Accumulation Of Advanced Oxidation Protein Products Research Articles

Advanced oxidation protein products upregulate ABCB1 expression and activity via HDAC2-Foxo3α-mediated signaling in vitro and in vivo

The unpredictable pharmacokinetics of non-renal cleared drugs in chronic kidney disease (CKD) patients is associated with the activity of drug transporters. However, the mechanisms underlying regulation of drug transporters are yet to be established. In this study, we demonstrated the involvement of a HDAC2-Foxo3α pathway in advanced oxidation protein products (AOPPs)-induced ATP-binding cassette subfamily B member 1 (ABCB1) expression and activity. The correlation of AOPPs accumulation with concentration of cyclosporine in plasma was evaluated in 194 patients with transplantation. Molecular changes in acetylation of various histones and related regulatory molecules were examined in HepG2 cell cultures treated with AOPPs. Accumulation of AOPPs in serum in relation to molecular changes in HDAC2-Foxo3α in vivo were evaluated in 5/6 nephrectomy (5/6 nx) and oral adenine (Adenine) CKD rat models. Interestingly, the cyclosporine level was negatively correlated with AOPPs in plasma. In addition, AOPPs markedly suppressed the expression of histone deacetylase 2 (HDAC2), inducing ABCB1 expression and activity in vitro and in vivo. Importantly, AOPPs modulated phosphorylation of Foxo3α and the upstream Akt protein. Our findings indicate that AOPPs regulate the expression and activity of ABCB1 via reducing HDAC2 expression and activating Foxo3α-dependent signaling. The collective results support the utility of AOPPs as a potential target for drug and/or dosage adjustment in CKD patients. Targeting of AOPPs presents a novel approach to regulate non-renal clearance.

Advanced oxidation protein products upregulate efflux transporter expression and activity through activation of the Nrf-2-mediated signaling pathway in vitro and in vivo

Clinical and benchtop studies suggest that chronic kidney disease (CKD) alters both renal and nonrenal clearance of drugs. Although studies have documented that the accumulating uremic toxins in the body under CKD conditions are humoral factors that alter the expression and/or activity of drug transporters, the specific process is poorly understood. In this study, we found that advanced oxidation protein products (AOPPs), which are a modified protein uremic toxin, could upregulate efflux transporters, including P-glycoprotein (ABCB1), multi-drug resistance-associated protein 2 (ABCC2) and breast cancer resistance protein (ABCG2) expression in CKD rat models and in HepG2 cells. Our research shows that renal function decline was associated with the accumulation of AOPPs in serum and the upregulation of efflux transporters in the liver in two rat models of CKD. In HepG2 cells, AOPPs significantly increased the expression of efflux transporters in a dose- and time-dependent manner and upregulated the mRNA expression, protein expression and activity of efflux transporters, but bovine serum albumin (BSA), a synthetic precursor of AOPPs, had no effect. This effect correlated with AOPPs activation of the nuclear factor E2-related factor 2 (Nrf-2)-mediated signaling pathway. Further investigation of the regulation of Nrf-2 by AOPPs revealed that ML385 and siNrf-2 abolished the upregulatory effects of AOPPs. These findings suggest that AOPPs upregulate ABCB1, ABCG2 and ABCC2 through Nrf-2 signaling pathways. Protein uremic toxins, such as AOPPs, may modify the nonrenal clearance of drugs in patients with CKD through effects on drug transporters.

Advanced Oxidation Protein Products Aggravate Tubulointerstitial Fibrosis Through Protein Kinase C-Dependent Mitochondrial Injury in Early Diabetic Nephropathy.

Diabetic nephropathy (DN) is the most common microvascular complications and the principal cause of mortality and morbidity rates in patients with diabetes. The expression of advanced oxidation protein products (AOPPs) has been found in vacuolated renal tubules in DN and correlated with patients' decreased renal function. The accumulation of AOPPs is regarded as an initiating factor in podocyte injuries via the protein kinase C (PKC) signaling, which plays a critical role in triggering oxidative stress and mitochondrial injuries in diseases including DN. Whether AOPPs could induce mitochondrial injuries and fibrosis in renal tubules remains largely unknown. Herein, we tested the hypothesis that the accumulation of AOPPs in diabetes incurs mitochondrial dysfunction and oxidative stress, causing renal tubulointerstitial fibrosis (TIF) via PKC signaling pathway. In vivo, intrarenal AOPPs accumulation correlated with oxidative stress, renal fibrosis, proteinuria, and declined renal function in DN patients and diabetic rats. AOPPs-induced mitochondrial injuries, apoptosis, and TIF were significantly mitigated by PKCη inhibition in diabetic rats. In vitro, high glucose (HG) stimulated AOPP expression and augmented PKC-mediated oxidative stress and fibrosis in HK-2 cells. Furthermore, we provide mechanistic evidence that inhibition of PKCη isoform alleviated mitochondrial injuries and function, attenuated apoptosis, and renal fibrosis in HG-cultured AOPPs-induced HK-2 cells. Innovation and Conclusion: We propose a novel mechanism that AOPPs-induced mitochondrial dysfunction and oxidative stress cause TIF in DN via activation of the PKCη isoform.

Advanced oxidation protein products promote NADPH oxidase-dependent β-cell destruction and dysfunction through the Bcl-2/Bax apoptotic pathway

The accumulation of plasma advanced oxidation protein products (AOPPs) has been linked with diverse disorders, including diabetes, chronic kidney disease, obesity, and metabolic syndrome. The aim of the present study was to evaluate the pathophysiological relevance of AOPPs in β-cell destruction and dysfunction. Exposure of cultured rat β-cells (INS-1) to AOPPs induced an increase in Bax expression, caspase-3 activity, and apoptosis as well as a decrease in Bcl-2 expression in a dose- and time-dependent manner. AOPP challenge rapidly increased the production of intracellular superoxide by activation of NADPH oxidases, demonstrated by p47phox translocation and interaction with p22phox and gp91phox, and this in turn led to apoptosis. AOPPs treatment resulted in β-cell apoptosis, AOPPs accumulation, and decreased insulin content in pancreas and plasma in unilateral nephrectomized rats. Chronic inhibition of NADPH oxidase by apocynin prevented β-cell apoptosis and ameliorated insulin deficiency in AOPP-challenged rats. This study demonstrates for the first time that accumulation of AOPPs promotes NADPH oxidase-dependent β-cell destruction and dysfunction by the Bcl-2/Bax–caspase apoptotic pathway. This finding may provide a mechanistic explanation for β-cell destruction and dysfunction in patients with diverse disorders.

Advanced oxidation protein products accelerate bone deterioration in aged rats

Advanced oxidation protein products (AOPPs) are novel markers of oxidation-mediated protein damage, and accumulation of AOPPs is involved in many pathophysiological conditions. Our previous studies demonstrated that the serum level of AOPPs negatively correlated with the age-related change in bone mineral density (BMD) in rats and that AOPPs inhibited rat osteoblast-like cell proliferation and differentiation in vitro. However, whether AOPPs are involved in senile osteoporosis is still largely unknown. The present study aimed to test the hypothesis that accumulation of AOPPs might accelerate bone deterioration in aged rats. Seventy 18-month-old male Sprague Dawley (SD) rats were randomized to intravenous injection of vehicle, native rat serum albumin (RSA), AOPPs-modified RSA (AOPPs-RSA) with or without oral administration of apocynin (a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor), or apocynin alone. After treatment for 8weeks or 16weeks, seven rats in each group were sacrificed. Bone and blood samples were harvested for BMD measurement, micro-computed tomographic (micro-CT) imaging, and biochemical analysis of circulating bone biomarkers. Compared to RSA- or vehicle-treated rats, AOPPs-RSA-treated animals displayed significantly decreased total vertebral BMD and deteriorated microstructure in both the tibias and the lumbar vertebral bodies, which were associated with down-regulated plasma bone-specific alkaline phosphatase concentration and up-regulated tartrate-resistant acid phosphatase 5b concentration. These AOPPs-induced perturbations in aged rats could be prevented by the oral administration of apocynin. However, no significant differences in BMD were detected in the femurs or the biomechanical parameters tested between the different treatment groups. These data suggest that accumulation of AOPPs accelerates bone deterioration in aged rats, likely via the activation of NADPH oxidase. This study provides new information toward understanding the pathogenic basis of senile osteoporosis and may provide targets for intervention.

Advanced oxidation protein products enhances soluble Fms-like tyrosine kinase 1 expression in trophoblasts: A possible link between oxidative stress and preeclampsia

Accumulation of advanced oxidation protein products (AOPPs) is prevalent in obesity, advanced maternal age, diabetes mellitus, and polycystic ovary syndrome. Alterations in the regulation and signaling of angiogenic pathways have been recognized as a link between these conditions and pre-eclampsia. To investigate the possible impact of AOPPs on soluble Fms-like tyrosine kinase 1 (sFlt-1) expression in trophoblasts. A trophoblast cell line (HRT-8/SVneo) was treated with various concentrations of AOPPs. The mRNA expression of sFlt-1, vascular endothelial growth factor (VEGF), and placental growth factor (PlGF) in trophoblasts were measured with the use of real-time polymerase chain reaction; and the secretion of sFlt-1, VEGF, and PlGF protein from trophoblasts were detected with the use of ELISA. Exposure of HRT-8/SVneo cells to AOPPs induced overexpression of sFlt-1 at mRNA and protein levels in a dose dependent manner. These effects could be inhibited by apocynin, an inhibitors of NADPH oxidase. Our data identified AOPPs as a class of important mediator in the regulation and signaling of angiogenic pathways of trophoblasts. Accumulation of AOPPs might contributes to the pathogenesis of preeclampsia by promoting sFlt-1 production in trophoblasts.

The receptor of advanced glycation end products plays a central role in advanced oxidation protein products-induced podocyte apoptosis

The accumulation of plasma advanced oxidation protein products (AOPPs) is prevalent in chronic kidney disease. We previously showed that accumulation of AOPPs resulted in podocyte apoptosis and their deletion by a cascade of signaling events coupled with intracellular oxidative stress. The transmembrane receptor that specifically transmits the AOPPs' signals to elicit cellular activity, however, remains unknown. Using co-immunoprecipitation and immunofluorescence, we found that AOPPs colocalized and interacted with the receptor of advanced glycation end products (RAGE) on podocytes. Blocking RAGE by anti-RAGE immunoglobulin G or its silencing by siRNA significantly protected podocytes from AOPPs-induced apoptosis both in vitro and in vivo and ameliorated albuminuria in AOPPs-challenged mice. AOPPs-induced activation of nicotinamide adenine dinucleotide phosphate oxidase and the excessive generation of intracellular superoxide were largely inhibited by anti-RAGE immunoglobulin G or RAGE siRNA. Moreover, blockade of RAGE decreased the activation of the p53/Bax/caspase-dependent proapoptotic pathway induced by AOPPs. Thus, AOPPs interact with RAGE to induce podocyte apoptosis and this, in part, may contribute to the progression of chronic kidney disease.

Accumulation of circulating advanced oxidation protein products is an independent risk factor for ischaemic heart disease in maintenance haemodialysis patients

Whether the burden of advanced oxidation protein products (AOPP) accumulation, a marker of oxidative stress, is affected by dialysis modality remains unclear. We compared the serum levels of AOPP in patients on haemodialysis (HD) and continuous ambulatory peritoneal dialysis (CAPD) and tested the hypothesis that an accumulation of AOPP was an independent risk factor for cardiovascular disease. This was a cross-section study. A total of 2095 patients (1539 HD, 556 CAPD) were recruited from the nine largest dialysis centres in China. Persons in medical centres for disease screening were selected as controls. Patients maintained on HD were dialyzed twice or thrice weekly. CAPD patients used lactate-buffered, glucose-containing solutions. The patients' data were abstracted from the medical record. The serum levels of AOPP were determined by spectrophotometric detection. The levels of AOPP were significantly elevated in both HD and CAPD patients compared to healthy controls. Accumulation of AOPP was more significant in HD compared to CAPD population. Meanwhile, AOPP accumulation was associated with the presence of ischaemic heart disease (IHD) only in HD, but not CAPD patients. A higher proportion of IHD was found in the HD population among those with higher levels of AOPP in each category of age and irrespective of the presence or absence of high triglyceride. Multivariate regression analysis indicated that accumulation of AOPP was an independent risk factor for IHD in HD population. Accumulation of AOPP was more significant in HD compared to CAPD patients. The level of AOPP was independently associated with IHD only in HD patients.

Advanced Oxidation Protein Products Activate Intrarenal Renin–Angiotensin Systemviaa CD36-Mediated, Redox-Dependent Pathway

Activation of intrarenal renin-angiotensin system (RAS) has a detrimental effect on the progression of chronic kidney diseases (CKDs), although the regulation of intrarenal RAS remains unclear. The aim of the present study was to evaluate the role of advanced oxidation protein products (AOPPs) in intrarenal RAS activation. AOPPs upregulated the expression of almost all components of RAS and increased activity of angiotensin-converting enzyme in cultured proximal tubular epithelial cells. The triggering effect of AOPP-albumin was 100-times stronger than that of unmodified albumin. The effect of AOPP-albumin was mainly mediated by a CD36-dependent, redox-sensitive signaling involving activation of protein kinase Cα, NADPH oxidase, and nuclear factor-κB/activation protein-1. Chronic AOPP-albumin loading in unilateral nephrectomy rats resulted in deposition of AOPPs in renal tubular cells accompanied with local RAS activation and functional perturbations such as increase in urinary albumin excretion. Accumulation of AOPPs was also detected in human renal tubular cells and correlated with expression of angiotensin II in renal biopsies from 19 patients with IgA nephropathy. This study demonstrated for the first time that AOPPs modified albumin functions as a strong trigger of intrarenal RAS via a CD36-mediated, redox-dependent pathway. Given the fact that accumulation of AOPPs is prevalent in diabetes and CKD, targeting AOPPs could be a strategy for the therapeutic intervention of CKD. Antioxid. Redox Signal. 18, 19-35.

Advanced Oxidation Protein Products and Advanced Glycation End Products in Children and Adolescents With Chronic Renal Insufficiency

Advanced oxidation protein products (AOPPs) represent dityrosine-containing cross-linked protein modifications formed mainly via myeloperoxidase reaction, supposed to accelerate the uremia-associated atherogenesis and renal fibrosis. DESIGN, SUBJECTS, AND MAIN OUTCOME MEASURES: In a cross-sectional study, we investigated the accumulation of AOPPs and advanced glycation end product (AGE)-specific fluorescence corrected for albumin in children and adolescents with chronic renal failure (CRF, n = 42), end-stage renal disease (ESRD, n = 12), kidney transplanted patients (Tx, n = 16), and age-matched healthy controls (n = 38). AOPP levels were 2.4-fold higher in the CRF and ESRD patients, and 1.6-fold higher in the transplanted subjects when compared with the controls (P < .001). In comparison with healthy controls, AGE levels rose 2-fold in the CRF, 7-fold in the ESRD, and 5-fold in the kidney transplanted children and adolescents, (P < .001). Patients with cardiovascular affliction presented with higher AGE levels than those without diagnosed cardiovascular disease (P < .02). In patients with stabilized renal function, AOPP and AGE levels did not change significantly during 12months. Pattern of accumulation of AOPP and AGE in children and adolescents with chronic renal disease differs. Accelerated rise in AOPP levels in some children and adolescents in predialysis stage of chronic renal insufficiency, inadequate to deterioration of renal function, might require further attention.

Advanced oxidation protein products inhibit differentiation and activate inflammation in 3T3‐L1 preadipocytes

Accumulation of advanced oxidation protein products (AOPPs) is prevalent in metabolic syndromes, a condition with impaired preadipocytes differentiation. In the present study, we tested the hypothesis that AOPPs disturb preadipocyte differentiation. Exposure of 3T3-L1 preadipocytes to increased levels of AOPPs inhibited accumulation of intracellular triglyceride and decreased the expression of the essential markers of matured adipocytes, such as adipocyte fatty-acid-binding protein (aP2), CAAT/enhancer-binding protein (C/EBP)-alpha, and peroxisome proliferator-activated receptor (PPAR)-gamma, in response to standard adipogenic induction. Inhibitory effects of AOPPs on preadipocytes differentiation was time sensitive, which occurred at the early stage of differentiation. In the presence of AOPPs, induction of preadipocytes differentiation resulted in upregulated expression of C/EBP homologous protein (CHOP) and CUG-Triplet repeat-binding protein (CUGBP), two important inhibitors of preadipocytes differentiation. In addition, treatment with AOPPs increased abundance of C/EBP-beta-liver enriched inhibitory protein (C/EBP-beta-LIP), a truncated C/EBP-beta isoform without adipogenic activity. Moreover, AOPPs-treated preadipocytes expressed a macrophage marker F4/80 and overexpressed tumor necrosis factor-alpha and interleukin-6 via nuclear factor-kappaB (NF-kappaB)-dependent pathway. However, blocking inflammation with NF-kappaB inhibitor failed to improve AOPPs-induced inhibition of preadipocytes differentiation. These data suggest that accumulation of AOPPs may inhibit differentiation of preadipocytes and activate inflammation in these cells. This information might have implication for understanding the impairment of preadipocytes differentiation and fat inflammation seen in metabolic syndrome.

Advanced oxidation protein products decrease expression of nephrin and podocin in podocytes via ROS-dependent activation of p38 MAPK

Accumulation of plasma advanced oxidation protein products (AOPPs) promotes progression of proteinuria and glomerulosclerosis. To investigate the molecular basis of AOPPs-induced proteinuria, normal Sprague-Dawley rats were treated with AOPPs-modified rat serum albumin. The expression of glomerular podocyte slit diaphragm (PSD)-associated proteins, nephrin and podocin, was significantly decreased coincident with the onset of albuminuria in rats treated with AOPPs. Chronic inhibition of NADPH oxidase by apocynin prevented down-regulation of nephrin and podocin and decreased albuminuria in AOPPs-challenged rats. This suggested that accumulation of AOPPs promotes proteinuria, possibly via down-regulating the expression of PSD-associated proteins.

Accumulation of advanced oxidation protein products induces podocyte apoptosis and deletion through NADPH-dependent mechanisms

The accumulation of plasma advanced oxidation protein products (AOPPs) is prevalent in diverse disorders such as diabetes, metabolic syndromes, and chronic kidney disease. To study whether accumulated AOPPs have an important role in the progression of proteinuria and glomerulosclerosis, we chronically treated normal Sprague-Dawley rats with AOPP-modified rat serum albumin. Podocyte apoptosis was significantly increased coincident with the onset of albuminuria and preceded significant losses of glomerular podocytes. Increasing the amount of AOPPs in the media of conditionally immortalized podocytes rapidly triggered the production of intracellular superoxide by activation of NADPH oxidase and this, in turn, led to an upregulation of p53, Bax, caspase 3 activity, and apoptosis. Chronic inhibition of NADPH oxidase by apocynin prevented podocyte apoptosis, ameliorated podocyte depletion, and decreased albuminuria in AOPP-challenged rats. Our study demonstrates that accumulation of AOPPs promotes NADPH oxidase-dependent podocyte depletion by a p53-Bax apoptotic pathway both in vivo and in vitro.

Advanced Oxidation Protein Products Promote Inflammation in Diabetic Kidney through Activation of Renal Nicotinamide Adenine Dinucleotide Phosphate Oxidase

The involvement of inflammatory processes has been recognized in development and/or progression of diabetic nephropathy. However, the mechanisms involved in the pathogenesis of renal inflammation have not been completely understood. In this study, we tested the hypothesis that accumulation of advanced oxidation protein products (AOPPs), which occurs in diabetes, may promote inflammatory responses in diabetic kidney. Streptozotocin-induced diabetic rats were randomized to iv injection of vehicle, native rat serum albumin (RSA), and AOPPs-modified RSA (AOPPs-RSA) in the presence or absence of oral administration of apocynin. A control group was followed concurrently. Compared with RSA- or vehicle-treated diabetic rats, AOPPs-RSA-treated animals displayed significant increase in renal macrophage infiltration and overexpression of monocyte chemoattractant protein-1 and TGF-beta1. This was associated with deteriorated structural and functional abnormalities of diabetic kidney, such as glomerular hypertrophy, fibronectin accumulation, and albuminuria. AOPP challenge significantly increased nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-dependent superoxide generation in renal homogenates and up-regulated membrane expression of renal NADPH oxidase subunits p47(phox) and gp91(phox). All these AOPPs-induced perturbations in diabetic kidney could be prevented by the NADPH oxidase inhibitor apocynin. These data suggest that chronic accumulation of AOPPs may promote renal inflammation in diabetes probably through activation of renal NADPH oxidase.

Advanced Oxidation Protein Products Accelerate Renal Fibrosis in a Remnant Kidney Model

Accumulation of plasma advanced oxidation protein products (AOPP) has been found in patients with chronic kidney disease. However, the biologic consequences of AOPP consumption on progression of renal disease still are unclear. For testing of the hypothesis that AOPP accelerate progression of chronic kidney disease, Sprague-Dawley rats were subjected to five-sixths nephrectomy (5/6 Nx) or to sham operation. Rats in each group were randomly assigned in three subgroups (n = 30 in each group) and treated with repeated intravenous injections of AOPP-modified rat serum albumin (RSA), unmodified RSA, or vehicle for indicated period. Compared with RSA- or vehicle-treated 5/6 Nx rats, AOPP RSA-treated 5/6 Nx rats displayed greater proteinuria, higher serum creatinine, and lower creatinine clearance. AOPP challenge resulted in more renal hypertrophy, higher macrophage influx, and greater renal fibrosis in the remnant kidney. Chronic administration of AOPP in sham-operated rats increased urinary protein excretion and renal macrophage infiltration, but histologic renal fibrosis was not observed during the study period. AOPP treatment enhanced AOPP level in renal tissue. This was associated with marked increase of thiobarbituric acid reactive substances, decrease of glutathione peroxidase activity, and upregulated expression of monocyte chemoattractant protein-1 and TGF-beta1 in renal cortex. These data indicate that AOPP might be a new and potentially important mediator of renal fibrosis in the remnant kidney. Chronic accumulation of AOPP promotes renal fibrosis probably via a redox-sensitive inflammatory pathway.