urinary levels of HMGB1 correlated with erythrocyte sedimentation rate (r=0.354, p=0.012), C-reactive protein (r=0.289, p=0.042), and Birmingham Vasculitis Activity Score (r=0.350, p=0.013). Renal tissue of active AAV patients showed HMGB1 was mainly expressed in the cytoplasm and the extracellular space. The percentage of HMGB1-negative nuclei in renal tissue of patients with active AAV was significantly higher than that in normal controls (60.6±20.2 % versus 2.7±0.6 %, p<0.01). CONCLUSION: Urinary levels of HMGB1 may be associated with the disease activity in AAV patients.

High-mobility group box-1 protein in adenine-induced chronic renal failure and the influence of gum arabic thereon.

Pathogenesis of adenine-induced chronic renal failure may involve inflammatory, immunological and/or oxidant mechanisms. Gum arabic (GA) is a complex polysaccharide that acts as an anti-oxidant which can modulate inflammatory and/or immunological processes. Therefore, we tested here the effect of GA treatment (15 % in the drinking water for 4 weeks) in plasma and urine of rats, on a novel cytokine that has been shown to be pro-inflammatory, viz, DNA-binding high-mobility group box-1 protein (HMGB1). Adenine (0.75 % in the feed, 4 weeks) significantly increased indoxyl sulphate, urea and creatinine concentrations in plasma, and significantly decreased the creatinine clearance. GA significantly abated these effects. The concentrations of HMGB1 in urine before the start of the experiment were similar in all four groups. However, 24 h after the last treatment, adenine treatment increased significantly the concentration of HMGB1 when compared with the control. GA treatment did not affect the HMGB1 concentration in urine. Moreover, the concentration of HMGB1 in plasma obtained 24 h after the last treatment in rats treated with adenine was drastically reduced compared with the control group. This may explain its significant rise in urine. In conclusion, HMGB1 can be considered a potentially useful biomarker in adenine induced CRF and its treatment.

Are urinary levels of high mobility group box 1 markers of active nephritis in anti-neutrophil cytoplasmic antibody-associated vasculitis?

The objective of this study is to evaluate urinary high mobility group box 1 (HMGB1) levels as markers for active nephritis in patients with anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) in comparison with urinary CD4(+) effector memory T cells and urinary monocyte chemoattractant protein-1 (MCP-1). Twenty-four AAV patients with active nephritis and 12 healthy controls (HC) were evaluated. In nine patients, samples were also obtained during remission. Urinary levels of HMGB1 were measured by Western blot. CD4(+) T cells and CD4(+) effector memory T cells (CD4(+) CD45RO(+) CCR7(-) ) were determined in urine and whole blood by flow cytometry. Measurement of urinary levels of MCP-1 and serum HMGB1 levels were performed by enzyme-linked immunosorbent assay (ELISA). AAV patients with active nephritis had higher median intensity of HMGB1 in urine than HC [10·3 (7·05-18·50) versus 5·8 (4·48-7·01); P = 0·004]. Both urinary HMGB1 and MCP-1 levels decreased significantly from active nephritis to remission. The urinary MCP-1/creatinine ratio correlated with Birmingham Vasculitis Activity Score (BVAS) (P = 0·042). No correlation was found between the HMGB1/creatinine ratio and 24-h proteinuria, estimated glomerular filtration rate (eGFR), MCP-1/creatinine ratio, BVAS and serum HMGB1. A positive correlation was found between urinary HMGB1/creatinine ratio and CD4(+) T cells/creatinine ratio (P = 0·028) and effector memory T cells/creatinine ratio (P = 0·039) in urine. Urinary HMGB1 levels are increased in AAV patients with active nephritis when compared with HC and patients in remission, and urinary HMGB1 levels are associated with CD4(+) T cells and CD4(+) effector memory T cells in urine. Measurement of urinary HMGB1 may be of additional value in identifying active glomerulonephritis in AAV patients.

Age-related differences in kidney injury biomarkers induced by cisplatin.

Acute kidney injury (AKI) occurs in a half of cisplatin (CDDP)-treated patients. Traditional biomarkers including blood urea nitrogen (BUN) and serum creatinine (SCr) are still used for detection of CDDP-induced AKI, but these biomarkers are not specific or sensitive. The aim of this study was to identify the specific and sensitive biomarkers against CDDP-induced renal injury between young (3-week-old) and old (20-week-old) rats. All animals were intraperitoneally injected once with CDDP (6 mg/kg). After 3 days, all animals were sacrificed and serum, urine, and kidney tissues were collected. Urinary and serum biomarkers as well as histological changes were measured. CDDP-induced proximal tubular damage was apparent from histopathological examination, being more severe in 3-week-old rats accompanied by increased number of TUNEL-positive apoptotic cells. This was associated with elevated urinary kidney injury molecule-1 (KIM-1), glutathione-S-transferase alpha (GST-α), vascular endothelial growth factor (VEGF), and tissue inhibitor of metalloproteinases-1 (TIMP-1). In contrast, the levels of neutrophil gelatinase-associated lipocalin (NGAL) and osteopontin were significantly increased in 20-week-old rats after CDDP treatment. These results indicate that the use of age-specific urinary biomarkers is necessary to diagnosis of CDDP-induced AKI. Especially, urinary KIM-1, GST-α, TIMP-1, and VEGF levels may help in the early diagnosis of young patients with CDDP-induced AKI.

The alarmin concept applied to human renal transplantation: evidence for a differential implication of HMGB1 and IL-33.

The endogenous molecules high mobility group box 1 (HMGB1) and interleukin-33 (IL-33) have been identified as alarmins, capable of mediating danger signals during tissue damage. Here, we address their possible role as innate-immune mediators in ischemia-reperfusion injury (IRI) following human kidney transplantation. We analysed serum and urinary HMGB1 and IL-33 levels, all determined by enzyme-linked immunosorbent assay, in a cohort of 26 deceased renal transplant recipients. Urinary HMGB1 and IL-33 levels were significantly increased as soon as 30 min after reperfusion, as compared to those before treatment. Moreover, both serum and urinary IL-33 (but not HMGB1) increase was positively correlated with cold ischemia time, from 30 min to 3 days post-transplantation. In vitro, human umbilical vein endothelial cells subjected to hypoxia conditions released both HMGB-1 and IL-33, while only the latter was further increased upon subsequent re-oxygenation. Finally, we postulate that leukocytes from renal recipient patients are targeted by both HMGB1 and IL-33, as suggested by increased transcription of their respective receptors (TLR2/4 and ST2L) shortly after transplantation. Consistent with this view, we found that iNKT cells, an innate-like T cell subset involved in IRI and targeted by IL-33 but not by HMGB1 was activated 1 hour post-transplantation. Altogether, these results are in keeping with a potential role of IL-33 as an innate-immune mediator during kidney IRI in humans.

Lupus nephritis - alarmins may sound the alarm?

A growing body of literature has documented the elevated levels of the alarmin HMGB1 in lupus skin and serum. Two recent reports highlight the increased expression of HMGB1 in lupus nephritis, within the diseased kidneys or in the urine. Taken together with previous reports, these findings suggest that the interaction of HMGB1 with a variety of receptors, including receptor for advanced glycation end products (RAGE) and Toll-like receptors, might play a role in the pathogenesis of lupus nephritis. These studies introduce urinary HMGB1 as a novel biomarker candidate in lupus nephritis. Whether alarmins would be effective in sounding the alarm at the incipience of renal damage remains to be ascertained.

Urine levels of HMGB1 in Systemic Lupus Erythematosus patients with and without renal manifestations.

INTRODUCTION: Lupus nephritis (LN) is a severe and frequent manifestation of systemic lupus erythematosus (SLE). Its pathogenesis has not been fully elucidated but immune complexes are considered to contribute to the inflammatory pathology in LN. High Mobility Group Box 1 (HMGB1) is a nuclear non-histone protein which is secreted from different types of cells during activation and/or cell death and may act as a pro-inflammatory mediator, alone or as part of DNA-containing immune complexes in SLE. Urinary excretion of HMGB1 might reflect renal inflammatory injury. To assess whether urinary HMGB1 reflects renal inflammation we determined serum levels of HMGB1 simultaneously with its urinary levels in SLE patients with and without LN in comparison to healthy controls (HC). We also analyzed urinary HMGB1 levels in relation with clinical and serological disease activity. METHODS: The study population consisted of 69 SLE patients and 17 HC. Twenty-one patients had biopsy proven active LN, 15 patients had a history of LN without current activity, and 33 patients had non-renal SLE. Serum and urine levels of HMGB1 were both measured by western blotting. Clinical and serological parameters were assessed according to routine procedures. In 17 patients with active LN a parallel analysis was performed on the expression of HMGB1 in renal biopsies. RESULTS: Serum and urinary levels of HMGB1 were significantly increased in patients with active LN compared to patients without active LN and HC. Similarly, renal tissue of active LN patients showed strong expression of HMGB1 at cytoplasmic and extracellular sites suggesting active release of HMGB1. Serum and urinary levels in patients without active LN were also significantly higher compared to HC. Urinary HMGB1 levels correlated with SLEDAI, and showed a negative correlation with complement C3 and C4. CONCLUSION: Levels of HMGB1 in urine of SLE patients, in particular in those with active LN, are increased and correlate with SLEDAI scores. Renal tissue of LN patients shows increased release of nuclear HMGB1 compared to control renal tissue. HMGB1, although at lower levels, is, however, also present in the urine of patients without active LN. These data suggest that urinary HMGB1 might reflect both local renal inflammation as well as systemic inflammation.