The Experts below are selected from a list of 2172 Experts worldwide ranked by ideXlab platform
Hai-qing Gao - One of the best experts on this subject based on the ideXlab platform.
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proteomic analysis of kidney and protective effects of grape seed Procyanidin B2 in db db mice indicate mfg e8 as a key molecule in the development of diabetic nephropathy
Biochimica et Biophysica Acta, 2013Co-Authors: Zhen Zhang, Qian Cai, Hai-qing Gao, Mei Cheng, Jun-fu Wang, Rui-hai Zhou, Lin ShenAbstract:Diabetic nephropathy, as a severe microvascular complication of diabetic mellitus, has become the leading cause of end-stage renal diseases. However, no effective therapeutic strategy has been developed to prevent renal damage progression to end stage renal disease. Hence, the present study evaluated the protective effects of grape seed Procyanidin B2 (GSPB2) and explored its molecular targets underlying diabetic nephropathy by a comprehensive quantitative proteomic analysis in db/db mice. Here, we found that oral administration of GSPB2 significantly attenuated the renal dysfunction and pathological changes in db/db mice. Proteome analysis by isobaric tags for relative and absolute quantification (iTRAQ) identified 53 down-regulated and 60 up-regulated proteins after treatment with GSPB2 in db/db mice. Western blot analysis confirmed that milk fat globule EGF-8 (MFG-E8) was significantly up-regulated in diabetic kidney. MFG-E8 silencing by transfection of MFG-E8 shRNA improved renal histological lesions by inhibiting phosphorylation of extracellular signal-regulated kinase1/2 (ERK1⁄2), Akt and glycogen synthase kinase-3beta (GSK-3β) in kidneys of db/db mice. In contrast, over-expression of MFG-E8 by injection of recombinant MFG-E8 resulted in the opposite effects. GSPB2 treatment significantly decreased protein levels of MFG-E8, phospho-ERK1/2, phospho-Akt, and phospho-GSK-3β in the kidneys of db/db mice. These findings yield insights into the pathogenesis of diabetic nephropathy, revealing MFG-E8 as a new therapeutic target and indicating GSPB2 as a prospective therapy by down-regulation of MFG-E8, along with ERK1/2, Akt and GSK-3β signaling pathway.
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proteomic analysis of aorta and protective effects of grape seed Procyanidin B2 in db db mice reveal a critical role of milk fat globule epidermal growth factor 8 in diabetic arterial damage
PLOS ONE, 2012Co-Authors: Qian Cai, Mei Cheng, Jian-hua Zhang, Jun-fu Wang, Rui-hai Zhou, Mei Yin, Zhen Zhang, Hai-qing GaoAbstract:Background Atherosclerosis is one of the major complications of type 2 diabetic patients (T2DM), leading to morbidity and mortality. Grape seed Procyanidin B2 (GSPB2) has demonstrated protective effect against atherosclerosis, which is believed to be, at least in part, a result of its antioxidative effects. The aim of this study is to identify the target protein of GSPB2 responsible for the protective effect against atherosclerosis in patients with DM. Methods and Results GSPB2 (30 mg/kg body weight/day) were administrated to db/db mice for 10 weeks. Proteomics of the aorta extracts by iTRAQ analysis was obtained from db/db mice. The results showed that expression of 557 proteins were either up- or down-regulated in the aorta of diabetic mice. Among those proteins, 139 proteins were normalized by GSPB2 to the levels comparable to those in control mice. Among the proteins regulated by GSPB2, the milk fat globule epidermal growth factor-8 (MFG-E8) was found to be increased in serum level in T2DM patients; the serum level of MFG-E8 was positively correlated with carotid-femoral pulse wave velocity (CF-PWV). Inhibition of MFG-E8 by RNA interference significantly suppressed whereas exogenous recombinant MFG-E8 administration exacerbated atherogenesis the db/db mice. To gain more insights into the mechanism of action of MFG-E8, we investigated the effects of MFG-E8 on the signal pathway involving the extracellular signal-regulated kinase (ERK) and monocyte chemoattractant protein-1 (MCP-1). Treatment with recombinant MFG-E8 led to increased whereas inhibition of MFG-E8 to decreased expression of MCP-1 and phosphorylation of ERK1/2. Conclusion Our data suggests that MFG-E8 plays an important role in atherogenesis in diabetes through both ERK and MCP-1 signaling pathways. GSPB2, a well-studied antioxidant, significantly inhibited the arterial wall changes favoring atherogenesis in db/db mice by down-regulating MFG-E8 expression in aorta and its serum level. Measuring MFG-E8 serum level could be a useful clinical surrogate prognosticating atherogenesis in DM patients.
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grape seed Procyanidin B2 inhibits human aortic smooth muscle cell proliferation and migration induced by advanced glycation end products
Bioscience Biotechnology and Biochemistry, 2011Co-Authors: Qian Cai, Hai-qing Gao, Jian-hua Zhang, Jun-fu Wang, Mei Yin, Zhen ZhangAbstract:Advanced glycation end product (AGE)-induced vascular smooth muscle cell (VSMC) proliferation is vital to the progression of diabetic vasculopathy. A grape seed Procyanidin extract has been reported to possess anti-oxidative and anti-inflammatory properties and to display a significant cardiovascular protective effect, but little is know about the underlying mechanism. The objective of this present study was to determine whether GSPB2 (grape seed Procyanidin B2), which is a dimeric Procyanidin and more biologically active, could inhibit AGE-induced VSMC proliferation by affecting the production of ubiquitin COOH-terminal hydrolase 1 (UCH-L1), the degradation of IκB-α and nuclear translocation of NF-κB in human aortic smooth muscle cells (HASMCs). Our data show that GSPB2 preincubation markedly inhibited AGE-induced proliferation and migration of HASMCs in a dose-dependent manner and upregulated the protein level of UCH-L1. Further studies revealed that the GSPB2 pretreatment markedly attenuated the degradation of IκB-α and nuclear translocation of NF-κB by modulating ubiquitination of IκB-α in AGE-exposed HASMCs. These results collectively suggest that AGE-induced HASMC proliferation and migration was suppressed by GSPB2 through regulating UCH-L1 and ubiquitination of IκB-α. GSPB2 may therefore have therapeutic potential in preventing and treating vascular complications of diabetes mellitus.
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Induction of lactadherin mediates the apoptosis of endothelial cells in response to advanced glycation end products and protective effects of grape seed Procyanidin B2 and resveratrol
Apoptosis : an international journal on programmed cell death, 2011Co-Authors: Qian Cai, Hai-qing Gao, Mei Cheng, Jian-hua Zhang, Jun-fu Wang, Rui-hai ZhouAbstract:One of characteristics of diabetes mellitus (DM) is endothelial cell (EC) dysfunction and apoptosis which contributes to the development of vasculopathy. Advanced glycation end products (AGEs) continuously produced in the setting of DM play an important role in causing EC dysfunction and apoptosis. However, the underlying molecular mechanism remains largely elusive. Lactadherin, a secreted glycoprotein of milk-fat globule, is expressed by multiple cell types of arterial wall including ECs. Our previous proteomic studies showed that the expression of lactadherin was significantly increased in the aorta of diabetic rats as compared with control rats and treatment with grape seed Procyanidin extracts significantly inhibited the lactadherin expression in diabetic rats. We hypothesized that lactadherin plays a critical role in AGEs-induced EC apoptosis; grape seed Procyanidin B2 (GSPB2) and resveratrol protect against AGEs-induced EC apoptosis through lactadherin regulation. Our results showed that AGEs upregulated lactadherin expression and lactadherin RNA interference significantly attenuated AGEs-induced EC apoptosis. Overexpression of lactadherin increased EC apoptosis with up-regulation of Bax/Bcl-2 ratio, cytochrome c release, caspase-9 and caspase-3 activation suggesting the involvement of mitochondria apoptosis pathway. Mechanistically, overexpression of lactadherin reduced the phosphorylation of GSK3beta at baseline. Our study also revealed nine proteins interacting with lactadherin in HUVEC and study of these candidate proteins could unveil further underlying molecular mechanisms. In summary, our study identified lactadherin as a key player responsible for AGEs-induced EC apoptosis and antioxidants GSPB2 and resveratrol protect against AGEs-induced EC apoptosis by inhibiting lactadherin. Targeting lactadherin with antioxidant could be translated into clinical application in the fighting against DM complications.
Qian Cai - One of the best experts on this subject based on the ideXlab platform.
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proteomic analysis of kidney and protective effects of grape seed Procyanidin B2 in db db mice indicate mfg e8 as a key molecule in the development of diabetic nephropathy
Biochimica et Biophysica Acta, 2013Co-Authors: Zhen Zhang, Qian Cai, Hai-qing Gao, Mei Cheng, Jun-fu Wang, Rui-hai Zhou, Lin ShenAbstract:Diabetic nephropathy, as a severe microvascular complication of diabetic mellitus, has become the leading cause of end-stage renal diseases. However, no effective therapeutic strategy has been developed to prevent renal damage progression to end stage renal disease. Hence, the present study evaluated the protective effects of grape seed Procyanidin B2 (GSPB2) and explored its molecular targets underlying diabetic nephropathy by a comprehensive quantitative proteomic analysis in db/db mice. Here, we found that oral administration of GSPB2 significantly attenuated the renal dysfunction and pathological changes in db/db mice. Proteome analysis by isobaric tags for relative and absolute quantification (iTRAQ) identified 53 down-regulated and 60 up-regulated proteins after treatment with GSPB2 in db/db mice. Western blot analysis confirmed that milk fat globule EGF-8 (MFG-E8) was significantly up-regulated in diabetic kidney. MFG-E8 silencing by transfection of MFG-E8 shRNA improved renal histological lesions by inhibiting phosphorylation of extracellular signal-regulated kinase1/2 (ERK1⁄2), Akt and glycogen synthase kinase-3beta (GSK-3β) in kidneys of db/db mice. In contrast, over-expression of MFG-E8 by injection of recombinant MFG-E8 resulted in the opposite effects. GSPB2 treatment significantly decreased protein levels of MFG-E8, phospho-ERK1/2, phospho-Akt, and phospho-GSK-3β in the kidneys of db/db mice. These findings yield insights into the pathogenesis of diabetic nephropathy, revealing MFG-E8 as a new therapeutic target and indicating GSPB2 as a prospective therapy by down-regulation of MFG-E8, along with ERK1/2, Akt and GSK-3β signaling pathway.
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proteomic analysis of aorta and protective effects of grape seed Procyanidin B2 in db db mice reveal a critical role of milk fat globule epidermal growth factor 8 in diabetic arterial damage
PLOS ONE, 2012Co-Authors: Qian Cai, Mei Cheng, Jian-hua Zhang, Jun-fu Wang, Rui-hai Zhou, Mei Yin, Zhen Zhang, Hai-qing GaoAbstract:Background Atherosclerosis is one of the major complications of type 2 diabetic patients (T2DM), leading to morbidity and mortality. Grape seed Procyanidin B2 (GSPB2) has demonstrated protective effect against atherosclerosis, which is believed to be, at least in part, a result of its antioxidative effects. The aim of this study is to identify the target protein of GSPB2 responsible for the protective effect against atherosclerosis in patients with DM. Methods and Results GSPB2 (30 mg/kg body weight/day) were administrated to db/db mice for 10 weeks. Proteomics of the aorta extracts by iTRAQ analysis was obtained from db/db mice. The results showed that expression of 557 proteins were either up- or down-regulated in the aorta of diabetic mice. Among those proteins, 139 proteins were normalized by GSPB2 to the levels comparable to those in control mice. Among the proteins regulated by GSPB2, the milk fat globule epidermal growth factor-8 (MFG-E8) was found to be increased in serum level in T2DM patients; the serum level of MFG-E8 was positively correlated with carotid-femoral pulse wave velocity (CF-PWV). Inhibition of MFG-E8 by RNA interference significantly suppressed whereas exogenous recombinant MFG-E8 administration exacerbated atherogenesis the db/db mice. To gain more insights into the mechanism of action of MFG-E8, we investigated the effects of MFG-E8 on the signal pathway involving the extracellular signal-regulated kinase (ERK) and monocyte chemoattractant protein-1 (MCP-1). Treatment with recombinant MFG-E8 led to increased whereas inhibition of MFG-E8 to decreased expression of MCP-1 and phosphorylation of ERK1/2. Conclusion Our data suggests that MFG-E8 plays an important role in atherogenesis in diabetes through both ERK and MCP-1 signaling pathways. GSPB2, a well-studied antioxidant, significantly inhibited the arterial wall changes favoring atherogenesis in db/db mice by down-regulating MFG-E8 expression in aorta and its serum level. Measuring MFG-E8 serum level could be a useful clinical surrogate prognosticating atherogenesis in DM patients.
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grape seed Procyanidin B2 inhibits human aortic smooth muscle cell proliferation and migration induced by advanced glycation end products
Bioscience Biotechnology and Biochemistry, 2011Co-Authors: Qian Cai, Hai-qing Gao, Jian-hua Zhang, Jun-fu Wang, Mei Yin, Zhen ZhangAbstract:Advanced glycation end product (AGE)-induced vascular smooth muscle cell (VSMC) proliferation is vital to the progression of diabetic vasculopathy. A grape seed Procyanidin extract has been reported to possess anti-oxidative and anti-inflammatory properties and to display a significant cardiovascular protective effect, but little is know about the underlying mechanism. The objective of this present study was to determine whether GSPB2 (grape seed Procyanidin B2), which is a dimeric Procyanidin and more biologically active, could inhibit AGE-induced VSMC proliferation by affecting the production of ubiquitin COOH-terminal hydrolase 1 (UCH-L1), the degradation of IκB-α and nuclear translocation of NF-κB in human aortic smooth muscle cells (HASMCs). Our data show that GSPB2 preincubation markedly inhibited AGE-induced proliferation and migration of HASMCs in a dose-dependent manner and upregulated the protein level of UCH-L1. Further studies revealed that the GSPB2 pretreatment markedly attenuated the degradation of IκB-α and nuclear translocation of NF-κB by modulating ubiquitination of IκB-α in AGE-exposed HASMCs. These results collectively suggest that AGE-induced HASMC proliferation and migration was suppressed by GSPB2 through regulating UCH-L1 and ubiquitination of IκB-α. GSPB2 may therefore have therapeutic potential in preventing and treating vascular complications of diabetes mellitus.
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Induction of lactadherin mediates the apoptosis of endothelial cells in response to advanced glycation end products and protective effects of grape seed Procyanidin B2 and resveratrol
Apoptosis : an international journal on programmed cell death, 2011Co-Authors: Qian Cai, Hai-qing Gao, Mei Cheng, Jian-hua Zhang, Jun-fu Wang, Rui-hai ZhouAbstract:One of characteristics of diabetes mellitus (DM) is endothelial cell (EC) dysfunction and apoptosis which contributes to the development of vasculopathy. Advanced glycation end products (AGEs) continuously produced in the setting of DM play an important role in causing EC dysfunction and apoptosis. However, the underlying molecular mechanism remains largely elusive. Lactadherin, a secreted glycoprotein of milk-fat globule, is expressed by multiple cell types of arterial wall including ECs. Our previous proteomic studies showed that the expression of lactadherin was significantly increased in the aorta of diabetic rats as compared with control rats and treatment with grape seed Procyanidin extracts significantly inhibited the lactadherin expression in diabetic rats. We hypothesized that lactadherin plays a critical role in AGEs-induced EC apoptosis; grape seed Procyanidin B2 (GSPB2) and resveratrol protect against AGEs-induced EC apoptosis through lactadherin regulation. Our results showed that AGEs upregulated lactadherin expression and lactadherin RNA interference significantly attenuated AGEs-induced EC apoptosis. Overexpression of lactadherin increased EC apoptosis with up-regulation of Bax/Bcl-2 ratio, cytochrome c release, caspase-9 and caspase-3 activation suggesting the involvement of mitochondria apoptosis pathway. Mechanistically, overexpression of lactadherin reduced the phosphorylation of GSK3beta at baseline. Our study also revealed nine proteins interacting with lactadherin in HUVEC and study of these candidate proteins could unveil further underlying molecular mechanisms. In summary, our study identified lactadherin as a key player responsible for AGEs-induced EC apoptosis and antioxidants GSPB2 and resveratrol protect against AGEs-induced EC apoptosis by inhibiting lactadherin. Targeting lactadherin with antioxidant could be translated into clinical application in the fighting against DM complications.
Gary Williamson - One of the best experts on this subject based on the ideXlab platform.
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Procyanidin B2 catabolism by human fecal microflora partial characterization of dimeric intermediates
Archives of Biochemistry and Biophysics, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Warren J Drynan, Denis Barron, Michael N CliffordAbstract:Abstract The catabolism by human fecal microflora of pure Procyanidin B2 ((–)-epicatechin-4β → 8-(–)-epicatechin) has been investigated using a static in vitro culture model. For the first time, 24 catabolites were detected by LC–MS ( n ) with M r greater than 290 indicating that they could not have formed from just one of the epicatechin units in the Procyanidin structure. Structures have been assigned on the basis of the fragmentation in the ion trap mass spectrometer and with regard to catabolic pathways known to occur in fecal microorganisms. Twenty of these ‘dimeric’ catabolites produced fragment ions characteristic of flavanols and/or proanthocyanidins. One catabolite was identified tentatively as either 6- or 8-hydroxy-Procyanidin B2. Thirteen were characterized as having been microbially reduced in at least one of the epicatechin units. Five contained an apparently unmodified epicatechin unit but in at least one case this was shown to consist of the B-ring of the “upper” epicatechin unit and the A-ring of the “lower”. These ‘dimeric’ catabolites were detected up to 9 h after the start of the incubation and collectively accounted for ∼20% of the substrate.
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a comparison of the in vitro biotransformation of epicatechin and Procyanidin B2 by human faecal microbiota
Molecular Nutrition & Food Research, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Warren J Drynan, Denis Barron, Michael N CliffordAbstract:The catabolism by human faecal microbiota of (-)-epicatechin (1) (2, 3-cis stereochemistry) and its dimer pure Procyanidin B2 (2), has been compared using a static in vitro culture model. The catabolites were characterised by LC-MS(n), UV absorption and relative retention time, and quantified relative to standards. No more than approximately 10% of Procyanidin B2 (2) was converted to epicatechin (1) by scission of the interflavan bond. Five phenolic acid catabolites (M(r) or =24 h incubation) were 5-(3'-hydroxy phenyl) valeric acid (9), 3-(3'-hydroxyphenyl) propionic acid (10) and phenyl acetic acid (12) (maximum yields 27.4+/-4.2, 38.2+/-4.2, 22.7+/-2.9%, respectively, from 1 and 9.4+/-1.2, 52.8+/-2.1, 28.8+/-1.6%, respectively, from 2). Substrate 2 was degraded twice as rapidly as 1. Evidence is presented for the production of previously unreported catabolites of 2 that retain the flavanol A-ring and the C4-->C8 interflavan bond. It was confirmed that catabolism favoured removal of the 4'-hydroxyl rather than the 3'-hydroxyl group and that both beta-oxidation and alpha-oxidation occurred.
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a comparison of the in vitro biotransformation of epicatechin and Procyanidin B2 by human faecal microbiota
Molecular Nutrition & Food Research, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Warren J Drynan, Denis Barron, Michael N CliffordAbstract:The catabolism by human faecal microbiota of (-)-epicatechin (1) (2, 3-cis stereochemistry) and its dimer pure Procyanidin B2 (2), has been compared using a static in vitro culture model. The catabolites were characterised by LC-MS n , UV absorption and relative retention time, and quantified relative to standards. No more than ∼10% of Procyanidin B2 (2) was converted to epicatechin (1) by scission of the interflavan bond. Five phenolic acid catabolites (M r < 290) were unique to 2, and ten phenolic acid catabolites (M r < 290) were common to both substrates. The dominant catabolites (≥24h incubation) were 5-(3'-hydroxy phenyl) valeric acid (9), 3-(3'-hydroxyphenyl) propionic acid (10) and phenyl acetic acid (12) (maximum yields 27.4 ± 4.2, 38.2 ± 4.2, 22.7 ± 2.9%, respectively, from 1 and 9.4±1.2, 52.8±2.1, 28.8±1.6%, respectively, from 2). Substrate 2 was degraded twice as rapidly as 1. Evidence is presented for the production of previously unreported catabolites of 2 that retain the flavanol A-ring and the C4 → C8 interflavan bond. It was confirmed that catabolism favoured removal of the 4'-hydroxyl rather than the 3'-hydroxyl group and that both β-oxidation and α-oxidation occurred.
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in vivo bioavailability absorption excretion and pharmacokinetics of 14c Procyanidin B2 in male rats
Drug Metabolism and Disposition, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Denis Barron, Florian Viton, Laurence J King, Jonathan E Brown, Michael N CliffordAbstract:Procyanidins are important biologically active compounds, but the pathway and extent of absorption and metabolism are controversial. We conducted a mass balance study to evaluate the total radioactivity excreted in urine and feces after oral administration of [(14)C]Procyanidin B2 to male rats (n = 5). Urine and feces were collected daily from 0 to 96 h. Absolute bioavailability of (14)C from [(14)C]Procyanidin B2 was calculated as approximately 82% using the values for total urinary (14)C. A pharmacokinetic study measured total radioactivity in the blood (n = 9). Blood samples were collected at designated time intervals (0.5-24 h) after administration. Three treatments were used: 1) intravenous, 2) oral higher dose (21 mg/kg b.wt.), and 3) oral lower dose (10.5 mg/kg). Blood concentration of total (14)C reached a maximum at approximately 6 h after ingestion of [(14)C]Procyanidin B2 (groups II and III), and area under the curve (AUC) was dependent on oral dose. After intravenous or oral administration the terminal half-lives were similar, whereas 8-fold larger values were obtained after oral dosing for total clearance and the apparent volumes of distribution. These pharmacokinetic differences explain the apparently lower (14)C bioavailability (8-11%) for [(14)C]Procyanidin calculated from blood [AUC((0-24))] values. After oral administration of [(14)C]Procyanidin B2, 63% was excreted via urine within 4 days. The data suggest that much of the parent compound administered orally is degraded by the gut microflora before absorption and that these microbial metabolites have a different distribution from the compounds circulating after the intravenous dose.
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first total synthesis of 14c labeled Procyanidin B2 a milestone toward understanding cocoa polyphenol metabolism eur j org chem 36 2008
European Journal of Organic Chemistry, 2008Co-Authors: Florian Viton, Gary Williamson, Cyrille Landreau, David Rustidge, Fabien Robert, Denis BarronAbstract:The cover picture shows the key steps of the first asymmetric total synthesis of Procyanidin B2, one of the major dietary polyphenols present in cocoa and chocolate. During the last decades, the health benefits of foods consumed for pure pleasure have received much recognition. Many biological studies have evidenced the beneficial health effects of Procyanidins. However, the absorption and metabolism of Procyanidins is still not fully understood, and some aspects are still controversial. In order to strengthen this knowledge, the first total synthesis of Procyanidin B2 was developed and applied to the preparation of a regioselectively radiolabeled analogue incorporating a 14C label at the 2-position of the upper C-ring moiety. This enantioselective synthesis was achieved in 14 “hot” steps, involving as key steps the Sharpless dihydroxylation of an elaborated alkene, a stereoselective intramolecular cyclization and the condensation of two (–)-epicatechin units. The radiolabelled Procyanidin B2 obtained through this reaction pathway will be used in bioavailability studies. Details are discussed in the article by F. Viton et al. on p. 6069 ff. The authors acknowledge Tonic Life Communications for the design of the cover page and the European Union 6th Framework project “FLAVO” for partial support of this research work.
Rui-hai Zhou - One of the best experts on this subject based on the ideXlab platform.
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proteomic analysis of kidney and protective effects of grape seed Procyanidin B2 in db db mice indicate mfg e8 as a key molecule in the development of diabetic nephropathy
Biochimica et Biophysica Acta, 2013Co-Authors: Zhen Zhang, Qian Cai, Hai-qing Gao, Mei Cheng, Jun-fu Wang, Rui-hai Zhou, Lin ShenAbstract:Diabetic nephropathy, as a severe microvascular complication of diabetic mellitus, has become the leading cause of end-stage renal diseases. However, no effective therapeutic strategy has been developed to prevent renal damage progression to end stage renal disease. Hence, the present study evaluated the protective effects of grape seed Procyanidin B2 (GSPB2) and explored its molecular targets underlying diabetic nephropathy by a comprehensive quantitative proteomic analysis in db/db mice. Here, we found that oral administration of GSPB2 significantly attenuated the renal dysfunction and pathological changes in db/db mice. Proteome analysis by isobaric tags for relative and absolute quantification (iTRAQ) identified 53 down-regulated and 60 up-regulated proteins after treatment with GSPB2 in db/db mice. Western blot analysis confirmed that milk fat globule EGF-8 (MFG-E8) was significantly up-regulated in diabetic kidney. MFG-E8 silencing by transfection of MFG-E8 shRNA improved renal histological lesions by inhibiting phosphorylation of extracellular signal-regulated kinase1/2 (ERK1⁄2), Akt and glycogen synthase kinase-3beta (GSK-3β) in kidneys of db/db mice. In contrast, over-expression of MFG-E8 by injection of recombinant MFG-E8 resulted in the opposite effects. GSPB2 treatment significantly decreased protein levels of MFG-E8, phospho-ERK1/2, phospho-Akt, and phospho-GSK-3β in the kidneys of db/db mice. These findings yield insights into the pathogenesis of diabetic nephropathy, revealing MFG-E8 as a new therapeutic target and indicating GSPB2 as a prospective therapy by down-regulation of MFG-E8, along with ERK1/2, Akt and GSK-3β signaling pathway.
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proteomic analysis of aorta and protective effects of grape seed Procyanidin B2 in db db mice reveal a critical role of milk fat globule epidermal growth factor 8 in diabetic arterial damage
PLOS ONE, 2012Co-Authors: Qian Cai, Mei Cheng, Jian-hua Zhang, Jun-fu Wang, Rui-hai Zhou, Mei Yin, Zhen Zhang, Hai-qing GaoAbstract:Background Atherosclerosis is one of the major complications of type 2 diabetic patients (T2DM), leading to morbidity and mortality. Grape seed Procyanidin B2 (GSPB2) has demonstrated protective effect against atherosclerosis, which is believed to be, at least in part, a result of its antioxidative effects. The aim of this study is to identify the target protein of GSPB2 responsible for the protective effect against atherosclerosis in patients with DM. Methods and Results GSPB2 (30 mg/kg body weight/day) were administrated to db/db mice for 10 weeks. Proteomics of the aorta extracts by iTRAQ analysis was obtained from db/db mice. The results showed that expression of 557 proteins were either up- or down-regulated in the aorta of diabetic mice. Among those proteins, 139 proteins were normalized by GSPB2 to the levels comparable to those in control mice. Among the proteins regulated by GSPB2, the milk fat globule epidermal growth factor-8 (MFG-E8) was found to be increased in serum level in T2DM patients; the serum level of MFG-E8 was positively correlated with carotid-femoral pulse wave velocity (CF-PWV). Inhibition of MFG-E8 by RNA interference significantly suppressed whereas exogenous recombinant MFG-E8 administration exacerbated atherogenesis the db/db mice. To gain more insights into the mechanism of action of MFG-E8, we investigated the effects of MFG-E8 on the signal pathway involving the extracellular signal-regulated kinase (ERK) and monocyte chemoattractant protein-1 (MCP-1). Treatment with recombinant MFG-E8 led to increased whereas inhibition of MFG-E8 to decreased expression of MCP-1 and phosphorylation of ERK1/2. Conclusion Our data suggests that MFG-E8 plays an important role in atherogenesis in diabetes through both ERK and MCP-1 signaling pathways. GSPB2, a well-studied antioxidant, significantly inhibited the arterial wall changes favoring atherogenesis in db/db mice by down-regulating MFG-E8 expression in aorta and its serum level. Measuring MFG-E8 serum level could be a useful clinical surrogate prognosticating atherogenesis in DM patients.
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Induction of lactadherin mediates the apoptosis of endothelial cells in response to advanced glycation end products and protective effects of grape seed Procyanidin B2 and resveratrol
Apoptosis : an international journal on programmed cell death, 2011Co-Authors: Qian Cai, Hai-qing Gao, Mei Cheng, Jian-hua Zhang, Jun-fu Wang, Rui-hai ZhouAbstract:One of characteristics of diabetes mellitus (DM) is endothelial cell (EC) dysfunction and apoptosis which contributes to the development of vasculopathy. Advanced glycation end products (AGEs) continuously produced in the setting of DM play an important role in causing EC dysfunction and apoptosis. However, the underlying molecular mechanism remains largely elusive. Lactadherin, a secreted glycoprotein of milk-fat globule, is expressed by multiple cell types of arterial wall including ECs. Our previous proteomic studies showed that the expression of lactadherin was significantly increased in the aorta of diabetic rats as compared with control rats and treatment with grape seed Procyanidin extracts significantly inhibited the lactadherin expression in diabetic rats. We hypothesized that lactadherin plays a critical role in AGEs-induced EC apoptosis; grape seed Procyanidin B2 (GSPB2) and resveratrol protect against AGEs-induced EC apoptosis through lactadherin regulation. Our results showed that AGEs upregulated lactadherin expression and lactadherin RNA interference significantly attenuated AGEs-induced EC apoptosis. Overexpression of lactadherin increased EC apoptosis with up-regulation of Bax/Bcl-2 ratio, cytochrome c release, caspase-9 and caspase-3 activation suggesting the involvement of mitochondria apoptosis pathway. Mechanistically, overexpression of lactadherin reduced the phosphorylation of GSK3beta at baseline. Our study also revealed nine proteins interacting with lactadherin in HUVEC and study of these candidate proteins could unveil further underlying molecular mechanisms. In summary, our study identified lactadherin as a key player responsible for AGEs-induced EC apoptosis and antioxidants GSPB2 and resveratrol protect against AGEs-induced EC apoptosis by inhibiting lactadherin. Targeting lactadherin with antioxidant could be translated into clinical application in the fighting against DM complications.
Michael N Clifford - One of the best experts on this subject based on the ideXlab platform.
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Procyanidin B2 catabolism by human fecal microflora partial characterization of dimeric intermediates
Archives of Biochemistry and Biophysics, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Warren J Drynan, Denis Barron, Michael N CliffordAbstract:Abstract The catabolism by human fecal microflora of pure Procyanidin B2 ((–)-epicatechin-4β → 8-(–)-epicatechin) has been investigated using a static in vitro culture model. For the first time, 24 catabolites were detected by LC–MS ( n ) with M r greater than 290 indicating that they could not have formed from just one of the epicatechin units in the Procyanidin structure. Structures have been assigned on the basis of the fragmentation in the ion trap mass spectrometer and with regard to catabolic pathways known to occur in fecal microorganisms. Twenty of these ‘dimeric’ catabolites produced fragment ions characteristic of flavanols and/or proanthocyanidins. One catabolite was identified tentatively as either 6- or 8-hydroxy-Procyanidin B2. Thirteen were characterized as having been microbially reduced in at least one of the epicatechin units. Five contained an apparently unmodified epicatechin unit but in at least one case this was shown to consist of the B-ring of the “upper” epicatechin unit and the A-ring of the “lower”. These ‘dimeric’ catabolites were detected up to 9 h after the start of the incubation and collectively accounted for ∼20% of the substrate.
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a comparison of the in vitro biotransformation of epicatechin and Procyanidin B2 by human faecal microbiota
Molecular Nutrition & Food Research, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Warren J Drynan, Denis Barron, Michael N CliffordAbstract:The catabolism by human faecal microbiota of (-)-epicatechin (1) (2, 3-cis stereochemistry) and its dimer pure Procyanidin B2 (2), has been compared using a static in vitro culture model. The catabolites were characterised by LC-MS(n), UV absorption and relative retention time, and quantified relative to standards. No more than approximately 10% of Procyanidin B2 (2) was converted to epicatechin (1) by scission of the interflavan bond. Five phenolic acid catabolites (M(r) or =24 h incubation) were 5-(3'-hydroxy phenyl) valeric acid (9), 3-(3'-hydroxyphenyl) propionic acid (10) and phenyl acetic acid (12) (maximum yields 27.4+/-4.2, 38.2+/-4.2, 22.7+/-2.9%, respectively, from 1 and 9.4+/-1.2, 52.8+/-2.1, 28.8+/-1.6%, respectively, from 2). Substrate 2 was degraded twice as rapidly as 1. Evidence is presented for the production of previously unreported catabolites of 2 that retain the flavanol A-ring and the C4-->C8 interflavan bond. It was confirmed that catabolism favoured removal of the 4'-hydroxyl rather than the 3'-hydroxyl group and that both beta-oxidation and alpha-oxidation occurred.
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a comparison of the in vitro biotransformation of epicatechin and Procyanidin B2 by human faecal microbiota
Molecular Nutrition & Food Research, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Warren J Drynan, Denis Barron, Michael N CliffordAbstract:The catabolism by human faecal microbiota of (-)-epicatechin (1) (2, 3-cis stereochemistry) and its dimer pure Procyanidin B2 (2), has been compared using a static in vitro culture model. The catabolites were characterised by LC-MS n , UV absorption and relative retention time, and quantified relative to standards. No more than ∼10% of Procyanidin B2 (2) was converted to epicatechin (1) by scission of the interflavan bond. Five phenolic acid catabolites (M r < 290) were unique to 2, and ten phenolic acid catabolites (M r < 290) were common to both substrates. The dominant catabolites (≥24h incubation) were 5-(3'-hydroxy phenyl) valeric acid (9), 3-(3'-hydroxyphenyl) propionic acid (10) and phenyl acetic acid (12) (maximum yields 27.4 ± 4.2, 38.2 ± 4.2, 22.7 ± 2.9%, respectively, from 1 and 9.4±1.2, 52.8±2.1, 28.8±1.6%, respectively, from 2). Substrate 2 was degraded twice as rapidly as 1. Evidence is presented for the production of previously unreported catabolites of 2 that retain the flavanol A-ring and the C4 → C8 interflavan bond. It was confirmed that catabolism favoured removal of the 4'-hydroxyl rather than the 3'-hydroxyl group and that both β-oxidation and α-oxidation occurred.
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in vivo bioavailability absorption excretion and pharmacokinetics of 14c Procyanidin B2 in male rats
Drug Metabolism and Disposition, 2010Co-Authors: Stavroula Stoupi, Gary Williamson, Denis Barron, Florian Viton, Laurence J King, Jonathan E Brown, Michael N CliffordAbstract:Procyanidins are important biologically active compounds, but the pathway and extent of absorption and metabolism are controversial. We conducted a mass balance study to evaluate the total radioactivity excreted in urine and feces after oral administration of [(14)C]Procyanidin B2 to male rats (n = 5). Urine and feces were collected daily from 0 to 96 h. Absolute bioavailability of (14)C from [(14)C]Procyanidin B2 was calculated as approximately 82% using the values for total urinary (14)C. A pharmacokinetic study measured total radioactivity in the blood (n = 9). Blood samples were collected at designated time intervals (0.5-24 h) after administration. Three treatments were used: 1) intravenous, 2) oral higher dose (21 mg/kg b.wt.), and 3) oral lower dose (10.5 mg/kg). Blood concentration of total (14)C reached a maximum at approximately 6 h after ingestion of [(14)C]Procyanidin B2 (groups II and III), and area under the curve (AUC) was dependent on oral dose. After intravenous or oral administration the terminal half-lives were similar, whereas 8-fold larger values were obtained after oral dosing for total clearance and the apparent volumes of distribution. These pharmacokinetic differences explain the apparently lower (14)C bioavailability (8-11%) for [(14)C]Procyanidin calculated from blood [AUC((0-24))] values. After oral administration of [(14)C]Procyanidin B2, 63% was excreted via urine within 4 days. The data suggest that much of the parent compound administered orally is degraded by the gut microflora before absorption and that these microbial metabolites have a different distribution from the compounds circulating after the intravenous dose.