Therapeutic progress of diabetic periodontitis / 口腔疾病防治

@#In China, the prevalence of diabetes is increasing each year. Diabetic periodontitis, as a major complication of diabetes, retains a strong bidirectional correlation with diabetes. Periodontitis increases the risk of diabetes, and hyperglycemia aggravates periodontal inflammation. In recent years, efficient treatments for diabetic periodontitis have been increasingly emphasized, and the prevention and control of diabetic periodontitis remain difficult. It is very challenging to find the best way to recognize risk factors and increase the diagnosis rate. Deeply understanding the local and systematic biological features of diabetic periodontitis could lead to the development of clinical preventive and therapeutic strategies. This article reviews the clinical and biological characteristics of diabetic periodontitis and its treatment progress from a systemic and local perspective, with the aim of extending the work on the prevention and treatment strategies for diabetic periodontitis.

Protective effect of pyridoxamine on RPE cells treated with AGEs / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To observe the effects of pyridoxamine(PM)on RAGE, ROS and apoptosis in RPE cells treated with advanced glycation end products(AGEs), and to investigate the protective effect of PM on RPE cells in diabetic retinopathy. <p>METHODS:Primary cultured human RPE cells, the third generation of cells were synchronized with serum-free Dulbecco-modified Eagle medium for 24h, and then grouped: 1)Control group: cultured with 100mg/L BSA for 48h; 2)AGEs-treated group: cultured with 200mg/L AGEs for 48h; 3)PM group: PM1 group: cultured with 16mg/L PM+200mg/L AGEs for 48h; PM2 group: cultured with 32mg/L PM+200mg/L AGEs for 48h. The expression of RAGE protein was detected by immunohistochemistry. The formation of ROS was observed by fluorescence microscopy. The apoptosis of cells was detected by TUNEL. <p>RESULTS:The expression of RAGE protein, ROS and apoptosis of RPE cells in PM group were significantly lower than those in AGEs-treated group, and decreased with the increase of PM concentration. <p>CONCLUSION:Pyridoxamine can inhibit the expression of RAGE and the production of ROS, reduce apoptosis, and have a protective effect on RPE cells.

Inhibition of puerarin on formation of advanced glycation end products in vivo and in vitro / 中草药

Objective: To study the influence of puerarin on advanced glycation end products (AGEs) formation in vivo and in vitro. Methods: C57BL/6 mice were fed with high fat diet and injected with streptozotocin (40 mg/kg) to establish diabetic model. After modeling, mice were randomly divided into blank control group, model group, aminoguanidine group (100 mg/kg), low-, mid-, and high-dose puerarin (50, 100, and 200 mg/kg) groups. After treatment of eight weeks, the levels of fasting blood-glucose (FBG), oral glucose tolerance test (OGTT), glycated serum protein (GSP), and AGEs in serum were detected. To establish non-enzymatic glycation reaction model in vitro, glucose and bovine serum albumin (BSA) were incubated with puerarin at different concent rations respectively for 144 h. The productivity of non-enzymatic advanced glycation end products was detected by aspectrophotometer. Results: Puerarin can significantly lower the level of FBG and oral glucose tolerance in diabetic mice, and inhibit the productivity of GSP and AGEs in serum. Besides, puerarin also significantly inhibits the productivity of AGEs in vitro in a concentration-dependent manner. Conclusion: Puerarin possesses significant inhibitory effect on the productivity of AGEs in vivo and in vitro.

Inhibitory effects of extracts from Moutan Cortex on formation of advanced glycation end-products (AGEs) in vitro and specificity screening of its potential active components / 中国中药杂志

In this study, bovine serum albumin (BSA)/methylglyoxal (MGO) non-enzymatic glycosylation reaction system was used for the evaluation of the inhibitory effects of Moutan Cortex extracts on the formation of AGEs. The HPLC-LC-ESI-MS/MS technology was adopted to test and indentify active components in Moutan Cortex against AGEs formation. The different concentrations of extracts (crude herb concentration 50, 100, 150, 200, 250 g•L⁻¹) from Moutan Cortexwas determined by fluorospectrophotometry, indicating an activity against AGEs formation in different concentrations of extracts, the inhibition ratio were (36.2±5.3)%, (43.5±6.2)%, (55.4±7.8)%, (68.6±6.7)%, (70.4±8.2)%, respectively after 6-day reaction in a dose dependent manner. Besides, the forming speed of AGEs tended to be steady after 24 h reaction. The HPLC technology was used to analyze chromatograms before and after the incubation of Moutan Cortex and methylglyoxal, identify changes in five chromatographic peaks and show decrease or increase in chromatographic peaks. These substances were trigalloyl glucose, tetragalloyl glucose, galloylpaeoniflorin, hexagalloyl glucose and benzoylpaeoniflorin after LC-ESI-MS/MS identification. Extracts from Moutan Cortex showed the remarkable inhibitory effects against formation of AGEs in BSA/glucose system. Furthermore, these potential active components might be associated with the efficacy of Moutan Cortex on treatment of diabetic nephropathy, which enriches basic studies for Moutan Cortex and provides ideas and reference basis for subsequent studies.

Liraglutide alleviates arteriosclerotic lesions and inhibits receptor for advanced glycation end-product expression in diabetic ApoE-/- mice / 中华内分泌代谢杂志

Objective To observe the effect of liraglutide on arteriosclerotic lesions in diabetic ApoE- / - mice via regulating receptor for advanced glycation end-products ( RAGE) expression of aorta. Methods Thirty male ApoE- / - mice (10 weeks) were randomly divided into 3 groups: control group, diabetes group, and diabetes +liraglutide group. All the mice were fed a high-fat diet. Diabetes was induced by 2 intraperitoneal injection of streptozotocin (STZ, 55 mg·kg-1 ·d-1 ). Mice were treated with subcutaneous injection of liraglutide (0. 4 mg· kg-1 ·d-1 ) in diabetes+liraglutide group. After 9 weeks, blood was drawn and the aorta were rapidly procured. The serum levels of advanced glycation end-products ( AGEs), soluble receptor for advanced glycation end-products (sRAGE ), stromal cell-derived factor-1α ( SDF-1α ), as well as total cholesterol and triacylglycerol were measured. Atherosclerotic plaque area was determined by Sudan Ⅳ staining. The expression of RAGE was determined using RT-PCR and western blot, respectively. Results Compared with control group, the serum AGEs, the RAGE protein and mRNA expression, the atherosclerotic lesions were significantly increased in the mice of diabetes group. Compared with diabetes group, the serum AGEs, the RAGE protein and mRNA expression, the atherosclerotic lesions were significantly decreased in the mice of diabetes+liraglutide group. Conclusion Liraglutide is confirmed as to have an anti-atherosclerotic effects and inhibits RAGE expression in diabetic ApoE- / - mice.

Effect of exogeneous adiponectin on hippocampal AGEs-ROS-ERS pathway in aged mice with postoperative cognitive dysfunction / 中华麻醉学杂志

Objective To evaluate the effect of exogeneous adiponectin on hippocampal advanced glycation end products (AGEs)-reactive oxygen species (ROS)-endoplasmic reticulum stress (ERS) pathway in aged mice with postoperative cognitive dysfunction (POCD).Methods Thirty-two healthy male C57BL/6 mice, aged 18 months, weighing 20-25 g, were randomly divided into 4 groups (n=8 each) using a random number table: control group (group C), POCD group, exogeneous adiponectin group (group APN), and vehicle group (group Veh).Splenectomy was performed to establish the POCD model in aged mice anesthetized with intraperitoneal pentobarbital sodium.In group APN, adiponectin 0.1 μg/g (in 2 μl of phosphate buffer solution) was injected into the lateral cerebral ventricle at 30 min before establishing the model.Phosphate buffer solution 2 μl was given at 30 min before establishing the model in group Veh.Cognitive function was assessed on day 7 after surgery.The mice were then sacrificed, and the hippocampus was harvested for determination of the area of AGE deposition (by immunohistochemistry), levels of ROS (by flow cytometry), and levels of glucose-regulated protein 78 (GRP78), C/EBP-homologous protein (CHOP), caspase-12 and ROS (using Western blot).Results Compared with group S, the freezing time in the contextual fear conditioning test was significantly shortened, the area of AGE deposition and levels of ROS, CHOP and caspase-12 were increased, and the level of GRP78 was decreased in POCD, APN and Veh groups.Compared with POCD and Veh groups, the freezing time in the contextual fear conditioning test was significantly prolonged, the area of AGE deposition and levels of ROS, CHOP and caspase-12 were decreased, and the level of GRP78 was increased in group APN.Conclusion Exogeneous adiponectin decreases the occurrence of POCD probably by blocking hippocampal AGEs-ROS-ERS pathway in aged mice.

Effect of urinary proteins and advanced glycosylation end products on ly-sosomes in renal tubular epithelial cells / 中国病理生理杂志

[ ABSTRACT] AIM:To investigate the effects of pathological products, urinary proteins and advanced glycosyla-tion end products ( AGE) produced in the progression of chronic kidney disease ( CKD) , on the structure and function of lysosomes in renal tubular epithelial cells ( TECs ) , and try to find a novel approach for preventing or delaying CKD. METHODS:The renal specimens of the untreated patients with minimal change nephrotic syndrome (MCNS), diabetic nephropathy (DN) or normal kidney were collected.The expression of lysosomal-associated membrane protein 1 (LAMP1) and cathepsin B ( CB) was studied in TECs by indirect immunofluorescent staining.Human renal tubular epithelial cell line HK-2 was incubated with 8 g/L urinary proteins or 100 mg/L AGE.The expression of LAMP1 and CB was investigated by indirect immunofluorescence and the activity of CB and cathepsin L ( CL) was measured by biochemical and enzymatic as-says.The degradation of DQ-ovalbumin was also determined.RESULTS: The lysosomal membrane permeabilization oc-curred in the TECs of MCNS and DN patients.After treatment with urinary proteins or AGE-BSA, the lysosomal membrane permeabilization of the HK-2 cells was increased.The activity of CB and CL and degradation of DQ-ovalbumin were de-creased as compared with normal control group.CONCLUSION:The digestive function of lysosome was decreased and ly-sosomal membrane permeabilization occurred in the TECs exposed to urinary proteins and AGE, which might be a key factor to induce the tubulointerstitial fibrosis.

Modulation of cell proliferation, survival and gene expression by RAGE and TLR signaling in cells of the innate and adaptive immune response: role of p38 MAPK and NF-KB

Objective: The aim of this study was to evaluate a possible synergism between AGE-RAGE and TLR4 signaling and the role of p38 MAPK and NF-kB signaling pathways on the modulation of the expression of inflammatory cytokines and proliferation of cells from the innate and adaptive immune response. Material and Methods: T lymphocyte (JM) and monocyte (U937) cell lines were stimulated with LPS and AGE-BSA independently and associated, both in the presence and absence of p38 MAPK and NF-kB inhibitors. Proliferation was assessed by direct counting and viability was assessed by a biochemical assay of mitochondrial function. Cytokine gene expression for RAGe, CCL3, CCR5, IL-6 and TNF-α was studied by RT-PCR and RT-qPCR. Results: RAGE mRNA expression was detected in both cell lines. LPS and AGE-BSA did not influence cell proliferation and viability of either cell line up to 72 hours. LPS and LPS associated with AGE induced expression of IL-6 and TNF-α in monocytes and T cells, respectively. Conclusions: There is no synergistic effect between RAGE and TLR signaling on the expression of IL-6, TNF-α , RAGE, CCR5 and CCL3 by monocytes and lymphocytes. Activation of RAGE associated or not with TLR signaling also had no effect on cell proliferation and survival of these cell types. .

Modulação da proliferação, apoptose e autofagia de linfocitos T por RAGE / Modulation of proliferation, apoptosis and autophagy of T lymphocytes by RAGE

RAGE (Receptor for advanced glycation endproducts) é um receptor de padrões moleculares (PRR) reconhecido principalmente por seu envolvimento na patogênese de complicações secundárias do Diabetes Mellitus. Sua ativação está relacionada à modulação da resposta imune-inflamatória, promovendo ativação, migração e maturação de leucócitos; e induzindo a liberação de citocinas inflamatórias. A sinalização via RAGE também está relacionada à regulação dos processos de sobrevivência, apoptose e autofagia celular. Pouco se sabe sobre o papel de RAGE na regulação desses processos em células da resposta imune adaptativa e, especificamente, em linfócitos T. Nosso objetivo foi avaliar a influência da ativação de RAGE na biologia de células T. Para isso, observamos seus efeitos na modulação da proliferação, apoptose e autofagia, e a contribuição da ativação de NF-κB na indução desses processos. Células de linhagem de linfócitos T (JM/Jurkat) foram estimuladas com os ligantes de RAGE BSA-AGE (100 e 200 µg/mL) e S100b (10 µg/mL). A estimulação também foi realizada mediante a pré-ativação do receptor de células T (TCR). Experimentos de ganho de função foram realizados por meio da transfecção transitória das células com um vetor plasmidial de RAGE. A proliferação celular foi determinada por ensaio de exclusão com azul de Trypan. A apoptose foi avaliada através da detecção da fragmentação do DNA pelo método TUNEL e da expressão de p53, Bax e Bcl-2 por Western blot. Ainda por Western blot, observamos a expressão de proteínas características do processo de autofagia, p62 e LC3; e de p50, subunidade do fator de transcrição NF-κB. O estímulo com BSAAGE e S100b não alterou a proliferação e a viabilidade, mas, em células transfectadas, esse estímulo tendeu a aumentar a proliferação e a viabilidade, sobretudo após a pré-ativação das células. O BSA-AGE e o S100b produziram efeitos distintos sobre a apoptose a depender da pré-ativação e do período experimental avaliado. O estímulo com os agonistas de RAGE atenuaram a inibição de LC3 e p53 induzida pela camptotecina. Efeito de "resgate" semelhante foi verificado na expressão de p50. O estímulo de RAGE parece favorecer a proliferação e a autofagia de linfócitos T. Os seus efeitos sobre a apoptose variam de acordo com o tipo, concentração e período de exposição ao ligante utilizado

RAGE (Receptor for advanced glycation endproducts) is a pattern recognition receptor (PRR) that is especially recognized for its involvement in the pathogenesis of secondary complications of Diabetes Mellitus. Its activation is related to the modulation of immune-inflammatory responses, by promoting activation, maturation and migration of leukocytes, as well as the induction of inflammatory cytokines. RAGE signaling is also related to the regulation of cellular processes like survival, apoptosis and autophagy. Little is known about the role of RAGE in the regulation of these processes in cells of the adaptive immune response, specifically in T lymphocytes. Our main purpose was to evaluate the influence of RAGE activation in T cell biology. We observed the role of this receptor in the modulation of proliferation, apoptosis and autophagy, and the contribution of NF-kB in the induction of these processes. A human cell line of T lymphocytes (JM/ Jurkat) were stimulated with the RAGE ligands: BSA-AGE (100 and 200 µg/mL) and S100B (10 µg/mL). The stimulation was also performed after pre-activation of the T cell receptor (TCR). Gain of function experiments were performed by transient transfection of cells with a plasmid vector to increase RAGE expression. Cell proliferation was determined by Trypan blue dye exclusion assay. Apoptosis was assessed by the detection of DNA fragmentation by the TUNEL method, and p53, Bax and Bcl-2 expression by Western blotting. Also by Western blotting, we observed the effect of RAGE on the expression of autophagy-related proteins, p62 and LC3, and on the expression of p50, subunit of the transcription factor, NF-κB. The stimulation with BSA- AGE and S100b did not alter the proliferation and viability, but, in transfected cells, this stimulation tended to increase proliferation and viability, especially after the cell pre-activation. BSA-AGE and S100b had distinct effects on apoptosis depending on the cell pre-activation and trial period evaluated. The stimulation with agonists of RAGE attenuated the inhibition of LC3 and p53 induced by camptothecin. Similar effect was observed in the expression of p50. The stimulation of RAGE seems to favor the proliferation and autophagy of T lymphocytes. Its effects on apoptosis may vary according to the type, concentration and period of exposure to the ligand used

Diabetic conditions modulate the adenosine monophosphate-activated protein kinase of podocytes

BACKGROUND: Adenosine monophosphate-activated protein kinases (AMPKs), as a sensor of cellular energy status, have been known to play an important role in the pathophysiology of diabetes and its complications. Because AMPKs are known to be expressed in podocytes, it is possible that podocyte AMPKs could be an important contributing factor in the development of diabetic proteinuria. We investigated the roles of AMPKs in the pathological changes in podocytes induced by high-glucose (HG) and advanced glycosylation end products (AGEs) in diabetic proteinuria. METHODS: We prepared streptozotocin-induced diabetic renal tissues and cultured rat and mouse podocytes under diabetic conditions with AMPK-modulating agents. The changes in AMPKalpha were analyzed with confocal imaging and Western blotting under the following conditions: (1) normal glucose (5mM, =control); (2) HG (30mM); (3) AGE-added; or (4) HG plus AGE-added. RESULTS: The density of glomerularphospho-AMPKalpha in experimental diabetic nephropathy decreased as a function of the diabetic duration. Diabetic conditions including HG and AGE changed the localization of phospho-AMPKalpha from peripheral cytoplasm to internal cytoplasm and peri- and intranuclear areas in podocytes. HG reduced the AMPKalpha (Thr172) phosphorylation of rat podocytes, and similarly, AGEs reduced the AMPKalpha (Thr172) phosphorylation of mouse podocytes. The distributional and quantitative changes in phospho-AMPKalpha caused by diabetic conditions were preventable using AMPK activators, metformin, and 5-aminoimidazole-4-carboxamide-1beta-riboside. CONCLUSION: We suggest that diabetic conditions induce the relocation and suppression of podocyte AMPKalpha, which would be a suggestive mechanism in diabetic podocyte injury.

Advanced glycosylation end products promote migration of podocytes through mTORC1/uPAR pathway / 中国病理生理杂志

[ ABSTRACT] AIM: To investigate the influence of advanced glycosylation end products-modified bovine serum albumin (AGE-BSA) on mammalian target of rapamycin complex 1 (mTORC1), urokinase-type plasminogen activator re-ceptor ( uPAR) , and cell mobility in the podocytes, and to further explore the probable relationship.METHODS: The conditionally immortalized mouse podocyte cell line was cultured in vitro.MTT assay and immunofluorescence were used to analyze the cell viability and cytoskeleton of the podocytes treated with the stimuli and intervention agents.The activity of mTORC1 and the expression level of uPAR in normal podocytes and podocytes treated with control BSA or AGE-BSA were detected by Western blotting.The migration ability of the podocytes was determined by would-healing assay.Rapamycin was added to inhibit the activity of mTORC1 along with the addition of AGE-BSA to observe the changes of uPAR and the motility of podocytes.RESULTS:No significant difference of the cell viability or cytoskeleton in the podocytes treated with the stimuli and intervention agents was observed.AGE-BSA up-regulated the activity of mTORC1 and the expression of uPAR, and induced the high mobility of the podocytes.Rapamycin obviously reduced the high expression level of uPAR and the increase in the migration ability of podocytes caused by AGE-BSA treatment.CONCLUSION: AGE-BSA might cause the high migration of podocytes through the mTORC1/uPAR signaling pathway.

Expression of high mobility group box 1 protein and the receptor for advanced glycation end products in patients with primary gouty arthritis / 中华风湿病学杂志

Objective To investigate the role of high mobility group box 1 protein(HMGB1) and the receptor for advanced glycation end products (RAGE) in the pathogenesis of primary gouty arthritis (GA).Methods Enzyme-linked immunosorbent assay(ELISA) was used to determine the level of plasma HMGB1 in 68 acute gout (AG),48 quiescent gout (QG) and 45 healthy control(HC).Real-time quantitative polymerase chain reaction (RT-qPCR) was employed to measure the expression of HMGB1 and RAGE mRNA in the peripheral blood mononuclear cells (PBMCs) in 68 AG,48 QG and 94 HC.One way ANOVA or Wilcoxon test and Spearman's correlations were used for statistical analysis.Results The level of plasma HMGB1,PBMCs HMGB1 and RAGE mRNA were significantly higher in GA than that in HC [(24±34) ng/ml,0.019±0.029,0.000 5±0.000 3] (P＜0.05),while the level of plasma HMGB1 and PBMCs HMGB1 mRNA were significantly higher in AG [(222±178) ng/ml,0.235±0.954,0.001 5±0.003 5] than that in QG [(107±176) ng/ml,0.044±0.117,0.001 3±0.000 9] (P＜0.05),and the level of PBMCs RAGE mRNA was higher in AG than that in QG (P＞0.05).In the GA patients,the level of plasma HMGB1 was positively correlated with white blood cell count,neutrophile granulocytes count,mononuclear cells and erythrocyte sedimentation rate (r=0.34,0.44,0.39,0.33; P＜0.05),while negatively correlated with apolipoprotein A1 (r=-0.28,P＜0.05); the level of PBMCs HMGB1 mRNA was positively correlated with RAGE mRNA,white blood cell counts,neutrophil counts,lymphocyte counts,serum total cholesterol level,low density lipoprotein level and apolipoprotein B100 level (r=0.29,0.36,0.26,0.28,0.29; P＜0.05),while negatively correlated with high density lipoprotein (r=-0.30,P＜0.01); the level of PBMCs RAGE mRNA was positively correlated with lymphocyte counts,total cholesterol and apolipoprotein B100 (r=0.35,0.35,0.44; P＜0.05).Conclusion HMGB1 and its signaling pathway may play important role in the pathogenesis of gouty arthritis,which may also be involved in the regulation of the lipid metabolism of gout.

Modulação da proliferação, morte celular e expressão gênica de mediadores inflamatórios pela ativação de RAGE e TLR4 em células da resposta imune inata e adaptativa: papel das vias de sinalização p38 MAPK e NF-kB / Modulation of cell proliferation, survival and gene expression of inflammatory mediators by RAGE and TLR signaling in cells of the innate and adaptive immune response: role of p38 MAPK and NF-KB

O diabetes está associado à maior susceptibilidade à infecções e sepsis, demonstrando uma influência desta condição sobre a resposta imune. A doença periodontal é um tipo de infecção crônica, modulada pela resposta imune que apresenta maior prevalência e severidade em pacientes diabéticos. Nosso objetivo foi avaliar um possível sinergismo entre os receptores RAGE e TLR4 na modulação da proliferação celular, atividade metabólica, apoptose e expressão de citocinas inflamatórias em células da resposta imune inata e adaptativa. Como objetivo secundário, avaliamos o papel das vias de sinalização p38 MAPK e NF-kB na expressão dos genes inflamatórios por estas células após estimulação de RAGE e TLR4. Linhagens de células humanas de linfócitos T (JM) e monócitos (U937) foram estimulados com LPS e AGE-BSA tanto de forma independente como associados. A estimulação foi realizada também na presença e na ausência de inibidores bioquímicos para p38 MAPK (SB203580) e NF-kB (Bay 11-7082). A proliferação celular foi determinada por ensaio de exclusão azul de trypan, a apoptose pela via intrínseca e atividade metabólica foi avaliada por um ensaio bioquímico da função mitocondrial, a expressão de citocinas foi estudada por RT-PCR e RTqPCR e a ativação das vias de sinalização de interesse pelos estímulos utilizados foi investigada através de Western blotting. LPS e AGE-BSA não influenciaram a proliferação e sobrevivência celular de monócitos e linfócitos T após 24, 48 e 72 h. LPS, isoladamente ou associado a AGE, induziu a expressão de IL-6 e TNF-α em monócitos e células T, respectivamente. A ativação de p38 MAPK, mas não de NF-kB, foi necessária para a indução por LPS e LPS/AGE de IL-6 e TNF-α por LPS, isolado ou associado a AGE. A expressão de RNAm de RAGE foi detectada em ambos os tipos de célulares. RNAm de CCL3 foi expresso de forma mais marcante em monócitos, particularmente após estimulação com LPS. Não houve sinergismo na sinalização entre RAGE e TLR4 na modulação da expressão de IL-6, TNF-α, RAGE, CCR5 e CCL3 por monócitos e células T

Diabetes is associated to increased susceptibility to infections and sepsis, indicating that this condition modulates the immune response. Periodontal disease is one type of chronic infection, which is modulated by the immune response and presents with increased prevalence and severity in diabetic patients. Our objective was to evaluate a possible synergism between RAGE and TLR4 signaling on the modulation of cell proliferation, metabolic activity, apoptosis and gene expression of inflammatory cytokines by cells of the innate and adaptive immune response. As a secondary objective, we assessed the role of p38 MAPK and NF-kB signaling pathways on the expression of the inflammatory genes by these cells after stimulation of RAGE and TLR4. Human cell lines of T lymphocytes (JM) and monocytes (U937) were stimulated with LPS and AGE-BSA both independently and associated. Stimulation was also performed in the presence and absence of biochemical inhibitors for p38 MAPK (SB203580) and NF-kB (Bay 11-7082). Cell proliferation was determined by trypan blue dye exclusion assay, apoptosis by the intrinsic pathway and metabolic activity were assessed by a biochemical assay of the mitochondrial function, cytokine gene expression was studied by RTPCR and RT-qPCR and the activation of the selected signaling pathways after RAGE and TLR4 activation was investigated by Western blotting. LPS and AGE-BSA did not influence cell proliferation and survival 24, 48 and 72 h after stimulation. LPS, alone or associated with AGE-BSA, induced expression of IL-6 and TNF- mRNA by monocytes and T cells, respectively. Activation of p38 MAPK, but not of NF-kB, was required for LPS and LPS/AGE-induced induction of IL-6 and TNF. RAGE mRNA expression was detected in both cell types. CCL3 mRNA expression levels were higher in monocytes upon stimulation with LPS. There was no synergism between RAGE and TLR4 signaling on the expression of IL-6, TNF-, RAGE, CCR5 and CCL3 by monocytes and T cells

Role of abnormal expression of receptor for advanced glycosylation end products in carcinogenesis, tumor invasion, and metastasis / 肿瘤

Receptor for advanced glycosylation end products (RAGE) has long been a focus in studies of diabetic vascular diseases. Recently, clinical evidence has shown that the expression level of RAGE is high in liver, stomach, pancreas and prostate cancer tissues whereas the expression level of RAGE is lower in chondrosarcoma and lung cancer tissues. The abnormal expression of RAGE is closely associated with tumorgenesis, angiogenesis, tumor invasion and metastasis. This review summarizes the abnormal expression of RAGE in different tumors and its role in carcinogenesis and development of tumor. Copyright© 2011 by TUMOR.

Beneficial effects of intensive therapy on arterial intima-media thickness and its risk factors in type 2 diabetic patients / 中华内分泌代谢杂志

Objective To analyze the changes of the intima-media thickness(IMT)of carotid and femoral arteries, serum advanced glycosylation end-products(AGEs),and AGEs soluble receptor(sRAGE)after intensively controlling blood glucose, blood pressure, and lipid. Methods One hundred and thirty-two type 2 diabetic patients were divided into 3 groups and followed for 5 years: 20 patients were treated with intensive control of blood glucose and blood pressure, 80 patients with intensive control of blood glucose, blood pressure, and lipid; and 32 patients with conventional therapy. AGEs, sRAGE, and IMT of carotid and femoral arteries were measured and compared among different groups. Results The IMT of carotid and femoral arteries and serum level of AGEs were significantly decreased after intensive treatment. The ratio of sRAGE and HbA1C(sRAGE/HbA1C)were negatively correlated with the mean of HbA1Cin the past five years(r=-0.417, P＜0.001)and the fluctuation of HbA1C(r=-0.309,P＜0.001). Multinomial regression analysis showed that AGEs were the important risk factors of IMT of femoral artery(β=0.152,P=0.068). Conclusion Intensive treatment is significant in controlling the growing IMT of carotid and femoral arteries, while decreasing serum level of AGEs.

Impact of interaction of advanced glycation end product and its receptor on podocytes apoptosis / 中华肾脏病杂志

Objective To study the effects of the interaction of advanced glycation end products (AGEs) and the receptor of AGEs (RAGE) on apoptosis of mice podocytes. Methods Podocytes were exposed to soluble AGEs such as bovine serum albumin (BSA), carboxymethyl-lysin (CML)-BSA, AGE-BSA and matrix-bound AGEs (AGE-modified collagen Ⅳ ), and to different concentrations of AGE, such as 10 mg/L, 50 mg/L, 100 mg/L. Apoptosis was assessed by TUNEL staining. Fluorescence-activated cell sorting (FACS) was used for the quantification of apoptotic andnecrotic podocytes after Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI) labeling. Apoptosis was described as the ratio of apoptotic cells to the total number cells under the high-power field, siRNA was transfected into podocytes through combining Dharmacon on Targetplus SMART pool siRNA reagents and Amaxa RNAi nucleofection kit. Results The apoptosis rate was higher in podoeytes exposed to either CML-BSA or AGE-BSA than that exposed to BSA. There was a two- to three-fold increase in apoptosis when podocytes were cultured in AGE-modified collagen Ⅳ as compared with native collagen Ⅳ. The apoptotic response of podocytes to AGE-BSA exposure occurred in a dose-dependent manner. Podocyte necrosis occurred only at the highest concentration of AGE-BSA(100 mg/L). AGE-BSA failed to induce apoptosis in podocytes transfected with RAGE siRNA. RAGE-specific gene knockdown did not significantly reduce the apoptosis of podocytes cultured in AGE-modified collagen IV. Conclusions The AGE-RAGE interaction plays a major role in the apoptosis of podocytes triggered by soluble AGEs, but not by matrix-bound AGEs. Reduction of AGE burden and RAGE expression may be important therapeutic approaches to prevent the progression of kidney disease.

N epsilon(Carboxymethyl)Lysine-Induced Mesangial Cell Activation / 대한신장학회잡지

BACKGROUND: Advanced glycation end products (AGE) are independent risk factors in the development and progression of diabetic nephropathy. Receptor for AGE(RAGE) is considered the main receptor involved in AGE-induced cell activation. Galectin-3, another AGE receptor, has recently been found upregulated in mesangial cells(MC) cultured under high glucose and in diabetic rat kidneys. N epsilon(carboxymethyl)lysine(CML) is a well characterized AGE but its role in MC activation is unknown. The present study examined the effects of CML on MC proliferation and extracellular matrix(ECM) secretion. METHODS: Synchronized rat MC were stimulated with different concentrations of CML-bovine serum albumin(BSA), control BSA, and transforming growth factor-beta(TGF-beta) for up to 72 hours. Cell proliferation was measured by [3H]-thymidine incorporation. Fibronectin, TGF-beta, plasminogen activator inhibitor(PAI)-1 secreted into the media and RAGE and galectin-3 expression in MC were measured by Western blot analysis and ELISA. RESULTS: 1,000 micro /mL of CML-BSA decreased [3H]-thymidine incorporation by MC at 48 hours and 10 ng/mL TGF-beta at 24 and 48 hours. CML-BSA 100 and 1,000 micro /mL, control BSA 1,000 micro /mL, and TGF-beta 10 ng/mL increased fibronectin secretion at 48 hours. CML-BSA up to 1,000 micro /mL did not affect TGF-beta or PAI-1 secretion. TGF-beta 10 ng/mL, however, significantly increased PAI-1 secretion. Cultured MC expressed both RAGE and galectin-3. CML-BSA 100 micro /mL upregulated galectin-3 expression. CONCLUSION: CML-BSA decreased MC proliferation and increased fibronectin secretion, suggesting that CML may lead to ECM accumulation and glomerulosclerosis in diabetic animals. MC express RAGE and galectin-3 constitutively and CML-induced galectin-3 upregulation may have a role in AGE-induced MC activation.

Advanced glycation end products-induced inflammatory reaction in human monocytes:cellular receptor pathway & intracellular signaling / 解放军医学杂志

Objective To investigate the cellular receptor pathway and the intracellular signaling of advanced glycation end products(AGE)-induced inflammatory reaction in monocytes. Methods Human peripheral monocytes were isolated from healthy volunteers. Cells were incubated with AGE modified by the addition of human serum albumin (AGE-HSA) either with pretreatment or no pretreatment of anti-AGE receptor (RAGE) IgG, NADPH oxidase inhibitor (apocynin)or a specific inhibitor of p38(SB 203580). The levels of interleukin-1?(IL-1?)and tumor necrosis factor-?(TNF-?) in the supernatants were assayed with enzyme-linked immunoadsorbent assay (ELISA). Reactive oxygen species (ROS) production was determined by MCLA chemiluminescence. Nuclear factor-?B translocation was assayed by immunochemical staining with anti-NF-?B/p65 and electrophoretic mobility shift assay(EMSA). Results AGE-HSA was found to induce activation of NF-?B, increase levels of IL-1? and TNF-? in the supernatants, and enhance production of ROS by monocytes. Pre-treatment of cells with anti-RAGE IgG or apocynin inhibited AGE-HSA to induce NF-?B translocation and IL-1? or TNF-? production. AGE stimulated ROS production could also be blocked by pre-treatment of cultured cells with anti-RAGE IgG or apocynin. Pre-treatment of cultured cells with SB 203580 inhibited both NF-?B activation and cytokines production, but showed no significant effect the cells to produce ROS. Conclusion AGE-HSA could induce IL-1? and TNF-? release as well as ROS production in human monocytes via a pathway mediated by RAGE. Activation of NADPH oxidase may be the upstream of the intracellular pathway. AGE-induced cytokines production was p38 pathway-dependent.

The serum level of advanced glycosylation end products and its expression in the vascular wall and peritoneum of uremic patients / 中华肾脏病杂志

Objective To determine the serum level of advanced glycosylation end products (AGEs) of uremia and its expression in the vascular wall and peritoneum of dialysis patient, meanwhile to explore its possible significance. Methods Using the ELISA method to determine the serum level of AGEs of uremia; using immunohistochemical method to examine the expression of AGEs in the vascular wall of uremia patients undergoing hemodialysis and the expression of AGEs and laminin in the peritoneum of uremic patients undergoing peritoneal dialysis. Results The AGEs level of uremic patients was obviously higher than that in the normal. No kind of dialysis could clear it well. AGEs accumulated obviously in the vassular wall of hemodialysis patients. After CAPD the expression of AGEs and laminin in the peritoneum was significantly strong. There was an obviously positive correlation between AGEs and laminin, so between these two and the time of peritoneal dialysis. Conclusions The high serum level of AGEs and its accumulation in the vascular wall may relate with the high incidence of cardic diseases of the uremia. Accumulation of AGEs and laminin in the peritoneum of patients undergoing peritoneal dialysis may be one of the reasons causing peritoneum sclerosis and dysultrafiltration

EFFECTS OF PRO-INFLAMMATORY MEDIATORS ON ADVANCED GLYCATION END PRODUCT RECEPTORS IN TYPE-B SYNOVIAL CELLS / 解放军医学杂志

To elucidate the effects of pro inflammatory mediators on mRNA expression of AGE receptors in type B synovial cells, type B synovial cells from normal subjects were isolated and cultured in vitro with human serum albumin modified with advanced glycation end products (AGE HSA), tumor necrosis factor ?(TNF ?)and interleukin 1?(IL 1?). The expression of RAGE mRNA and AGE R3 mRNA was examined by RT PCR techniques. TNF ?, AGE HSA and IL 1? up regulated the expression of AGE R3 mRNA in type B synovial cells in a dose and time dependent manner. In contrast, TNF ? and AGE HSA down regulated the expression of RAGE mRNA in a dose and time dependent manner. IL 1? had no effect on RAGE mRNA expression. The regulatory responses induced by AGE HSA were blocked by a neutralizing polyclonal anti human TNF ? antibody, suggesting that the effects of AGE HSA were mediated by TNF ?. The proinflammatory mediators may regulate the gene expression of AGE receptors in type B synovial cells, and the regulatory role of these receptors is different in response to the proinflammatory mediators.