GABRIELA CASTILHO
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conferenceObject N-acetylcystein Reduces Lipid Peroxidation and Advanced Glycation Related to Prevention of Macrophage Endoplasmic Reticulum Stress Induced by Albumin Isolated from Rats With Chronic Kidney Disease(2014) MACHADO, Juliana T.; IBORRA, Rodrigo T.; FUSCO, Fernanda B.; CASTILHO, Gabriela; PINTO, Raphael S.; MACHADO-LIMA, Adriana; NAKANDAKARE, Edna R.; SHIMIZU, Maria Heloisa M.; SEGURO, Antonio Carlos; CATANOZI, Sergio; PASSARELLI, MarisaconferenceObject Palmitic Interesterified Fat Induces Atherosclerosis and Inflammatory Cytokine Secretion in LDL Receptor Knockout Mice(2014) AFONSO, Milessa S.; LAVRADOR, Maria Silvia F.; KOIKE, Marcia; BOMBO, Renata P.; NUNES, Valeria S.; CATANOZI, Sergio; CASTILHO, Gabriela; PASSARELLI, Marisa; NAKANDAKARE, Edna R.; LOTTENBERG, Ana Maria- Advanced glycated albumin impairs HDL anti-inflammatory activity and primes macrophages for inflammatory response that reduces reverse cholesterol transport(2012) OKUDA, Ligia S.; CASTILHO, Gabriela; ROCCO, Debora D. F. M.; NAKANDAKARE, Edna R.; CATANOZI, Sergio; PASSARELLI, MarisaObjective: We investigated the effect of advanced glycated albumin (AGE-albumin) on macrophage sensitivity to inflammation elicited by S100B calgranulin and lipopolysaccharide (LPS) and the mechanism by which HDL modulates this response. We also measured the influence of the culture medium, isolated from macrophages treated with AGE-albumin, on reverse cholesterol transport (RCT). Methods and results: Macrophages were incubated with control (C) or AGE-albumin in the presence or absence of HDL, followed by incubations with S100B or LPS. Also, culture medium obtained from cells treated with C- or AGE-albumin, following S100B or LPS stimulation was utilized to treat naive macrophages in order to evaluate cholesterol efflux and the expression of HDL receptors. In comparison with C-albumin, AGE-albumin, promoted a greater secretion of cytokines after stimulation with S100B or LPS. A greater amount of cytokines was also produced by macrophages treated with AGE-albumin even in the presence of HDL Cytokine-enriched medium, drawn from incubations with AGE-albumin and S100B or LPS impaired the cholesterol efflux mediated by apoA-I (23% and 37%, respectively), HDL2 (43% and 47%, respectively) and HDL3 (20% and 8.5%, respectively) and reduced ABCA-1 protein level (16% and 26%, respectively). Conclusions: AGE-albumin primes macrophages for an inflammatory response impairing the RCT. Moreover, AGE-albumin abrogates the anti-inflammatory role of HDL, which may aggravate the development of atherosclerosis in DM.
- N-acetylcysteine prevents endoplasmic reticulum stress elicited in macrophages by serum albumin drawn from chronic kidney disease rats and selectively affects lipid transporters, ABCA-1 and ABCG-1(2014) MACHADO, Juliana T.; IBORRA, Rodrigo T.; FUSCO, Fernanda B.; CASTILHO, Gabriela; PINTO, Raphael S.; MACHADO-LIMA, Adriana; NAKANDAKARE, Edna R.; SEGURO, Antonio C.; SHIMIZU, Maria H.; CATANOZI, Sergio; PASSARELLI, MarisaIn chronic kidney disease (CKD) nontraditional risk factors, such as oxidative stress and advanced glycation end products (AGE) contribute to cardiovascular disease. Particularly, disturbances in reverse cholesterol transport favor the development of atherosclerosis. We analyzed the influence of N-acetylcysteine (NAC) in CKD rats on plasma concentration of lipid peroxides (TBARS) and AGE and on the impact of serum albumin in the development of macrophage endoplasmic reticulum stress (ERS) and cholesterol efflux, namely apo A-I and HDL2-mediated cholesterol removal and ABCA-1 and ABCG-1 protein level. CKD was induced by 5/6 nephrectomy in 2-month old male Wistar rats. Controls (Sham) were false operated. Animals were treated or not with NAC (600 mg/L of water). After 60 days serum albumin was isolated by FPLC and purified by alcoholic extraction. J774 macrophages were incubated with serum albumin (1 mg/mL; 18 h) from all groups, and the expression of ERS markers (protein disulfide isomerase - PDI, Grp78 and Grp94), ABCA-1 and ABCG-1 determined by immunoblot. HDL2 or apo A-I were used for cholesterol efflux assays. Protein and lipid composition of total HDL from Sham and CKD was determined and these particles tested on their abilities to accept cell cholesterol. Comparisons were done by one-way ANOVA and Newman Keuls post test. After 60 days of CKD, body weight was 10% lower in CKD compared to Sham (p < 0.01). This was prevented by NAC. Urea, creatinine, total cholesterol (TC), triglycerides (TG) (mg/dL), proteinuria (mg/24 h) (Sham, n = 31; Sham + NAC, n = 20; CKD, n = 74; CKD + NAC, n = 32), total AGE and pentosidine (n = 8; fluorescence arbitrary unit) and TBARS (n = 7; nmoL/mL) were higher in CKD (122 +/- 8; 0.9 +/- 0.07; 151 +/- 6; 83 +/- 4; 46 +/- 2.5; 32,620 +/- 673; 16,700 +/- 1,370; 6.6 +/- 0.5, respectively) and in CKD + NAC (91.4 +/- 5; 0.6 +/- 0.02; 126 +/- 7.5; 73 +/- 6; 51 +/- 3.5; 24,720 +/- 1,114; 10,080 +/- 748; 4.5 +/- 0.5, respectively) in comparison to Sham (41 +/- 0.9; 0.4 +/- 0.03; 76 +/- 2.7; 51.5 +/- 3; 14 +/- 0.9; 21,750 +/- 960; 5,314 +/- 129; 2.0 +/- 0.2, respectively; p < 0.001) and Sham + NAC (40 +/- 0.9; 0.3 +/- 0.02; 76 +/- 2.6; 68 +/- 4; 18.4 +/- 1.5; 20,040 +/- 700; 5,050 +/- 267; 1.8 +/- 0.2, respectively; p < 0.001). TC, urea, creatinine, total AGE, pentosidine and TBARS were respectively, 17%, 25%, 33%, 24%, 40% and 28% (p < 0.01) lower in CKD + NAC, than in CKD. Glycemia was higher in Sham + NAC (107 +/- 4.6) and CKD + NAC (107 +/- 2.6) than in Sham (96 +/- 1.8; p < 0.05) and CKD (98 +/- 1.6; p < 0.01), respectively. In macrophages (n = 6), CKD albumin increased PDI (3 and 6 times, p < 0.01) and Grp94 (66% and 80%, p < 0.01) in comparison to Sham and CKD + NAC-albumin treated cells, respectively. ABCA-1 expression was lower (87% and 70%, p < 0.001) in macrophage treated with Sham + NAC and CKD albumin respectively in comparison to Sham albumin; ABCG-1 was higher (4 and 7 times, p < 0.001) in macrophages treated with Sham + NAC and CKD + NAC albumin, respectively in comparison to Sham and CKD albumin. Apo A-I mediated cholesterol efflux was lower (59% and 70%, p < 0.0001) in macrophage treated with Sham + NAC and CKD albumin respectively in comparison to Sham albumin, however, the HDL2 mediated cholesterol efflux was higher (54% and 25%, p < 0. 0001) in macrophage treated with Sham + NAC albumin, in comparison to Sham and CKD + NAC albumin, respectively. CKD-HDL was enriched in total protein and lipids compared to Sham-HDL but preserved its capacity to remove cholesterol from macrophages. NAC reduces plasma lipid peroxidation and AGE and abrogates ERS induced by CKD-albumin. Despite diminishing ABCA-1, NAC increases ABCG-1 that counteracts the reduction in apo A-I-mediated cholesterol efflux. NAC may contribute to attenuate the deleterious effects of CKD-albumin on lipid accumulation in macrophages helping to prevent atherogenesis in CKD.